Rozprawy doktorskie na temat „Afferent pathways”

The current knowledge of the visual pathways in primates includes the patterns of projection from the retina through the dorsal lateral geniculate nucleus (dLGN) to the striate cortex (V1) and the extra-striate projections towards the dorsal and ventral streams. Cells with short wavelength sensitive cone (S-cone) inputs in the dLGN have been studied extensively in New World marmosets but not in Old World macaques. This thesis presents results from studies in the macaque monkey which are more relevant to humans since humans are closer in evolution to Old World than New World monkeys.<br>The spatial, temporal, chromatic and orientation preferences of neurons in the dLGN of the macaque were investigated by electrophysiological methods. The physiological findings of cells with S-cone inputs were compared to cells with opponent inputs from the long and medium wavelength sensitive cones (L-cones & M-cones, respectively). The cells receiving S-cone inputs (blue-yellow or B-Y cells) preferred lower spatial frequencies than the cells with opponent L-cone and M-cone inputs (red-green or R-G cells). Orthodromic latencies from optic chiasm stimulation were measured where possible to distinguish differences in conduction velocity between the cell groups. Although the B-Y cells usually had longer latencies than R-G cells, there wasconsiderable overlap between the cell groups.<br>The recorded cells were localised through histological reconstruction of dLGN sections stained for Nissl substance. The distribution of B-Y cells within the dLGN was compared to the distribution of R-G cells. The majority of B-Y cells were located within the intercalated koniocellular layers as well as the koniocellular bridges (extensions of the koniocellular layers into the adjacent parvocellular layers). The B-Y cells were also largely segregated within the middle dLGN layers (K3, P3, K4 & P4). The R-G cells were mainly concentrated within the parvocellular layers (P3, P4, P5 & P6) and were evenly distributed throughout the middle and outer layers of the dLGN.<br>The study also included recordings from the extra-striate middle temporal area (MT) to determine whether a fast S-cone input exists from the dLGN to area MT which bypasses V1. The pattern of cone inputs to area MT neurons was investigated before and during inactivation of V1. The inactivation was done through reversible cooling with a Peltier thermocouple device or focal inactivation with y-amino butyric acid (GABA) iontophoresis. Precise inactivation of V1 to the topographically matching visual fields of the recording sites in area MT revealed a preservation of all three coneinputs in many cells. The subcortical sources of these preserved inputs are discussed with their relevance to blindsight, which is the limited retention of visual perception after V1 damage. Analysis of the latencies of area MT cells revealed a rough segregation into latencies faster or slower than 70 ms. Cells both with and without a significant change in response during V1 inactivation were present in each group. The findings reported in this thesis indicate that some of the preserved inputs in area MT during V1 inactivation may be carried by a direct input from the dLGN which bypasses V1.

A study was performed on decerebrate ferrets to define the contribution of vagal afferent non-myelinated fibres to the baroreceptor heart rate reflex produced by bolus i.v. injection of phenylephrine, using capsaicin as a selective C fibre blocker. Capsaicin blocked pulmonary chemoreflex substantially without any effects on bradycardia evoked by electrical stimulation of vagal efferent fibres to the heart. The significance of the contribution to bradycardia in response to marked increases in blood pressure by vagal C fibres are discussed in relation to findings in electrophysiological studies. A further study was performed on decerebrate ferrets and chloralose anaesthetised lambs. Baroreflex sensitivity was assessed by the relationship between cardiac interval changes and a rise in systolic blood pressure produced by bolus injection of phenylephrine and descending aorta occlusion. Moderate hypothermia (30-34<SUP>o</SUP>C) enhanced the baroreflex heart rate reflex substantially and was without effect on the sensitivity of pulmonary J receptor reflex pathways involved in the heart rate control. Action of vagal efferent fibres in altering heart rate was increased by moderate cooling. Such an effect may be partially responsible for the enhanced heart rate component of baroreflex response. Other possible mechanisms of enhanced baroreflex sensitivity are discussed. The consequence of enhanced vagal efferent fibre on heart was studied by electrical stimulation of the peripheral end of cervical vagus nerves in decerebrate ferrets and anaesthetised lambs. Moderate cooling substantially increases cardiac arrhythmias, such as sinus bradycardia, sinoatrial block, sinus arrest and A-V block. In addition vagal stimulation resulted in lethal ventricular arrythmia during infusion of noradrenaline. The possible mechanisms underlying the collapse and sudden death following rescue are discussed.

Hypertension affects 1 billion individuals worldwide and is the major contributing factor to cardiovascular disease. However, the WHO considers current antihypertensive drug therapies inadequate, highlighting a need for a novel approach to hypertension management. Baroreceptors are a promising drug target, and express an unusual glutamate receptor coupled to phospholipase D (PLD-GluR). The PLD-GluR has not been isolated and characterised, which is an important step towards its use as a drug target. A good source of the PLD-GluR is muscle spindle primary afferent nerve terminals, the largest mechanoreceptor in the body. This study thus focuses upon the identification and progress towards isolation of the PLD-GluR from muscle spindle primary afferent nerve terminals. A novel dissection method for high yield extraction of muscle spindles from a high density source, the rat deep masseter muscle, was developed for Western blotting and mass spectrometry screens of all GluRs. Western blots showed spindle homogenate contained a low molecular weight mGluR5 isoform and GluK2. Immunofluorescence showed mGluR5 was expressed on putative nociceptors, not mechanosensory nerve terminals. However, spindle mechanosensory nerve terminals labelled brightly for GluK2, as did baroreceptor nerve terminals. Furthermore, GluK2 appears to be the only GluR subunit on these mechanoreceptors, although mass spectrometry and affinity chromatography could not successfully isolate this receptor. Finally, piezo2 has recently been suggested as the major mechanotransducer protein. However, no evidence was found for piezo2 expression in adult spindle mechanosensory nerve terminals in adult rats or mice. As previous studies have largely focussed on adolescent mice, this could represent a developmental difference. Conversely, a number of candidate mechanosensory proteins, such as TRPs, were identified by a targeted mass spectrometry approach. This provides good candidates for future research. Collectively, this study indicates both spindle and baroreceptor mechanosensory nerve terminals express GluK2, suggesting it is at least a component of the PLD-GluR, and therefore potentially represents a novel drug target for treating hypertension.

Animals perceive the external world through their sensory systems, which consists in: i) sensory receptors that detect external stimuli; ii) neuronal pathways that convey the sensory information to the brain; and iii) central neurons organized in relay nuclei that process this information. To study the selective innervation of hindbrain regions by sensory afferents, we mapped the fine-grained topographical representation of sensory projections at the central level: sensory ganglia located more anteriorly project more medially than do ganglia located more posteriorly, and this relates to the time of sensory ganglia differentiation. This somatotopic arrangement is laid out very early, prompting the question of the origin of the signals involved in the induction and maintenance of this patterning. Up to date, several studies have tried to unveil how peripheral ganglia “send” afferent projections to “reach” their entry points in the hindbrain exploring whether these neurons search for axon guidance cues coming from the vicinity tissues or whether they intrinsically know where to go. This view seems to consider differentiated neurons as a population of cells arising in the middle of nowhere that should cross through axonal navigation, a mesenchyme sort of dark forest. By SPIM in vivo imaging we demonstrate that once placodal-derived neurons of dorsal posterior cranial ganglia differentiate, they never lose contact with neural ectoderm. First, delaminated neuroblasts differentiate in close contact with the neural tube, and afferents entrance points are established by plasma membrane interactions between primary differentiated peripheral sensory neurons and neural tube border cells, with the cooperation of neural crest cells. Then, neural crest cells and repulsive slit1/robo2 guidance cues guide later-differentiated axons and mediate sensory ganglion coalescence, axonal branching and fasciculation.<br>Los animales perciben el mundo externo a través de los sistemas sensoriales. Estos consisten en: i) los receptores sensoriales que detectan estímulos externos; ii) las vías neuronales que transmiten la información sensorial al cerebro; y iii) las neuronas centrales organizadas en núcleos que procesan la información. Con el objetivo de estudiar la inervación selectiva de las regiones del rombencéfalo por las aferentes sensoriales, hemos mapado la representación topográfica de las proyecciones a nivel central: los ganglios situados más anterior proyectan más medialmente que los situados más posterior, y esto depende del momento de diferenciación de estos ganglios. Esta organización somatotópica se establece muy tempranamente, lo que comporta la pregunta del origen de las señales implicadas en la inducción y mantenimiento de este patterning. Hasta ahora, muchos estudios han tratado de desvelar como los ganglios periféricos “mandan” las proyecciones aferentes para “alcanzar” sus puntos de entrada en el rombencéfalo y explorado si estas neuronas buscan claves de guidaje axonal provenientes de los tejidos adyacentes o si saben intrínsecamente dónde deben ir. Esta visión parece considerar las neuronas diferenciadas como una población de células que se originan en el medio de la nada y que deben navegar gracias a sus axones un mesénquima como si fuera un inhóspito bosque. Gracias a los métodos de imagen in vivo por SPIM hemos demostrado que una vez que las neuronas de los ganglios craneales posteriores han delaminado y diferenciado, nunca pierden contacto con el ectodermo neural. Primero, los neuroblastos se diferencian en íntimo contacto con el tubo neural, estableciéndose los puntos de entrada aferentes gracias a las interacciones entre membranas plasmáticas de las neuronas sensoriales y de las células del borde del tubo neural, con la cooperación de las células de la cresta neural. Luego, las células de la cresta neural y las señales de repulsión slit1/robo2 guían los axones que se han diferenciados más tarde y median la coalescencia de los ganglios, las bifurcaciones axonales y la fasciculación.

[Truncated abstract] One of the problems in researching tinnitus is that it has often been assumed that the physiological mechanisms underlying the tinnitus percept cannot be objectively measured. Nonetheless, it is generally accepted that the percept results from altered spontaneous neural activity at some site along the auditory pathway, although it is still debated whether it is produced by: synchronisation of activity of adjacent neurones; a change in the temporal pattern of activity of individual neurones; or an increase in the spontaneous firing rate per se. Similarly, it is possible that the recently coined “auditory neuropathy” is produced by under-firing of the primary afferent synapse, although several other mechanisms can also produce the symptoms described by this disorder (normal cochlear mechanical function but absent, or abnormal, synchronous neural firing arising from the cochlea and auditory brainstem, known as the auditory brainstem response, or ABR). Despite an absent ABR, some subjects can detect pure tones at near-normal levels, although their ability to integrate complex sounds, such as speech, is severely degraded in comparison with the pure-tone audiogram. The aim of the following study was to investigate the normal mechanisms underlying neural firing at the primary afferent synapse, and its regulation, to determine the possible mechanisms underlying over-firing (tinnitus) or under-firing (auditory neuropathy) of primary afferent neurones.

The work presented in this thesis concerns neurophysiology and pharmacology of the oesophageal afferent pathways involved in oesophago-pharyngeal reflexes and oesophageal nociception. Disturbances of reflexes governing contractile and relaxation responses of the upper oesophageal sphincter (DOS) are likely to be implicated in the pathophysiology of conditions involving excessive oesophago-pharyngeal regurgitation, impaired oesophageal clearance, and an abnormal belch reflex. Visceral hypersensitivity, a heightened perception of gastrointestinal sensation is frequently observed in functional gastrointestinal disorders and provides compelling evidence that it plays an important role in the pathogenesis of functional heartburn and non-erosive reflux disease. The work in this thesis explores the neurophysiology, pharmacology and pathophysiology of oesophago-DOS reflexes in humans by experimentally inducing DOS relaxations in healthy controls and patients with reflux laryngitis, and by recording DOS motor responses during spontaneous oesophago-pharyngeal regurgitation. Nociception was assessed by measuring oesophageal sensitivity to electrical stimulation and oesophageal acid perfusion in healthy controls, which was then compared with several heartburn populations (functional heartburn, erosive and non-erosive reflux disease). Additional studies were performed to evaluate the potential role of intraluminal impedance in defining antegrade bolus flow through the pharyngo-oesophageal segment during swallowing as a prelude to the adaptation of the technique to find a more accurate method for the detection of oesophago-pharyngeal regurgitation. The main findings from this work are as follows. 1) Mucosal lignocaine-sensitive afferents mediate the distension-induced oesophago-DOS relaxation reflex and lignocaine insensitive, presumably muscular mechanoceptors, mediate the distension-induced oesophago-DOS contractile reflex. The latter reflex is also upregulated by oesophageal acidification indicative of a possible protective mechanism. 2) Prolonged studies in patients with proven oesophago-pharyngeal regurgitation demonstrated that the most common mechanism of oesophago-pharyngeal regurgitation is a transient, non-swallow related, relaxation of the DOS. 3) Experimental evaluation of the oesophago-DOS relaxation reflex revealed that it is upregulated in patients with reflux laryngitis, suggesting that the aberrant afferent signalling in the oesophagus may be a contributory factor mediating oesophago-pharyngeal regurgitation. 4) Measurement of oesophageal sensory thresholds in response to electrical stimulation and acid perfusion revealed that all patients, irrespective of the presence or absence of mucosal injury, exhibit acid-induced hypersensitisation. 5) The viscro-somatic referral pattern of acid- and electrically-induced chest pain is increased in patients with functional heartburn and non-erosive reflux disease. These findings support the hypothesis that central sensitisation of nociceptive pathways may contribute to symptom reporting in these heartburn populations.

Esta tese de doutorado está dividida em três capítulos. O primeiro capítulo teve sua fundamentação em resultados da literatura, que mostraram que a aplicação de trens de alta freqüência de estimulação elétrica na perna pode gerar forças maiores do que seria esperado pela estimulação direta de axônios motores, por um mecanismo central gerado em motoneurônios por ativação de sinapses de aferências sensoriais. O objetivo foi investigar se estímulos vibratórios aplicados ao tendão de Aquiles são também capazes de gerar torques extras de grande magnitude no grupo muscular do tríceps sural. Para tanto, o torque isométrico gerado pelo tornozelo foi medido em resposta a diferentes padrões de associação entre estímulos elétricos e mecânicos aplicados ao grupo muscular do tríceps sural. As salvas de estímulos vibratórios puderam gerar torques extras substanciais, que eram acompanhados por um aumento da excitabilidade motoneuronal, visto que foi observado um concomitante aumento na amplitude pico a pico das ondas F captadas no músculo sóleo. Esses resultados mostraram que estímulos vibratórios aplicados juntamente a uma estimulação elétrica basal podem gerar níveis de força consideráveis, devido ao recrutamento de motoneurônios na medula espinhal. Os resultados indicam que a associação entre vibração e estimulação elétrica pode ser benéfica para diversas intervenções terapêuticas e programas de exercícios baseados em vibração. O comando para a geração de torques extras pela vibração provavelmente ativa motoneurônios na medula espinhal seguindo o princípio do tamanho, o que é uma característica desejável para paradigmas de estimulação. O segundo capítulo teve como ponto de partida o fato de que, quando indivíduos exercem um leve toque com a ponta dedo indicador sobre uma superfície estacionária, observa-se uma melhora na estabilidade postural. Nosso estudo investigou se uma melhora adicional na estabilidade postural pode ser atingida pela aplicação de um baixo nível de ruído mecânico (ruído vibratório) à superfície de contato com o dedo. Tal hipótese foi baseada na teoria da ressonância estocástica, que consiste na idéia de que a detecção e transmissão de um sinal fraco ou subliminar podem ser otimizadas pela presença de um nível particular de ruído. Os sujeitos permaneceram em postura ortostática sobre uma plataforma de força, enquanto tocavam com a ponta do dedo indicador direito um aparato usado para produzir oscilações vibratórias na superfície de contato do dedo. Parâmetros caracterizando as oscilações posturais foram computados a partir de medidas provenientes dos sinais do centro de pressão (adquiridos por uma plataforma de força). Os resultados mostraram que a aplicação de um ruído vibratório à superfície de contato do dedo causou uma melhora significativa na estabilidade postural quando comparada à condição em que a superfície de contado permanecia estacionária. Os resultados podem ter implicações clínicas relevantes em relação ao projeto de dispositivos de auxílio a pessoas com déficits sensório-motores, como bengalas, visando aumentar a estabilidade postural. Por fim, o terceiro capítulo investigou os efeitos da aplicação de ruído elétrico nos músculos da perna sobre o controle neuromuscular (i.e. variabilidade da força de flexão plantar exercida isometricamente) e sobre o controle postural humano (i.e. medida das oscilações posturais por uma plataforma de força). Os resultados mostraram pela primeira vez que um ruído elétrico abaixo do limiar sensorial pode causar uma melhora no controle neuromuscular durante uma tarefa de manter um determinado nível de força isométrica o mais constante possível, por mecanismo provavelmente associado à ressonância estocástica. Além disso, foi encontrada uma correlação significativa entre as reduções obtidas na variabilidade do torque e as reduções observadas em algumas medidas do centro de pressão, indicando que a variabilidade do torque de flexão plantar obtida com os sujeitos sentados pode ser um bom indicativo da amplitude das oscilações posturais para certas intervenções. Em conjunto, os resultados indicam um potencial de uso clínico de estimulações com ruído elétrico para a melhora do controle neuromuscular de pessoas com diferentes acometimentos sensório-motores.<br>This doctorate thesis is divided into three chapters. The first chapter was inspired on previous results from the literature which showed that high-frequency trains of electrical stimulation applied over the lower limb muscles can generate forces higher than would be expected by direct activation of motor axons, by a mechanism generated within the central nervous system by synaptic input from sensory afferents to motoneurons. The objective was to investigate if vibration applied to the Achilles tendon is also able to generate large magnitude extra torques in the triceps surae muscle group. The isometric ankle torque was measured in response to different patterns of coupled electrical and mechanical stimuli applied to the triceps surae muscle group. The vibratory bursts could generate substantial extra torques that were accompanied by increased motoneuron excitability, since a parallel increase in the peak-to-peak amplitude of soleus F waves was observed. These results showed that vibratory stimuli applied with a background electrical stimulation generate considerable force levels due to the spinal recruitment of motoneurons. So, the association of vibration and electrical stimulation could be beneficial for many therapeutic interventions and vibration-based exercise programs. The command for the vibration-induced extra torques presumably activates spinal motoneurons following the size principle, which is a desirable feature for stimulation paradigms. The second chapter was based on the knowledge that when a subjects index fingertip touches lightly a rigid and stationary surface there is an improvement of his/her postural stability. Our study investigated whether a further increase in postural steadiness may be achieved by applying a low level mechanical noise (vibratory noise) to the fingertip contact surface. Such a hypothesis is based on the stochastic resonance theory, which says that the detection and transmission of a weak or sub-threshold input signal may be enhanced by the presence of a particular level of noise. The subjects stood as quietly as possible over a force plate while touching with their right index fingertip a surface that could be either quiescent or vibrated. Body sway parameters were computed from measurements of the center of pressure (acquired from the force plate). The results showed that the delivery of the vibratory noise to the fingertips contact surface caused a significant improvement on postural stability when compared with the condition in which the surface was stationary. The results could be helpful for the design of rehabilitation aids such as canes commonly used to improve stability in people with sensory deficits. Finally, in the third chapter, the effects of electrical noise applied to the leg muscles on neuromuscular control (i.e. isometric plantar flexion force variability) and on human postural control (i.e. measures of postural sway) were investigated. The results showed for the first time that a sub-threshold electrical noise may improve neuromuscular control during a task in which an isometric force level is maintained as constant as possible, presumably by a stochastic resonance mechanism. Furthermore, a significant correlation was found between the reductions in torque variability and the improvements in postural sway, indicating that plantarflexion torque variability acquired while subjects are in a seated position may provide a good estimate of their postural sway under the same intervention. Taken together, the results indicate the potential for the clinical usage of noise-based stimulations to enhance neuromuscular control in a population with sensory-motor impairments.

Spinal cord injuries (SCI) sever communication between supraspinal centers and the central pattern generator (CPG) responsible for locomotion. Because the CPG is intact and retains the ability to initiate locomotor activity, it can be accessed electrically and pharmacologically. The goal of this thesis was to identify and characterize a novel spinal cord surface site along the sacral dorsal column (sDC) for electrically evoking locomotor-like activity in the neonatal rat spinal cord. Stimulation of the sDC robustly activated rhythmic left-right alternation in flexor-related ventral roots that was dependent on the activation of high-threshold C fiber afferents. The C fibers synapsed onto spinal neurons, which project to the lumbar segments as part of a pathway dependent on purinergic, adrenergic, and cholinergic receptor activation. In ventral roots containing only somatic efferents, rhythmic activity was rarely recruited. However, in ventral roots containing both autonomic and somatic efferents, sacral dorsal column stimulation recruited autonomic efferent rhythms, which subsequently recruited somatic efferent motor rhythms. The efferent rhythms revealed a half-center organization with very low stimulation frequencies, and the evoked alternating bursts entrained to the stimuli. Similar entrainment was seen when sDC stimuli were applied during ongoing neurochemically-induced locomotor rhythms. The rhythmic patterns evoked by sDC stimulation operated over a limited frequency range, with a discrete burst structure of fast-onset, frequency-independent peaks. In comparison, neurochemically-induced locomotor bursts operated over a wide frequency range and had slower time to peaks that varied with burst frequency. The overall findings support the discovery of an autonomic efferent pattern generator that is recruited by sacral visceral C fiber afferents. It is hoped that this research will advance the understanding of afferent activation of the lumbar central pattern generator and potentially provide insight useful for future development and design of neuroprosthetic devices.

The central nervous system is largely responsible for receiving sensory information from the environment and determining motor output. Yet, centrally-derived behavior and sensation depends on the optimal maintenance of the cells, tissues, and organs that feed and support these functions. Most of visceral regulation occurs without conscious oversight, making the spinal cord a key site for integration and control. How the spinal cord modulates output to our organs, or sensory information from them, is poorly understood. The overall aim of this dissertation was to better understand spinal processing of both visceral sensory information to and sympathetic output from the spinal cord. I first established and validated a HB9-GFP transgenic mouse model that unambiguously identified sympathetic preganglionic neurons (SPNs), the spinal output neurons for the sympathetic nervous system. Using this model, I investigated the electrophysiological similarities and diversity of SPNs, and compared their active and passive membrane properties to those in other animal models. My results indicate that while many of the same characteristics are shared, SPNs are a heterogeneous group that can be differentiated based on their electrophysiological properties. Since descending monoaminergic pathways have particularly dense projections to sympathetic regions of the spinal cord, I next examined the modulatory role that the monoamines have on spinal sympathetic output. While each neuromodulator tested had a unique signature of action, serotonin and norepinephrine appeared to increase the excitability of individual SPNs, while dopamine had more mixed actions. Since many autonomic reflexes are integrated by the spinal cord, I also questioned whether these reflexes would be similarly modulated. I therefore developed a novel in vitro spinal cord and sympathetic chain preparation, which allowed for the investigation of visceral afferent-mediated reflexes and their neuromodulation by monoamines. This preparation exposed a dichotomy of action, where sympathetic and somatic motor output is generally enhanced by the monoamines, but reflexes mediated by visceral input are depressed. Utilizing the spinal cord and sympathetic chain preparation, I also investigated how the spinal cord modulates visceral sensory information. One of the most powerful means of selectively inhibiting afferent information from reaching the spinal cord is presynaptic inhibition. I hypothesized that both spinal visceral afferents and descending monoaminergic systems would depress transmission of visceral afferents to the spinal cord. My results demonstrated that activity in spinal visceral afferents can lead to spinally generated presynaptic inhibition, and that in addition to depressing synaptic transmission to the spinal cord, the monoamines also depress the intrinsic circuitry that generates this activity-dependent presynaptic inhibition. Taken together, my results indicate that descending monoaminergic pathways act to limit the amount of visceral sensory information reaching the central nervous system and increase sympathetic output, resulting in an uncoupling of output from visceral sensory input and transitioning to a feed-forward, sympathetically dominant control strategy. This combination offers complex modulatory strategies for descending systems.

Cough is a common symptom of respiratory disease. Assessment of antitussives has relied mainly on animal studies and clinical trials in which recording of natural cough is difficult. This thesis describes the use of ultrasonically nebulized distilled water (UNOW) to induce cough in man. Investigation of the chemosensitivity of this response identified that extremes of pH, a chloride concentration below 75mmol/l, but not changes in osmolarity induce cough which reflects afferent rapidly adapting recep~or sensitivity in animal studies. Inhaled beta-adrenergic and anticholinergic bronchodilators, which inhibit cough in asthma, markedly reduced UNOW-induced cough in both healthy and asthmatic volunteers. Bronchoconstriction with inhaled leukotriene 04, which constricts both asthmatic and non-asthmatic airways, also caused coughing. Inhibition of bronchoconstriction either specifically or non-specifically resulted in inhibition of cough. Nedocromil sodium and the diuretic, frusemide, but not the commonly prescribed opiate, codeine, exhibited antitussive activity. Cough was also induced by inhalation of the C-fibre stimulants, capsaicin and prostaglandin E2 (PGE2), which was characterised by studies of adaptation, cross-adaptation and antitussives. UNOW and PGE2, but not capsaicin, exhibited rapid adaptation of cough. Crossadaptation, however, did not occur suggesting distinct mechanisms of cough mediation. Nedocromil inhibited capsaicin-induced cough but not PGE2-induced cough, while fenoterol did not affect either challenge. Oxitropium, which inhibited UNOW-induced cough, did not reduce cough associated with upper respiratory tract infection. Cough can be induced by a variety of inhaled stimuli. These can identify differences in response which may signal a number of pathways leading to cough. Antitussive activity may also be specific to individual challenges. This diversity in response reflects the complex neurological organisation of cough and may be related to pathological causes of cough.

There is a considerable need for treatments in MS for preventing progressive neurological disability. Assessment of the afferent visual pathway shows potential in investigating new therapies in MS. Mesenchymal Stem Cells exhibit properties of potential therapeutic relevance in progressive MS. A phase I/IIA trial of adult autologous mesenchymal stem cells as a potential therapy for Multiple Sclerosis [MSCIMS] was designed as an open label, pre (up to 20 months) vs. post treatment (up to 10 months) (single intravenous administration of autologous bone marrow derived mesenchymal stem cells) comparison study in ten secondary progressive MS patients. Primary end points were adverse events and secondary end points were efficacy measures. All 10 patients had previous history of clinical optic neuritis: this was in order to enable longitudinal structural and functional assessments of the disease-affected afferent visual pathway. Piecewise linear mixed models were used to assess the change in gradients over time at the point of intervention. All 10 patients tolerated the trial assessments and intervention. No significant or serious adverse events were seen. Improvement after treatment was seen in visual acuity and visual evoked response latency, along with an increase in optic nerve cross-sectional area. The results suggest that autologous mesenchymal stem cells are safe and could possibly promote endogenous repair mechanisms such as remyelination, although a definitive conclusion of this cannot be made from this small study. While MSCIMS was a proof of concept study only, based on the encouraging experience derived from it, there would seem to be potential value in future, larger placebo controlled, double-blinded, randomised therapeutic phase IIb/III trials that could (i) more definitively investigate stem cells as a therapy and (ii) use the visual pathway disease model for investigating the efficacy of potential neuroprotective and reparative therapeutic agents.

Includes bibliographical references (leaves 136-156)<br>156 leaves : ill. ; 30 cm.<br>Title page, contents and abstract only. The complete thesis in print form is available from the University Library.<br>Two novel in vitro preparations were developed from which recordings were made from colonic afferents in the rat and mouse.<br>Thesis (Ph.D.)--University of Adelaide, Dept. of Medicine, 2001

"April 2003"<br>Bibliography: leaves 124-144.<br>xvii, 144 leaves : ill. ; 30 cm.<br>Thesis (Ph.D.)--University of Adelaide, School of Molecular and Biomedical Sciences, Discipline of Physiology, 2003

"April 2003"<br>Bibliography: leaves 124-144.<br>xvii, 144 leaves : ill. ; 30 cm.<br>Title page, contents and abstract only. The complete thesis in print form is available from the University Library.<br>Thesis (Ph.D.)--University of Adelaide, School of Molecular and Biomedical Sciences, Discipline of Physiology, 2003

Thesis (Ph. D.)--Florida State University, 2005.<br>Advisor: Dr. Richard Hyson, Florida State University, Program in Neuroscience. Title and description from dissertation home page (viewed Sept. 19, 2005). Document formatted into pages; contains ix, 64 pages. Includes bibliographical references.

Thesis (M. Sc.)--York University, 2001. Graduate Programme in Kinesiology and Health Science.<br>Typescript. Includes bibliographical references. Also available on the Internet. MODE OF ACCESS via web browser by entering the following URL: http://wwwlib.umi.com/cr/yorku/fullcit?pMQ71585.

博士<br>國立成功大學<br>醫學工程研究所碩博士班<br>94<br>Normal lower urinary tract function involves spinal and supraspinal pathways that control urine storage and release. In rats, urine release is mediated by contraction of the bladder detrusor accompanied coordinated activation of two types (tonic activity and bursting) of external urethral sphincter (EUS). This study used cystometry, EUS electromyography and nerve activity to examine the origin of EUS bursting and EUS activity as well as pudendal nerve evoked by electrical stimulation at pudendal and pelvic nerves (pudendal-EUS, pudendal-pudendal nerve and pelvic-EUS reflexes). Furthermore, the changes in the reflexes induced by bladder distension and by administration of agonists or antagonists for glutamatergic or serotonergic receptors were examined in rats with intact spinal cord (SC) rats or spinal cord injury (SCI) following administration of glutamatergic antagonist receptors (MK801 or LY215490) or 5HT-1A receptor agonist (8-OH-DPAT) and antagonist (WAY100635). 　In intact-SC rats, stimulation of the entire pudendal nerve elicited short latency (8-12 ms) responses in the EUS and short (3-8 ms) and long latency responses (16-20 ms) in contralateral pudendal nerve. The long latency pudendal-pudendal reflex was reduced by 36.7 % in area during bladder distension. However, there was no significant change in the area of pudendal-EUS reflex during bladder distension. Peak amplitudes of both reflexes were reduced 32% by bladder distension. The effects of glutamatergic receptor antagonists on the reflexes were also examined. MK801 (0.3-5 mg/kg, i.v.) markedly depressed the pudendal-pudendal reflex, but LY215490 (3 mg/kg, i.v.) had a minimal inhibitory effect. Both glutamatergic receptor antagonists significantly suppressed the pudendal-EUS reflex. 　The pelvic-EUS reflex consisted of an early response (ER, latency, 18-22 ms) and a late, long duration response (LR, latency greater than 100 ms) that consisted of bursts of activity at 20-160 ms inter-burst intervals in intact-SC rats. The LR was markedly enhanced when the bladder was distended. The ER and LR was significantly decreased 75% and 35%, respectively, by MK801 (0.3 mg/kg, i.v.), but only decreased 18% and 14%, respectively, by LY215490 (3 mg/kg, i.v.). 8-OH-DPAT (1 mg/kg, i.v.) enhanced spontaneous EUS activity and the pelvic-EUS reflexes. WAY100635 (0.3-1 mg/kg, i.v.) reversed the effect of 8-OH-DPAT and suppressed EUS activity and the pelvic-EUS reflex. 　The effects of 8-OH-DPAT and WAY100635 were examined in the rats following acute and chronic (2-5 weeks) SCI. Tonic EUS activity remained but bursting was abolished during bladder distension in the rats with acute T8-9 or L3-4 and chronic L3-4 SCI. Both tonic and bursting EUS activity were completely abolished in acute and chronic L6-S1 SCI rats. Only the ER remained in acute and chronic T8-9 and L3-4 SCI rats, but was absent in L6-S1 rats. 8-OH-DPAT (1 mg/kg, i.v.) facilitated tonic activity, EUS bursting and LR in T8-9 chronic SCI rats. However, only tonic activity was enhanced (5-10%) by 8-OH-DPAT in T8-9 acute SCI rats. WAY100635 (1 mg/kg, i.v.) reversed the effect of 8-OH-DPAT. Neither drug had any effect in acute or chronic L3-4 or L6-S1 SCI rats. These results indicate that the EUS is innervated by multiple pathways and that glutamatergic excitatory transmission is important in the neural mechanisms underlying bladder-sphincter coordination in the rat. Glutamatergic and serotonergic mechanisms are important in the reflex pathways underlying bladder-sphincter coordination in rats. Spinal serotonergic mechanisms facilitate tonic and bursting EUS activity. The circuitry for generating EUS bursting seems to be located in the spinal cord between T8-9 and L3-4.

"November 2005"<br>Bibliography: leaves 346-266.<br>366 leaves : ill. (some col.), col. plates ; 30 cm.<br>Title page, contents and abstract only. The complete thesis in print form is available from the University Library.<br>Thesis (Ph.D.)--University of Adelaide, School of Molecular and Biomedical Sciences, Discipline of Physiology, 2005