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Key points Inputs impinging on layer 5 pyramidal neurons perform essential operations as these cells represent one of the most important output carriers of the cerebral cortex. However, the contribution of astrocytes, a type of glial cell, to these operations is poorly documented. Here we found that optogenetic activation of astrocytes in the vicinity of layer 5 in the mouse primary visual cortex induces spiking in local pyramidal neurons through Nav1.6 ion channels and prolongs the responses elicited in these neurons by stimulation of their distal inputs in cortical layer 1. This effect partially involved glutamatergic signalling but relied mostly on the astrocytic calcium-binding protein S100β, which regulates the concentration of calcium in the extracellular space around neurons. These findings show that astrocytes contribute to the fundamental computational operations of the cortex by acting on the ionic environment of neurons. Abstract The most complex cerebral functions are performed by the cortex, whose most important output is carried out by its layer 5 pyramidal neurons. Their firing reflects integration of the sensory and contextual information that they receive. There is evidence that astrocytes influence cortical neuron firing through the release of gliotransmitters such as ATP, glutamate or GABA. These effects have been described at the network and at the synaptic levels, but it is still unclear how astrocytes influence neuron input-output transfer function at the cellular level. Here, we used optogenetic tools coupled with electrophysiological, imaging and anatomical approaches to test whether and how astrocytic activation affected processing of distal inputs to layer 5 pyramidal neurons (L5PNs). We show that optogenetic activation of astrocytes near L5PN cell body prolonged firing induced by distal inputs to L5PNs and potentiated their ability to trigger spikes. The observed astrocytic effects on L5PN firing involved glutamatergic transmission to some extent but relied mostly on release of S100β, an astrocytic Ca2+ -binding protein that decreases extracellular Ca2+ once released. This astrocyte-evoked decrease in extracellular Ca2+ elicited firing mediated by activation of Nav1.6 channels. Our findings suggest that astrocytes contribute to the cortical fundamental computational operations by controlling the extracellular ionic environment.

The most complex cerebral functions are performed by the cortex which most important output is carried out by its layer 5 pyramidal neurons. Their firing reflects integration of sensory and contextual information that they receive. There is evidence that astrocytes influence cortical neurons firing through the release of gliotransmitters such as ATP, glutamate or GABA. These effects were described at the network and at the synaptic levels, but it is still unclear how astrocytes influence neurons input-output transfer function at the cellular level. Here, we used optogenetic tools coupled with electrophysiological, imaging and anatomical approaches to test whether and how astrocytic activation affected processing and integration of distal inputs to layer 5 pyramidal neurons (L5PN). We show that optogenetic activation of astrocytes near L5PN cell body prolonged firing induced by distal inputs to L5PN and potentiated their ability to trigger spikes. The observed astrocytic effects on L5PN firing involved glutamatergic transmission to some extent but relied on release of S100β, an astrocytic Ca2+-binding protein that decreases extracellular Ca2+ once released. This astrocyte-evoked decrease of extracellular Ca2+ elicited firing mediated by activation of Nav1.6 channels. Our findings suggest that astrocytes contribute to the cortical fundamental computational operations by controlling the extracellular ionic environment.Key Points SummaryIntegration of inputs along the dendritic tree of layer 5 pyramidal neurons is an essential operation as these cells represent the most important output carrier of the cerebral cortex. However, the contribution of astrocytes, a type of glial cell to these operations is poorly documented.Here we found that optogenetic activation of astrocytes in the vicinity of layer 5 in the mouse primary visual cortex induce spiking in local pyramidal neurons through Nav1.6 ion channels and prolongs the responses elicited in these neurons by stimulation of their distal inputs in cortical layer 1.This effect partially involved glutamatergic signalling but relied mostly on the astrocytic calcium-binding protein S100β, which regulates the concentration of calcium in the extracellular space around neurons.These findings show that astrocytes contribute to the fundamental computational operations of the cortex by acting on the ionic environment of neurons.

Stimuli that induce rhythmic firing in trigeminal neurons also increase astrocytic coupling and reveal networks that define the boundaries of this particular population. Rhythmic firing depends on astrocytic coupling which in turn depends on S100β. In many nervous functions that rely on the ability of neuronal networks to generate a rhythmic pattern of activity, coordination of firing is an essential feature. Astrocytes play an important role in some of these networks, but the contribution of astrocytic coupling remains poorly defined. Here we investigate the modulation and organization of astrocytic networks in the dorsal part of the trigeminal main sensory nucleus (NVsnpr), which forms part of the network generating chewing movements. Using whole-cell recordings and the dye coupling approach by filling a single astrocyte with biocytin to reveal astrocytic networks, we showed that coupling is limited under resting conditions, but increases importantly under conditions that induce rhythmic firing in NVsnpr neurons. These are: repetitive electrical stimulation of the sensory inputs to the nucleus, local application of NMDA and decrease of extracellular Ca2+ . We have previously shown that rhythmic firing induced in NVsnpr neurons by these stimuli depends on astrocytes and their Ca2+ -binding protein S100β. Here we show that extracellular blockade of S100β also prevents the increase in astrocytic coupling induced by local application of NMDA. Most of the networks were small and remained confined to the functionally distinct area of dorsal NVsnpr. Disrupting coupling by perfusion with the nonspecific gap junction blocker, carbenoxolone or with GAP26, a selective inhibitor of connexin 43, mostly expressed in astrocytes, abolished NMDA-induced rhythmic firing in NVsnpr neurons. These results suggest that astrocytic coupling is regulated by sensory inputs, necessary for neuronal bursting, and organized in a region specific manner.

Dendritic defects occur in neurodegenerative diseases accompanied by axonopathy, yet the mechanisms that regulate these pathologic changes are poorly understood. Using Thy1-YFPH mice subjected to optic nerve axotomy, we demonstrate early retraction of retinal ganglion cell (RGC) dendrites and selective loss of mammalian target of rapamycin (mTOR) activity, which precede soma loss. Axonal injury triggered rapid upregulation of the stress-induced protein REDD2 (regulated in development and DNA damage response 2), a potent inhibitor of mTOR. Short interfering RNA-mediated REDD2 knockdown restored mTOR activity and rescued dendritic length, area and branch complexity in a rapamycin-dependent manner. Whole-cell recordings demonstrated that REDD2 depletion leading to mTOR activation in RGCs restored their light response properties. Lastly, we show that REDD2-dependent mTOR activity extended RGC survival following axonal damage. These results indicate that injury-induced stress leads to REDD2 upregulation, mTOR inhibition and dendrite pathology causing neuronal dysfunction and subsequent cell death.

KEY POINTS: GABA is an essential molecule for sensory information processing. It is usually assumed to be released by neurons. Here we show that in the dorsal horn of the spinal cord, astrocytes respond to glutamate by releasing GABA. Our findings suggest a novel role for astrocytes in somatosensory information processing. ABSTRACT: Astrocytes participate in neuronal signalling by releasing gliotransmitters in response to neurotransmitters. We investigated if astrocytes from the dorsal horn of the spinal cord of adult red‐eared turtles (Trachemys scripta elegans) release GABA in response to glutamatergic receptor activation. For this, we developed a GABA sensor consisting of HEK cells expressing GABA(A) receptors. By positioning the sensor recorded in the whole‐cell patch‐clamp configuration within the dorsal horn of a spinal cord slice, we could detect GABA in the extracellular space. Puff application of glutamate induced GABA release events with time courses that exceeded the duration of inhibitory postsynaptic currents by one order of magnitude. Because the events were neither affected by extracellular addition of nickel, cadmium and tetrodotoxin nor by removal of Ca(2+), we concluded that they originated from non‐neuronal cells. Immunohistochemical staining allowed the detection of GABA in a fraction of dorsal horn astrocytes. The selective stimulation of A∂ and C fibres in a dorsal root filament induced a Ca(2+) increase in astrocytes loaded with Oregon Green BAPTA. Finally, chelating Ca(2+) in a single astrocyte was sufficient to prevent the GABA release evoked by glutamate. Our results indicate that glutamate triggers the release of GABA from dorsal horn astrocytes with a time course compatible with the integration of sensory inputs.

The basic pattern of rhythmic jaw movements produced during mastication is generated by a neuronal network located in the brainstem and referred to as the masticatory central pattern generator (CPG). This network composed of neurons mostly associated to the trigeminal system is found between the rostral borders of the trigeminal motor nucleus and facial nucleus. This review summarizes current knowledge on the anatomical organization, the development, the connectivity and the cellular properties of these trigeminal circuits in relation to mastication. Emphasis is put on a population of rhythmogenic neurons in the dorsal part of the trigeminal sensory nucleus. These neurons have intrinsic bursting capabilities, supported by a persistent Na(+) current (I(NaP)), which are enhanced when the extracellular concentration of Ca(2+) diminishes. Presented evidence suggest that the Ca(2+) dependency of this current combined with its voltage-dependency could provide a mechanism for cortical and sensory afferent inputs to the nucleus to interact with the rhythmogenic properties of its neurons to adjust and adapt the rhythmic output. Astrocytes are postulated to contribute to this process by modulating the extracellular Ca(2+) concentration and a model is proposed to explain how functional microdomains defined by the boundaries of astrocytic syncitia may form under the influence of incoming inputs.

De nombreuses pratiques utilisees pour traiter les douleurs temporomandibulaires et les syndromes apparentes sont fondees sur le modele du « cercle vicieux » qui suppose un lien reciproque entre la douleur chronique et le niveau d’activite musculaire. Le bilan des donnees cliniques et experimentales ne supporte pas ce modele et suggere que la douleur diminue, plutot qu’elle n’augmente, la force de contraction maximale. Selon le modele propose d’adaptation a la douleur, cet effet s’obtient par une diminution de l’activite des muscles agonistes et une augmentation de celle des antagonistes. A la lumiere de ces donnees, de nouvelles approches doivent etre envisagees.

Brainstem networks generating the respiratory rhythm in lampreys are still not fully characterized. In this study, we described the patterns of respiratory activities and we identified the general location of underlying neural networks. In a semi-intact preparation including the brain and gills, rhythmic discharges were recorded bilaterally with surface electrodes placed over the vagal motoneurons. The main respiratory output driving rhythmic gill movements consisted of short bursts (40.9+/-15.6 ms) of discharge occurring at a frequency of 1.0+/-0.3 Hz. This fast pattern was interrupted by long bursts (506.3+/-174.6 ms) recurring with an average period of 37.4+/-24.9 s. After isolating the brainstem by cutting all cranial nerves, the frequency of the short respiratory bursts did not change significantly, but the slow pattern was less frequent. Local injections of a glutamate agonist (AMPA) and antagonists (6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) or D,L-amino-5-phosphonopentanoic acid (AP5)) were made over different brainstem regions to influence respiratory output. The results were similar in the semi-intact and isolated-brainstem preparations. Unilateral injection of AP5 or CNQX over a rostral rhombencephalic region, lateral to the rostral pole of the trigeminal motor nucleus, decreased the frequency of the fast respiratory rhythm bilaterally or stopped it altogether. Injection of AMPA at the same site increased the rate of the fast respiratory rhythm and decreased the frequency of the slow pattern. The activity recorded in this area was synchronous with that recorded over the vagal motoneurons. After a complete transverse lesion of the brainstem caudal to the trigeminal motor nucleus, the fast rhythm was confined to the rostral area, while only the slow activity persisted in the vagal motoneurons. Our results support the hypothesis that normal breathing depends on the activity of neurons located in the rostral rhombencephalon in lampreys, whereas the caudal rhombencephalon generates the slow pattern.

Proper function of all excitable cells depends on ion homeostasis. Nowhere is this more critical than in the brain where the extracellular concentration of some ions determines neurons' firing pattern and ability to encode information. Several neuronal functions depend on the ability of neurons to change their firing pattern to a rhythmic bursting pattern, whereas, in some circuits, rhythmic firing is, on the contrary, associated to pathologies like epilepsy or Parkinson's disease. In this review, we focus on the four main ions known to fluctuate during rhythmic firing: calcium, potassium, sodium, and chloride. We discuss the synergistic interactions between these elements to promote an oscillatory activity. We also review evidence supporting an important role for astrocytes in the homeostasis of each of these ions and describe mechanisms by which astrocytes may regulate neuronal firing by altering their extracellular concentrations. A particular emphasis is put on the mechanisms underlying rhythmogenesis in the circuit forming the central pattern generator (CPG) for mastication and other CPG systems. Finally, we discuss how an impairment in the ability of glial cells to maintain such homeostasis may result in pathologies like epilepsy and Parkinson's disease.

Little is known about the effects of undernutrition on the specific muscles and neuronal circuits involved in mastication. The aim of this study was to document the effects of neonatal low-protein diet on masticatory efficiency. Newborn rats whose mothers were fed 17 % (nourished (N),n60) or 8 % (undernourished (U),n56) protein were compared. Their weight was monitored and their masticatory jaw movements were video-recorded. Whole-cell patch-clamp recordings were performed in brainstem slice preparations to investigate the intrinsic membrane properties andN-methyl-d-aspartate-induced bursting characteristics of the rhythmogenic neurons (N,n43; U,n39) within the trigeminal main sensory nucleus (NVsnpr). Morphometric analysis (N,n4; U,n5) were conducted on masseteric muscles serial cross-sections. Our results showed that undernourished animals had lower numbers of masticatory sequences (P=0·049) and cycles (P=0·045) and slower chewing frequencies (P=0·004) (N,n32; U,n28). Undernutrition reduced body weight but had little effect on many basic NVsnpr neuronal electrophysiological parameters. It did, however, affect sag potentials (PP=0·005) that influence firing pattern. Undernutrition delayed the appearance of bursting and reduced the propensity to burst (P=0·002), as well as the bursting frequency (P=0·032). Undernourished animals showed increased and reduced proportions of fibre type IIA (PPPPPP

Communication between neurons rests on their capacity to change their firing pattern to encode different messages. For several vital functions, such as respiration and mastication, neurons need to generate a rhythmic firing pattern. Here we show in the rat trigeminal sensori-motor circuit for mastication that this ability depends on regulation of the extracellular Ca(2+) concentration ([Ca(2+)]e) by astrocytes. In this circuit, astrocytes respond to sensory stimuli that induce neuronal rhythmic activity, and their blockade with a Ca(2+) chelator prevents neurons from generating a rhythmic bursting pattern. This ability is restored by adding S100β, an astrocytic Ca(2+)-binding protein, to the extracellular space, while application of an anti-S100β antibody prevents generation of rhythmic activity. These results indicate that astrocytes regulate a fundamental neuronal property: the capacity to change firing pattern. These findings may have broad implications for many other neural networks whose functions depend on the generation of rhythmic activity.

There is increasing evidence that a subpopulation of neurons in the dorsal principal sensory trigeminal nucleus are not simple sensory relays to the thalamus but may form the core of the central pattern generating circuits responsible for mastication. In this paper, we used whole cell patch recordings in brain stem slices of young rats to show that these neurons have intrinsic bursting abilities that persist in absence of extracellular Ca2+. Application of different K+ channel blockers affected duration and firing rate of bursts, but left bursting ability intact. Bursting was voltage dependent and was abolished by low concentrations of Na+ channel blockers. The proportion of bursting neurons increased dramatically in the second postnatal week, in parallel with profound changes in several electrophysiological properties. This is the period in which masticatory movements appear and mature. Bursting was associated with the development of an afterdepolarization that depend on maturation of a persistent sodium conductance ( INaP). An interesting finding was that the occurrence of bursting and the magnitude of INaP were both modulated by the extracellular concentration of Ca2+. Lowering extracellular [Ca2+] increased both INaP and probability of bursting. We suggest that these mechanisms underlie burst generation in mastication and that similar processes may be found in other motor pattern generators.

Mastication results from the interaction of an intrinsic rhythmical neural pattern and sensory feedback from the mouth, muscles and joints. The pattern is matched to the physical characteristics of food, but also varies with age. There are large differences in masticatory movements among subjects. The intrinsic rhythmical pattern is generated by an assembly of neurons called a central pattern generator (CPG) located in the pons and medulla. The CPG receives inputs from higher centers of the brain, especially from the inferio-lateral region of the sensorimotor cortex and from sensory receptors. Mechanoreceptors in the lips and oral mucosa, in muscles, and in the periodontal ligaments around the roots of the teeth have particularly powerful effects on movement parameters. The central pattern generator includes a core group of neurons with intrinsic bursting properties, as well as a variety of other neurons that receive inputs from oral and muscle spindle afferents. Reorganization of subpopulations of neurons within the CPG underlies changes in movement pattern. In addition to controlling motoneurons supplying the jaw, tongue, and facial muscles, the CPG also modulates reflex circuits. It is proposed that these brainstem circuits also participate in the control of human speech. Learning outcomes: Readers will be able to: (1) describe the general location and function of the central pattern generator for mastication, (2) identify the primary nuclei involved in the central pattern generator for mastication, (3) describe the general interactions among the central pattern generators of speech, mastication, respiration, and locomotion, and (4) compare/relate the brainstem systems controlling mastication and speech.

Three series of experiments were carried out to characterize interneurons located within the trigeminal motor nucleus of young rats aged 5-24 days. Cholera toxin injections were made bilaterally into the masseter and, sometimes, digastric muscles to label motoneurons. In the first set of experiments, thick slices were taken from the pontine brainstem and cholera toxin-positive and cholera toxin-negative neurons located inside the trigeminal motor nucleus were filled with biocytin through whole-cell recording patch electrodes. Positively identified motoneurons (cholera toxin+) of various shapes and sizes always had a thick, unbranched axon that entered the motor root following a tight zigzag course. Many cholera toxin-negative neurons were also classified as motoneurons after biocytin filling based on this particularity of their axon. These are probably either fusimotor motoneurons or motoneurons supplying other jaw muscles. The cholera toxin-negative neurons classified as interneurons differed markedly from motoneurons in that they had thin, usually branched axons that supplied the ipsilateral reticular region surrounding the trigeminal motor nucleus (peritrigeminal area), the main trigeminal sensory nucleus, the trigeminal mesencephalic nucleus, the medial reticular formation of both sides, and the contralateral medial peritrigeminal area. Most often, their dendrites were arranged in bipolar arbors that extended beyond the borders of the trigeminal motor nucleus into the peritrigeminal area. Immunohistochemistry against glutamate, GABA and glycine was used to further document the nature and distribution of putative interneurons. Immunoreactive neurons were uniformly distributed throughout the rostro-caudal extent of the trigeminal motor nucleus. Their concentration seemed greater toward the edges of the nucleus and they were scarce in the digastric motoneuron pool. Glutamate- outnumbered GABA- and glycine-immunoreactive neurons. There was no clear segregation between the three populations. In the final experiment, 1,1'-dioctadecyl-3,3,3',3'-tetra-methylindocarbocyanine perchlorate crystals were inserted into one trigeminal motor nucleus in thick slices and allowed to diffuse for several weeks. This procedure marked commissural fibers and interneurons in the contralateral trigeminal motor nucleus. Together these results conclusively support the existence of interneurons in the trigeminal motor nucleus.

A population of neurons in the trigeminal principal sensory nucleus (NVsnpr) fire rhythmically during fictive mastication induced in the in vivo rabbit. To elucidate whether these neurons form part of the central pattern generator (CPG) for mastication, we performed intracellular recordings in brainstem slices taken from young rats. Two cell types were defined, nonbursting (63%) and bursting (37%). In response to membrane depolarization, bursting cells, which dominated in the dorsal part of the NVsnpr, fired an initial burst followed by single spikes or recurring bursts. Non-bursting neurons, scattered throughout the nucleus, fired single action potentials. Microstimulation applied to the trigeminal motor nucleus (NVmt), the reticular border zone surrounding the NVmt, the parvocellular reticular formation or the nucleus reticularis pontis caudalis (NPontc) elicited a postsynaptic potential in 81% of the neurons tested for synaptic inputs. Responses obtained were predominately excitatory and sensitive to glutamatergic antagonists DNQX and/or APV. Some inhibitory and biphasic responses were also evoked. Bicuculline methiodide or strychnine blocked the IPSPs indicating that they were mediated by GABA(A) or glycinergic receptors. About one-third of the stimulations activated both types of neurons antidromically, mostly from the masseteric motoneuron pool of NVmt and dorsal part of NPontc. In conclusion, our new findings show that some neurons in the dorsal NVsnpr display both firing properties and axonal connections which support the hypothesis that they may participate in masticatory pattern generation. Thus, the present data provide an extended basis for further studies on the organization of the masticatory CPG network.

We propose that mastication is the result of the interaction of an intrinsic rhythmical pattern of neural commands and sensory feedback generated by the interaction of the effector system (muscles, bones, joints, teeth, tongue, lips and cheeks) with food. The main sources of variability in the pattern of human masticatory cycles are the subjects themselves, their age, the type of food being eaten and the position of the cycle within a series of movements. The intrinsic rhythmical pattern of mastication is generated by an assembly of neurons, a Central Pattern Generator (CPG), located at the ponto-bulbar junction of the brainstem. This simple neural program is modified by inputs that descend from higher centres of the brain and by feedback from sensory receptors. We describe the features of the masticatory CPG together with the interaction between the CPG and inputs from muscle spindles and periodontal pressoreceptors, which have especially powerful effects on movement parameters.

In the present study we used the expression of the c-Fos-like protein as a "functional marker" to map populations of brainstem neurons involved in the generation of mastication. Experiments were conducted on urethane-anesthetized and paralyzed rabbits. In five animals (experimental group), rhythmical bouts of fictive masticatory-like motoneuron activity (cumulative duration 60-130 min) were induced by electrical stimulation of the left cortical "masticatory area" and recorded from the right digastric motoneuron pool. A control group of five animals (non-masticatory) were treated in the same way as the experimental animals with regard to surgical procedures, anesthesia, paralysis, and survival time. To detect the c-Fos-like protein, the animals were perfused, and the brainstems were cryosectioned and processed immunocytochemically. In the experimental group, the number of c-Fos-like immunoreactive neurons increased significantly in several brainstem areas. In rostral and lateral areas, increments occurred bilaterally in the borderzones surrounding the trigeminal motor nucleus (Regio h); the rostrodorsomedial half of the trigeminal main sensory nucleus; subnucleus oralis-gamma of the spinal trigeminal tract; nuclei reticularis parvocellularis pars alpha and nucleus reticularis pontis caudalis (RPc) pars alpha. Further caudally-enhanced labeling occurred bilaterally in nucleus reticularis parvocellularis and nucleus reticularis gigantocellularis (Rgc) including its pars-alpha. Our results provide a detailed anatomical record of neuronal populations that are correlated with the generation of the masticatory motor behavior.

Neurons of the dorsal nucleus reticularis pontis caudalis (nPontc) fire rhythmically during fictive mastication, while neurons of the ventral half tend to fire tonically (Westberg et al., 2001). This paper describes the changes in the pattern of rhythmical mastication elicited by stimulation of the sensorimotor cortex during inhibition or excitation of neurons in this nucleus and adjacent parts of nucleus reticularis gigantocellularis (Rgc) in the anaesthetized rabbit. Masticatory movements and electromyographic (EMG) activity of the masseter and digastric muscles produced by cortical stimulation were recorded before, during and after injections of a local anaesthetic (lidocaine) or excitatory amino acid N-methyl-d-aspartate (NMDA) into nPontc and Rgc through a microsyringe with attached microelectrode to record neuronal activity. Lidocaine inhibited local neurons and modified the motor program, and the effects varied with the site of injection. Most injections into the ventral half of nPontc increased cycle duration, digastric burst duration and burst area. The action of lidocaine in dorsal nPontc was more variable, although burst duration and area were often decreased. The effects on the muscle activity were always bilateral. Lidocaine block of the rostromedial part of Rgc had no effect on movements or on EMGs. Injections of NMDA excited local neurons and when injected into ventral nPontc, it completely blocked mastication. Dorsal injections either had no effect or increased cycle frequency, while decreasing burst duration and area. No increases in EMG burst duration or area were observed with NMDA. Our findings suggest that neurons of ventral nPontc tonically inhibit other parts of the central pattern generator during mastication, while dorsal neurons have mixed effects. We incorporated these findings into a new model of the masticatory central pattern generator.

The trigeminal principal sensory nucleus (NVsnpr) contains both trigemino-thalamic neurons and interneurons projecting to the reticular formation and brainstem motor nuclei. Here we describe the inputs and patterns of firing of NVsnpr neurons during fictive mastication in anaesthetized and paralysed rabbits to determine the role that NVsnpr may play in patterning mastication. Of the 272 neurons recorded in NVsnpr, 107 changed their firing patterns during repetitive stimulation of the left or right sensorimotor cortex to induce fictive mastication. Thirty increased their firing tonically. Seventy-seven became rhythmically active, but only 31 fired in phase with mastication. The others discharged in bursts at more than twice the frequency of trigeminal motoneurons. Most rhythmic masticatory neurons were concentrated in the dorsal part, and those which fired during the jaw closing phase of the cycle were confined to the anterior pole of the nucleus. Most of these cells had inputs from muscle spindle afferents, whereas most of those firing during jaw opening had inputs from periodontal receptors. Non-masticatory rhythmical neurons had receptive fields on the lips and face. The majority of rhythmical masticatory units were modulated during fictive mastication evoked by both the left and right cortices and only four changed their phase of firing when switching from one cortex to the other. When coupled with the finding that NVsnpr neurons exhibit spontaneous bursting in vitro[Sandler et al. (1998) Neuroscience, 83, 891], the results described here suggest that neurons of dorsal NVsnpr may form the core of the central pattern generator for mastication.

Sleep bruxism (SB) is reported by 8% of the adult population and is mainly associated with rhythmic masticatory muscle activity (RMMA) characterized by repetitive jaw muscle contractions (3 bursts or more at a frequency of 1 Hz). The consequences of SB may include tooth destruction, jaw pain, headaches, or the limitation of mandibular movement, as well as tooth-grinding sounds that disrupt the sleep of bed partners. SB is probably an extreme manifestation of a masticatory muscle activity occurring during the sleep of most normal subjects, since RMMA is observed in 60% of normal sleepers in the absence of grinding sounds. The pathophysiology of SB is becoming clearer, and there is an abundance of evidence outlining the neurophysiology and neurochemistry of rhythmic jaw movements (RJM) in relation to chewing, swallowing, and breathing. The sleep literature provides much evidence describing the mechanisms involved in the reduction of muscle tone, from sleep onset to the atonia that characterizes rapid eye movement (REM) sleep. Several brainstem structures ( e.g., reticular pontis oralis, pontis caudalis, parvocellularis) and neurochemicals ( e.g., serotonin, dopamine, gamma aminobutyric acid [GABA], noradrenaline) are involved in both the genesis of RJM and the modulation of muscle tone during sleep. It remains unknown why a high percentage of normal subjects present RMMA during sleep and why this activity is three times more frequent and higher in amplitude in SB patients. It is also unclear why RMMA during sleep is characterized by co-activation of both jaw-opening and jaw-closing muscles instead of the alternating jaw-opening and jaw-closing muscle activity pattern typical of chewing. The final section of this review proposes that RMMA during sleep has a role in lubricating the upper alimentary tract and increasing airway patency. The review concludes with an outline of questions for future research.

Numerous evidence suggests that interneurons located in the lateral tegmentum at the level of the trigeminal motor nucleus contribute importantly to the circuitry involved in mastication. However, the question of whether these neurons participate actively to genesis of the rhythmic motor pattern or simply relay it to trigeminal motoneurons remains open. To answer this question, intracellular recordings were performed in an in vitro slice preparation comprising interneurons of the peritrigeminal area (PeriV) surrounding the trigeminal motor nucleus (NVmt) and the parvocellular reticular formation ventral and caudal to it (PCRt). Intracellular and extracellular injections of anterograde tracers were also used to examine the local connections established by these neurons. In 97% of recordings, electrical stimulation of adjacent areas evoked a postsynaptic potential (PSP). These PSPs were primarily excitatory, but inhibitory and biphasic responses were also induced. Most occurred at latencies longer than those required for monosynaptic transmission and were considered to involve oligosynaptic pathways. Both the anatomical and physiological findings show that all divisions of PeriV and PCRt are extensively interconnected. Most responses followed high-frequency stimulation (50 Hz) and showed little variability in latency indicating that the network reliably distributes inputs across all areas. In all neurons but one, excitatory postsynaptic potentials (EPSPs) or inhibitory postsynaptic potentials (IPSPs) were also elicited by stimulation of NVmt, suggesting the existence of excitatory and inhibitory interneurons within the motor nucleus. In a number of cases, these PSPs were reproduced by local injection of glutamate in lieu of the electrical stimulation. All EPSPs induced by stimulation of PeriV, PCRt, or NVmt were sensitive to ionotropic glutamate receptor antagonists 6-cyano-7-dinitroquinoxaline and d,l-2-amino-5-phosphonovaleric acid, while IPSPs were blocked by bicuculline and strychnine, antagonists of GABAA and glycine receptors. Examination of PeriV and PCRt intrinsic properties indicate that they form a fairly uniform network. Three types of neurons were identified on the basis of their firing adaptation properties. These types were not associated with particular regions. Only 5% of all neurons showed bursting behavior. Our results do not support the hypothesis that neurons of PeriV and PCRt participate actively to rhythm generation, but suggest instead that they are driven by rhythmical synaptic inputs. The organization of the network allows for rapid distribution of this rhythmic input across premotoneuron groups.

Primary afferent neurons innervating muscle spindles in jaw-closing muscles have cell bodies in the trigeminal mesencephalic nucleus (NVmes) that are electrically coupled and receive synapses. Each stem axon gives rise to a peripheral branch and a descending central branch. It was previously shown that some spikes generated by constant muscle stretch fail to enter the soma during fictive mastication. The present study examines whether the central axon is similarly controlled. These axons were functionally identified in anaesthetized and paralysed rabbits, and tonic afferent firing was elicited by muscle stretch. For the purpose of comparison, responses were recorded extracellularly both from the somatic region and from the central axon in the lateral brainstem. Two types of fictive masticatory movement patterns were induced by repetitive stimulation of the masticatory cortex and monitored from the trigeminal motor nucleus. Field potentials generated by spike-triggered averaging of action potentials from the spindle afferents were employed to determine their postsynaptic effects on jaw-closing motoneurons. Tonic firing of 32% NVmes units was inhibited during the jaw-opening phase, but spike frequency during closing was almost equal to the control rate during both types of fictive mastication. A similar inhibition occurred during opening in 83% of the units recorded along the central branch. However, firing frequency in these was significantly increased during closing in 94%, probably because of the addition of antidromic action potentials generated by presynaptic depolarization of terminals of the central branch. These additional spikes do not reach the soma, but do appear to excite motoneurons. The data also show that the duration and/or frequency of firing during the bursts varied from one pattern of fictive mastication to another. We conclude that the central axons of trigeminal muscle spindle afferents are functionally decoupled from their stem axons during the jaw-closing phase of mastication. During this phase, it appears that antidromic impulses in the central axons provide one of the inputs from the masticatory central pattern generator (CPG) to trigeminal motoneurons.

The essential elements controlling trigeminal motoneurons during feeding lie between the trigeminal and facial motor nuclei. These include populations of neurons in the medial reticular formation and pre-motoneurons in the lateral brainstem that reorganize to generate various patterns. Orofacial sensory feedback, antidromic firing in spindle afferents and intrinsic properties of motoneurons also contribute to the final masticatory motor output.

Kolta, Arlette. In vitro investigation of synaptic relations between interneurons surrounding the trigeminal motor nucleus and masseteric motoneurons. J. Neurophysiol. 78: 1720–1725, 1997. Because of their many inputs and bilateral projections, interneurons surrounding the trigeminal motor nucleus (MotV) are thought to be very important in control of jaw movements and reflexes. However, their interactions with the trigeminal motoneurons are almost unknown. In the present study an in vitro slice preparation was used to investigate this relationship in rat. The zone bordering MotV has been subdivided into four regions: the supra-, juxta-, and intertrigeminal areas (SupV, JuxtV, and IntV, respectively) and the parvocellular reticular formation ventral and caudal to MotV. Stimulation of all areas evoked short-latency excitatory postsynaptic potentials (EPSPs) in masseteric motoneurons. Frequently the EPSPs masked inhibitory postsynaptic potentials (IPSPs) or were followed by long-lasting inhibitory potentials. Only responses obtained from stimulation of JuxtV and IntV seemed devoid of inhibitory components. The EPSPs were mediated through kainate/α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors, whereas the IPSPs appear to be due to γ-aminobutyric acid and glycine. EPSPs and IPSPs were also recorded in SupV premotor interneurons after stimulation of IntV and MotV, respectively, thus suggesting that reciprocal connections exist between premotor areas and also between premotor interneurons of SupV and inhibitory interneurons located within MotV. It is concluded that the preparation used here will doubtless prove useful for further investigation of the circuitry involved in the bilateral coordination of the jaw.

Field EPSPs were recorded from the CA1 region of hippocampal slices under conditions in which components of the responses other than those generated by AMPA-type glutamate receptors were blocked. Laminar profile analysis indicated that the resultant potentials had separable phases: an early and fast stage followed by a late and slow stage. The location of the fast response was sensitive to stimulation position in the stratum radiatum; i.e., distal stimulation elicited maximum, negative going potentials in the distal stratum radiatum while proximal stimulation recordings were maximal in the proximal segment. The distribution of the late component was largely independent of stimulation electrode position in stratum radiatum. Current source density analysis revealed that the late response had a source in the most distal dendrites (stratum moleculare) and a sink near the cell body layer. It was not accompanied by evident changes in membrane conductance and had a decay time constant similar to the membrane time constant. Stimulation of the afferents to the basal dendrites of the pyramidal cells also resulted in a late response which again had a source in stratum moleculare. These results strongly suggest that the magnitude and waveform of the late component of the AMPA receptor mediated field potential reflects the biophysical properties of the most distal branches of the dendritic arborization. The laminar analyses also show that the late potential is minimal in the mid-stratum radiatum and thus suggest that this site is most appropriate for investigations concerned with the waveform of the fast component of AMPA receptor mediated synaptic response.

Mastication is a vital function that ensures that ingested food is broken down into pieces and prepared for digestion. This review outlines the masticatory behavior in terms of the muscle activation patterns and jaw movements and gives an overview of the organization and function of the trigeminal neuronal circuits that are known to take part in the generation and control of oro-facial motor functions. The basic pattern of rhythmic jaw movements produced during mastication is generated by a Central Pattern Generator (CPG) located in the pons and medulla. Neurons within the CPG have intrinsic properties that produce a rhythmic activity, but the output of these neurons is modified by inputs that descend from the higher centers of the brain, and by feedback from sensory receptors, in order to constantly adapt the movement to the food properties.

Mastication is a vital function that ensures that ingested food is broken down into pieces and prepared for digestion. This review outlines the masticatory behavior in terms of the muscle activation patterns and jaw movements and gives an overview of the organization and function of the trigeminal neuronal circuits that are known to take part in the generation and control of oro-facial motor functions. The basic pattern of rhythmic jaw movements produced during mastication is generated by a Central Pattern Generator (CPG) located in the pons and medulla. Neurons within the CPG have intrinsic properties that produce a rhythmic activity, but the output of these neurons is modified by inputs that descend from the higher centers of the brain, and by feedback from sensory receptors, in order to constantly adapt the movement to the food properties.

Increasing evidence suggests that the dorsal part of the principal sensory nucleus of the trigeminal nerve (NVsnpr) contains a significant core of the central pattern generator (CPG) circuitry required for mastication (Tsuboi et al., 2003). Like many trigeminal brainstem neurons, those of NVsnpr are rhythmically active in phase with fictive mastication in vivo (Tsuboi et al., 2003) and project directly to the trigeminal motoneurons (Kolta et al., 2000), but in contrast with the others, they are the only neurons with intrinsic bursting abilities (Sandler et al., 1998; Brocard et al., 2006) within the minimal area of the brainstem necessary to produce rhythmic activity in trigeminal nerves (Bourque and Kolta, 2001). Development of bursting in NVsnpr neurons closely follows the development of mastication. It is mediated by a persistent Na(+) current (I(NaP)) that is expressed only within a certain membrane potential range and that is modulated by the extracellular Ca(2+) concentration ([Ca(2+)](e)), the lower the concentration, the larger the magnitude of I(NaP). Under physiological [Ca(2+)](e), bursting can also be induced in vitro by repetitive electrical stimulation of the trigeminal sensory tract, which projects massively to NVsnpr or by local applications of N-methyl-d-aspartic acid. Both types of stimuli also depolarize glial cells recorded in NVsnpr and increase coupling between them. Glial cells play a determinant role in setting [Ca(2+)](e) and hence are in a key position to influence NVsnpr neuronal firing pattern.

The act of mastication requires the coordinated activity of jaw, facial, and tongue muscles. The form of movements varies between foods and changes gradually during a single masticatory sequence, in parallel with the physical properties of the food bolus. The basic pattern of mastication is produced by a brainstem central pattern generator (CPG) when tonically activated by inputs from sensory afferents, higher brain regions, or both. Although not required to produce the basic pattern, sensory feedback is largely responsible for adjusting the motor output to changes in food properties, while cortical inputs are thought to play a role in anticipation and determination of the initial patterns of movements.

Background: The phenotype of large diameter sensory afferent neurons changes in several models of neuropathic pain. We asked if similar changes also occur in ‘‘functional’’ pain syndromes. Methodology/Principal Findings: Acidic saline (AS, pH 4.0) injections into the masseter muscle were used to induce persistent myalgia. Controls received saline at pH 7.2. Nocifensive responses of Experimental rats to applications of Von Frey Filaments to the masseters were above control levels 1–38 days post-injection. This effect was bilateral. Expression of c-Fos in the Trigeminal Mesencephalic Nucleus (NVmes), which contains the somata of masseter muscle spindle afferents (MSA), was above baseline levels 1 and 4 days after AS. The resting membrane potentials of neurons exposed to AS (n = 167) were hyperpolarized when compared to their control counterparts (n = 141), as were their thresholds for firing, high frequency membrane oscillations (HFMO), bursting, inward and outward rectification. The amplitude of HFMO was increased and spontaneous ectopic firing occurred in 10% of acid-exposed neurons, but never in Controls. These changes appeared within the same time frame as the observed nocifensive behaviour. Ectopic action potentials can travel centrally, but also antidromically to the peripheral terminals of MSA where they could cause neurotransmitter release and activation of adjacent fibre terminals. Using immunohistochemistry, we confirmed that annulospiral endings of masseter MSA express the glutamate vesicular transporter VGLUT1, indicating that they can release glutamate. Many capsules also contained fine fibers that were labelled by markers associated with nociceptors (calcitonin gene-related peptide, Substance P, P2X3 receptors and TRPV1 receptors) and that expressed the metabotropic glutamate receptor, mGluR5. Antagonists of glutamatergic receptors given together with the 2nd injection of AS prevented the hypersensitivity observed bilaterally but were ineffective if given contralaterally. Conclusions/Significance: Low pH leads to changes in several electrical properties of MSA, including initiation of ectopic action potentials which could propagate centrally but could also invade the peripheral endings causing glutamate release and activation of nearby nociceptors within the spindle capsule. This peripheral drive could contribute both to the transition to, and maintenance of, persistent muscle pain as seen in some ‘‘functional’’ pain syndromes.

Activity of splanchnocranial and orofacial muscles regulates a number of behaviors and plays an important articulatory role in vocalization and speech. This chapter reviews experimental findings on oromotor central pattern generators (CPGs) and associated distributed neural networks in selected animals and humans. These generators control such primary behaviors as respiration, sucking, licking and mastication. A special emphasis is given to features that are relevant for the development and production of speech.

Increasing evidence suggests that the dorsal part of the principal sensory nucleus of the trigeminal nerve (NVsnpr) contains a significant core of the central pattern generator (CPG) circuitry required for mastication ( Tsuboi et al., 2003 ). Like many trigeminal brainstem neurons, those of NVsnpr are rhythmically active in phase with fictive mastication in vivo ( Tsuboi et al., 2003 ) and project directly to the trigeminal motoneurons ( Kolta et al., 2000 ), but in contrast with the others, they are the only neurons with intrinsic bursting abilities ( Sandler et al., 1998 , Brocard et al., 2006 ) within the minimal area of the brainstem necessary to produce rhythmic activity in trigeminal nerves ( Bourque and Kolta, 2001 ). Development of bursting in NVsnpr neurons closely follows the development of mastication. It is mediated by a persistent Na + current ( I NaP ) that is expressed only within a certain membrane potential range and that is modulated by the extracellular Ca 2+ concentration ([Ca 2+ ] e ), the lower the concentration, the larger the magnitude of I NaP . Under physiological [Ca 2+ ] e , bursting can also be induced in vitro by repetitive electrical stimulation of the trigeminal sensory tract, which projects massively to NVsnpr or by local applications of N -methyl- d -aspartic acid. Both types of stimuli also depolarize glial cells recorded in NVsnpr and increase coupling between them. Glial cells play a determinant role in setting [Ca 2+ ] e and hence are in a key position to influence NVsnpr neuronal firing pattern.

Mastication can be triggered by repetitive stimulation of the cortex or of sensory inputs, but is patterned by a brain stem central pattern generator (CPG). This CPG may include the dorsal part of the principal trigeminal sensory nucleus (NVsnpr), where neurons burst repetitively when the extracellular concentration of Ca2+ ([Ca2+]e) drops. We examined the effects of repetitive stimulation of sensory afferents of the trigeminal tract on activity of NVsnpr neurons recorded extracellularly in vitro under physiologic [Ca2+]e (1.6 mM). Spontaneously active cells had either a tonic ( n = 145) or a bursting ( n = 46) firing pattern. Afferent stimulation altered burst duration and/or burst frequency in bursting cells and firing frequency in most tonic cells. In 28% of the latter, the firing pattern switched to rhythmic bursting. This effect could be mimicked by local application of N-methyl-d-aspartate and blocked by APV but not DNQX. Detailed analysis showed that rhythm indices (RIs) of 35 tonic neurons that were negative (nonrhythmic) before stimulation became significantly rhythmic (RI ≥ 0.01) after stimulation. Mean and median bursting frequency of these units were 8.32 ± 0.72 (SE) Hz and 6.25 Hz (range, 2.5–17.5 Hz). In seven instances, two units were recorded simultaneously, and cross-correlation analysis showed that firing of six pairs was rhythmic and synchronized after stimulation. Optimal stimulation parameters for eliciting rhythmic bursting consisted in 500-ms trains of pulses delivered at 40–60 Hz. Together, our results show that repetitive stimulation of sensory afferents in vitro can elicit masticatory-like rhythmic bursting in NVsnpr neurons at physiological [Ca2+]e.

The major motor functions of the trigeminal (V) motor system include biting, chewing (mastication), swallowing, respiration and speech, and several reflexes. This article provides a review of the highly coordinated processes that ensure the proper function of the jaw muscles, with a particular focus on the genesis of rhythmical mastication by the central nervous system and the role of sensory feedback in shaping the parameters of movement. The article also notes significant differences between the V and spinal motor systems.

The trigeminal motor nucleus contains the somata of motoneurons innervating the jaw muscles, but also those of interneurons that we have characterized morphologically and immunohistochemically previously. Here we compare their basic physiological characteristics and synaptic inputs from the peri-trigeminal area (PeriV) to those of motoneurons using whole-cell recordings made with pipettes containing biocytin in brainstem slices of rats that had a tracer injected into their masseters. Values for input resistance, spike duration and overall duration of afterhyperpolarization (AHP) were greater for interneurons than for motoneurons. Some interneurons (44%) and motoneurons (33%) had an outward rectification during depolarization. Hyperpolarization-induced inward rectification was seen predominantly in interneurons (85% vs. 31% for motoneurons). Few interneurons (15%) showed depolarization and time-dependent firing frequency accommodation, while half (52%) of the motoneurons did. Rebound excitation at the offset of hyperpolarization was more common in interneurons than in motoneurons (62% vs. 34%). Both populations received synaptic inputs from PeriV. These inputs were predominantly excitatory and were mediated by non-N-methyl-d-aspartate glutamatergic receptors. Response latencies and rise times of the evoked potentials were longer in interneurons than in motoneurons, suggesting that some of the inputs to interneurons could be polysynaptic and/or occurring at distal dendritic locations. Miniature synaptic events could be seen in about half of the neurons in both populations. These results suggest that interneurons can be clearly distinguished from motoneurons on the basis of some electrophysiological properties like the input resistance and spike and AHP durations, and the kinetics of their synaptic inputs from adjacent areas.

Mammalian mastication results from the interaction of an intrinsic rhythmical neural pattern and sensory feedback generated by the interaction of the effecter system (muscles, bones, joints, teeth, soft tissues) with food. The main variables that explain variation in the pattern of human mastication are the subjects themselves, their age, the type of food being eaten, and time during a sequence of movements. The intrinsic pattern of mastication is generated by a central pattern generator (CPG) located in the pons and medulla. The output of the CPG is modified by inputs that descend from higher centers of the brain and by feedback from sensory receptors. Intraoral touch receptors, muscle spindles in the jaw-closing muscles, and specialized mechanoreceptors in the periodontal ligament have especially powerful effects on movement parameters.