Scott H. Chandler - US grants

The specific aim of this proposal is to elucidate the brain stem neuronal mechanisms responsible for the production and coordination of spontaneously occurring and cortically induced rhythmical jaw movements (RJM) resembling mastication in the guinea pig. We propose to systematically search the brain stem reticular formation for neuronal sites which participate in the production of RJMs. This proposal is divided into three main sections. Part one will utilize brain stem microstimulation techniques and field potential analysis within specific motoneuronal pools of the trigeminal motor nucleus to determine if a particular brain stem site has an effect on the motor pool. Part two will involve extracellular recording from reticular neurons of the brain stem during RJMs. Computer analysis and spike triggered averaging techniques will be employed to determine the functional connectivity that exists between the brain stem neuron, motoneuron and masticatory area of the cortex. Finally, intracellular recording from specific identified trigeminal motoneurons innervating the muscles which control opening, closing and lateral movements of the mandible will be obtained during microstimulation of brain stem sites shown to effect the trigeminal motor nucleus. This will allow us to determine the precise synaptic effect these areas exert on the specific motoneurons. The long term goal of the research is to understand how the central nervous system initiates, coordinates, and executes the rhythmical stereotypic movements of the mandible during mastication. Involuntary, rhythmical movements of the jaws occurs in humans during sleep (bruxism). These patients tend to develop myofacial pain. Uncontrolled rhythmical movements of oral-facial structures also occurs during long term administration and withdrawal from anti-psycotic drugs (tardive dyskinesia). The etiology of these movement disorders is unknown. The results of these studies will shed light on the neuronal mechanism(s) responsible for these involuntary rhythmical jaw movements.

The specific aims of this proposal are to elucidate the brain stem neurophysiological and neuropharmacological mechanisms controlling rhythmical jaw movements (RJMs), resembling mastication, in the guinea pig. We propose to characterize the pharmacological mechanisms controlling jaw opener (digastric) motoneuron and premotoneuron excitability during RJMs elicited by electrical stimulation of the cortex (or pyramidal tract), and during RJMs elicited by systemic administration of a dopamine agonist, apomorphine (APO). The project is divided into three parts. Parts one and two will focus on recording extracellularly from digastric motoneurons (DIG) or pre-motoneurons, respectively, during cortically induced RJMs while simultaneously applying to the cell, through micropipettes, small quantities of putative neurotransmitter (NT) agonists and antagonists. Finally, part three will investigate the neurophysiological and neuropharmacological mechanisms controlling these same cell populations during RJMs induced by APO administration. The long-term goal is to understand both the mechanisms underlying the central nervous system control of normal rhythmic jaw movements that occur during activities such as feeding and drinking, as well as the abnormal, involuntary rhythmic jaw movements occurring in disorders such as tardive dyskinesia and bruxism. Tardive dyskinesia is a movement disorder characterized by uncontrolled rhythmic jaw movements of oral-facial structures that result from long term administration of neuroleptic drugs. Bruxism, the grinding or clenching of teeth in sleep, is another uncontrolled jaw movement behavior that often leads to the myofacial pain-dysfunction syndrome. The etiology of the generation of these abnormal jaw movements in unknown, although brusixm is thought to be related to stress, and tardive dyskinesia to a disruption of dopamine activity in the basal ganglia. The results of the proposed experiments in the guinea pig will provide insights into the neurophysiological and neuropharmacological mechanisms underlying the production of involuntary rhythmic jaw movements in humans.

The purpose of this pilot project is to develop an in vitro isolated brain stem preparation in the guinea pig in which neurophysiological and neuropharmacological studies can be performed on trigeminal reflex systems as well as on brainstem neuronal networks responsible for the generation of suckling and/or mastication. Specifically, we will establish the suitability of the in vitro preparation for the characterization of motoneuron membrane properties, synaptic and pharmacological mechanisms control trigeminal jaw opener and closer motoneuron excitability during either resting conditions, specific neuronal pathway activation or chemical stimulation. Furthermore, the ability of this preparation to produce coordinated rhythmical patterns of neuronal discharge which resemble mastication or suckling during bath application of excitatory amino acids will be determined. The project is divided into three parts. Part I will focus on recording intracellularly form trigeminal motoneurons during resting conditions for periods of time sufficient to characterize motoneuron membrane properties as well as generate synaptic potentials to peripheral reflex activation. This data will serve as normative data for further pharmacological studies. Part II will assess the role of excitatory amino acids in mediating monosynaptic and polysynaptic activation of trigeminal motoneurons during peripheral reflex activation. The ability of excitatory amino acid antagonists to antagonize reflex transmission to motoneurons will be determined. Part III will assess the ability of brainstem circuits to generate coordinated rhythmical discharges which resemble those patterns that occur during mastication of suckling during either central electrical stimulation of specific brainstem loci or pharmacological activation by excitatory amino acids. The long-term goal of this research is to understand both the mechanism underlying the central nervous system control of normal rhythmic jaw movements that occur during activities such as feeding and drinking, as well as the abnormal, involuntary rhythmic jaw movements occurring in disorders such as tardive dyskinesia and bruxism. The etiology of the generation of these abnormal jaw movements is unknown. The results of the proposed experiments will provide insights into the neurophysiological and neuropharmacological mechanisms underlying the production of normal and abnormal rhythmic jaw movements in humans.

The long-term objective of this proposal is to determine the neutral basis for the coordination and control of jaw and tongue movements during feeding and drinking. Two experimentally generated rhythmic oral-facial behaviors have been developed in the anesthetized guinea pig: (1) cortically evoked and, (2) apomorphie induced RJMs. Jaw and tongue muscle contraction patterns and mandibular movement trajectories of these RJM behaviors have characteristics that resemble, respectively, those found during mastication and lapping the awake animal. Recent investigations have indicated the importance of neutral networks in the pontomedullary reticular formation for the control and coordination of oral-facial motor behaviors. The purpose of the proposed studies is to investigate the organization and functional characeristics of these networks. The first area of investigation is the parvocellular nucleus (PVC). The PVC is in the lateral region of the reticular formation and is the location of trigeminal and hypoglossal premotoneurons. The second area of investigation involves nuclei in the medial region of the reticular formation. It is hypothesized that this region contains a neutral network responsible for rhythm generation in mastication and drinking. Methods to be used in these studies include intracellular recording in trigeminal and hypoglossal motoneurons, extracellular recording of reticular formation neurons, correlation of activity of neurons in the PVC with those in the medial nuclei, and correlation of PVC neutral activity with synaptic potentials evoked in motoneurons innervating jaw and tongue muscles. Two important clincial entities recognized in dentistry are the myofacial pain dysfunction syndrome and bruxism. Both have neuromuscular etiologies and can lead to pathologies of hard and soft tissues in the oral-facial area. Furthermore, aberrant habitual movements of the jaws and tongue are thought play a major role in generating morphological malformations that require orthodontic correction. The importance of jaow and tongue control in prosthodontics is becoming more evident. Dyskinesias involving the jaw and tongue are also manifest in tardive dyskinesia, senility, stroke and in comatose patients. There is, however, very little understanding of central nervous system mechanisms responsible for the coordination and control of oral-facial movements, the impairment of which is the central feature of all of these disorders. In spite its significance, the study of the mechanisms of motor control of oral-facial behaviors has lagged far behind those involving locomotion, respiration and eye movements. The results of experiments proposed here will provide insights into the neutral mechanisms underlying the central nervious sytem control of jaw and tongue coordination during rhythmic jaw movement behaviors. Such information is necessary for the development fo more effective clinical treatment paradigms for oral-facial motor disorders.

The proper functioning of oral-facial motor behaviors is necessary for the survival of humans. The ingestive behaviors of mammals begins with suckling and progresses to drinking and feeding. Presently, there are very few studies addressing how the brain is organized to produce these behaviors. Even less is known about the etiology of various oral-motor dysfunctions such as tardive dyskinesia. bruxism, and myofacial pain dysfunction syndromes. The long-term goals of this research are to understand both the mechanism underlying the central nervous system control of normal jaw movements that occur during activities such as feeding and drinking, as well as the abnormal jaw movements that occur during various disorders. The specific aims of this proposal are to continue investigations, at the cellular level, into the neuronal processes controlling jaw movements in the guinea pig. We will combine intracellular or whole cell patch clamp recording methods with retrograde-tract tracing techniques to identify specific populations of jaw-opener and closer motoneurons, and trigeminal premotoneurons in thin and thick brain slices. The project is divided into two main parts. In part I, we will determine for identified jaw-opener and closer motoneurons and premotoneurons, I) the presence of specific intrinsic membrane conductances and their contribution to neuronal burst discharge, 2) whether these conductances are substrates for modulation by two monoamines (5-HT and NE). Part II will focus on modulation of excitatory amino acid mediated synaptic transmission by monoamines. We will determine if there are fundamental similarities and differences in modulation by serotonin and norepinephrine between NMDA and non-NMDA components of synaptic transmission to trigeminal motoneurons from mesencephalic of V afferents (Mes V). We will examine these relationships by recording the compound EPSP or EPSC and the single fiber EPSP or EPSC evoked from Mes V nucleus or single Mes V neuron stimulation, respectively. The results of the proposed studies will provide insights into the cellular mechanisms controlling discharge of distinct populations of neurons involved in production of jaw movements and will serve as a cellular foundation for neuronal models on masticatory rhythm and burst pattern generation.

DESCRIPTION (provided by applicant): The proper functioning of oral-facial motor systems is necessary for the survival of humans. The ingestive behaviors of mammals begins with suckling and progresses to drinking and chewing. Presently, there are very few studies addressing how the brain is organized to produce these behaviors. Even less is known about the etiology of various oral-motor disorders such as tardive dyskinesia, bruxism, and myofacial pain dysfunction syndromes. The long-term goals of this research are to understand both the mechanisms underlying the central nervous system control of normal jaw movements that occur during activities such as drinking and chewing, as well as the abnormal jaw movements that occur during various disorders. The specific aims of this proposal are to continue investigations, at the cellular level, into the neuronal mechanisms controlling trigeminal neuronal membrane excitability and burst discharge that are associated with the critical transition from primitive suckling behavior to adult-like mastication in the rat. We will combine whole cell patch clamp recording methods of neurons within the trigeminal nuclei responsible for oral-motor activity (mesencephalic V neurons and trigeminal interneurons) in brain slices with microstimulation, neurochemical, and mathematical modeling techniques to more fully elucidate 1) the mechanisms controlling excitability, and 2) the local chemical and electrical microcircuitry of these neurons. The project is divided into two Specific Aims. In Specific Aim I we will determine the locations of, and ionic mechanisms underlying, intrinsic burst generation in Mes V and trigeminal interneurons in the vicinity of the trigeminal motor nucleus and test the hypothesis that the underlying conductances are substrates for modulation by metabotropic glutamate receptor (mGluR) and serotonergic (5-HT) receptor activation. Specific Aim II focuses on characterizing the local excitatory and inhibitory chemical and electrical synaptic interactions that occur between distinct trigeminal neurons, and the modulation of these interactions by activation of mGluR and 5-HT receptors. The results of the proposed studies will provide insights into the local microcircuitry and the cellular mechanisms controlling discharge of distinct populations of trigeminal neurons involved in production of jaw movements at distinct developmental time points, and will serve as a cellular foundation for creation of neuronal models of masticatory rhythm and burst pattern generation.

DESCRIPTION (provided by applicant): Amyotrophic lateral sclerosis (ALS) is a progressive, fatal neurodegenerative disease that ends with degeneration in both upper and lower motoneurons. Approximately 10-15% of the diagnosed ALS cases are inherited, (familiar ALS) (FALS), while the remaining are sporadic. Clinically, the disease is characterized by muscle weakness, atrophy and fasciculations in the limbs and difficulty in swallowing and chewing. Patients typically die within 3-5 years of diagnosis due to respiratory failure. The long-term goals of this research are to determine the cellular mechanisms that lead to the progressive loss of motoneuronal function and pathogenesis of ALS, and to establish targets that can be used to develop a multifaceted therapeutic approach to delaying the progression of the degeneration. Our immediate goal, using a mouse model of ALS (SOD1 mice) and electrophysiological, as well as mitochondrial and calcium imaging methods, is to test our working hypothesis that presymptomatic alterations of intrinsic voltage-gated calcium and/or potassium channels in trigeminal motoneurons and presynaptic trigeminal proprioceptive primary afferent neurons occur simultaneously, and contribute to the hyperexcitability previously observed in SOD1 mutant mice. This information is important because simultaneous changes in intrinsic ion channel function in pre- and postsynaptic target neurons could 1) be complex conjoint signals to initiate the disease process, and 2) produce an increase in pre- or postsynaptic membrane excitability that leads to the observed spasticity and fasciculations observed in ALS patients, as well as 3) trigger the processes that lead to calcium excitotoxicity in vulnerable target neurons (trigeminal). Parallel studies on ALS resistant abducens motoneurons will provide valuable information on the mechanism(s) responsible for the differential vulnerability of motoneurons to the disease process. Our experiments will use a unique brainstem slice preparation that contains in close proximity, trigeminal and abducens motoneurons, as well as sensory Mes V neuronal cell bodies. Direct comparisons of changes in potassium and calcium channel properties, calcium concentration changes and assessment of mitochondrial function between different neuron types in control and SOD1 mutant animals will be obtained and provide valuable information on the pathogenesis of the ALS. PUBLIC HEALTH RELEVANCE: Amyotrophic lateral sclerosis (ALS) is a fatal neuro-degenerative disease clinically characterized by progressive loss of muscle force and difficulty in swallowing and chewing, for which there is no cure. Neuronal ion channels produce the electrical signals necessary for proper sensory-motor function and abnormalities in these channels can lead to a variety of disorders. Detection of presymptomatic changes in ion channel function using animal models for ALS could lead to identification of physiological targets that can be used in development of therapeutic strategies to prolong life of those with ALS.