Unrelated course of subthalamic nucleus and globus pallidus neuronal activities across vigilance states in the rat

The pallido-subthalamic pathway powerfully controls the output of the basal ganglia circuitry and has been implicated in movement disorders observed in Parkinson's disease (PD). To investigate the normal functioning of this pathway across the sleep-wake cycle, single-unit activities of subthalamic nucleus (STN) and globus pallidus (GP) neurons were examined, together with cortical electroencephalogram and nuchal muscular activity, in non-anaesthetized head-restrained rats. STN neurons shifted from a random discharge in wakefulness (W) to a bursting pattern in slow wave sleep (SWS), without any change in their mean firing rate. This burst discharge occurred in the 1-2 Hz range, but was not correlated with cortical slow wave activity. In contrast, GP neurons, with a mean firing rate higher in W than in SWS, exhibited a relatively regular discharge whatever the state of vigilance. During paradoxical sleep, both STN and GP neurons increased markedly their mean firing rate relative to W and SWS. Our results are not in agreement with the classical 'direct/indirect' model of the basal ganglia organization, as an inverse relationship between STN and GP activities is not observed under normal physiological conditions. Actually, because the STN discharge pattern appears dependent on coincident cortical activity, this nucleus can hardly be viewed as a relay along the indirect pathway, but might rather be considered as an input stage conveying corticothalamic information to the basal ganglia.     

Serotonin (1A) receptor ligands act on norepinephrine neuron firing through excitatory amino acid and GABA(A) receptors: a microiontophoretic study in the rat locus coeruleus.

It was previously shown that the excitatory effect of the 5-HT(1A) agonist 8-OH-DPAT on firing activity of locus coeruleus (LC) norepinephrine (NE) neurons and the inhibitory action of the 5-HT(1A) antagonist WAY 100,635 are dependent on the presence of 5-HT neurons, whereas the inhibitory action of the 5-HT(2) agonist DOI is not. Using in vivo extracellular unitary recordings performed in anesthetized rats, iontophoretic applications of the excitatory amino acid antagonist kynurenate attenuated the enhancement in firing produced by glutamate and kainate. In contrast, GABA applications decreased the firing activity of NE neurons which was attenuated by the enhancement produced by glutamate and kainate. In contrast, GABA applications decreased the firing activity of NE neurons which was attenuated by the GABA(A) receptor antagonist bicuculline. 8-OH-DPAT (10-60 microg kg(-1), i.v.) produced a dose-dependent enhancement in the firing activity of NE neurons that was abolished in the presence of kynurenate application. The selective 5-HT(1A) receptor antagonist WAY 100,635 (100 microg kg(-1), i.v.) suppressed NE firing which was reversed by the selective 5-HT(2A) antagonist MDL 100,907 (200 microg kg(-1), i.v.). In the presence of bicuculline, the inhibitory effect of WAY 100,635 was blunted. These results suggest that WAY 100,635 mainly attenuates NE neuron firing by blocking inhibitory 5-HT(1A) receptors on glutamatergic neurons, thereby enhancing glutamate release and activating excitatory amino acid receptors, possibly of the kainate subtype, on 5-HT terminals. The ensuing increased 5-HT release would then act on excitatory 5-HT(2A) receptors on GABA neurons that would ultimately mediate the inhibition of NE neurons. The prevention of the excitatory action of 8-OH-DPAT on NE neuron firing by kynurenate is also consistent with this neurocircuitry.     

Intraspinal amino acid neurotransmitter activities are involved in the generation of rhythmic sympathetic nerve discharge in newborn rat spinal cord

Endogenous neurotransmitter activities underlying the sympathetic nerve discharge (SND) generated by newborn rat spinal cord in vitro were investigated using glutamatergic, glycinergic, and GABAergic antagonists. Under control conditions, the SND power spectrum had two major frequency components: synchronous bursting SND (bSND) with power dominant at < 0.1 Hz and quasiperiodic SND (qSND) oscillating at 1-2 Hz. Using high Mg2+ solution (12-24 mM) to block Ca2+-dependent synaptic transmission reversibly abolished SND. An interruption of glutamatergic neurotransmission by CNQX (non-NMDA receptor blocker) or L-AP4 (reducing the synaptic release of glutamate) failed to affect qSND, but consistently reduced bSND. Application of kynurenate, a broad-spectrum ionotropic glutamate receptor blocker, only caused an unstable SND but did not reduce SND. In contrast, strychnine (Stry, glycine receptor antagonist) consistently reduced qSND in a dose-dependent manner. Bicuculline (Bic, GABA(A) receptor antagonist) induced a synchronous bSND of irregular rhythm, which could be further regularized by adding Stry. Bic-induced bSND was reversibly abolished by CNQX or L-AP4. In conclusion, intraspinal glycinergic, GABAergic, and glutamatergic activities are involved in the generation of the spinal cord-derived SND in newborn rats. Intraspinal GABAergic interneurons may tonically inhibit the glutamatergic bursting neurons that generate a synchronous bSND. Activities of these glutamatergic bursting neurons may also be modulated by intraspinal glycinergic interneurons.     

Motor cortical control of internal pallidal activity through glutamatergic and GABAergic inputs in awake monkeys

The internal segment of the globus pallidus (GPi) receives motor-related cortical signals mainly through the striatum, the external segment of the globus pallidus (GPe) and the subthalamic nucleus (STN). The GPi sends its outputs outside the basal ganglia and plays a key role in motor control. Extracellular unit recordings were performed in awake monkeys to explore how glutamatergic STN inputs and GABAergic striatal and GPe inputs control spontaneous activity and how these inputs contribute to motor cortex stimulation-induced responses of GPi neurons. The typical responses of GPi neurons to cortical stimulation consisted of an early excitation, an inhibition and a late excitation. Local applications of the NMDA receptor antagonist 3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid and/or the AMPA/kainate receptor antagonist 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulphonamide in the vicinity of recorded GPi neurons reduced the firing rate, and abolished or attenuated both early and late excitations following cortical stimulation. Local application of the GABA_A receptor antagonist gabazine increased the firing rate, induced oscillatory firings and diminished the cortically induced inhibition. Muscimol or gabazine injection into the STN or GPe also altered the firing rate, and attenuated the late excitation of GPi neurons. The gabazine injection into the STN occasionally induced dyskinesia with significantly decreased GPi activity. These data suggest that the early and late excitations are glutamatergic and induced by the cortico-STN-GPi and cortico-striato-GPe-STN-GPi pathways, respectively. The inhibition is GABAergic and induced by the cortico-striato-GPi pathway. In addition, these inputs are the main factors governing the spontaneous activity of GPi neurons.     

Evidence for a tonic GABA-ergic inhibition of excitatory respiratory-related afferents to presympathetic neurons in the rostral ventrolateral medulla

The effect of blockade of ionotropic GABA and glutamate receptors in the rostral ventrolateral medulla (RVLM) on the relationship between phrenic nerve, splanchnic sympathetic nerve and lumbar sympathetic nerve activities was examined in urethane anesthetized, paralyzed and vagotomized Sprague-Dawley rats. Bilateral microinjection of the GABA-A receptor antagonist, bicuculline (4 mM, 100 nl), into the RVLM dramatically, and almost exclusively, increased the post-inspiratory related discharge in both splanchnic sympathetic nerve and lumbar sympathetic nerve activities and elicited hypertension with fluctuations of arterial pressure phase locked to the discharge of the phrenic nerve. Subsequent bilateral microinjection of kynurenate, a non-selective ionotropic excitatory amino acid receptor antagonist (50 mM, 100 nl), into the RVLM significantly attenuated the sympathoexcitation and hypertension evoked by injection of bicuculline. This was accompanied by an abolition of the post-inspiratory related burst discharge of splanchnic sympathetic nerve and lumbar sympathetic nerve activities. These data suggest that the GABAergic inputs to RVLM tonically inhibit glutamatergic inputs from central respiratory neurons that normally act to increase the firing of presympathetic neurons in the RVLM. Inputs from post-inspiratory neurons appear to be an especially potent excitatory synaptic drive to the presympathetic neurons in the absence of the GABAergic inhibition.     

Effect of microiontophoretic application of dopamine on subthalamic nucleus neuronal activity in normal rats and in rats with unilateral lesion of the nigrostriatal pathway

The subthalamic nucleus (STN) receives dopamine inputs from the substantia nigra but their implication in the pathophysiology of parkinsonism is still debated. Extracellular microrecordings were used to study the effect of microiontophoretic injection of dopamine and the D1 receptor agonist SKF 38393 on the activity of STN neurons in normal and 6-hydroxydopamine-lesioned rats under urethane anaesthesia. Dopamine and SKF induced an increase in the firing rate of the majority of STN neurons in both normal and 6-OHDA rats. In rats with 6-OHDA lesions, the percentage of firing rate increase did not differ from that of controls. When GABA, glutamate and dopamine were all applied to the same individual STN neurons, GABA induced an inhibitory effect and glutamate and dopamine caused an excitatory effect in both groups. This excitatory response was suppressed by the application of GABA. Systemic administration of apomorphine provoked a decrease in the firing rate of STN neurons in rats with 6-OHDA lesions. These results show that dopamine exerts an excitatory influence on STN neurons, suggesting that the inhibitory effect induced by the systemic injection of apomorphine is due to the GABAergic inputs from the globus pallidus as predicted by the current model of basal ganglia organization. In addition, we show that dopamine, GABA and glutamate can act on the same STN neuron and that GABA can reverse the excitatory effect of dopamine and glutamate, suggesting the predominant influence of GABAergic inputs to the subthalamic nucleus.     

Postural and anticonvulsant effects of inhibition of the rat subthalamic nucleus.

The subthalamic nucleus (STN) plays a crucial role as a regulator of basal ganglia outflow by providing excitatory glutamatergic input into the two output nuclei of the basal ganglia, substantia nigra pars reticulata (SNpr), and entopeduncular nucleus. This study examined the effects of suppressing activity in the STN of the awake, behaving rat. Specifically, we evaluated the effects of unilateral and bilateral focal inhibition of STN on posture, locomotion, and susceptibility to limbic motor seizures. Unilateral microinjection of a GABA(A) receptor agonist (muscimol, 200 pmol) into STN produced a site-dependent contralaterally directed postural asymmetry without locomotor activation. This effect differed from responses produced by the same dose of muscimol placed into SNpr, which included locomotor activation in addition to contralaterally directed postural asymmetry. Locomotor activation and postural asymmetry were obtained also after blockade of glutamate transmission in SNpr by the unilateral application of kynurenate (100 nmol). Our observation that STN inhibition did not induce the locomotor activation characteristic of SNpr inhibition suggests that there are glutamatergic inputs to SNpr, other than those from STN, that are responsible for controlling locomotion. Bilateral, but not unilateral, injection of muscimol (200 pmol) into STN protected against limbic motor seizures evoked either by intravenous bicuculline or by focal application of bicuculline into anterior piriform cortex (area tempestas). These results demonstrate that focal inhibition of STN reproduces the postural asymmetry and anticonvulsant actions that are obtained with the inhibition of SNpr. This provides behavioral support for the concept that STN contributes a crucial tonic excitatory (glutamatergic) drive to the rat SNpr.     

Some pharmacological differences between hippocampal excitatory and inhibitory synapses in transmitter release: an in vitro study.

The effects of adenosine, carbachol, and baclofen on synaptic transmission between neurons in cultured rat hippocampal explants were studied using the tight-seal whole cell clamp technique. In the culture, stimulations of neurites cause postsynaptic currents (PSCs) in nearby neurons under voltage-clamp condition. In the presence of 20 microM bicuculline, most PSCs were considered as glutamatergic excitatory postsynaptic currents (EPSCs), because they were blocked by glutamate antagonist, kynurenate at 1 mM. In the presence of 1 mM kynurenate, PSCs seemed to be inhibitory postsynaptic currents mediated by gamma-aminobutyric acid (GABA), because they were blocked by GABA antagonist, bicuculline at 20 microM. Adenosine at 100 microM and carbachol at 10 microM suppressed these EPSCs to about 35% of control. However, adenosine and carbachol at the same concentration did not suppress the IPSCs. Baclofen at 10 microM suppressed both EPSCs and IPSCs significantly (EPSCs: to about 40% of control, IPSCs: to about 30% of control). In contrast, membrane currents elicited by ionophoretically applied glutamate and GABA were not suppressed by 100 microM adenosine, 10 microM carbachol, and 10 microM baclofen. From these results, it is suggested that the pharmacological sensitivities of transmitter release from presynaptic terminals are different between glutamatergic excitatory synapses and GABAergic inhibitory synapses in hippocampal cultures.     

Cultured neurons as model systems for biochemical and pharmacological studies on receptors for neurotransmitter amino acids

By the use of primary cultures of neurons consisting of cerebral cortex interneurons or cerebellar granule cells it is possible to study biochemical and pharmacological aspects of receptors for GABA and glutamate. Cerebellar granule cells have been shown to express both high- and low-affinity GABA receptors. The latter ones develop, however, only when the neurons are treated with GABA or GABA receptor agonists. It is suggested that the high-affinity receptors play a role in the neurotrophic activity of GABA, whereas the low-affinity GABA receptors are involved in the mediation of the inhibitory action of GABA on evoked release of glutamate, which is the neurotransmitter in cerebellar granule cells. Also glutamate receptors have been studied with regard to the 2 types of neurons. Both cerebral cortex neurons (GABAergic) and cerebellar granule cells (glutamatergic) possess glutamate receptors, which mediate an L-glutamate-induced transmitter release. The pharmacological properties of these glutamate receptors are, however, distinctly different for the 2 types of neurons. While cerebral cortex neurons express both quisqualate-, N-methyl-D-aspartate- and kainate-receptors, the cerebellar granule cells have a receptor which is activated only by L-glutamate and L-aspartate.     

Alpha 2 adrenergic receptors on GABAergic,putative sleep-promoting basal forebrain neurons

The basal forebrain plays an important role in the modulation of cortical activity and sleep-wake states. Yet its role must be multivalent as lesions reportedly diminish cortical fast activity and also cortical slow activity along with slow wave sleep (SWS). Basal forebrain cholinergic vs. GABAergic cell groups could differentially influence these processes. By labelling recorded neurons with Neurobiotin (Nb) using the juxtacellular technique and identifying them by immunostaining, we previously found that whereas all cholinergic cells increased their firing, the majority of GABAergic neurons decreased their firing in association with evoked cortical activation in urethane-anaesthetized rats. Here, we examined the possibility that such GABAergic, cortical activation 'off' cells might bear alpha 2 adrenergic receptors (alpha2AR) through which noradrenaline (NA) could inhibit them during cortical activation. First using simple dual-immunostaining for glutamic acid decarboxylase (GAD) and the alpha2AAR, we found that the majority (approximately 60%) of GAD-immunopositive (GAD+) neurons through the magnocellular preoptic nucleus (MCPO) and substantia innominata (SI) were labelled for the alpha2AAR. Second, in urethane-anaesthetized rats, we examined whether Nb-labelled, GAD+ cortical activation 'off' neurons that discharged maximally in association with cortical slow wave activity, were immunopositive for alpha2AAR. We found that all the Nb+/GAD+'off' cells were labelled for the alpha2AAR. Such cells could be inhibited in association with cortical activation and waking when noradrenergic locus coeruleus (LC) neurons discharge and be disinhibited with cortical slow waves and SWS when these neurons become inactive. We thus propose that alpha2AR-bearing GABAergic basal forebrain neurons constitute sleep-active and sleep-promoting neurons.     

Glutamatergic input governs periodicity and synchronization of bursting activity in oxytocin neurons in hypothalamic organotypic cultures

During suckling, oxytocin (OT) neurons display a bursting electrical activity, consisting of a brief burst of action potentials which is synchronized throughout the OT neuron population and which periodically occurs just before each milk ejection in the lactating rat. To investigate the basis of such synchronization, we performed simultaneous intracellular recordings from pairs of OT neurons identified retrospectively by intracellular fluorescent labelling and immunocytochemistry in organotypic slice cultures derived from postnatal rat hypothalamus. A spontaneous bursting activity was recorded in 65% of OT neurons; the remaining showed only a slow, irregular activity. Application of OT triggered bursts in nonbursting neurons and accelerated bursting activity in spontaneously bursting cells. These cultures included rare vasopressinergic neurons showing no bursting activity and no reaction to OT. Bursts occurred simultaneously in all pairs of bursting OT neurons but, as in vivo, there were differences in burst onset, amplitude and duration. Coordination of firing was not due to electrotonic coupling because depolarizing one neuron in a pair had no effect on the membrane potential of its partner and halothane and proprionate did not desynchronize activity. On the other hand, bursting activity was superimposed on volleys of excitatory postsynaptic potentials (EPSPs) which occurred simultaneously in pairs of neurons. EPSPs, and consequently action potentials, were reversibly blocked by the non-NMDA glutamatergic receptor antagonist CNQX. Taken together, these data, obtained from organotypic cultures, strongly suggest that a local hypothalamic network governs synchronization of bursting firing in OT neurons through synchronous afferent volleys of EPSPs originating from intrahypothalamic glutamatergic inputs.     

Activation of cortical interneurons during sleep: an anatomical link to homeostatic sleep regulation?

Although slow wave activity in the EEG has been linked to homeostatic sleep regulation, the neurobiological substrate of sleep homeostasis is not well understood. Whereas cortical neurons typically exhibit reduced discharge rates during slow wave sleep (SWS), a subpopulation of GABAergic interneurons, which express the enzyme neuronal nitric oxide synthase (nNOS), has recently been found to be activated during SWS. The extent of activation of these nNOS neurons is proportional to homeostatic sleep 'drive'. These cells are an exception among cortical interneurons in that they are projection neurons. We propose that cortical nNOS neurons are positioned to influence neuronal activity across widespread brain areas. They could thus provide a long-sought anatomical link for understanding homeostatic sleep regulation.     

Sympathoinhibition after angiotensin receptor blockade in the rostral ventrolateral medulla is independent of glutamate and gamma-aminobutyric acid receptors.

Bilateral blockade of angiotensin (Ang) receptors in the rostral ventrolateral medulla (RVLM) causes a profound fall in arterial pressure. In this study, we tested whether this effect is due to an interaction between Ang receptors and either glutamatergic or gamma-aminobutyric acidergic (GABAergic) synaptic inputs to RVLM sympathoexcitatory neurons. In urethane-anaesthetised rats, bilateral microinjections of the Ang receptor antagonists [Sar1,Thr8]Ang II or [Sar1,Ile8]Ang II into the RVLM pressor region caused large decreases in arterial pressure, heart rate and renal sympathetic nerve activity (RSNA). These responses were not significantly altered following bilateral microinjections into the RVLM of the glutamate receptor antagonist kynurenic acid (4.5 nmol). Furthermore, bilateral injections of kynurenic acid plus the GABA(A) receptor antagonist bicuculline (200 pmol) into the RVLM increased the baseline arterial pressure and RSNA, but did not alter the percentage decreases in these variables evoked by bilateral microinjections of [Sar1,Ile8]Ang II. However, the level of arterial pressure and RSNA following bilateral injections of kynurenic acid, bicuculline and [Sar1,Ile8]Ang II were similar to the levels before injection of any of these compounds. The effectiveness of the microinjections of kynurenic acid and bicuculline into the RVLM was demonstrated by the observation that they virtually abolished the somato-sympathoexcitatory and baroreceptor-sympathoinhibitory reflexes, which are mediated by glutamatergic and GABAergic synapses, respectively, in the RVLM. These results indicate that (1) blockade of Ang receptors greatly reduces the firing rate of RVLM sympathoexcitatory neurons via a mechanism that is independent of glutamatergic or GABAergic neurotransmission, and (2) in the absence of inputs mediated by ionotropic glutamate, GABA(A) and Ang receptors, there are other mechanisms which generate a level of tonic activity in RVLM sympathoexcitatory neurons sufficient to maintain a normal level of sympathetic vasomotor activity.     

Basal forebrain glutamatergic modulation of cortical acetylcholine release

The mediation of cortical ACh release by basal forebrain glutamate receptors was studied in awake rats fitted with microdialysis probes in medial prefrontal cortex and ipsilateral basal forebrain. Repeated presentation of a stimulus consisting of exposure to darkness with the opportunity to consume a sweetened cereal resulted in a transient increase in cortical ACh efflux. This stimulated release was dependent on basal forebrain glutamate receptor activity as intrabasalis perfusion with the ionotropic glutamate receptor antagonist kynurenate (1.0 mM) markedly attenuated darkness/cereal-induced ACh release. Activation of AMPA/kainate receptors by intrabasalis perfusion of kainate (100 microM) was sufficient to increase cortical ACh efflux even under basal (nonstimulated) conditions. This effect of kainate was blocked by coperfusion with the antagonist DNQX (0.1-5.0 mM). Stimulation of NMDA receptors with intrabasalis perfusion of NMDA (50 or 200 microM) did not increase basal cortical ACh efflux. However, perfusion of NMDA in rats following exposure to the darkness/cereal stimulus resulted in a potentiation of both the magnitude and duration of stimulated cortical ACh efflux. Moreover, intrabasalis perfusion of the higher dose of NMDA resulted in a rapid increase in cortical ACh efflux even before presentation of the darkness/cereal stimulus, suggesting an anticipatory change in the excitability of basal forebrain cholinergic neurons. These data demonstrate that basal forebrain glutamate receptors contribute to the stimulation of cortical ACh efflux in response to behavioral stimuli. The specific roles of basal forebrain glutamate receptor subtypes in mediating cortical ACh release differ and depend on the level of activity of basal forebrain cholinergic neurons.     

Activation of subthalamic alpha 2 noradrenergic receptors induces motor deficits as a consequence of neuronal burst firing

The subthalamic nucleus (STN) plays a key role in the pathophysiology of Parkinson's disease. This was demonstrated by the fact that STN neurons express more bursts in animal models of the disease and by the ability of STN inactivation to alleviate motor deficits. However, the origin of the bursts and the causal link between STN bursts and motor deficits remain unknown. The present study aimed to investigate the role of noradrenergic receptor modulation on the firing activity of STN neurons and the impact of this modulation on locomotor activity in sham and 6-hydroxydopamine-lesioned rats. Using selective agonists and antagonists of α1- and α2-adrenergic receptors (AR), we show that local infusion of clonidine, an α2-AR agonist, induced a switch from tonic to bursty pattern without changing the firing rate. This change in the pattern was prevented by the local infusion of idazoxan, an α2-AR antagonist. Furthermore, clonidine injection into the STN reduced locomotor activity in sham and 6-hydroxydopamine-lesioned rats. In contrast, local injection of phenylephrine, an α1-AR agonist, increased the firing rate of STN neurons without changing the firing pattern. In parallel, phenylephrine did not change locomotor activity. This is the first evidence showing the implication of α1-ARs in the modulation of firing rate and α2-ARs in the modulation of the firing pattern of STN neurons. Furthermore, our data provide also evidence that activation of the STN α2-ARs plays a key role in the genesis of subthalamic burst activity, which may be at the origin of motor deficits.     

Differential effects of zinc on glutamatergic and GABAergic neurotransmitter systems in the hippocampus

Approximately 10% of total zinc in the brain exists in synaptic vesicles of glutamatergic neurons; however, the function of vesicular zinc is poorly understood. The presynaptic action of zinc against excitatory and inhibitory neurotransmission was studied in rat hippocampus using in vivo microdialysis. When the hippocampal CA3 region was perfused with 10-300 microM ZnCl(2), the level of glutamate in the perfusate was decreased, whereas the level of gamma-aminobutyric acid (GABA) was increased. Chelation of endogenous zinc with CaEDTA increased the glutamate level in the perfusate but decreased the GABA level, suggesting that zinc released into the synaptic cleft acts differentially on glutamatergic and GABAergic neurons in the CA3 region. The increase of GABA level by zinc was antagonized by 2,3-dioxo-6-nitro-1,2.3,4-tetrahydrobenzo(f)quinoxaline-7-sulphonamide (NBQX), an antagonist of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)/kainate receptors, but not affected by MK801, an antagonist of N-methyl-D-aspartate (NMDA) receptors, and verapamil, a blocker of voltage-dependent calcium channels. The present study suggests that zinc enhances GABA release via potentiation of AMPA/kainate receptors in the CA3 region, followed by a decrease in presynaptic glutamate release in the same region. Zinc seems to be an inhibitory neuromodulator of glutamate release.     

Subthalamic locomotor region is involved in running activity originating in the rat ventromedial hypothalamus

We have previously shown the involvement of the ventromedial nucleus of the hypothalamus (VMH) in inducing running behavior. Stimulation of kainate (KA)-type glutamate receptors in the unilateral VMH of the rat exclusively elicited stereotyped running behavior. However, the neural pathways or functional connections of the VMH neurons involved in the running activity are yet to be elucidated further. In this study we examined whether the subthalamic locomotor region (SLR) is involved in the expression of the running activity originating in the VMH. The multiunit activity (MUA) in the ipsilateral SLR was significantly increased by KA injection into the VMH of urethane-anesthetized animals. Concomitant injection of 6,7-dinitroquioxalline-2,3-dione (DNQX, a KA-type glutamate receptor antagonist) with KA blocked this change in the MUA. Unilateral pre-injection of either kynurenate (non-selective glutamate receptor antagonist), D-2-amino-5-phosphonovalerate (AP5, an NMDA-type glutamate receptor antagonist) or DNQX into the SLR blocked the expression of the running activity induced by KA injection into the ipsilateral VMH. Results from the present study suggest that communication between KA-sensitive efferents from the VMH to glutamatergic pathways acting via NMDA and non-NMDA receptors in the SLR may underlie expression of running behavior originating in the VMH.     

Kainate and NMDA toxicity for cultured developing and adult rat spiral ganglion neurons: further evidence for a glutamatergic excitatory neurotransmission at the inner hair cell synapse.

In the inner ear, the excitatory amino acid glutamate is a proposed neurotransmitter acting at the synapse between hair cells and afferent auditory neurons. Using cultures of 5-day-old rat auditory neurons, we show that the afferent auditory neuronal population can be divided, on the basis of its sensitivity to the neuronotoxic effect of glutamate and its analogs, in at least 3 subpopulations, one responding to N-methyl-D-aspartate (NMDA), one responding to kainate and a third minor one unresponsive to NMDA, kainic acid and glutamate. No toxic effect of quisqualate is observed. The use of specific antagonists (kynurenate and 2-amino-5-phosphonovalerate (DAP-5) demonstrates the specificity of the receptors to the excitatory amino acids on the afferent auditory neurons. Afferent auditory neurons from adult rats can also be cultured and in these preparations only the large neurons are sensitive to glutamate, kainate and NMDA while the small neurons are not responsive, suggesting that a glutamatergic neurotransmission occurs only at this synapse between the inner hair cells and the large radial afferent auditory neurons. We also show that, in vitro, the organ of Corti releases, in response to an increased potassium concentration and in the presence of calcium, a toxic activity for the afferent auditory neurons that is antagonized by kynurenate and DAP-5. Pathophysiological implications are discussed.     

Inhibitory Effects of L-Glutamate on Central Processes of Crustacean Leg Motoneurons

In crustaceans, glutamatergic excitation at the neuromuscular synapse has been extensively studied. Fewer reports exist of the central and possibly inhibitory actions of glutamate on neurons. The present study analyses the response of intracellularly identified motoneurons, which innervate the proximal leg muscles, to local glutamate pressure applications in the neuropil, in an in vitro thoracic preparation of the crayfish Procambarus clarkii. L-Glutamate application always inhibited motoneuron activity, with a decrease in input resistance. The resulting depolarization or hyperpolarization could usually be reversed within 10 mV of the resting potential. The response persisted in neurons pharmacologically isolated with Cd²⁺ or tetrodotoxin. The reversal potential of the response to glutamate was displaced in a low-chloride solution. Similar responses were obtained with GABA. Application of GABA blocked the glutamate response in a competitive manner. Both responses were suppressed by β-guanidino-propionic acid, a competitive antagonist for GABA receptors. This indicates that glutamate activates a chloride-GABA receptor-channel. Micromolar concentrations of picrotoxin reduced both the L-glutamate and the GABA inhibitory responses, thereby unmasking a smaller, picrotoxin-resistant effect of glutamate (but not of GABA), which was excitatory and sensitive to 6,7-dinitroquinoxaline-2,3-dione (DNQX). These results suggest dual and opposite roles for motoneuron glutamatergic connections–a peripheral (well known) net excitatory one and a central net inhibitory one. Direct inhibition of motoneurons by L-glutamatergic neurons is to be expected.     

Synaptic and intrinsic mechanisms shape synchronous oscillations in hippocampal neurons in culture

We have detected spontaneous, synchronous calcium oscillations, associated with variations in membrane potential, in hippocampal neurons maintained in primary culture. The oscillatory activity is synaptically driven, as it is blocked by tetrodotoxin, by the glutamate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and by toxins inhibiting neurotransmitter release from presynaptic nerve endings. Neuronal oscillations do not require for their expression the presence of a polyneuronal network and are not primarily influenced by the gamma-aminobutyric acid (GABA(A)) receptor antagonist picrotoxin, suggesting that they entirely rely on glutamatergic neurotransmission. Synaptic and intrinsic conductances shape the synchronized oscillations in hippocampal neurons. The concomitant activation of N-methyl-D-aspartate (NMDA) receptors and voltage-activated L-type calcium channels allows calcium entering from the extracellular medium and sustaining the long depolarization, which shapes every single calcium wave.