Tesis sobre el tema "Neurologia"

En aquesta tesi hem estudiat el paper de sistema autofágic-lisosomal durant el desenvolupament de les malalties amb cossos de Lewy, especialment la demència amb cossos de Lewy (DCL). Tot i que aquesta demència representa, després de la malaltia d’Alzheimer (MA), la segona causa més important de la demència, el seu diagnòstic clínic és molt complex i fins al 80% de tots els casos segueixen sent diagnosticats erròniament. La causa per aquesta dificultat diagnòstica es troba a nivell neuropatològic, ja que la DCL comparteix canvis característics amb l’EA, d’una banda, i a el ser una sinucleinopatía, amb la malaltia de Parkinson (MP), per una altra. La majoria dels pacients amb DCL es diagnostiquen com MA i, conseqüentment, reben els tractaments corresponents, els quals causen reaccions adverses severes en un 50% d’ells. Per això, la caracterització molecular de la DCL identificant mecanismes específics per a aquesta és de cabdal importància, perquè només coneixent aquests mecanismes serà possible desenvolupar les teràpies necessàries. Fins al moment es disposa de dades limitades sobre el paper de sistema autofàgic-lisosomal en la patogènesi de la DCL. En aquest context, el gen més estudiat és GBA, mutacions en el qual s’havien descrit recentment com a factors de risc per DCL i MP. Per això, en el primer estudi vam analitzar l’expressió de tres transcrits de GBA en cervells i sang de pacients amb DCL i MP. Trobem la disminució dels nivells de GBA en cervell, al còrtex de DCL i en el nucli caudat d’MP amb demència. També en sang es va detectar l’expressió de GBA disminuïda. La correlació entre paràmetres clínics i els nivells d’expressió de GBA va revelar que els nivells més baixos de GBA es corresponien a les formes més agressives de DCL i MP. Per identificar les possibles causes de la disminució de GBA, hem analitzat l’expressió de diferents transcrits de dos dels seus gens reguladors, LIMP2 i TFEB, en cervell. Com troballa principal, identifiquem una marcada sobreexpressió de TFEB, especialment en l’escorça temporal amb patologia Lewy pura. Aquest increment de TFEB estava acompanyat d’una sobreexpressió important d’un dels transcrits de SCARB2 en la mateixa àrea cerebral. En l’escorça temporal trobem també una correlació entre els nivells d’expressió de TFEB i GBA així com SCRAB2tv2 i GBA, corresponent els nivells més elevats de TFEB o SCARB2 amb la disminució més pronunciada de GBA. Com el sistema lisosomal està estretament relacionat amb el sistema autofàgic, hem analitzat l’expressió de sis gens implicats en punts específics de l’autofàgia. Encara BECN1, ATG3 i ATG5 s’expressaven de forma diferencial, la troballa més important d’aquest tercer estudi va ser que canvis transcripcionals en els gens autofàgics no estan primàriament involucrats en el desenvolupament de les malalties amb cossos de Lewy. En conclusió, els estudis que composen aquesta tesi van revelar que el sistema lisosomal juga un paper important en la patogènesi de la DCL. En canvi, el sistema autofàgic no pateix canvis importants sinó només canvis específics per a cada cas. Especialment l’escorça temporal, àrea d’afectació primerenca en la malaltia, es caracteritza per la disminució de GBA acompanyada de l’increment del seu receptor SCARB2 i el regulador transcricional TFEB. Així, GBA disminuïda pot activar l’expressió d’TFEB com a resposta cel·lular per restaurar la proteostasi lisosomal, però l’activació de TFEB resultaria a el mateix temps a la sobreexpressió d’altres gens lisosomals, incloent SCARB2.
En esta tesis hemos estudiado el papel del sistema autofágico-lisosomal durante el desarrollo de las enfermedades con cuerpos de Lewy, especialmente la demencia con cuerpos de Lewy (DCL). Aunque esta demencia representa, después de la enfermedad de Alzheimer (EA), la segunda causa más importante de la demencia, su diagnóstico clínico es muy complejo y hasta el 80% de todos los casos siguen siendo diagnosticados erróneamente. La causa para esta dificultad diagnóstica se encuentra a nivel neuropatológico, ya que la DCL comparte cambios característicos con la EA, por una parte, y al ser una sinucleinopatía, con la enfermedad de Parkinson (EP), por otra. La mayoría de los pacientes con DCL se diagnostican como EA y, consecuentemente, reciben los tratamientos correspondientes, los cuales causan reacciones adversas severas en un 50% de ellos. Por eso, la caracterización molecular de la DCL identificando mecanismos específicos para ésta es de primordial importancia, porque solamente conociendo dichos mecanismos será posible desarrollar las terapias necesarias. Hasta el momento se dispone de datos limitados sobre el papel del sistema autofágico-lisosomal en la patogénesis de la DCL. En este contexto, el gen más estudiado es GBA, mutaciones en el cual se habían descrito recientemente como factores de riesgo para DCL y EP. Por eso, en el primer estudio analizamos la expresión de tres transcritos de GBA en cerebros y sangre de pacientes con DCL y EP. Encontramos la disminución de los niveles de GBA en cerebro, en el córtex de DCL y en el núcleo caudado de EP con demencia. También en sangre se detectó la expresión de GBA disminuida. La correlación entre parámetros clínicos y los niveles de expresión de GBA reveló que los niveles más bajos de GBA se correspondían a las formas más agresivas de DLB y EP. Para identificar las posibles causas de la disminución de GBA, analizamos la expresión de diferentes transcritos de dos de sus genes reguladores, LIMP2 y TFEB, en cerebro. Como hallazgo principal, identificamos una marcada sobreexpresión de TFEB, especialmente en la corteza temporal con patología Lewy pura. Este incremento de TFEB estaba acompañado de una sobreexpresión importante de uno de los transcritos de SCARB2 en la misma área cerebral. En la corteza temporal encontramos también una correlación entre los niveles de expresión de TFEB y GBA así como SCRAB2tv2 y GBA, correspondiendo los niveles más elevados de TFEB o SCARB2 con la disminución más pronunciada de GBA. Como el sistema lisosomal está estrechamente relacionado con el sistema autofágico, analizamos la expresión de seis genes implicados en puntos específicos de la autofagia. Aunque BECN1, ATG3 y ATG5 se expresaban de forma diferencial, el hallazgo más importante de este tercer estudio fue que cambios transcripcionales en los genes autofágicos no están primariamente involucrados en el desarrollo de enfermedades con cuerpos de Lewy. En conclusión, los estudios que componen esta tesis revelaron que el sistema lisosomal juega un papel importante en la patogénesis de la DCL. En cambio, sistema autofágico no sufre cambios importantes sino solo cambios específicos para cada caso. Especialmente la corteza temporal, área de afectación temprana en la enfermedad, se caracteriza por la disminución de GBA acompañada del incremento de su receptor SCARB2 y el regulador transcricional TFEB. Así, GBA disminuida puede activar la expresión de TFEB como respuesta celular para restaurar la proteostasis lisosomal, pero la activación de TFEB resultaría al mismo tiempo en la sobreexpresión de otros genes lisosomales, incluyendo SCARB2.
In this thesis we have studied the role of the autophagy-lysosomal system during the development of Lewy body diseases, especially of dementia with Lewy bodies (DLB). Although this dementia represents the second most important cause of dementia after Alzheimer’s disease (AD), its clinical diagnosis is very complex and up to 80% of all cases continue to be misdiagnosed. The cause for this diagnostic difficulty is the important neuropathological overlap between DLB and AD, since DLB shares characteristic changes with AD, on one hand, and as a synucleinopathy, with Parkinson’s disease (PD), on the other. Most of DLB patients are diagnosed as AD and, consequently, receive the corresponding treatments which may cause severe adverse reactions. Therefore, the molecular characterization of DLB identifying disease specific mechanisms is of primary importance, because only unveiling these mechanisms will permit the development of the necessary therapies. To date, limited data are available on the role of the autophagy-lysosomal system in the pathogenesis of DLB. In this context, the most studied gene is GBA, mutations in which have been recently described as risk factors for DLB and PD. Therefore, in the first study we analyzed the expression of three GBA transcripts in the brains and blood of patients with DLB and PD. We found a decrease in GBA levels in the brain, in DLB cortices and in the caudate nucleus of PD with dementia. Decreased GBA expression was also detected in blood. The correlation between clinical parameters and GBA expression levels revealed that the lowest GBA levels corresponded to the most aggressive forms of DLB and EP. To identify the possible causes of GBA diminution in brain, we analyzed the expression of different transcripts of two of its regulatory genes, LIMP2 and TFEB, in the same brain areas as for GBA. As the main finding, we identified a marked overexpression of TFEB, especially in the temporal cortex with pure Lewy pathology. This TFEB increase was accompanied by a significant overexpression of one of the SCARB2 transcripts in the same brain area. In the temporal cortex we also found a correlation between the expression levels of TFEB and GBA as well as SCRAB2tv2 and GBA, the highest levels of TFEB or SCARB2 corresponding to the most pronounced decrease in GBA. As the lysosomal system is closely related to the autophagy system, we analyzed the expression of six genes involved in specific points of autophagy. Although BECN1, ATG3, and ATG5 showed a slight differential expression between groups, the most important finding from this third study was that transcriptional changes in autophagy genes are not primarily involved in the development of Lewy body diseases. In conclusion, the studies composing this thesis revealed that the lysosomal system plays an important role in the pathogenesis of DLB, but the autophagy system does not show major changes undergoing only case-specific changes. Especially the temporal cortex, an area of early involvement in the disease, is characterized by a decrease in GBA accompanied by an increase in its receptor SCARB2 and the transcriptional regulator TFEB. Thus, decreased GBA can activate the expression of TFEB as a cellular response to restore lysosomal proteostasis, but the activation of TFEB would at the same time result in the overexpression of other lysosomal genes, including SCARB2.
Universitat Autònoma de Barcelona. Programa de Doctorat en Cirurgia i Ciències Morfològiques

Brain derived neurotrophic factor (BDNF) is a chemokine which levels are regulated by neuronal activity and could act as a sensor in front of distinct physiologic stimulus, activating the transcription of specific group of genes. In this work we show that BDNF induces the expression of BMP7 in neurons through TrkB receptor and MAPK/ERK pathways, an induction mechanism that is mediated in part by the release of the transcriptional repression exerted by p53 family proteins. BMP members in mammals are expressed in the growing nervous system where emerged as crucial regulators of dorsoventral patterning of the neural tube, neural cell fate determination, and cell death as well as terminal neural cell differentiation. In the earlier cerebral cortex development (at embryonic day 13, E13) BMPs predominantly induce cell death and inhibit the proliferation, as a mechanism for the regulation of cell number and phenotype within the developing cortex. Subsequently they exert sequential actions promoting neuronal differentiation at E16 and increasingly with time, they promote astrocytic differentiation and inhibit oligodendrocytes generation. This thesis demonstrates that BMP7 injection at midgestation alters the laminar distribution of pyramidal neurons in the cerebral cortex while GABAergic neurons distribution was not affected. We observed that abnormal high levels of BMP7 during cerebral cortex development induce the premature radial glia maturation into astrocytes impairing the radial migration of upper layers pyramidal neurons that remained accumulated in lower cortical regions. We also observed that altered BMP7 levels during midgestation lead to corpus callosum malformation. Although corpus callosum agenesis can be due to multiple causes, our analysis show that the correct pattern of BMP7 expression is necessary for the proper maturation of intermediate structures such as the glial wedge, the induseum griseum and the subcallosal sling, that provide essential guidepost signals for the proper corpus callosum development. Based on these results, it is proposed a physiologic model where the expression of BDNF induced by the initial electrical activity in the perinatal period would induce in turn, an increase in BMP7 expression. Both chemokines may act co-ordinately maturating neurons and glial cells at the end of neurogenic period. The alteration of BDNF and BMP7 spatio-temporal expression patterns could dramatically affect the proper cerebral cytoarchitecture and consequently the cerebral functioning. Indeed, different traumas occurred during embryonic and perinatal development are associated with an imbalance in BDNF and BMP7 levels. To check this hypothesis we reproduced an embryonic sublethal hypoxia, a pathological condition that can be associated to altered BDNF and BMP7 expression. Moreover, perinatal reduction of oxygen input can dramatically affect the cerebral cortex developmental program. As a result, many behavioural and learning disorders in infants have been associated to this pathological condition. We observed that this condition reduces BMP7 expression and signalling in the cerebral cortex promoting the differentiation of cortical progenitors into the oligodendrocytes in detrimental to the astroglial fate in vitro and in vivo. So, our findings indicate that changes on BMP7 expression in the tightly regulated developmental program of the central nervous system might importantly modify the cellular fate choice of cortical progenitors. When this change occurs during the critic perinatal developmental period, it could compromise the normal brain functionality in the affected individual.
“FUNCIÓN Y REGULACIÓN DE LA PROTEINA MORFOGENÉTICA DE HUESO (BMP7) EN EL DESARROLLO DE LA CORTEZA” TEXTO: "Brain derived neurotrophic factor" (BDNF) es una citoquina regulada por la actividad neuronal y puede actuar como sensor en respuesta a distintos estímulos fisiológicos, activando grupos específicos de genes. En este trabajo demuestro que BDNF induce la expresión de BMP7 en neuronas a través del receptor TrkB y la vía de señalización MAPK/ERK. Un mecanismo de inducción mediado en parte por la liberación de la represión transcripcional ejercida por la familia de proteínas p53. La inyección intraventricular de BMP7 durante la corticogenesis altera la distribución de las neuronas piramidales en la corteza cerebral. BMP7 induce la maduración prematura de la glia radial hacia astrocito alterando la migración radial de las neuronas piramidales de capas altas, que quedan anormalmente acumuladas en capas corticales inferiores. Niveles anormales de BMP7 durante fases gestacionales intermedias provocan malformación del cuerpo calloso (CC). Aunque la agénesis del CC puede ser debida a múltiples causas, nuestros análisis muestran que BMP7 es necesario para la formación de poblaciones de la línea media (glial wedge, induseum griseum y subcallosal sling) que participan en mecanismos de guía axonal necesarios para el desarrollo del CC. Proponemos un modelo fisiológico donde la expresión de BDNF inducida por el aumento de actividad eléctrica perinatal induciría a su vez un aumento de los niveles de BMP7. Ambas citoquinas actuarían conjuntamente madurando de una manera sincrónica las poblaciones neuronales y gliales de la corteza cerebral. La modificación del patrón de expresión espacio-temporal de ambas citoquinas podría afectar la composición celular y por tanto la correcta funcionalidad de la corteza cerebral. De hecho, diferentes traumas producidos durante el desarrollo embrionario y perinatal, donde se observa alteración de los niveles de BDNF y BMP7, están asociados a distintos desordenes neurológicos. En este trabajo reproducimos una hipoxia embrionaria sub-letal y observamos que los niveles de expresión y señalización de BMP7 están reducidos en animales hipóxicos. Esta reducción en los niveles de BMP7 promueve la diferenciación de los progenitores corticales hacia un fenotipo oligodendroglial en detrimento del fenotipo astroglial. Por tanto, BMP7 es vital para la correcta determinación de diferentes progenitores neurales durante el desarrollo cortical.

In individui con disturbi cerebrali sospetti o confermati, lo screening cognitivo tramite test performance-based convoglia informazioni di rilievo sia in termini diagnostici e prognostici che in ambito interventistico. Inoltre, tali test vengono spesso impiegati come misure di outcome nell’ambito di studi clinici aventi come oggetto la cognizione. Pertanto, gli screener cognitivi devono possedere solide caratteristiche statistiche e di usabilità. Tuttavia, nel panorama Italiano, gli screener cognitivi non sempre possiedono tali caratteristiche. Inoltre, in Italia, le pratiche di screening cognitivo da remoto sono state storicamente poco considerate, nonostante si prestino facilmente ad essere erogate a distanza tramite media di facile impiego, come il telefono. Tuttavia, avendo il potenziale di abbattere barriere geografiche, logistiche, socio-demografiche ed economiche, la disponibilità di screener cognitivi telefonici implicherebbe rilevanti vantaggi sia in ambito clinico che per l’implementazione e la portata a termine di studi clinici. In ragione di quanto suddetto, nelle prime due Sezioni della presente Dissertazione, in seguito ad una panoramica generale dei principi sottesi alle procedure di screening, viene fornita un’esauriente cornice per lo studio statistico degli screener cognitivi – affrontando 1) le loro applicazioni a fini di screening di popolazione e di case-finding nella pratica e ricerca cliniche, nonché le questioni relative a 2) proprietà clinimetriche (i.e., caratteristiche psicometriche e diagnostiche), 3) taratura (con un focus sul metodo dei Punteggi Equivalenti) e 4) usabilità trasversale/longitudinale. Inoltre, la Sezione 2 riporta una discussione dettagliata e critica 1) sul razionale, 2) su vantaggi e svantaggi e 3) sulle applicazioni cliniche/di ricerca degli screener cognitivi telefonici – focalizzandosi anche 4) sugli aspetti statistici relativi alla loro standardizzazione. Nella terza Sezione, in seguito ad una sinossi aggiornata in merito allo status quo Italiano relativo alle caratteristiche statistiche degli screener di persona e telefonici, viene riassunta una serie di studi recentemente pubblicati e focalizzantesi sull’aggiornamento della taratura e/o sul miglioramento delle proprietà clinimetriche e dell’usabilità di tre screener Italiani di persona – i.e., 1) la Frontal Assessment Battery (FAB), 2) il Montreal Cognitive Assessment (MoCA) e 3) la sezione cognitiva dell’Edinburgh Cognitive and Behavioural ALS Screen (ECAS). In seguito, la Sezione 3 include la descrizione di alcuni studi di standardizzazione Italian di screener telefonici – i.e., la Telephone Interview for Cognitive Status (TICS), 2) l’ALS Cognitive Behavioural Screen-Phone Version (ALS-CBS-PhV) e 3) la telephone-based Frontal Assessment Battery (t-FAB). Successivamente, la Sezione 4 riporta per intero due studi al momento non ancora pubblicati – il primo, relativo alla valutazione dell’usabilità clinica della TICS Italiana, il secondo, riportante la standardizzazione di uno screening telefonico per i disturbi linguistici, i.e. il Telephone Language Screener (TLS). Infine, la quinta e conclusiva Sezione riporta uno sguardo sinottico al contenuto della presente Dissertazione, le prospettive future derivanti da essa e una serie di considerazioni critiche in merito alle questioni aperte sul tema dello screening cognitivo in Italia.
In individuals with suspected or confirmed brain disorders, cognitive screening via performance-based tests conveys pivotal information both towards diagnosis and prognosis and within interventional settings. Moreover, such tests are often employed as outcomes measures within clinical studies addressing cognition. Hence, cognitive screeners need to come with sound statistical and feasibility features. However, within the Italian scenario, cognitive screeners do not always present with such characteristics. Moreover, remote cognitive screening procedures have been historically underdeveloped in Italy, despite easily lending themselves to be delivered from a distance via practicable media, such as the telephone. However, by bridging down geographical, logistical, socio-demographic and economic barriers, the availability of telephone-based cognitive screeners would both entail improvements in healthcare settings and ease the implementation and accomplishment of clinical studies. Given the above premises, within the first two Sections of the present Dissertation, after outlining the principles underlying screening procedures in general, a comprehensive, practical framework for the statistical study of cognitive screeners is delivered – addressing 1) their applications for population-screening and case-findings aims within both clinical practice and research, as well as the issues of 2) clinimetrics (i.e., psychometrics and diagnostics), 3) norm derivation (with a focus on the Equivalent Score method) and 4) cross-sectional/longitudinal feasibility. In addition, Section 2 delivers an extensive, critical discussion on the 1) rationale, 2) benefits and shortcomings and 3) clinical/research applications of telephone-based cognitive screeners – also focusing on 4) the statistical issues related to their standardization. Within the third Section, after providing an up-to-date synopsis of the Italian status quo as to the statistical features of both in-person and telephone-based cognitive screeners, a number of recently published investigations are summarized that focused on updating norms and/or improving the clinimetrics and feasibility of three Italian, in-person cognitive screeners – i.e., 1) the Frontal Assessment Battery (FAB), 2) the Montreal Cognitive Assessment (MoCA) and 3) the cognitive section of the Edinburgh Cognitive and Behavioural ALS Screen (ECAS). Subsequently, Section 3 displays the description of recently published, Italian standardization studies of telephone-based cognitive screeners – i.e., 1) the Telephone Interview for Cognitive Status (TICS), 2) the ALS Cognitive Behavioural Screen-Phone Version (ALS-CBS-PhV) and 3) the telephone-based Frontal Assessment Battery (t-FAB). Then, Section 4 reports in full two currently unpublished studies – the first, addressing the assessment of the clinical usability of the Italian TICS, the second, providing the Italian standardization of a telephone-based screener for language disorders, i.e. the Telephone Language Screener (TLS). A synoptic glance at the content of the present Dissertation, the future perspectives yielding from it and a number of critical considerations related to outstanding issues on the topic of cognitive screening in Italy are finally delivered within the conclusive, fifth Section.

Un problema important en el maneig de l’Estat Epiléptico (EE) és el diagnòstic precoç d’aquest. Se sap que l’activitat epilèptica causa un augment de la demanda metabòlica a nivell cerebral, que s’acompanya d’un augment temporal de la perfusió cerebral, per això investiguem si la TC cerebral de perfusió (TC-P) podia ser una eina de diagnòstic útil per a pacients amb EE. Realitzem un estudi de pacients amb EE, diagnosticats per semiologia clínica i EEG, en els quals es va realitzar de manera prospectiva un TC-P en fase crítica. Es va realitzar una anàlisi visual i quantitatiu dels mapes de perfusió. Es van calcular els índexs de asimetría-(IA) per al flux cerebral regional-(rCBF), el volum-(rCBV), el temps a l’pic-(TTP) i el temps de trànsit mig-(MTT). En 9 dels casos es va realitzar una TC-P de seguiment un cop resolt el EE i es van comparar amb les TC-P realitzades en fase crítica. A més, incloem un grup de control en els quals també es va realitzar un TC-P. Incloem 19 pacients. L’anàlisi visual dels mapes de perfusió durant la fase crítica, va mostrar àrees de hiperperfusión en el 78,9% dels pacients. L’anàlisi quantitativa va mostrar un augment significatiu dels valors de rCBF (p=0.002) i rCBV (p=0.004), i una disminució de TTP (p <0.001) MTT (p=0.001) en les àrees corticals de el costat afectat versus el costat no afectat. Dels 9 pacients amb una TC-P de seguiment, 8 van mostrar disminució de la intensitat, rCBV (p=0.035) i rCBF (p=0.024) en les àrees de hiperperfusión. La sensibilitat de la detecció de hiperperfusión per al diagnòstic d’EE va ser 78.95%, i l’especificitat de l’90%. L’anàlisi quantitativa comparativa dels índexs de asimetria per rCBF, rCBV i MTT entre les TC-P crítiques i el grup control va mostrar diferències significatives per a tots els paràmetres (rCBF p=0.001; rCBV p=0.002; TTP p=0.001 i MTT p=0.001) En aquest estudi vam demostrar que la TC-P pot proporcionar informació diagnòstica valuosa en pacients amb EE i complementar a l’EEG. D’altra banda, la identificació de factors clínics que puguin predir l’evolució dels pacients en EE és important. L’escala STESS-(Status-epilepticus-Severity-Score) és una eina per predir la mortalitat en el EE. No obstant això, aquesta escala no té en compte la situació funcional prèvia de l’malalt. Per això en el nostre segon estudi vam voler valorar si l’Rankin modificat-(mRS) podria ser un factor pronòstic en el EE i si afegint aquesta variable a l’STESS es podria millorar la predicció de el pronòstic en aquests pacients. Es va realitzar un registre retrospectiu dels pacientes≥16 anys que van presentar un EE. Realitzem corbes ROC i models de regressió logística per estimar les puntuacions d’una nova escala “”mSTESS””(modified STESS) i comparem amb els resultats de l’STESS. Es van incloure 136 pacients. La capacitat de l’STESS per predir la mortalitat va ser de l’74,3%, mentre que la capacitat de l’mRS per predir la mortalitat va ser de l’65,2%. El model de regressió logística i les corbes ROC van permetre classificar el mRS en tres grups: 0 (mRS=0); 1 (mRS=1 a 3) i 2 (mRS> 3). Aquests valors van ser afegits als altres ítems de l’STESS, resultant una nova escala, el mSTESS, amb puntuacions entre 0 i 8 punts. La capacitat de l’mSTESS per predir la mortalitat va ser de l’80,1%. Un mSTESS> 4 va establir una precisió general de 81.8% per predir la mortalitat, que va ser considerablement més gran que la precisió general de l’STESS≥3 (59.6%). Concloem que el mRS basal s’associa amb un alt risc de mortalitat, i que el mSTESS, podria ser una escala més precisa per predir el pronòstic de pacients amb EE.
Un problema importante en el manejo del Estado Epiléptico (EE) es el diagnóstico precoz del mismo. Se sabe que la actividad epiléptica causa un aumento de la demanda metabólica en la corteza cerebral, que se acompaña de un aumento temporal de la perfusión cerebral, por ello investigamos si la TC cerebral de perfusión (TC-P) podía ser una herramienta de diagnóstico útil para pacientes con EE. Realizamos un estudio de pacientes con EE, diagnosticados por semiología clínica y EEG, en los que se realizó de forma prospectiva una TC-P en fase crítica. Se realizó un análisis visual y cuantitativo de los mapas de perfusión. Se calcularon los índices de asimetría-(IA) para el flujo cerebral regional-(rCBF), el volumen-(rCBV), el tiempo al pico-(TTP) y el tiempo de tránsito medio-(MTT). En 9 de los casos se realizó una TC-P de seguimiento una vez resuelto el EE y se compararon con las TC-P realizadas en fase crítica. Además, incluimos un grupo de control en los que también se realizó un TC-P. Incluimos 19 pacientes. El análisis visual de los mapas de perfusión durante la fase crítica, mostró áreas de hiperperfusión en el 78,9% de los pacientes. El análisis cuantitativo mostró un aumento significativo de los valores de rCBF (p=0.002) y rCBV (p=0.004), y una disminución de TTP (p <0.001) MTT (p=0.001) en las áreas corticales del lado afectado versus el lado no afectado. De los 9 pacientes con una TC-P de seguimiento, 8 mostraron disminución de la intensidad, rCBV (p=0.035) y rCBF (p=0.024) en las áreas de hiperperfusión. La sensibilidad de la detección de hiperperfusión para el diagnóstico de EE fue 78.95%, y la especificidad del 90%. El análisis cuantitativo comparativo de los índices de asimetría para rCBF, rCBV y MTT entre las TC-P críticas y el grupo control mostró diferencias significativas para todos los parámetros (rCBF p=0.001; rCBV p=0.002; TTP p=0.001 y MTT p=0.001) En este estudio demostramos que la TC-P puede proporcionar información diagnóstica valiosa en pacientes con EE y complementar al EEG. Por otro lado, la identificación de factores clínicos que puedan predecir la evolución de los pacientes en EE es importante. La escala STESS (Status-Epilepticus-Severity-Score) es una herramienta para predecir la mortalidad en el EE. Sin embargo, esta escala no tiene en cuenta la situación funcional previa del enfermo. Por ello en nuestro segundo estudio quisimos valorar si el Rankin modificado (mRS) podría ser un factor pronóstico en el EE y si añadiendo esta variable al STESS se podría mejorar la predicción del pronóstico en estos pacientes. Se realizó un registro retrospectivo de los pacientes≥16 años que presentaron un EE. Realizamos curvas ROC y modelos de regresión logística para estimar las puntaciones de una nueva escala “mSTESS”(modified STESS) y comparamos con los resultados del STESS. Se incluyeron 136 pacientes. La capacidad del STESS para predecir la mortalidad fue del 74,3%, mientras que la capacidad del mRS para predecir la mortalidad fue del 65,2%. El modelo de regresión logística y las curvas ROC permitieron clasificar el mRS en tres grupos: 0 (mRS=0); 1 (mRS=1 a 3) y 2 (mRS> 3). Estos valores fueron añadidos a los otros ítems del STESS, resultando una nueva escala, el mSTESS, con puntajes entre 0 y 8 puntos. La capacidad del mSTESS para predecir la mortalidad fue del 80,1%. Un mSTESS> 4 estableció una precisión general de 81.8% para predecir la mortalidad, que fue considerablemente mayor que la precisión general del STESS≥3 (59.6%). Concluimos que el mRS basal se asocia con un alto riesgo de mortalidad, y que el mSTESS, podría ser una escala más precisa para predecir el pronóstico de pacientes con EE.
One challenge in SE management is establishing the diagnosis, particularly in patients with nonconvulsive seizures. It is known that epileptic activity causes an increase in the metabolic demand of the affected cerebral cortex, and this is accompanied by a transient increase in blood perfusion of the region, Therefore, we wanted to evaluate if the PCT cerebral perfusion could a be of value for diagnosing a SE. We included SE patients, diagnosed by EEG and clinical semiology, who prospectively underwent a PCT study in the ictal phase. Visual and quantitative analysis of the perfusion maps were performed. Asymmetry index-(AI) between affected and unaffected hemispheres were calculated for regional-cerebral blood flow-(rCBF), regional-cerebral blood volume-(rCBV), time to peak-(TTP), and mean transit time-(MTT). Nine patients underwent a follow-up PCT after SE resolution, and the corresponding maps were compared to the ictal maps. In addition, we included a control group, who also underwent acute PCT during the study period. We included 19 patients. On visual analysis of parametric perfusion maps during the ictal phase, regional cortical hyperperfusion was depicted in 78.9% of patients. Quantitative analysis showed significantly increased rCBF (p=0.002) and rCBV (p=0.004) values, and decreased TTP (p<0.001) MTT (p=0.001) in cortical areas of the affected versus the unaffected side. In the 9 patients with a follow-up PCT, 8 showed decreased intensity, rCBV (p=0.035), and rCBF (p=0.024) in the hyperperfusion areas. The sensitivity of hyperperfusion detection for the diagnosis of SE was 78.95%, specificity 90%. Comparative quantitative analysis of asymmetry indices for rCBF, rCBV, and MTT between ictal PCT and control patients showed significant differences for all parameters (rCBF p=0.001; rCBV p=0.002; TTP p=0.001 and MTT p=0.001). In this study we demonstrate that PCT may provide valuable diagnostic information in patients with SE and complement the diagnostic value of EEG. On the other hand, identifying the clinical factors that predict the outcome of patients with SE is important. The Status Epilepticus Severity Score (STESS) is a score for predicting mortality in SE. However, this scale does not consider the previous functional situation of the patient. Therefore, in our second study we wanted to assess if the baseline modified Rankin Scale (mRS) might be a prognostic factor for assessing the short-tem outcomes of SE and whether its addition to STESS can improves the prediction of mortality. We recruited consecutive patients with SE >16 years. We developed ROC curves and a logistic regression model to estimate the scores of the new score, designated as modified STESS (mSTESS) and subsequently compared it with the STESS. We included 136 patients. The capacity of STESS to predict mortality was 74.3% (IC:63.8-81.8%), while the capacity of the mRS to predict mortality was 65.2% (IC:54.2-76.2%). The logistic regression model and ROC curves enabled the classification of mRS as follows: 0 (mRS=0); 1 (mRS=1 to 3) and 2 (mRS>3). These values, when added to the other items of the STESS, resulted in the mSTESS with scores between 0 and 8 points. The capacity of the mSTESS to predict mortality was 80.1%. A mSTESS>4 established an overall accuracy of 81.8% for predicting mortality, which was considerably higher than the overall accuracy of STESS≥3 (59.6%). In this second study, we conclude that the baseline mRS was associated with high mortality risk and we propose to use mSTESS to improve the prediction of mortality risk in SE.

Orientadores: Jose Wilson Magalhães Bassani, Antonio Carlos Guimarães de Almeida
Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia Eletrica e de Computação
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Resumo: Durante o burst epileptiforme e a depressão alastrante (DA), são observados um aumento da [K+]o (concentração extracelular de potássio) e uma diminuição da [Ca2+]o (concentração extracelular de cálcio), evidenciando a participação deste mecanismo não-sináptico nestes padrões oscilatórios anormais. Essas variações nas [K+]o e [Ca2+]o elevam a excitabilidade neuronal. No entanto, não está claro se a alta [K+]o é um fator primário na geração destas atividades neuronais ou se apenas desempenha um papel secundário neste processo. Para melhor compreender a dinâmica não-linear destes padrões, as condições experimentais de alta [K+]o e zero [Ca2+]o foram replicadas em um modelo ampliado de Golomb, referente à região CA1 da formação hipocampal. Importantes mecanismos regulatórios de concentração iônica, como a bomba Na+/K+, a difusão iônica e o sistema de buffer da glia, foram acrescentados ao modelo de Golomb. Dentro destas condições, foi possível simular atividades elétricas neuronais tipicamente apresentadas no burst epileptiforme em sua fase ictal. A DA foi iniciada pela interrupção da atividade da bomba Na+/K+. O bloqueio da bomba Na+/K+ por meio da hipóxia celular é uma manobra experimental para se obter a DA, conhecida também como depressão alastrante hipóxica - DAH. A teoria de bifurcação e o método fast-slow analysis foram utilizados para estudar a interferência do K+ extracelular na excitabilidade celular. Este estudo indicou que o sistema perde a sua estabilidade com o aumento da [K+]o, transitando para um elevado estado de excitabilidade. Este crescimento da [K+]o provoca bifurcações no comportamento dinâmico neuronal, que determinam transições entre diferentes estágios dessas atividades elétricas. No primeiro estágio, o aumento da [K+]o propicia a deflagração do burst epileptiforme e da DA via bifurcações sela-nó e de Hopf supercrítica, respectivamente. Ao longo da atividade neuronal, o nível de excitabilidade é mantido por meio de um crescimento contínuo da [K+]o, que deprime as correntes de K+ em um processo de realimentação positiva. Neste estágio, em relação ao burst epileptiforme, a amplitude e a freqüência dos disparos dos potenciais de ação são alteradas via bifurcação de Hopf supercrítica. No último estágio, com a depressão das correntes de K+, a bomba de Na+/K+ tem uma participação importante no término da atividade neuronal. O burst epileptiforme e a DA são finalizados por meio das bifurcações sela-órbita homoclínica e sela-nó, respectivamente. Portanto, este trabalho sugere que o K+ extracelular pode desempenhar um papel fundamental na dinâmica não-linear do burst epileptiforme e da sua transição para a DA.
Abstract: During the epileptiform burst and the spreading depression (SD), it is observed an increase of [K+]o (extracellular potassium concentration) and a decrease of [Ca2+]o (extracellular calcium concentration), pointing out the participation of this nonsynaptic mechanism in these abnormal oscillatory patterns. These ionic variations raise the neuronal excitability. However, whether the high [K+]o is a primary factor in the beginning of these neuronal activities or just plays a secondary role into this process is unclear. To better understand the nonlinear dynamics of these patterns, the experimental conditions of high [K+]o and zero [Ca2+]o were replicated in an extended Golomb model in which we added important regulatory mechanisms of ion concentration as Na+/K+ pump, ion diffusion and glial buffering. Within these conditions, it was possible to simulate epileptiform burst within the ictal phase. The SD was elicited by the interruption of the Na+/K+ pump activity. The blockage of Na+/K+ pump by cellular hypoxia is an experimental procedure to elicit SD, known as hypoxic spreading depression - HSD. The bifurcation theory and the method of fast-slow analysis were used to study the interference of extracellular K+ in the cellular excitability. This analysis indicates that the system loses its stability at a high [K+]o, transiting to an elevated state of neuronal excitability. This raise of [K+]o provokes bifurcations in the neuronal dynamic behavior, that determine transitions between different stages of these electrical activities. In the initial stage, the increase of [K+]o creates favorable conditions to trigger the epileptiform burst and the SD by saddle-node and supercritical Hopf bifurcations, respectively. During the neuronal activity, the level of excitability is maintained by a continuous growth of [K+]o that depresses K+ currents in a positive feedback way. At this stage, concerning epileptiform burst, the amplitude and frequency of action potentials are changed by supercritical Hopf bifurcation. At the last stage, with the depression of K+ currents, the Na+/K+ pump plays an important role in the end of neuronal activity. The epileptiform burst and SD activities terminate by saddle-homoclinic orbit and saddle-node bifurcations, respectively. Thus, this work suggests that [K+]o may play a fundamental role in the nonlinear dynamics of the epileptiform burst and the transition to SD.
Doutorado
Engenharia Biomedica
Doutor em Engenharia Elétrica

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
A lesão medular não é um evento incomum na medicina humana e veterinária e resulta em disfunções neurológicas de graus variados. Apesar dos esforços no tratamento, lesões medulares frequentemente causam sequelas permanentes, desde déficit proprioceptivo isolado até paralisia completa de membros. A membrana confeccionada à base de látex natural extraído da seringueira Hevea brasiliensis tem sido utilizada com sucesso para regeneração de tecidos, e seu potencial regenerador foi reconhecido como sendo de uma fração proteica conhecida como proteína P1. Este novo biomaterial foi eficaz em vários testes de regeneração em animais de experimentação e humanos, e, ainda que tenha mostrado potencial regenerador em teste de lesão de nervo periférico, a proteína ainda não foi testada em Sistema Nervoso Central. Objetivou-se avaliar o potencial regenerador da fração proteica extraída do látex natural (P1) na hemissecção da medula espinhal em ratos. Foram utilizados 18 ratos machos adultos submetidos à hemissecção medular no segmento medular T13 subdivididos em dois grupos: GHP (tratados com proteína P1 em gel de ácido hialurônico) e GHSP (tratado somente com ácido hialurônico). Foram realizadas análises neuroclínicas, bem como avaliação funcional da marcha em plataforma de acrílico. Após oito semanas os animais foram submetidos à eutanásia e os segmentos medulares envolvidos foram avaliados por histoquímica pela coloração Tricrômico de Masson e Luxol Fast Blue, além de avaliação da reação astroglial por imunoistoquímica. Não houve diferença significativa entre os dois grupos para as avaliações neuroclínicas, não sendo observado melhora clínica satisfatória para as variáveis analisadas, porém, em relação à análise histológica foi notada diferença importante entre os grupos, evidenciando menor reação cicatricial e degeneração do tecido medular restante, além de presença de hipercelularidade organizada e menor reação astroglial nos animais tratados com a fração protéica P1. Apesar da não evidência de melhora nos parâmetro neuroclínicos com o uso da proteína P1, houve alterações histológicas notórias e relevantes nos animais em que foi utilizada, além de ser benéfica como indutora de neuroproteção.
Spinal cord injury is not an uncommon event in human and veterinary medicine and results in neurological dysfunctions of varying degrees. Despite efforts in treatment, spinal cord injuries often cause permanent sequelae, ranging from isolated proprioceptive deficit to complete limb paralysis. The membrane made from natural latex extracted from the rubber tree Hevea brasiliensis has been successfully used for tissue regeneration and its regenerative potential has been recognized as being of a protein fraction known as P1 protein. This new biomaterial has been effective in several regeneration tests in experimental animals and humans, and although it has shown a potential regenerator in a peripheral nerve injury test, the protein has not yet been tested in the Central Nervous System. The objective of this study was to evaluate the regenerative potential of the protein fraction extracted from natural latex (P1) in the hemisection of the spinal cord in rats. Eighteen adult male rats submitted to medullary hemisection in the T13 medullary segment were subdivided into two groups: GHP (treated with protein P1 in hyaluronic acid gel) and GHSP (treated with hyaluronic acid only). Neuroclinical analyzes were performed as well as functional evaluation of gait on acrylic platform. After eight weeks the animals were submitted to euthanasia and the involved spinal segments were evaluated by histochemistry by Masson and Luxol Fast Blue Trichrome staining, as well as evaluation of the astroglial reaction by immunohistochemistry. There was no significant difference between the two groups for the neuroclinical evaluations, and no satisfactory clinical improvement was observed for the analyzed variables; however, in relation to the histological analysis, an important difference between the groups was observed, evidencing a smaller cicatricial reaction and degeneration of the remaining spinal cord tissue, besides the presence of organized hypercellularity and less astroglial reaction in the animals treated with the protein fraction P1. Although there was no evidence of improvement in the neuroclinical parameters with the use of the P1 protein, there were significant and relevant histological changes in the animals in which it was used, besides being beneficial as inducer of neuroprotection.

The survey was conducted in order to verify the acquisition of skills and competencies in Neurology for medical training, required by the National Curriculum Guidelines. Introduced methodological innovation in Neurology discipline, encouraging the students to produce a vídeo on Stroke as a pedagogical activity. For this the students performed literature search and preparation of the script of the vídeos considered relevant aspects of the clinical picture, diagnosis, as well as ethical and social issues surrounding this type of pathology. The choice of vídeo contente was due to the high incidence and prevalence of the condition, being relevant to the medical population been abled in diagnosis, treatment and empower people about prevention and care strategies. After the presentation of the videos, students completed questionnaires about the use of this methodology for learning, to develop skills and competencies in medical training. The information obtained was analyzed and showed that the vídeo production favored the learning contentand contributed to the development of skills, competencies and attitudes in Neurology for medical training.
A pesquisa foi realizada com o objetivo de verificar a aquisição de habilidades e competências em Neurologia, para a formação médica, requeridas pelas Diretrizes Curriculares Nacionais. Introduzimos inovação metodológica na disciplina de Neurologia, estimulando os discentes a produzirem um vídeo sobre Acidente Vascular Cerebral como atividade pedagógica. Para isso os estudantes realizaram pesquisa bibliográfica e na elaboração do roteiro dos vídeos consideraram aspectos relevantes do quadro clinico, diagnóstico, bem como questões éticas e sociais que envolvem esse tipo de patologia. A escolha do conteúdo dos vídeos foi decorrente da alta incidência e prevalência da patologia, sendo relevante para a comunidade médica estar habilitada no diagnóstico, tratamento e empoderando a população sobre estratégias de prevenção e cuidados. Após a apresentação dos vídeos, os estudantes responderam questionários sobre o uso dessa metodologia para a aprendizagem, para o desenvolvimento de habilidades e competências na formação médica. As informações obtidas foram analisadas e apontaram que a produção de vídeo favoreceu a aprendizagem do conteúdo e contribuiu para o desenvolvimento de habilidades, competências e atitudes em Neurologia para a formação médica. As oficinas foram o Produto de Intervenção proposto com o objetivo de envolver os integrantes do processo educacional, proporcionando o compartilhar de saberes e o incentivo para a implementação das estratégias de ensino-aprendizagem coerentes com as competências gerais das Diretrizes Curriculares Nacionais do Curso de Graduação em Medicina.

Dissertação de Mestrado Integrado em Medicina Veterinária
As hérnias discais resultam de um deslocamento parcial ou total do disco intervertebral (DIV), o qual pode ter como causa a degeneração do DIV ou, mais raramente, um trauma. Quando o DIV altera as forças mecânicas normais exercidas sobre a coluna vertebral resultam, frequentemente, na deslocação de material degenerado do disco para o exterior e consequente compressão da medula espinhal. As hérnias discais podem ocorrer em qualquer espécie animal, no entanto, ocorrem mais frequentemente em cães, sobretudo em raças condrodistróficas. Os sinais clínicos são variáveis, podendo ser confundidos com outras lesões medulares. O diagnóstico precoce, assim como um tratamento adequado são essenciais para que a resolução da lesão seja bem sucedida. O tratamento das hérnias discais pode ser médico ou cirúrgico dependendo da gravidade dos sinais clínicos. O tratamento médico consiste em manter o animal em repouso absoluto numa jaula, de modo a restingir os seus movimentos durante três a quatro semanas, devendo o exercício físico ser posteriormente reintroduzido de uma forma gradual, enquanto o tratamento cirúrgico deve ser realizado até um período de 48h após o início dos sinais clínicos. Qualquer uma das situações deve ser sempre combinada com um protocolo de reabilitação física, o qual é definido consoante os sinais clínicos apresentados e os problemas a resolver. Nesta dissertação serão apresentados e discutidos quatro casos clínicos de hérnias discais e algumas das possíveis razões para o insucesso na reabilitação física das mesmas.
ABSTRACT - Physical rehabilitation of postsurgical neurological patient - Discal herniation results from a partial or a total displacement of the intervertebral disc (IVD), which may be caused by degeneration of the IVD or more rarely by a trauma. When IVD degenerate, normal mechanical forces exerted on the spinal column, often result in degenerated disc material from being squeezed out and, therefore, in spinal cord compression. The disc herniation can occur in any animal species, but they are more common in dogs, especially in condrodistrophic races. The clinical signs are variable and can be confused with other spinal injuries. Early diagnosis and the appropriate treatment are essential for a successful lesion resolution. The treatment of discal herniation may be medical or surgical depending on the severity of clinical signs. Medical treatment consists on keeping the animal in a cage, to restricte their movements for three to four weeks, so exercise can later be reintroduced in a gradual manner, while surgical treatment should be performed within 48h after onset of clinical signs. Either situation should always be combined with a physical therapy protocol, which is defined according to the presented clinical signs and problems to solve. In this dissertation will be presented and discussed four clinical cases of discal herniation and some of the possible reasons for the failure of the physical rehabilitation of the same.

Esta pesquisa buscou investigar como se constitui a demanda para a Neurologia a partir dos professores no contexto da Educação Básica. Os referenciais teóricos foram construídos a partir de dois eixos: um olhar crítico para o campo da neuroeducação e seu crescimento a partir dos anos 1990; propõe-se o termo neurocolonização, onde os saberes das neurociências seriam imprescindíveis para a Educação. O outro eixo envolveu o debate sobre os processos de medicalização, formulados a partir de uma leitura panorâmica e de uma revisão de duas genealogias – a de Michel Foucault (2010) e a de Jurandir Freire Costa (1979). A discussão teórica também incluiu um olhar para a interface Psiquiatria-Educação e para a fronteira-território que se constitui entre Psiquiatria e Neurologia. Realizou-se um levantamento dos encaminhamentos feitos à Neurologia na cidade de Novo Hamburgo/RS no segundo semestre de 2015. Junto às cartas de referência analisadas neste levantamento foram encontrados seis documentos assinados por professores. Cinco das seis professoras signatárias destes documentos foram entrevistadas, tematizando a construção de si e os percursos profissionais; os encontros com os trabalhadores de saúde; as hipóteses e expectativas em torno do caso da criança que decidiram solicitar encaminhamento, e como percebem a influência das neurociências sobre o seu trabalho. A análise dos encaminhamentos aponta para uma frequência de situações relacionadas ao campo da Educação superior aos casos de cefaleia e epilepsia/convulsões As entrevistas oferecem indícios da constituição da demanda para a Neurologia apoiada em diferentes elementos: a) o recurso ao saber especialista, demarcado especificamente no dispositivo ‘consulta’; b) o deslizamento dos discursos das neurociências-pesquisa – que se remetem a uma criança qualquer - em direção ao que se constrói na prática clínica e também na prática pedagógica, uma relação entre sujeitos reais; c) a não-aderência a um sistema diagnóstico ou a um campo de problemas, podendo-se falar de crianças e adolescentes descabentes, que povoam as margens das classificações diagnósticas e alimentam um circuito tautológico entre Saúde e Educação; d) hipóteses formuladas pelos professores centradas no modo da família criar sua prole e na expectativa de que o neurologista interfira nestas relações, numa tentativa de normatizar ou padronizar as condutas entre família e escola; e) pouca ênfase à importância dos saberes das neurociências ou dos exames complementares para a tomada de decisão nestes encaminhamentos. Por outro lado, práticas desmedicalizantes também foram reconhecidas a partir da sensibilidade do olhar das ensinantes, ressignificando as diferenças e desarmando os automatismos patologizantes

La adaptación que ocurre en el sistema auditivo es un fenónemo que todos experimentamos cuando dejamos de oir sonidos irrelevantes, constantes o incluso molestos. La adaptación para sonidos conocidos aumenta también la sensibilidad y la percepción para estímulos nuevos o poco conocidos. Por tanto, la similaridad entre la historia previa de estimulación y la subsiguiente también puede influenciar la adaptación. La adaptación a la estimulación repetida es un fenómeno que se ha visto en diferentes modalidades sensoriales o especies de animales. El curso temporal de la adaptación en corteza auditiva primaria (A1) se ha estudiado principalmente en intervalos entre estímulos muy rápidos (<400ms) y diferentes mecanismos han sido sugeridos (inhibición sináptica, disbalance excitación-inhibición, inhibición lateral, disminución de la excitación, o inhibición aumentada), aunque los mecanismos intrínsecos neuronales casi no han sido considerados. Por otro lado, numerosos estudios han mostrado el efecto que tiene la anestesia sobre la excitabilidad cortical, pudiendo, por tanto, afectar al estudio de la adaptación. Por último, la adaptación podría estar influenciada por estructuras subcorticales (como el colículo inferior o el tálamo) aunque la influencia intracortical también se ha demostrado. El primer estudio presentado en este trabajo tiene como objetivo caracterizar, en la rata despierta en movimiento, el curso temporal de la adaptación auditiva en las neuronas únicas de A1 aisladas con tetrodos. Con este propósito, se estudió cómo el intervalo entre estímulos, la duración o la intensidad de la estimulación previa afectaba a la amplitud de respuesta y su latencia de respuesta. También se estudió el curso temporal durante la estimulación sostenida y el fenómeno de la postadaptación. La comprensión de cómo la actividad neuronal codifica la información sensorial sigue siendo una cuestión fundamental en el campo de la atención auditiva. Así, la codificación de la información temporal es un aspecto clave en A1. El análisis de la “información mutua” de la respuesta neuronal nos permite cuantificar el contenido de la información de la actividad neuronal. Por otro lado, la variabilidad de la respuesta neuronal podría ser un parámetro clave para la codificación de los estímulos relevantes durante una tarea. También, la respuesta neuronal sostenida se ha sugerido que podría aportar información adicional en el animal en comportamiento. Hasta el momento, se desconoce cómo las neuronas únicas de A1 codifican la categoría temporal de los estímulos auditivos. Con este objetivo se registró la actividad de neuronas únicas en A1, por medio de tetrodos, en el animal en comportamiento. Las ratas debían discriminar si dos sonidos idénticos estaban separados por 150 o 300 ms y se ha estudiado el contenido de la información, la variabilidad y las respuestas post-estímulo de la actividad neuronal en el estado atentivo y el pasivo del animal.

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
A modalidade de rodeio denominada Prova do Laço de Bezerro tem sido questionada sobre a ocorrência de possíveis lesões nas vértebras cervicais ocasionadas pela tração da corda no pescoço dos bezerros. Neste trabalho avaliou-se 15 bezerros mestiços, machos ou fêmeas, entre cinco e seis meses de idade experimentalmente submetidos a prova do laço. Os animais foram laçados três vezes por semana, em dias alternados, durante cinco semanas, somando o total geral de 225 laçadas. A prova experimental foi realizada de forma semelhante à prova oficial, pelo mesmo cavaleiro profissional da modalidade. Os bezerros foram avaliados mediante exame clínico geral e neurológico ao início da primeira, durante a terceira e ao término da quinta semana experimental. Radiografias simples e contrastadas das vértebras cervicais foram efetuadas ao início da primeira e ao término da quinta semana de experimento. Os métodos de laçadas foram acompanhados e classificados qualitativamente em fortes ou fracos. Não foram encontradas alterações clínicas e radiográficas nos animais durante o experimento. O rigor da laçada foi considerado forte em 77% dos casos. O fato de não terem sido encontradas alterações clínicas e radiográficas indicam que a ocorrência de lesões cervicais em bezerros submetidos a prova de laço não é tão alta como o propalado, entretanto, trata-se de procedimento rude e agressivo. Número mais expressivo de experimentos semelhantes a este deverá ser conduzido tanto sob condições controladas como em provas reais para confirmar os dados da presente pesquisa.
The modality of roundup Calf Roping has been questioned on the occurrence of possible injuries in the cervical vertebrae caused by the rope tension in calfs neck. In this work 15 calves, male or female, ages varying from five and six months experimentally submitted to calf roping were evaluated. The procedure was carried through three times per week, in alternated days, during five weeks, adding the total of 225 lassoed. The experimental test was carried through of similar form to the official test, for the same professional knight of the modality. The calves had been evaluated by means of general and neurological clinical examination to the beginning of the first one, during third and to the ending of the fifth experimental week. Simple and contrasted x-rays of the cervical vertebrae had been made to the beginning of the first one and the ending of the fifth week of experiment. The lassoed methods had been observed and classified qualitatively in weak or strong. Clinical and radiographic alterations in the animais during the experiment had not been found. The severity of the lassoed was considered strong in 77% of the cases. The fact not to have been found clinical and radiographic abnormalities indicates that the occurrence of cervical injuries in calves submitted to calf roping is not as high as divulged, however, is about aggressive and rude procedure. Similar experiments must be made in such a way to be lead under controlled conditions as in real tests to confirm the data of the present research.

This thesis will describe two functional Magnetic Resonance Imaging (fMRI) experiments and one Voxel-Based Morphometry (VBM) study, each investigating how the human brain identifies objects and their associated properties. In particular, we used three different categories of objects – living (animals), nonliving (tools and nontools) and faces (famous and non-famous) – to examine the type of knowledge attribute in question: one perceptual (movement) and two semantic attributes (typical object location and biographic knowledge). We know from neuropsychological literature that the most anterior portions of the temporal cortices critically support human conceptual knowledge. Unfortunately, the Anterior Temporal Lobe (ATL) is a challenging region for fMRI due to susceptibility artifacts, especially at high fields. For these reasons we established an optimized fMRI protocol (described in the second Chapter) by adjusting key acquisition parameters like phase-encoding gradient polarity, slice thickness, echo time, and slice angle. The protocol gave reliable Blood-Oxygen-Level Dependence (BOLD) signal sensitivity in the ATL. Clinical data describe patients with specific semantic impairments at the level of category (living, nonliving) as well as disproportionate deficits for a modality or type of knowledge (e.g., visual/perceptual knowledge or manipulation knowledge). Functional neuroimaging studies on semantic organization with normal subjects found an “action network†specific for tools rather than living items. In the first experiment (Chapter 3) we devised an fMRI paradigm to investigate the processing of movement (action) and place (encyclopedic) features, and their influence on category-specific activations. Within the “movement network†statistical analyses did not show any significant interaction between categories. These findings suggest that the visuomotor “action network†is not specific for tools because it is also activated when the action related knowledge is elicited for other categories, such as animals. The second and the third experiment (Chapter 4) focus on the processing of faces. Neuropsychological literature attributes semantic and lexical retrieval deficits in patients to ATL lesions. In Part I of Chapter 4, we report data from a VBM study on patients with known lesions in the temporal lobe. Unfortunately, as far as we know, data on patients and functional neuroimaging in healthy individuals has not clarified the differential role of this area in the two mental operations because semantic and lexical processes usually occur simultaneously and automatically. In Part II, we devised an event-related fMRI activation paradigm that allowed us to study the identification (i.e., association of semantic biographical information) of celebrities, with and without the ability to retrieve the proper name. While semantic retrieval reliably activated the ATL, only more posterior areas in the left temporal and temporal-parietal junction were significantly modulated by covert lexical retrieval. These results support findings from patients with ATL lesions and suggest that their anomia is due to semantic rather than lexical retrieval impairment.

The highly complex nervous system is built upon an intricate network of neurons. In order to make a functional network, the establishment of precise connections is crucial. Neuronal networks are established early during development when neurons send out axons that navigate through complex environments to connect to their target. Chemotropic attractant or repellent cues, cell adhesion molecules, morphogens and a wide range of factors secreted or expressed by guidepost cells enable axon guidance. The leading tip of the axon, the GC is important to sense the environment and integrate extracellular signals to navigate precisely. The axonal GC has a large repertoire of mRNAs that are dynamic in nature. Local regulation of transcripts in navigating axons is suspected to ensure precise pathfinding. However, mechanisms involving regulation of expression of these transcripts within GCs are largely unknown. This thesis investigates whether microRNAs, one of the quintessential posttranscriptional regulators, can regulate axon guidance by fine-tuning mRNA expression within subcellular compartments. To explore microRNA roles in axon guidance, Xenopus laevis visual system was used as a model. Profiling axons of retinal ganglion cells revealed the presence of miRNAs within axons. The most abundant axonal miRNAs, the miR-181 family and miR-182, exhibit distinct roles in regulating axon guidance in vivo. Loss of function analyses suggests that both miRNA families are required for accurate axonal targeting but involve different mechanisms. Thus, specific axonal microRNAs locally regulate mRNAs contributing to error-free pathfinding.

Functional Magnetic Resonance Imaging (fMRI) has consistently highlighted aberrant functional connectivity across brain regions of autism spectrum disorder (ASD) patients. However, the manifestation and neural substrates of these alterations are highly heterogeneous and often conflicting. Moreover, their neurobiological under- pinnings and etiopathological significance remain largely unknown. A deeper understanding of the complex pathophysiological cascade leading to impaired connectivity in ASD can greatly benefit from the use of model organisms where individual pathophysiological or phenotypic components of ASD can be recreated and investigated via approaches that are either off limits or confounded by clinical heterogeneity. In this work, we first describe the intrinsic organization of the mouse brain at the macroscale as seen through resting-state fMRI (rsfMRI). The analysis of a large rsfMRI dataset revealed the presence of six distinct functional modules related to known brainwide functional partitions, including a homologue of the human default-mode network (DMN). Consistent with human studies, interconnected functional hubs were identified in several sub-regions of the DMN, in the thalamus, and in small foci within integrative cortical structures such as the insular and temporal association cortices. We then study the effects of mutations in contactin associated protein-like 2 (Cntnap2), a neurexin-related cell-adhesion protein, on functional connectivity. Homozygous mutations in this gene are strongly linked to autism and epilepsy in humans, and using rsfMRI, we showed that homozygous mice lacking Cntnap2 exhibit aberrant functional connectivity in prefrontal and midline functional hubs, an effect that was associated with reduced social investigation, a core “autism trait†in mice. Notably, viral tracing revealed reduced frequency of prefrontal-projecting neural clusters in the cingulate cortex of Cntnap2−/− mutants, suggesting a possible contribution of defective mesoscale axonal wiring to the observed functional impairments. Macroscale cortico-cortical white-matter organization appeared to be otherwise preserved in these animals. These findings revealed a key contribution of ASD-associated gene CNTNAP2 in modulating macroscale functional connectivity, and suggest that homozygous loss-of-function mutations in this gene may predispose to neurodevelopmental disorders and autism through a selective dysregulation of connectivity in integrative prefrontal areas. Finally, we discuss the role mouse models could play in generating and testing mechanistic hypotheses about the elusive origin and significance of connectional aberrations observed in autism and recent progress towards this goal.

Conselho Nacional de Desenvolvimento Científico e Tecnológico
A retrospective study including dogs with neurological disease was conducted at the Service of Neurology (SN) of the Veterinary Teaching Hospital, Universidade Federal de Santa Maria (UFSM) between 2006 and 2013, with the objective to identify and characterize the age, breed and sex, neurological diseases and classify them according to the anatomical region and DINAMIT-V acronym. Were evaluated 1.277 neurological records of dogs and obtained the information for inclusion in the study in 1.184 of them being the etiological diagnosis in 525 (44,4%) and presumptive in 659 dogs (55,6%). The most common breeds were dachshunds (27,5%), followed by mixed breed. The most common sites were the spinal cord between T3-L3 (40,9%) and thalamus-cortex (17,5%). Most dogs were diagnosed with degenerative disorders (49%), being disc disease intervertebral more observed, followed inflammatory/infectious disease (16,6%). It can be concluded that the higher prevalence of neurological disorders in dogs involve the spinal cord and thalamus-cortex, with the most frequent being degenerative and the data obtained may assist future studies associated with frequency and distribution of the main neurological diseases in dogs.
Foi realizado um estudo retrospectivo de cães atendidos no Serviço de Neurologia (SN) do Hospital Veterinário Universitário (HVU) da Universidade Federal de Santa Maria (UFSM), entre 2006 e 2013, com o objetivo de identificar e caracterizar a idade, raça e sexo, as doenças neurológicas e classificá-las de acordo com a região anatômica e acrônimo DINAMIT-V. Foram avaliadas 1.277 fichas neurológicas de cães e obtidas as informações para inclusão no estudo em 1.184 delas, sendo o diagnóstico etiológico confirmado em 525 cães (44,4%) e presuntivo em 659 (55,6%). As raças mais frequentes foram dachshunds (29,2%), seguido das sem raça definida. Os locais mais comuns foram a medula espinhal entre T3-L3 (40,9%) e tálamo-córtex (17,5%). A maioria dos cães foram diagnosticados com doenças degenerativas (49%) , sendo a doença do disco intervertebral a mais observada, seguido das doenças inflamatórias/infecciosas (16,6%). Pode-se concluir que a maior prevalência das doenças neurológicas de cães envolvem a medula espinhal e tálamo-cortex, sendo as degenerativas as mais frequentes e os dados obtidos poderão auxiliar em futuros estudos sobre a frequência e distribuição das principais doenças neurológicas em cães.

L’objectiu principal d’aquesta tesis doctoral ha estat estudiar l’estat del Se en sang i LCR de pacients amb errors congènits del metabolisme amb tractament dietètic i altres trastorns neuropediàtrics de diferent etiologia. En el primer treball ens plantejàvem l’establiment de valors de referència per a determinats elements traça en sang total. La necessitat venia de la inexistència d’estudis sobre l’estat d’aquests elements en poblacions pediàtriques afectades de diferents errors congènits del metabolisme. En el segon treball ens plantejàvem l’establiment de valors de referència per a seleni en LCR. La necessitat venia de la inexistència d’estudis sobre l’estat d’aquest element en poblacions pediàtriques i de la possible associació entre una alteració de l’estat de Seleni i algunes disfuncions neurològiques. Un cop establerts els valors de referència de Se en LCR, el tercer treball consistia en valorar l’estat de Se en pacients amb malalties neurològiques per intentar detectar alguna deficiència o excés de Se en els líquids d’aquests pacients. A part de valorar l’estat de Se, preteníem valorar també altres paràmetres bioquímiques com HVA, 5-MTHF, 5-HIAA, proteïnes, activitat de GPx i pterines, per tal de veure si les correlacions observades en l’anterior treball es mantenien. Finalment, l’últim treball es centrava en l’estudi d’un grup de pacients amb uns valors de Se en LCR permanentment elevats sense tenir-ne una explicació clara. Aquests pacients (que s’havien identificat en el tercer treball) estaven afectats de la síndrome de Kearns-Sayre (malaltia mitocondrial que es manifesta per deleció del DNA mitocondrial). En aquest nou treball ens plantejàvem l’estudi d’uns paràmetres bioquímics que ens permetessin detectar la disfunció del plexe coroïdal en pacients afectats d’aquesta síndrome. Així doncs i a títol de discussió, una de les nostres principals necessitats pel control regular dels pacients amb malalties metabòliques i tractament dietètic era la monitorització de diferents elements traça en diferents fluids biològics. Tradicionalment s’havia descrit deficiències d’alguns d’aquests elements en pacients que tenen com a base del seu tractament les dietes restrictives. De tots els elements, el Se semblava un candidat ideal, ja que prèviament s’havien descrit deficiències en algunes malalties, com la fenilcetonúria.La importància del Se radica en que és un element essencial per al correcte funcionament de les selenoproteïnes. Per tant, ens semblava d’interès monitoritzar les concentracions de Se tant en sang (com a marcador de dèficit nutricional) com en LCR (com a marcador potencial de mecanismes fisiopatològics en malalties d’àmbit neuropediàtric). Les fites més importants del nostre treball han estat tres: 1. L’estandardització del procediment en sang ha permès establir la tècnica de forma rutinària per al control de trastorns nutricionals al nostre centre. Des d’un punt de vista científic, la conclusió més important és que el dèficit de seleni és comú a les malalties metabòliques amb tractament dietètic i que per tant el seu estudi i suplementació són necessàries per a l’òptim control de les malalties. 2. Respecte a l’estudi del seleni a nivell central, les aportacions han estat més rellevants ja que era un camp poc estudiat. Hem establert per primera vegada valors normals de Se en LCR per a una població pediàtrica i confirmat que el cervell es un òrgan que regula mot bé l’homeòstasi d’aquest element traça, ja que els valors eren independents dels observats en sang. 3. Especialment rellevant creiem que és el perfil que ens va permetre valorar la funció del plexe coroïdal en la síndrome de KSS i el diagnòstic directe d’un cas. Aquest resultats no tan sols tenen un valor per a la pràctica diagnòstica, sinó que obren la porta a la investigació d’altres malalties en les que s’afecta el plexe coroïdal.
PhD STUDENT NAME Mireia Tondo Colomer TESIS TITLE Blood and cerebrospinal fluid selenium status in paediatric patients affected with neurological diseases. SUMMARY Our objective was to study blood and CSF Se status in patients with IEM with dietary treatment and other neurological disorders with different clinical aetiology. In this first study our objective was the establishment of reference values for some trace elements in whole blood. At that time, studies of several trace element status for a paediatric population with different inborn errors of metabolism were scarce. To achieve this aim, we used an Induced Couple Plasma Mass Spectrometer (ICP-MS), which allowed multiple analyses of different trace elements using a small sample volume. Leftover samples form patients who came to our hospital for minor surgery were used as reference population. In this second study our objective was the establishment of cerebrospinal fluid selenium reference values. At that time, no previous studies of Se central nervous system status for a paediatric population existed in the scientific literature. CSF Se concentrations had to be considered in order to find out any possible association between Se status and some neurological functions. Moreover, the implications of Se in the synthesis and function of more than 20 selenoproteins indicated that central nervous system Se status was important. After the establishment of CSF Se reference values for a paediatric population, the next step was to evaluate Se CSF concentrations in paediatric patients with different neurological disorders. Previous studies demonstrated the tight regulation of Se homeostasis in brain as well as the evidence of the role of Se in CNS disorder pathophysiology. In spite of growing evidence of the role of Se in CNS, no studies of Se CNS status in a large cohort of paediatric patients affected with different neurological disorders had been carried out. Finally, our last study derived from the one before which allowed us to identify a group of patients with clearly high CSF Se values. The main common feature was that all patients had a mitochondrial DNA deletion syndrome (Kearns Sayre Syndrome). This group of patients was not included in the previous work and was treated separately.

Mestrado em Tradução Especializada
O presente relatório foi realizado no âmbito do Mestrado de Tradução Especializada da Universidade de Aveiro com base numa tradução de um artigo especializado na área da Neurologia, mais concretamente a harmonização da medicina moderna com a medicina tradicional no combate às doenças neurodegenerativas, especificamente a doença de Alzheimer. As línguas em estudo são o inglês e o português e o projeto contém todos os processos tradutológicos envolvidos na sua elaboração, bem como apresenta o uso de ferramentas online e softwares especializados da área e os problemas aquando da realização. Também inclui um glossário terminológico que reúne todo o vocabulário científico encontrado.
This report was carried out throughout my second year in the Master’s degree program in Specialized Translation at the University of Aveiro. It focuses on the translation of a specialized article in the field of Neurology, more specifically on the harmonization of modern medicine and traditional medicine against neurodegenerative diseases, especially Alzheimer’s disease. The working languages are English and Portuguese and it contains all the translation processes involved in its conception, as well as all the online tools and specialized softwares utilized for this purpose while outlining the problems that emerged during its elaboration. A terminological glossary, which brings together all the scientific vocabulary encountered, is also included.

La Podocalixina es una proteína que pertenece a la familia de proteínas CD34/Sialomucinas, altamente glicosilada y sializada. Ampliamente distribuida por el organismo, presentando diversas funciones según en el tipo celular en el que se encuentre y del grado de sialización que posea. Durante el desarrollo de esta tesis se ha demostrado que la Podocalixina está ampliamente distribuida en el Sistema Nervioso Central en desarrollo y en adulto, estando altamente sializada en el tejido neural, y en Sistema Nervioso Periférico. Una de las alteraciones más dramáticas producidas por la ausencia de Podocalixina en neuronas es el incremento en el crecimiento y ramificación de las neuritas y axones. Asimismo, mostramos que la ausencia de Podocalixina da como resultado un menor número de contactos sinápticos en el Sistema Nervioso Central y en el Sistema Neuromuscular, lo que sugiere que la Podocalixina es requerida para la correcta formación y/o estabilización de sinapsis. Nuestro estudio también demuestra que los efectos de la Podocalixina en ramificación neurítica y sinaptogénesis no requieren del ácido siálico presente en la proteína, sino que dependen de la propia Podocalixina per se. Estos proceso del desarrollo pueden ser mediados por la vía NHERF1-2/Ezrina/RhoA-G, que une la Podocalixina al citoesqueleto de actina. El estudio de las nuevas neuronas generadas en el giro dentado del hipocampo adulto nos permite aportar fuertes evidencias en el desarrollo e integración aberrante de las nuevas neuronas generadas en el adulto en ausencia de Podocalixina, mostrando por primera vez que la Podocalixina es esencial para el establecimiento del desarrollo de estas neuronas dentro de la red neuronal activa. La Podocalixina regula negativamente la dendritogénesis y espinogénesis de las nuevas neuronas generadas en el adulto, mientras que es un regulador positivo de la sinaptogénesis. Los estudios que se han llevado a cabo han demostrado que la Podocalixina también está expresada en OPCs y en oligodendrocitos maduros. Tras el estudio in vivo de los animales deficientes en Podocalixina, vimos que la migración de OPCs estaba alterada, puesto que en estadios embrionarios los OPCs no salían de las zonas de producción, quedando desprovistas de OPCs las zonas de destino final. Además, nuestros datos también muestran evidencia de que el ácido siálico presente en la Podocalixina es esencial en la migración de OPCs dependiente de Podocalixina. Los OPCs tienen una respuesta migratoria a CXCL12, inhibida por antagonistas de CXCR4 o anticuerpos frente a CXCL12 (Dziembowska, Tham et al. 2005). Esta respuesta inhibitoria de la migración direccional hacia la fuente de CXCL12 también queda bloqueada en ausencia de Podocalixina o por la adición del ectodominio de la Podocalixina, con o sin ácido siálico. La migración de los OPCs está afectada por el ácido siálico presente en la Podocalixina, pero la migración direccional en respuesta a CXCL12 viene mediada por la propia proteína Podocalixina, independientemente del ácido siálico presente en ella. Para estudiar la función de una proteína es muy importante conocer con que proteínas interacciona. El conjunto de proteínas que hemos visto que interaccionan con la Podocalixina lo podemos subdividir en proteínas implicadas en diversos mecanismos celulares: proteínas que interaccionan con el citoesqueleto, proteínas involucradas en la formación/mantenimiento de sinapsis, proteínas que intervienen en la maquinaria de exocitosis neuronal, proteínas asociadas a mielina y proteínas de señalización intracelular. Estas interacciones con la Podocalixina se pueden producir directamente o mediante la interacción con otra proteína/complejo proteico. Estudios muy preliminares dan evidencia de la alteración del sistema nigro-estriatal en los ratones deficientes en Podocalixina.
Role of Podocalyxin protein in the Nervous System Podocalyxin is a protein belonging to the CD34/Sialomucin family, highly glycosylated and sialyalted. Widely distributed throughout the body, presents various functions depending on the cell type and the degree of sialylation. The development of this thesis has shown that Podocalyxin is highly sialylated and widely distributed in the Central and Peripheral Nervous System. One of the most dramatic changes caused by the absence of Podocalyxin in neurons is the increase in the growth and branching of axons and neurites. Furthermore, neurons present less synaptic contacts, suggesting that Podocalyxin is required for proper formation and/or stabilization of synapses. Our study also shows that the effects of Podocalyxin in neuritic branching and synaptogenesis do not requiere the sialylation of the protein, unlike in dentritic length, where it is needed. This process of development can be mediated by NHERF1-2/Ezrin/RhoA-G, linking Podocalyxin to the actin cytoskeleton. We provide strong evidences on aberrant development and integration of adult-born neurons in the hippocampal dentate gyrus lacking Podocalyxin, showing for the first time that Podocalyxin is essential to establish the tempo of development of adult-generated neurons within an active neuronal network. As a result of our study, we can say that Podocalyxin negatively regulates all essential steps of integration of adult-born neurons, serving as a key intrinsic determinant that balances stimulating effects of extrinsic mechanism and maintains the correct development and integration of adult-born neurons in order. Podocalyxin negatively regulates dendritogenesis and spinogenesis of these neurons, while it is a positive regulator of synaptogenesis. Podocalyxin is also expressed in oligodendrocytes precursor cells (OPCs) and in mature oligodendrocytes, and it is essential for the migration of OPCs. The sialic acid present on Podocalyxin is essential in this process. OPCs present a directional migratory response to CXCL12, that is inhibited by the absence of Podocalyxin. To study the function of a protein it is very important to know which proteins interact. The set of proteins that we have described interplaying with Podocalyxin can be subdivided into: cytoskeletal-interacting, synapse formation/maintenance, exocytotic, myelin-associated and intracellular signaling proteins. Very preliminary studies give evidence of impaired nigro-striatal system in Podocalyxin KO mice.

L’àrea de treball de la present tesi doctoral se situa en el context de les malalties genètiques del metabolisme energètic mitocondrial. Són malalties rares i hereditàries que afecten al conjunt de sistemes que fa servir l’organisme per incorporar i transformar els substrats en energia utilitzable per al correcte funcionament cel·lular. El coenzim Q10 és un component lipídic de totes les membranes cel·lulars que realitza un paper essencial a la cadena respiratòria mitocondrial, però també participa en moltes altres funcions cel·lulars, tan dins dels mitocondris com fora. En l’àmbit pediàtric, la deficiència de coenzim Q10 s’associa a estats de malaltia amb expressions fenotípiques heterogènies, i les causes que la expliquen poden ser primàries o bé secundàries (és a dir, per alteració dels gens implicats en la via de síntesi d’aquesta molècula –deficiència primària–, o per alteració d’altres gens no directament relacionats en la via biosintètica del coenzim Q10 –deficiència secundària). Aquesta deficiència bioquímica implica una disfunció del sistema de la fosforilació oxidativa mitocondrial, i normalment es manifesta de forma multiorgànica, alterant en major o menor grau els diferents òrgans, segons els nivells energètics que requereixen els teixits i d’altres factors no massa coneguts. L’objectiu principal d’aquesta tesi ha estat la millora del diagnòstic de pacients amb deficiències de coenzim Q10, a través de l’estudi sistemàtic d’aquest en diverses espècimens biològics i en associació amb dades clíniques, bioquímiques, histoquímiques, enzimàtiques i moleculars. A través de l’estudi i valoració de grans grups de pacients, s’ha pogut intuir la dinàmica d’aquesta molècula en certs tipus de malalties. Hem pogut descriure tres malalties que s’associen a una deficiència de coenzim Q10 de forma secundària, permetent que pacients afectats puguin beneficiar-se de la suplementació oral amb coenzim Q10, la qual ha demostrat millores clíniques de l’estat de pacients afectats amb patologies mitocondrials. També, hem realitzat avenços metodològics i tècnics, a nivell bioquímic i d’anàlisi de dades, que permetran abordar les classificacions actuals dels pacients amb malalties mitocondrials, en les quals és complicat assolir un diagnòstic molecular definitiu degut a la seva immensa complexitat.
Mitochondrial diseases are genetic rare diseases which affect the energetic cellular system to obtain the required energy for basic survival. Coenzyme Q10 is a lipidic antioxidant located in all eukaryotic cellular membranes that is essential for mitochondrial respiratory chain activity, amongst other important roles not strictly related to mitochondrial function. Coenzyme Q10 deficiency is a biochemical trait defined by low coenzyme Q10 levels in tissues, which can manifest in five main classical phenotypes (from isolated nephropathies to fatal infantile multisystemic disease). The ethiology can be primary (when the genetic defect is in a gene affecting the coenzyme Q10 biosynthetic pathway) or secondary (when the altered gene is not directly related to the coenzyme Q10 biosynthesis), and this partially explains the high heterogeneity observed in these patients. The patophysiology is explained because there is a mitochondrial respiratory chain malfunction that affects the oxidative phosphorylation system and unbalances the antioxidant protection, consequently changing normal cellular behaviour. The main objective of this work has been to improve the diagnosis of patients with coenzyme Q10 deficiency, through the systematic analysis of various biological samples in association with clinical, biochemical, histochemical, enzymatic and molecular data. Through the study and evaluation of big cohorts of patients, we could establish that secondary coenzyme Q10 deficiencies are commoner than primary. Furthermore, we have reported an association of three different diseases with secondary coenzyme Q10 deficient states (GLUT-1 deficiency syndrome, pyrivate dehydrogenase deficiency, mucopolysaccharidosis type III), diseases that could benefit from coenzyme Q10 supplementation, which has demonstrated to produce clinical amelioration in mitochondrial patients. Finally, methodological improvements for coenzyme Q10 deficiency diagnosis were done through two different approaches. One is the analysis of coenzyme Q10 in urinary sediment to assess coenzyme Q10 levels of renal system cells, and the other one is the development of a statistical algorithm which shows the potential of coenzyme Q10 as a mitochondrial activity biomarker.

Spatial embedding and inherited metric constraints are a fundamental trait of biological neuronal circuits. However their role in shaping connectivity and dynamics has been often disregarded, with models of neuronal networks paying much more attention to the distribution of connections in the quest for understanding network's behavior. In this thesis we aim at filling this gap by studying the importance of metric features in the complex connectivity- dynamics-noise interplay that shapes spontaneous neuronal activity. This thesis combines experiments in rat dissociated neuronal cultures with theoretical analyses to better comprehend the relevance of spatial embedding. We developed a new theoretical model grounded on Ising Models to assess metric effects in neuronal cultures' behavior, and in the context of percolation approaches. Once metric effects were settled, we illustrated their relevance in shaping spontaneous activity by perturbing the structural connectivity blueprint of neuronal cultures. This was achieved by patterning the substrate where neurons grow, and by using topographical molds that dictated the connectivity of the network. Next, and since the initiation of bursting activity is governed in great manner by a complex amplification mechanism that involves metric correlations and noise, we focused on the metric-driven amplification of spontaneous single-neuron noise to derive an analytical model that predicts the frequency of bursting events in neuronal cultures. We then further investigated in an experimental context the contribution of noise to the observed activity patterns, and by implementing a moderate electrical stimulation protocol that increases the level of activity noise in cultures. Finally, the latter study was completed with experiments regarding the specific role of inhibition in neuronal networks, to provide a wider understanding of the mechanisms that govern the initiation and propagation of activity fronts in cortical cultures.
L'objectiu d'aquesta tesis és investigar els mecanismes que generen l'activitat espontània i estimulada en xarxes neuronals, més concretament en cultius corticals dissociats, i fent un especial èmfasi en l’efecte de les correlacions mètriques. En aquest marc, l’activitat col·lectiva consisteix en episodis esporàdics de dispars quasi sincronitzats entre totes les neurones del cultiu, anomenats “esclats de xarxa”. Tres elements principals en determinen les característiques: connectivitat entre neurones, dinàmica intrínseca neuronal, i soroll (activacions neuronals aleatòries). La investigació s’ha centrat en cinc línies de recerca: l’estudi de correlacions mètriques en cultius neuronals; el desenvolupament d’un model teòric per descriure i predir l’esclat de xarxa; l’anàlisi de la propagació dels fronts d’activitat experimentals sota pertorbacions estructurals de la connectivitat del cultiu; l’estudi de l’efecte de la inhibició en la iniciació i propagació dels esclats ‘in vitro’; i l’estudi de la resposta experimental dels cultius sota una estimulació elèctrica moderada de baixa freqüència. En la primera línia de recerca hem comprovat que les correlacions mètriques dominen el comportament dinàmic del cultiu, fins al punt d’emmascarar la contribució de la distribució del nombre de connexions. En la segona línia hem desenvolupat un model analític que prediu semi- quantitativament la freqüència dels esclats observada experimentalment. La tercera línia s’ha centrat en l’efecte de pertorbacions estructurals en la connectivitat; la dinàmica resultant ha mostrat una gran riquesa en patrons d’activitat, esclats de xarxa a diferents escales, i propagació altament específica de cada cultiu. La quarta línia de recerca ha demostrat que les xarxes sense inhibició disminueixen la seva freqüència d’esclat respecte a les xarxes control, que la velocitat de propagació de l’activitat incrementa lleugerament quan s’ha bloquejat la inhibició, i que els punts on s’inicien ens esclats varien respecte als controls. I, finalment, la cinquena línia de recerca ha constatat que l’aplicació d’un camp elèctric feble augmenta el soroll d’activitat de la xarxa, generant un increment en la freqüència dels esclats de xarxa.

The aim of this work is to investigate the human nociceptive system at the peripheral level. Researchers are still debating how the pain perception arises from this very intricate network. The human perception is the most elusive part of our knowledge since different subsystems are involved. The external information such as noxious stimuli must be processed at the peripheral level and through signal cascades and transduction this signal must reach the brain. At the brain level the information is processed and some decisions are taken, such as the well-known fight-or-flight response. In the introduction, the author describes how the human nociceptive system works and in which way the noxious stimulus is converted into a signal understandable by the brain. Several cortical and subcortical areas are involved in this signal processing and going deeper in this assembly line the information becomes more abstracted. The whole pathway is fundamental for pain perception, however some diseases start at the peripheral level. This in turn makes wrong signals reaching the brain. The brain is then processing information that are not real and the responses do not suit with the needs. Therefore, the peripheral system must be investigated and understood firstly, since some central diseases may have a peripheral component as well. With this purpose in mind the microneurography technique has been used. This technique has got some complexity and a computer-aided system must be implemented. The hardware aims to filter out the noisy signal and perform recording and stimulation of the neural fibers. The software is instead used to make the stimulation and recording as automatic as possible in a way that researchers do not have to deal with a lot of parameters and steps to carry out this powerful but also time consuming technique. Some software are already available in the market however even if they work fine with slow conduction fibers such as C-fibers they cannot cope with faster neurons (e.g. Aδ fibers). The aim of this work is to create a software (i.e. SPiike) able to stimulate and record every type of fibers implementing advanced analysis technique as well. Furthermore, considering that some in vivo experiments have been pursued within the project to check the functionality of the software, more specifically in rats and mice, the comparison between human nociceptors and mouse nociceptors is depicted in this section. In the method section, the experimental approach is described step by step. This is composed by several systems that work together for the stimulation, recording and analysis of the neural fibers. The control and acquisition module is composed by the software and a data acquisition board that trigger the stimulator and record the filtered signal. The stimulation module is composed by a stimulator that can be tuned as wish through dedicated knobs. Then the stimulus is delivered to the animal model (or the human patient) and the signal is recorded though a microelectrode inserted into the sciatic nerve. The amplification module is filtering out the noisy signal and is feeding a audio monitor for helping the researcher during the insertion of the electrode inside the nerve and it provides support during the whole experiment giving insights on fiber discharges. In this section the whole setup is described in details as well as the devices needed for the recording. Furthermore, the software development that is the core of this project is described as well, with all the considerations that must be considered during coding. Indeed, the flow chart must be followed methodically in order to minimize bugs and errors that may arise in the final product. Thus a description of the compiler and the Matlab IDE is given along with system and software requirements for the making of the SPiike software. Eventually the explanation of embedded functionalities and capabilities of SPiike is depicted in the final part of this section. This software is indeed able to stimulate slow conducting fibers as well as faster ones, and enhanced analysis techniques such as supervised machine learning are implemented. In the results section, the graphical user interface of the Spiike software is reveled. It resembles the one of another software already available in the market, with a filtered signal and a raster plot embedded on it. However, this software is more user-friendly and it accounts with icons and drop-down menus that enhance the experience of the users during the use of the tool, making their interactions smooth and intuitive. The SPiike software is subdivide into two different tools, a recording module and a analysis module. The former allows the stimulation and recording of neural fibers with a stimulation frequency up to 1000Hz and some online analysis can be conducted to have insights on fibers type and behavior. The analysis module is instead a more powerful analysis environment that can retrieve the dataset recorded with the other module or with the LabChart software. Advanced analysis techniques are implemented in this module, this is meant to speed up fiber classification and analysis. Conclusion and discussion provide a overview on some results. These will be compared to those obtainable through other software available in the market. In this section, pros and cons of the new implemented software, SPiike, will be described as well.

Orientador: Jose Jorge Facure
Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas
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Resumo: O uso do computador vem crescendo assustadoramente, tornando-se uma ferramenta indispensável, quase que em todas as áreas, a educação não foge a esta regra, estamos ainda longe de não mais precisarmos de professores I mas em futuro próximo os professores serão mais orientadores e monitores , do que a função que exercem atualmente. Este trabalho visa mostrar a evolução deste crescimento da informática dentro do ensino médico, enfocar qual a base pedagógica , como eram os primeiros programas. Nesta dissertação os resultados mostram que o uso do computador como método didático, desperta mais a atenção do aluno, em virtude do uso da nova tecnologia (100% dos alunos responderam que a atenção prestada no seminário foi total), e aumenta o interesse dele pelo seminário (89% responderam que o seminário foi muito interessante). O ensino através do computador traz muitas vantagens, como por exemplo; dá acesso a uma imensidão de informações em um tempo muito mais curto; dá ao estudante a oportunidade de aprender segundo o seu ritmo; existe uma maior uniformidade e veracidade das informações por ex.(o aluno de medicina de São Paulo teria a mesma aula que o do Acre) . O estudante teria uma maior vivencia com os problemas que enfrentaria na sua vida prática com questões tipo caso-problema em que ele teria que tomar as decisões de conduta; e num futuro próximo teríamos cirurgias com realidade virtual onde treinaríamos os residentes de cirurgia antes que eles começassem a operar realmente. O uso do computador como instrumento didático evolui muito, desde as máquinas de ensinar proposta por SKINER ,que eram muito primitivas e ofereciam poucos recursos até os micros atuais com grande capacidade de memória ,e de alta velocidade, oferecendo recursos de áudio , imagens e até vídeos. As perspectivas do uso do computador no ensino médico são ilimitadas, e progressivamente vai se tornar peça fundamental no ensino médico .0 computador não veio para substituir o professor. O computador veio para ser mais uma ferramenta importante nas mãos do professor, ele apenas tem que saber utiliza-Ia da melhor forma possível.
Abstract: The use of computers has being increasing at very large scales, with the outcome that they have now become a vital aid in almost all areas. Education is no exception to this fact. Teachers are far from being banned, but in a near future they are bound to become more of over ceers and monitors possible no longer exerting the same range of duties of nowadays. This work is aimed at presenting the evolution of computing in medical teaching, giving a comprehensive approach to its didactic basis and how the earliest programs were. The results presented in this dissertation show that the use of computers as a didactic method strikes more of the student's attention, owing to the use of the new technology (100% of the students questioned answered that the attention paid to the seminar was total) and increase the interest for the seminar itself (89% answered that the seminar was very interesting). Teaching through computers introduces marry advantages, for instance, it provides access to a large variety of information in a much shorter time and allows students to learn according to thair own pace. Moreover, it better standardizes information and provides a possibility of confirmation (ex: A student of medicine from São Paulo would have the same teaching as the one from Acre). Students would acquire a better understanding of life experience with the real live situations they would encounter, approaching case-problems issues in which they would have to take decisions of conduct. In a near future we would have virtual reality surgeries where surgery residents could be trained before being able to actually start operating. The use of computers as a didactic support to teaching has developed a lot, from the teaching machines propose by SKINNER, which were far toa simple and offered few resources to the high speed and memory capacity micro-computers of nowadays that contain audio, image and even video resources. The prospects for the use of computers in this area are unlimited and they will gradually become the cornerstone of medical teaching. The computer has not com to substitute the teacher. Yet, it has come to act as one more important tool in the teacher's hand, who has simply got to learn how to use it in the best possible way.
Mestrado
Mestre em Neurociencias

Orientador: Donizete Cesar Honorato
Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas
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Resumo: O presente estudo tem por objetivo analisar uma população de pacientes, vítimas de acidentes de trabalho com traumatismo craniencefálico, registrados no HC - Unicamp. Tendo em vista que grande parte destes pacientes, em fase produtiva, não retoma à sua vida social, econômica e familiar com a mesma qualidade de vida anterior, a caracterização deste triste quadro serve-nos para estabelecer uma proposta de assistência a estes pacientes, visando sua qualidade de vida e saúde. Nossa proposta baseia-se no conceito de que essa assistência, enfocada no paciente e em suas reais necessidades, deve ser mais abrangente. Nesse sentido, deve haver uma integração que envolva a equipe multiprofissional de saúde, centrada, sobretudo, nos serviços. do pessoal de enfermagem, a família e os órgãos governamentais, oferecendo o respaldo financeiro que é indispensável. Só a participação e o suporte de cada um destes elementos, de forma integrada, podem contribuir para a luta que o paciente trava na busca por sua reabilitação
Abstract: The present study has -the objective of analysing a group of patients victims of work accident with brain trauma, registered at HC - Unicamp. Having in mind that a great amount of these patients, in productive phase, do not return to their social, economical and familiar life, with the same former quality of life, the characteristics of this sad picture make us establish an assistance proposal to these patients, looking for their quality life and health. Our proposal is based on the concept that this assistance, focused on tpe patient and his real necessities, must be more extended. In this sense, there must be an integtration that involves the health multiprofessional team, centered, above all, on the nursing staff services, financial the family and the government bodies, offering the backing that is extremely necessary. Only the participation and support of each of these elements, in an integrated way, can contribute on the struggle that holds out the patient in his search ofhis recover
Doutorado
Ciencias Biomedicas
Doutor em Ciências Médicas

O Transtorno Bipolar (TB) é uma patologia prevalente, grave, crônica, e que apresenta um curso longitudinal muito pior que se imaginava décadas atrás. Além da alternância entre períodos de depressão, mania e eutimia, a recorrência e a progressão do TB conferem gravidade e frequência crescentes aos episódios. A neuroprogressão foi um termo cunhado para definir a aceleração do processo de doença e seus fatores subjacentes, como alterações de biomarcadores periféricos, funções cognitivas, neuroimagem e funcionalidade, que emergem em graus variáveis dependendo da fase de evolução. Todas estas evidências justificaram a classificação do TB em estágios clínicos teóricos distintos, que ainda carecem de validação empírica. Um dos mais recentes deles propõe 1 estágio latente e 4 estágios clínicos distintos (Kapczinski et al, 2009) (1). O presente trabalho avaliou, portanto, as diferentes estratégias farmacológicas para a manutenção da eutimia em uma amostra de pacientes com TB em diferentes estágios (artigo 1), e a associação do atraso no tratamento do primeiro episódio com fatores de pior prognóstico e com os estágios do TB (artigo 2). Em conjunto, os resultados mostraram que a abordagem farmacológica instituída pelo psiquiatra clínico, necessária para manter o paciente em eutimia, é diferente conforme a classificação em estágios. O número de fármacos prescrito também está relacionado ao declínio da funcionalidade. Além disso, o atraso no início do tratamento do TB é diretamente proporcional ao estágio de evolução da doença, e está relacionado a fatores de pior prognóstico, como o número de episódios. Os achados fornecem evidência para modificar certas intervenções no TB: a primeira, que diretrizes de tratamento poderiam considerar os estágios, visando um tratamento mais personalizado para guiar a eficácia do tratamento; e por último, que o esforço em diagnosticar e tratar o TB com precisão e rapidez pode ser uma das medidas para frear a neuroprogressão.
Bipolar Disorder (TB) is a severe, prevalent, and chronic disease, that exhibits worse longitudinal course than previously thought. Beyond the switching phases of depression, mania and euthymia, recurrence and progression of TB increases severity and frequency of episodes. The neuroprogression was a term created to define the acceleration of the disease and its underlying factors, such as changes in peripheral biomarkers, in cognitive performance, in neuroimaging and functioning, that emerge in different degrees depending on the stage of evolution. All those evidences justify the classification of TB in different clinical stages, which still lack empirical validation. One of most recently proposed staging model describes 1 latent stage and 4 clinical stages (Kapczinski et al, 2009) (1). The present study therefore evaluated the different pharmacological strategies for the maintenance of euthymia in a sample of TB patients at different stages (Article 1), and the association of first-episode tretament delay with poor prognosis and BD stages (Article 2). Together, the results showed that maintenance pharmacological treatment in a naturallistic setting is different according to the staging classification. The number of drugs prescribed is also associated to functioning. Moreover, the treatment delay is positively correlated to the stage of the disease, and is related to poor outcomes (i.e. number of episodes). The findings provide evidence to modify certain interventions in TB: first, that treatment guidelines might consider staging to provide tailored approaches and to guide treatment efficacy; and secondly, that the effort to accurately diagnose and treat TB can be one of the measures to restrain neuroprogression.

This thesis will describe two functional Magnetic Resonance Imaging (fMRI) experiments and one Voxel-Based Morphometry (VBM) study, each investigating how the human brain identifies objects and their associated properties. In particular, we used three different categories of objects – living (animals), nonliving (tools and nontools) and faces (famous and non-famous) – to examine the type of knowledge attribute in question: one perceptual (movement) and two semantic attributes (typical object location and biographic knowledge). We know from neuropsychological literature that the most anterior portions of the temporal cortices critically support human conceptual knowledge. Unfortunately, the Anterior Temporal Lobe (ATL) is a challenging region for fMRI due to susceptibility artifacts, especially at high fields. For these reasons we established an optimized fMRI protocol (described in the second Chapter) by adjusting key acquisition parameters like phase-encoding gradient polarity, slice thickness, echo time, and slice angle. The protocol gave reliable Blood-Oxygen-Level Dependence (BOLD) signal sensitivity in the ATL. Clinical data describe patients with specific semantic impairments at the level of category (living, nonliving) as well as disproportionate deficits for a modality or type of knowledge (e.g., visual/perceptual knowledge or manipulation knowledge). Functional neuroimaging studies on semantic organization with normal subjects found an “action network” specific for tools rather than living items. In the first experiment (Chapter 3) we devised an fMRI paradigm to investigate the processing of movement (action) and place (encyclopedic) features, and their influence on category-specific activations. Within the “movement network” statistical analyses did not show any significant interaction between categories. These findings suggest that the visuomotor “action network” is not specific for tools because it is also activated when the action related knowledge is elicited for other categories, such as animals. The second and the third experiment (Chapter 4) focus on the processing of faces. Neuropsychological literature attributes semantic and lexical retrieval deficits in patients to ATL lesions. In Part I of Chapter 4, we report data from a VBM study on patients with known lesions in the temporal lobe. Unfortunately, as far as we know, data on patients and functional neuroimaging in healthy individuals has not clarified the differential role of this area in the two mental operations because semantic and lexical processes usually occur simultaneously and automatically. In Part II, we devised an event-related fMRI activation paradigm that allowed us to study the identification (i.e., association of semantic biographical information) of celebrities, with and without the ability to retrieve the proper name. While semantic retrieval reliably activated the ATL, only more posterior areas in the left temporal and temporal-parietal junction were significantly modulated by covert lexical retrieval. These results support findings from patients with ATL lesions and suggest that their anomia is due to semantic rather than lexical retrieval impairment.

The highly complex nervous system is built upon an intricate network of neurons. In order to make a functional network, the establishment of precise connections is crucial. Neuronal networks are established early during development when neurons send out axons that navigate through complex environments to connect to their target. Chemotropic attractant or repellent cues, cell adhesion molecules, morphogens and a wide range of factors secreted or expressed by guidepost cells enable axon guidance. The leading tip of the axon, the GC is important to sense the environment and integrate extracellular signals to navigate precisely. The axonal GC has a large repertoire of mRNAs that are dynamic in nature. Local regulation of transcripts in navigating axons is suspected to ensure precise pathfinding. However, mechanisms involving regulation of expression of these transcripts within GCs are largely unknown. This thesis investigates whether microRNAs, one of the quintessential posttranscriptional regulators, can regulate axon guidance by fine-tuning mRNA expression within subcellular compartments. To explore microRNA roles in axon guidance, Xenopus laevis visual system was used as a model. Profiling axons of retinal ganglion cells revealed the presence of miRNAs within axons. The most abundant axonal miRNAs, the miR-181 family and miR-182, exhibit distinct roles in regulating axon guidance in vivo. Loss of function analyses suggests that both miRNA families are required for accurate axonal targeting but involve different mechanisms. Thus, specific axonal microRNAs locally regulate mRNAs contributing to error-free pathfinding.

Functional Magnetic Resonance Imaging (fMRI) has consistently highlighted aberrant functional connectivity across brain regions of autism spectrum disorder (ASD) patients. However, the manifestation and neural substrates of these alterations are highly heterogeneous and often conflicting. Moreover, their neurobiological under- pinnings and etiopathological significance remain largely unknown. A deeper understanding of the complex pathophysiological cascade leading to impaired connectivity in ASD can greatly benefit from the use of model organisms where individual pathophysiological or phenotypic components of ASD can be recreated and investigated via approaches that are either off limits or confounded by clinical heterogeneity. In this work, we first describe the intrinsic organization of the mouse brain at the macroscale as seen through resting-state fMRI (rsfMRI). The analysis of a large rsfMRI dataset revealed the presence of six distinct functional modules related to known brainwide functional partitions, including a homologue of the human default-mode network (DMN). Consistent with human studies, interconnected functional hubs were identified in several sub-regions of the DMN, in the thalamus, and in small foci within integrative cortical structures such as the insular and temporal association cortices. We then study the effects of mutations in contactin associated protein-like 2 (Cntnap2), a neurexin-related cell-adhesion protein, on functional connectivity. Homozygous mutations in this gene are strongly linked to autism and epilepsy in humans, and using rsfMRI, we showed that homozygous mice lacking Cntnap2 exhibit aberrant functional connectivity in prefrontal and midline functional hubs, an effect that was associated with reduced social investigation, a core “autism trait” in mice. Notably, viral tracing revealed reduced frequency of prefrontal-projecting neural clusters in the cingulate cortex of Cntnap2−/− mutants, suggesting a possible contribution of defective mesoscale axonal wiring to the observed functional impairments. Macroscale cortico-cortical white-matter organization appeared to be otherwise preserved in these animals. These findings revealed a key contribution of ASD-associated gene CNTNAP2 in modulating macroscale functional connectivity, and suggest that homozygous loss-of-function mutations in this gene may predispose to neurodevelopmental disorders and autism through a selective dysregulation of connectivity in integrative prefrontal areas. Finally, we discuss the role mouse models could play in generating and testing mechanistic hypotheses about the elusive origin and significance of connectional aberrations observed in autism and recent progress towards this goal.

Impairment due to narcolepsy strongly limits job performance, but there are no standard criteria to assess disability in people with narcolepsy and a scale of disease severity is still lacking. We explored: 1. the interobserver reliability among Italian Medical Commissions making disability and handicap benefit decisions for people with narcolepsy, searching for correlations between the recognized disability degree and patients’ features; 2. the willingness to report patients to the driving licence authority; 3. possible sources of variance in judgement. Fifteen narcoleptic patients were examined by four Medical Commissions in simulated sessions. Raw agreement and interobserver reliability among Commissions were calculated for disability and handicap benefit decisions and for driving licence decisions. Levels of judgement differed on percentage of disability (p<0.001), severity of handicap (p=0.0007) and the need to inform the driving licence authority (p=0.032). Interobserver reliability ranged from Kappa = - 0.10 to Kappa = 0.35 for disability benefit decision and from Kappa = - 0.26 to Kappa = 0.36 for handicap benefit decision. The raw agreement on driving licence decision ranged from 73% to 100% (Kappa not calculable). Spearman’s correlation between percentages of disability and patients’ features showed correlations with age, daytime naps, sleepiness, cataplexy and quality of life. This first interobserver reliability study on social benefit decisions for narcolepsy shows the difficulty of reaching an agreement in this field, mainly due to variance in interpretation of the assessment criteria. The minimum set of indicators of disease severity correlating with patients’ self assessments encourages a disability classification of narcolepsy.

Impairment due to narcolepsy strongly limits job performance, but there are no standard criteria to assess disability in people with narcolepsy and a scale of disease severity is still lacking. We explored: 1. the interobserver reliability among Italian Medical Commissions making disability and handicap benefit decisions for people with narcolepsy, searching for correlations between the recognized disability degree and patients’ features; 2. the willingness to report patients to the driving licence authority; 3. possible sources of variance in judgement. Fifteen narcoleptic patients were examined by four Medical Commissions in simulated sessions. Raw agreement and interobserver reliability among Commissions were calculated for disability and handicap benefit decisions and for driving licence decisions. Levels of judgement differed on percentage of disability (p<0.001), severity of handicap (p=0.0007) and the need to inform the driving licence authority (p=0.032). Interobserver reliability ranged from Kappa = - 0.10 to Kappa = 0.35 for disability benefit decision and from Kappa = - 0.26 to Kappa = 0.36 for handicap benefit decision. The raw agreement on driving licence decision ranged from 73% to 100% (Kappa not calculable). Spearman’s correlation between percentages of disability and patients’ features showed correlations with age, daytime naps, sleepiness, cataplexy and quality of life. This first interobserver reliability study on social benefit decisions for narcolepsy shows the difficulty of reaching an agreement in this field, mainly due to variance in interpretation of the assessment criteria. The minimum set of indicators of disease severity correlating with patients’ self assessments encourages a disability classification of narcolepsy.

Obstructive sleep apnoea/hypopnoea syndrome (OSAHS) is the periodic reduction or cessation of airflow during sleep. The syndrome is associated whit loud snoring, disrupted sleep and observed apnoeas. Surgery aims to alleviate symptoms of daytime sleepiness, improve quality of life and reduce the signs of sleep apnoea recordered by polysomnography. Surgical intervention for snoring and OSAHS includes several procedures, each designed to increase the patency of the upper airway. Procedures addressing nasal obstruction include septoplasty, turbinectomy, and radiofrequency ablation (RF) of the turbinates. Surgical procedures to reduce soft palate redundancy include uvulopalatopharyngoplasty with or without tonsillectomy, uvulopalatal flap, laser-assisted uvulopalatoplasty, and RF of the soft palate. More significant, however, particularly in cases of severe OSA, is hypopharyngeal or retrolingual obstruction related to an enlarged tongue, or more commonly due to maxillomandibular deficiency. Surgeries in these cases are aimed at reducing the bulk of the tongue base or providing more space for the tongue in the oropharynx so as to limit posterior collapse during sleep. These procedures include tongue-base suspension, genioglossal advancement, hyoid suspension, lingualplasty, and maxillomandibular advancement. We reviewed 269 patients undergoing to osas surgery at the ENT Department of Forlì Hospital in the last decade. Surgery was considered a success if the postoperative apnea/hypopnea index (AHI) was less than 20/h. According to the results, we have developed surgical decisional algorithms with the aims to optimize the success of these procedures by identifying proper candidates for surgery and the most appropriate surgical techniques. Although not without risks and not as predictable as positive airway pressure therapy, surgery remains an important treatment option for patients with obstructive sleep apnea (OSA), particularly for those who have failed or cannot tolerate positive airway pressure therapy. Successful surgery depends on proper patient selection, proper procedure selection, and experience of the surgeon. The intended purpose of medical algorithms is to improve and standardize decisions made in the delivery of medical care, assist in standardizing selection and application of treatment regimens, to reduce potential introduction of errors. Nasal Continuous Positive Airway Pressure (nCPAP) is the recommended therapy for patients with moderate to severe OSAS. Unfortunately this treatment is not accepted by some patient, appears to be poorly tolerated in a not neglible number of subjects, and the compliance may be critical, especially in the long term if correctly evaluated with interview as well with CPAP smart cards analysis. Among the alternative options in Literature, surgery is a long time honoured solution. However until now no clear scientific evidence exists that surgery can be considered a really effective option in OSAHS management. We have design a randomized prospective study comparing MMA and a ventilatory device (Autotitrating Positive Airways Pressure – APAP) in order to understand the real effectiveness of surgery in the management of moderate to severe OSAS. Fifty consecutive previously full informed patients suffering from severe OSAHS were enrolled and randomised into a conservative (APAP) or surgical (MMA) arm. Demographic, biometric, PSG and ESS profiles of the two group were statistically not significantly different. One year after surgery or continuous APAP treatment both groups showed a remarkable improvement of mean AHI and ESS; the degree of improvement was not statistically different. Provided the relatively small sample of studied subjects and the relatively short time of follow up, MMA proved to be in our adult and severe OSAHS patients group a valuable alternative therapeutical tool with a success rate not inferior to APAP.

Obstructive sleep apnoea/hypopnoea syndrome (OSAHS) is the periodic reduction or cessation of airflow during sleep. The syndrome is associated whit loud snoring, disrupted sleep and observed apnoeas. Surgery aims to alleviate symptoms of daytime sleepiness, improve quality of life and reduce the signs of sleep apnoea recordered by polysomnography. Surgical intervention for snoring and OSAHS includes several procedures, each designed to increase the patency of the upper airway. Procedures addressing nasal obstruction include septoplasty, turbinectomy, and radiofrequency ablation (RF) of the turbinates. Surgical procedures to reduce soft palate redundancy include uvulopalatopharyngoplasty with or without tonsillectomy, uvulopalatal flap, laser-assisted uvulopalatoplasty, and RF of the soft palate. More significant, however, particularly in cases of severe OSA, is hypopharyngeal or retrolingual obstruction related to an enlarged tongue, or more commonly due to maxillomandibular deficiency. Surgeries in these cases are aimed at reducing the bulk of the tongue base or providing more space for the tongue in the oropharynx so as to limit posterior collapse during sleep. These procedures include tongue-base suspension, genioglossal advancement, hyoid suspension, lingualplasty, and maxillomandibular advancement. We reviewed 269 patients undergoing to osas surgery at the ENT Department of Forlì Hospital in the last decade. Surgery was considered a success if the postoperative apnea/hypopnea index (AHI) was less than 20/h. According to the results, we have developed surgical decisional algorithms with the aims to optimize the success of these procedures by identifying proper candidates for surgery and the most appropriate surgical techniques. Although not without risks and not as predictable as positive airway pressure therapy, surgery remains an important treatment option for patients with obstructive sleep apnea (OSA), particularly for those who have failed or cannot tolerate positive airway pressure therapy. Successful surgery depends on proper patient selection, proper procedure selection, and experience of the surgeon. The intended purpose of medical algorithms is to improve and standardize decisions made in the delivery of medical care, assist in standardizing selection and application of treatment regimens, to reduce potential introduction of errors. Nasal Continuous Positive Airway Pressure (nCPAP) is the recommended therapy for patients with moderate to severe OSAS. Unfortunately this treatment is not accepted by some patient, appears to be poorly tolerated in a not neglible number of subjects, and the compliance may be critical, especially in the long term if correctly evaluated with interview as well with CPAP smart cards analysis. Among the alternative options in Literature, surgery is a long time honoured solution. However until now no clear scientific evidence exists that surgery can be considered a really effective option in OSAHS management. We have design a randomized prospective study comparing MMA and a ventilatory device (Autotitrating Positive Airways Pressure – APAP) in order to understand the real effectiveness of surgery in the management of moderate to severe OSAS. Fifty consecutive previously full informed patients suffering from severe OSAHS were enrolled and randomised into a conservative (APAP) or surgical (MMA) arm. Demographic, biometric, PSG and ESS profiles of the two group were statistically not significantly different. One year after surgery or continuous APAP treatment both groups showed a remarkable improvement of mean AHI and ESS; the degree of improvement was not statistically different. Provided the relatively small sample of studied subjects and the relatively short time of follow up, MMA proved to be in our adult and severe OSAHS patients group a valuable alternative therapeutical tool with a success rate not inferior to APAP.

STUDY OBJECTIVE: Cyclic Alternating Pattern (CAP) is a fluctuation of the arousal level during NREM sleep and consists of the alternation between two phases: phase A (divided into three subtypes A1, A2, and A3) and phase B. A1 is thought to be generated by the frontal cortex and is characterized by the presence of K complexes or delta bursts; additionally, CAP A1 seems to have a role in the involvement of sleep slow wave activity in cognitive processing. Our hypothesis was that an overall CAP rate would have a negative influence on cognitive performance due to excessive fluctuation of the arousal level during NREM sleep. However, we also predicted that CAP A1 would be positively correlated with cognitive functions, especially those related to frontal lobe functioning. For this reason, the objective of our study was to correlate objective sleep parameters with cognitive behavioral measures in normal healthy adults. METHODS: 8 subjects (4 males; 4 females; mean age 27.75 years, range 2334) were recruited for this study. Two nocturnal polysomnography (night 2 and 3 = N2 and N3) were carried out after a night of adaptation. A series of neuropsychological tests were performed by the subjects in the morning and afternoon of the second day (D2am; D2pm) and in the morning of the third day (D3am). Raw scores from the neuropsychological tests were used as dependent variables in the statistical analysis of the results. RESULTS: We computed a series of partial correlations between sleep microstructure parameters (CAP, A1, A2 and A3 rate) and a number of indices of cognitive functioning. CAP rate was positively correlated with visuospatial working memory (Corsi block test), Trial Making Test Part A (planning and motor sequencing) and the retention of words from the Hopkins Verbal Learning Test (HVLT). Conversely, CAP was negatively correlated with visuospatial fluency (Ruff Figure Fluency Test). CAP A1 were correlated with many of the tests of neuropsychological functioning, such as verbal fluency (as measured by the COWAT), working memory (as measured by the Digit Span – Backward test), and both delay recall and retention of the words from the HVLT. The same parameters were found to be negatively correlated with CAP A2 subtypes. CAP 3 were negatively correlated with the Trial Making Test Parts A and B. DISCUSSION: To our knowledge this is the first study indicating a role of CAP A1 and A2 on behavioral cognitive performance of healthy adults. The results suggest that high rate of CAP A1 might be related to an improvement whereas high rate of CAP A2 to a decline of cognitive functions. Further studies need to be done to better determine the role of the overall CAP rate and CAP A3 on cognitive behavioral performances.

STUDY OBJECTIVE: Cyclic Alternating Pattern (CAP) is a fluctuation of the arousal level during NREM sleep and consists of the alternation between two phases: phase A (divided into three subtypes A1, A2, and A3) and phase B. A1 is thought to be generated by the frontal cortex and is characterized by the presence of K complexes or delta bursts; additionally, CAP A1 seems to have a role in the involvement of sleep slow wave activity in cognitive processing. Our hypothesis was that an overall CAP rate would have a negative influence on cognitive performance due to excessive fluctuation of the arousal level during NREM sleep. However, we also predicted that CAP A1 would be positively correlated with cognitive functions, especially those related to frontal lobe functioning. For this reason, the objective of our study was to correlate objective sleep parameters with cognitive behavioral measures in normal healthy adults. METHODS: 8 subjects (4 males; 4 females; mean age 27.75 years, range 2334) were recruited for this study. Two nocturnal polysomnography (night 2 and 3 = N2 and N3) were carried out after a night of adaptation. A series of neuropsychological tests were performed by the subjects in the morning and afternoon of the second day (D2am; D2pm) and in the morning of the third day (D3am). Raw scores from the neuropsychological tests were used as dependent variables in the statistical analysis of the results. RESULTS: We computed a series of partial correlations between sleep microstructure parameters (CAP, A1, A2 and A3 rate) and a number of indices of cognitive functioning. CAP rate was positively correlated with visuospatial working memory (Corsi block test), Trial Making Test Part A (planning and motor sequencing) and the retention of words from the Hopkins Verbal Learning Test (HVLT). Conversely, CAP was negatively correlated with visuospatial fluency (Ruff Figure Fluency Test). CAP A1 were correlated with many of the tests of neuropsychological functioning, such as verbal fluency (as measured by the COWAT), working memory (as measured by the Digit Span – Backward test), and both delay recall and retention of the words from the HVLT. The same parameters were found to be negatively correlated with CAP A2 subtypes. CAP 3 were negatively correlated with the Trial Making Test Parts A and B. DISCUSSION: To our knowledge this is the first study indicating a role of CAP A1 and A2 on behavioral cognitive performance of healthy adults. The results suggest that high rate of CAP A1 might be related to an improvement whereas high rate of CAP A2 to a decline of cognitive functions. Further studies need to be done to better determine the role of the overall CAP rate and CAP A3 on cognitive behavioral performances.

Objective: To study circadian rhythms (sleep-wake, body core temperature and melatonin circadian rhythms) in patients in vegetative state (VS) in basal condition and after nocturnal blue light exposure. Methods: Eight patients in VS underwent two experimental sessions of 48 consecutive hours polysomnography with body core temperature (BCT) measurement separated by a 1-week interval. For a week between the two experimental sessions, patients underwent nocturnal blue light exposure (470 nm; 58 μW/cm2 for 4 hours from 11.30 p.m. to 3.30 a.m.). Brain MRI, Level of Cognitive Functioning Scale (LCF) and Disability Rating Scale (DRS) were assessed just before polysomnography. Results: In all patients LCF and DRS confirmed vegetative state. All patients showed a sleep-wake cycle. All patients showed spindle or spindle-like activities. REM sleep was detected in only 7 patients. Patients displayed a greater fragmentation of nocturnal sleep due to frequent awakenings. Mean nocturnal sleep efficiency was significantly reduced (40±22 vs. 74±17) in VS patients respect to controls. A significantly increasing of phase 1 and a significantly reduction of phase 2 and phase 3 were observed too. A modification of diurnal sleep total time and of diurnal duration of REM sleep were found after 1-week nocturnal blue light exposure. All patients displayed a normal BCT 24-h rhythm in basal condition and after nocturnal blue light exposure. A reduction of mean nocturnal melatonin levels in basal condition were observed in VS patients. Melatonin suppression after blue light exposure was observed in only 2 patients in VS. Conclusions: We found disorganized sleep-wake cycle and a normal BCT rhythm in our patients in VS. A reduction of mean nocturnal melatonin levels in basal condition were observed too.

Objective: To study circadian rhythms (sleep-wake, body core temperature and melatonin circadian rhythms) in patients in vegetative state (VS) in basal condition and after nocturnal blue light exposure. Methods: Eight patients in VS underwent two experimental sessions of 48 consecutive hours polysomnography with body core temperature (BCT) measurement separated by a 1-week interval. For a week between the two experimental sessions, patients underwent nocturnal blue light exposure (470 nm; 58 μW/cm2 for 4 hours from 11.30 p.m. to 3.30 a.m.). Brain MRI, Level of Cognitive Functioning Scale (LCF) and Disability Rating Scale (DRS) were assessed just before polysomnography. Results: In all patients LCF and DRS confirmed vegetative state. All patients showed a sleep-wake cycle. All patients showed spindle or spindle-like activities. REM sleep was detected in only 7 patients. Patients displayed a greater fragmentation of nocturnal sleep due to frequent awakenings. Mean nocturnal sleep efficiency was significantly reduced (40±22 vs. 74±17) in VS patients respect to controls. A significantly increasing of phase 1 and a significantly reduction of phase 2 and phase 3 were observed too. A modification of diurnal sleep total time and of diurnal duration of REM sleep were found after 1-week nocturnal blue light exposure. All patients displayed a normal BCT 24-h rhythm in basal condition and after nocturnal blue light exposure. A reduction of mean nocturnal melatonin levels in basal condition were observed in VS patients. Melatonin suppression after blue light exposure was observed in only 2 patients in VS. Conclusions: We found disorganized sleep-wake cycle and a normal BCT rhythm in our patients in VS. A reduction of mean nocturnal melatonin levels in basal condition were observed too.

Si tratta di uno studio osservazionale analitico di coorte prospettico volto a rilevare disfunzioni neuropsicologiche nei pazienti affetti da epilessia frontale notturna, attraverso una batteria di test che esplora i seguenti domini cognitivi: intelligenza generale, memoria, linguaggio, funzioni esecutive, attenzione, vigilanza, tempi di reazione, percezione della qualità della vita ed eventuale presenza di sintomi psichiatrici. Lo studio ha un follow up medio di 20 anni e riporta, per la prima volta in letteratura, l’evoluzione clinica dei soggetti che hanno avuto un esordio dell’epilessia in età evolutiva. Fino ad ora, l’epilessia frontale notturna è stata associata a disfunzioni cognitive nei soli casi di famiglie affette e nelle quali è stato possibile rilevare il difetto genetico. Questo studio ha rilevato la prevalenza di disturbi cognitivi e psichici in un campione di 24 soggetti affetti, mediante la somministrazione di una batteria di test specifica. I risultati sono stati analizzati con il programma statistico SPSS. Tutti i soggetti presentano abilità cognitive inferiori alla media in uno o più test ma il quoziente intellettivo risulta normale nei tre quarti del campione. Il ritardo mentale è più frequente e più grave nei soggetti idiopatici rispetto a quelli con alterazioni morfologiche frontali rilevate alla risonanza magnetica. Sono risultati più frequenti i disturbi della memoria, soprattutto quella a lungo termine e del linguaggio rispetto a quelli di tipo disesecutivo. Tutti i soggetti, che non hanno ottenuto un controllo delle crisi, manifestano una percezione della qualità della vita inferiore alla media. E’ stata valutata l’influenza delle variabili cliniche (età di esordio dell’epilessia, frequenza e semeiologia delle crisi, durata della malattia e terapia antiepilettica), le anomalie elettroencefalografiche e le anomalie rilevate alla risonanza magnetica. Le variabili che sono in rapporto con un maggiore numero di disfunzioni neuropsicologiche sono: l’elevata frequenza di crisi all’esordio, l’associazione con crisi in veglia, la presenza di crisi parziali secondariamente generalizzate e l’assunzione di una politerapia. I disturbi psichici prevalgono nei soggetti con anomalie elettroencefalografiche frontali sinistre. I dati neuropsicologici suggeriscono una disfunzione cognitiva prevalentemente fronto-temporale e, assieme ai dati clinici ed elettroencefalografici, sembrano confermare l’origine mesiale e orbitale frontale delle anomalie epilettiche nell’epilessia frontale notturna.

Si tratta di uno studio osservazionale analitico di coorte prospettico volto a rilevare disfunzioni neuropsicologiche nei pazienti affetti da epilessia frontale notturna, attraverso una batteria di test che esplora i seguenti domini cognitivi: intelligenza generale, memoria, linguaggio, funzioni esecutive, attenzione, vigilanza, tempi di reazione, percezione della qualità della vita ed eventuale presenza di sintomi psichiatrici. Lo studio ha un follow up medio di 20 anni e riporta, per la prima volta in letteratura, l’evoluzione clinica dei soggetti che hanno avuto un esordio dell’epilessia in età evolutiva. Fino ad ora, l’epilessia frontale notturna è stata associata a disfunzioni cognitive nei soli casi di famiglie affette e nelle quali è stato possibile rilevare il difetto genetico. Questo studio ha rilevato la prevalenza di disturbi cognitivi e psichici in un campione di 24 soggetti affetti, mediante la somministrazione di una batteria di test specifica. I risultati sono stati analizzati con il programma statistico SPSS. Tutti i soggetti presentano abilità cognitive inferiori alla media in uno o più test ma il quoziente intellettivo risulta normale nei tre quarti del campione. Il ritardo mentale è più frequente e più grave nei soggetti idiopatici rispetto a quelli con alterazioni morfologiche frontali rilevate alla risonanza magnetica. Sono risultati più frequenti i disturbi della memoria, soprattutto quella a lungo termine e del linguaggio rispetto a quelli di tipo disesecutivo. Tutti i soggetti, che non hanno ottenuto un controllo delle crisi, manifestano una percezione della qualità della vita inferiore alla media. E’ stata valutata l’influenza delle variabili cliniche (età di esordio dell’epilessia, frequenza e semeiologia delle crisi, durata della malattia e terapia antiepilettica), le anomalie elettroencefalografiche e le anomalie rilevate alla risonanza magnetica. Le variabili che sono in rapporto con un maggiore numero di disfunzioni neuropsicologiche sono: l’elevata frequenza di crisi all’esordio, l’associazione con crisi in veglia, la presenza di crisi parziali secondariamente generalizzate e l’assunzione di una politerapia. I disturbi psichici prevalgono nei soggetti con anomalie elettroencefalografiche frontali sinistre. I dati neuropsicologici suggeriscono una disfunzione cognitiva prevalentemente fronto-temporale e, assieme ai dati clinici ed elettroencefalografici, sembrano confermare l’origine mesiale e orbitale frontale delle anomalie epilettiche nell’epilessia frontale notturna.