Indiana University-Purdue University Indianapolis (IUPUI) / Voltage-gated sodium channels (VGSCs) are dynamic membrane spanning proteins which mediate the rapid influx of Na+ during the upstroke of the action potential (AP). In addition to the large inward Na+ currents responsible for the upstroke of the AP, some VGSC isoforms produce smaller, subthreshold Na+ currents, which can influence the excitable properties of neurons. An example of such a subthreshold current is resurgent Na+ current (INaR). These unusual currents are active during repolarization of the membrane potential, where the channel is normally refractory to activity. INaR exhibit slow gating kinetics and unusual voltage-dependence derived from a novel mechanism of channel inactivation which allows the channel to recover through an open configuration resulting in membrane depolarization early in the falling phase of the AP, ultra-fast re-priming of channels, and multiple AP spikes. Although originally identified in fast spiking central nervous system (CNS) neurons, INaR has recently been observed in a subpopulation of peripheral dorsal root ganglion (DRG) neurons. Because INaR is believed to contribute to spontaneous and high frequency firing of APs, I have hypothesized that increased INaR may contribute to ectopic AP firing associated with inherited and acquired disorders of excitability. Specifically, this dissertation explores the mechanisms which underlie the electrogenesis of INaR in DRG neurons and determines whether the biophysical properties of these unique currents were altered by mutations that cause inherited muscle and neuronal channelopathies or in an experimental model of nerve injury. The results demonstrate that (1) multiple Na+ channel isoforms are capable of producing INaR in DRG neurons, including NaV1.3, NaV1.6, and NaV1.7, (2) inherited muscle and neuronal channelopathIy mutations that slow the rate of channel inactivation increase INaR amplitude, (3) temperature sensitive INaR produced by select skeletal muscle channelopthy mutations may contribute to the triggering of cold-induced myotonia, and (4) INaR amplitude and distribution is significantly increased two weeks post contusive spinal cord injury (SCI). Taken together, results from this dissertation provide foundational knowledge of the properties and mechanism of INaR in DRG neurons and indicates that increased INaR likely contributes to the enhanced membrane excitability associated with multiple inherited and acquired disorders of excitability.

A esclerose múltipla (EM) é doença desmielinizante do sistema nervoso central com vários sinais e sintomas, dentre eles a paralisia muscular. Essa paralisia foi estudada neste trabalho utilizando um modelo animal (camundongo) de EM, a Encefalomielite Autoimune Eperimental (EAE). Comparando a controles, os animais EAE, além da paralisia muscular, apresentaram diminuição da massa corporal e do músculo extensor longo dos dedos (ELD). Isolado com ou sem o nervo isquiático, o ELD mostrou decréscimo das contrações indiretas e diretas, tanto isoladas como tetânicas, e maior resistência à fadiga muscular. No ELD, o bloqueador neuromuscular pancurônio mostrou potência semelhante sobre contrações indiretas isoladas, em animais EAE e controles, enquanto as contrações indiretas tetânicas foram mais resistentes ao pancurônio, em animais EAE. A área de secção transversa das fibras musculares foi menor e houve troca (shift) de tipos de fibras de IIB para IIA, em animais EAE. Concluiu-se que as alterações nos animais EAE são tanto da transmissão neuromuscular como do músculo. / Multiple sclerosis (MS) is a demielinating disease of the Central Nervous System with a number of signs and symptoms, among them muscle paralysis. In this work this paralysis was studied using an animal (mouse) model of MS: the experimental autoimmune encephalomyelitis (EAE). Compared to controls, the EAE animals, besides muscle paralysis, had a lower body and extensor digitorum longus (EDL) muscle mass. Isolated with or without the sciatic nerve, the indirect and the direct isolated contractions of the EDL were smaller and the muscle fatigue was lower, in EAE animals. The neuromuscular blocker pancuronium was equally potent on indirect isolated contractions, in control and EAE animals, while the tetanic contractions were more resistant to pancuronium, in EAE animals. Compared to controls, the area of the transverse section of the muscle fibers was smaller and there was a shift from the IIB to the IIA type of fiber in EAE animals. It is concluded that the alterations in EAE animals result from changes in the process of neuromuscular transmission and in the muscle.

[Truncated abstract] The goal of this research was to investigate some of the ways that neurons located in the primary motor cortex (MI) code for skilled movement. The task-related and temporally correlated spike activity that occurred during the performance of a goal-directed reaching and retrieval task invloving multiple motion elements and limbs was evaluated in cats. The contributions made by different neuronal subtypes loctaed in MI (which were identified based upon extracellular spiking features0 to the coding of movement was also investigated. Spike activity was simulateously recorded from microelectrodes that were chronically implanted into the motor cortex of both cerebral hemispheres. Task-related neurons modulated their activity during the reaching and retrieval movements of one forelimb, or the postural reactions of the contralateral forelimb and ipsilateral hindlimb. Spike durations and baseline firing rates of neurons were used to distinguish between putative excitatory (Regular Spiking; RS) and inhibitory (Fast Spiking; FS) neurons in the cortex. Frame by frame video analysis of the task was used to subdivide each task trial into stages (e.g. premovement, reach, withdraw and feed) and relate modulations in neural activity to the individual task stages. Task-related neurons were classified as either narrowly tuned or broadly tuned depending on whether their activity modulated during a single task stage or more than one stage respectively. Recordings were made from 163 task-related neurons, and temporal correlations in the spike activity of simultaneously recorded neurons were identified using shuffle corrected cross-correlograms on 662 different neuronal pairs.... The results of this research suggest that temporally correlated activity may reflect the activation of intracortical and callosal connections between a variety of efferent zones involved in task performance, playing a role in the coordination of muscles and limbs during motor tasks. The differences in the patterns of task-related activity, and in the incidence of significant neuronal interactions that were observed between the RS and FS neuronal populations implies that they make different contributions to the coding of movement in MI.

Neste trabalho foi preparada uma série de onze brometo de 2-[(4-X-benzamido) etil]benzildimetilamônio, compostos I.1-I.11, série I, estruturalmente análogos à procainamida, não descritos na literatura. Os valores do coeficiente de partição, log Papp7.40, destes compostos foram determinados pelo método shake-flask e foram utilizados como parâmetro lipofílico experimental. Os valores de deslocamento químico do grupamento carbonila, δ 13 C=O, foram determinados e foram determinados cm um espectrómetro de ressonância magnética nuclear a 75 MHz. Adicionalmente, outros parâmetros físico-químicos foram retirados da literatura: π, σp, Τ, R e MR4 ou obtidos por cálculo: log Pcalc e πexp. Os valores da concentração inibitória média (IC50) capaz de reduzir a contração máxima a 50% no período de 15 minutos, foram determinados em preparações nervo frênico-músculo diafragma de camundongos. Para verificar a natureza e a contribuição relativa dos parâmetros físico-químicos frente ao bloqueio da transmissão neuromuscular foi feito uma análise de QSAR, obtendo-se equações, usando análise de regressão linear. As análises de QSAR sugerem uma dependência positiva da lipofilicidade para o bloqueio da transmissão neuromuscular expresso por pIC50, segundo o modelo proposto expresso pela equação: (Ver arquivo PDF). / In this work, a set of eleven 2-[(4-X-benzamido)ethyl]benzyl dimethylammonium bromide structurally related to procainamide was synthesized. The apparent partition coefficient values were determined by means of \"shake-flask method and were taken as lipophilic parameters. The carbonyl chemical shifts values were determined in methanol-d4 and taken as electronic parameters. Additionally, physicochemical parameters were either taken from literature: π, σp, Τ, R e MR4 or calculated: log Pcalc and πexp. The median inhibitory concentration values (IC50) able to reduce maximal contraction to 50% at 15 minutes was determined in phrenic nerve-diafragm muscle preparation of mices. In order to verify the nature and relative contribution of the physicochemical parameters to neuromuscular blockage, QSAR equations were derived using regression analysis. The obtained QSAR model, expressed by the equation below, suggest that lipophilicity term plays an important role to neuromuscular blockage. (See files PDF).

Avec le besoin de développer un organe artificiel remplaçant le doigt humain dans le cas d'un déficit et la nécessité de comprendre le fonctionnement de ce système physiologique, un modèle physique inverse du système doigt, permettant de chercher les activations neuronales à partir du mouvement, est nécessaire. Malgré le grand nombre d'études dans la modélisation de la main humaine, presque il n'existe aucun modèle physique inverse du système doigt majeur qui s'intéresse à chercher les activations neuronales. Presque tous les modèles existants se sont intéressés à la recherche des forces et des activations musculaires. L'objectif de la thèse est de présenter un modèle neuromusculo-squelettique du système doigt majeur humain permettant d'obtenir les activations neuronales, les activations musculaires et les forces musculaires des tous les muscles agissants sur le système doigt d'après l'analyse du mouvement. Le but de ce type des modèles est de représenter les caractéristiques essentielles du mouvement avec le plus de réalisme possible. Notre travail consiste à étudier, modéliser et à simuler le mouvement du doigt humain. L'innovation du modèle proposé est le couplage entre la biomécanique et les aspects neurophysiologiques afin de simuler la chaine inverse complet du mouvement en allant des données dynamiques du doigt aux intentions neuronales qui contrôlent les activations musculaires. L'autre innovation est la conception d'un protocole expérimental spécifique qui traite à la fois les données sEMG multicanal et les données cinématiques d'après une procédure de capture de mouvement. / With the need to develop an artificial organ replacing the human finger in the case of a deficiency and the need to understand how this physiological system works, an inverse physical model of the finger system for estimating neuronal activations from the movement, is necessary. Despite the large number of studies in the human hand modeling, almost there is no inverse physical model of the middle finger system that focuses on search neuronal activations. Al most all existing models have focused on the research of the muscle forces and muscle activations. The purpose of the manuscript is to present a neuromusculoskeletal model of the human middle finger system for estimating neuronal activations, muscle activations and muscle forces of all the acting muscles after movement analysis. The aim of such models is to represent the essential characteristics of the movement with the best possible realism. Our job is to study, model and simulate the movement of the human finger. The innovation of the proposed model is the coupling between the biomechanical and neurophysiological aspects to simulate the complete inverse movement chain from dynamic finger data to neuronal intents that control muscle activations. Another innovation is the design of a specific experimental protocol that treats both the multichannel sEMG and kinematic data from a data capture procedure of the movement.

Abstract The introduction of physiologically and therapeutically effective drugs in anaesthetic treatment has meant a significant change. Their discovery has thus contributed to a minimisation of their negative effects on a living organism. As a result, anaesthesiology has become more controllable and safer. These new drugs translate into more comfort for the patient in the course of anaesthesiology as well as his or her rapid recovery resulting in a shorter period of hospitalisation. The research of new physiologically and therapeutically effective substances is a pre-requisition of a potential higher standard of medical care. Older substances, e.g. Alkuronium and Gallamin, which, due to their undesirable effects, have come out of use, have lost their significance. Nowadays, these substances have been generally replaced by substances with an intermedial effect, e.g. Rokuronium and Cisatrakurium, which, compared with Alkuronium and Gallamin, show a minimum of undesirable effects. One of the foremost objectives of the current pharmaceutical research is to find a replacement of Sukcinylcholin, which, in spite of its known side effects, has had a non-substitutable position in urgent intubation. As a result, its use is limited to out-patient application and a certain selection of patient categories. Let us hope that we shall see a replacement of Sukcinylcholin being introduced in treatment in the near future. This would mean a final solution of problems arising from its side effects limiting its scope of application.

Indiana University-Purdue University Indianapolis (IUPUI) / The sympathetic nervous system strongly modulates the contractile and electrical function of the heart. The anatomical underpinnings that enable a spatially and temporally coordinated dissemination of sympathetic signals within the cardiac tissue are only incompletely characterized. In this work we took the first step of unraveling the in situ 3D microarchitecture of the cardiac sympathetic nervous system. Using a combination of two-photon excitation fluorescence microscopy and computer-assisted image analyses, we reconstructed the sympathetic network in a portion of the left ventricular epicardium from adult transgenic mice expressing a fluorescent reporter protein in all peripheral sympathetic neurons. The reconstruction revealed several organizational principles of the local sympathetic tree that synergize to enable a coordinated and efficient signal transfer to the target tissue. First, synaptic boutons are aligned with high density along much of axon-cell contacts. Second, axon segments are oriented parallel to the main, i.e., longitudinal, axes of their apposed cardiomyocytes, optimizing the frequency of transmitter release sites per axon/per cardiomyocyte. Third, the local network was partitioned into branched and/or looped sub-trees which extended both radially and tangentially through the image volume. Fourth, sub-trees arrange to not much overlap, giving rise to multiple annexed innervation domains of variable complexity and configuration. The sympathetic network in the epicardial border zone of a chronic myocardial infarction was observed to undergo substantive remodeling, which included almost complete loss of fibers at depths >10 µm from the surface, spatially heterogeneous gain of axons, irregularly shaped synaptic boutons, and formation of axonal plexuses composed of nested loops of variable length. In conclusion, we provide, to the best of our knowledge, the first in situ 3D reconstruction of the local cardiac sympathetic network in normal and injured mammalian myocardium. Mapping the sympathetic network connectivity will aid in elucidating its role in sympathetic signal transmisson and processing.

Indiana University-Purdue University Indianapolis (IUPUI) / Although peripheral motoneurons are phenotypically endowed with robust regenerative capacity, functional recovery is often suboptimal following peripheral nerve injury (PNI). Research to date indicates that the greatest success in achieving full functional recovery will require the use of a combinatorial approach that can simultaneously target different aspects of the post-injury response. In general, the concept of a combinatorial approach to neural repair has been established in the scientific literature but has yet to be successfully applied in the clinical situation. Emerging evidence from animal studies supports the use of electrical stimulation (ES) and testosterone as one type of combinatorial treatment after crush injury to the facial nerve (CN VII). With the facial nerve injury model, we have previously demonstrated that ES and testosterone target different stages of the regeneration process and enhance functional recovery after facial nerve crush injury. What is currently unknown, but critical to determine, is the impact of a combinatorial treatment strategy of ES and testosterone on functional recovery after crush injury to the sciatic nerve, a mixed sensory and motor spinal nerve which is one of the most serious PNI clinical problems. The results of the present study indicate that either treatment alone or in combination positively impact motor recovery. With regard to molecular effects,single and combinatorial treatments differentially alter the expression of regeneration-associated genes following sciatic nerve crush injury relative to facial nerve injury. Thus, our data indicate that not all injuries equally respond to treatment. Furthermore, the results support the importance of treatment strategy development in an injury-dependent manner and based upon the functional characteristics of spinal vs. cranial nerves.