Cellular effects of swim stress in the dorsal raphe nucleus

Swim stress regulates forebrain 5-hydroxytryptamine (5-HT) release in a complex manner and its effects are initiated in the serotonergic dorsal raphe nucleus (DRN). The purpose of this study was to examine the effects of swim stress on the physiology of DRN neurons in conjunction with 5-HT immunohistochemistry. Basic membrane properties, 5-HT(1A) and 5-HT(1B) receptor-mediated responses and glutamatergic excitatory postsynaptic currents (EPSCs) were measured using whole-cell patch clamp techniques. Rats were forced to swim for 15min and 24h later DRN brain slices were prepared for electrophysiology. Swim stress altered the resting membrane potential, input resistance and action potential duration of DRN neurons in a neurochemical-specific manner. Swim stress selectively elevated glutamate EPSC frequency in 5-HT DRN neurons. Swim stress non-selectively reduced EPSC amplitude in all DRN cells. Swim stress elevated the 5-HT(1B) receptor-mediated inhibition of glutamatergic synaptic activity that selectively targeted 5-HT cells. Non-5-HT DRN neurons appeared to be particularly responsive to the effects of a milder handling stress. Handling elevated EPSC frequency, reduced EPSC decay time and enhanced a 5-HT(1B) receptor-mediated inhibition of mEPSC frequency selectively in non-5-HT DRN cells. These results indicate that swim stress has both direct, i.e., changes in membrane characteristics, and indirect effects, i.e., via glutamatergic afferents, on DRN neurons. These results also indicate that there are distinct local glutamatergic afferents to neurochemically specific populations of DRN neurons, and furthermore that these distinct afferents are differentially regulated by swim stress. These cellular changes may contribute to the complex effects of swim stress on 5-HT neurotransmission and/or the behavioral changes underlying the forced swimming test model of depression.     

Immunohistochemical characterisation of Fos-positive cells in brainstem monoaminergic nuclei following intracranial self-stimulation of the medial forebrain bundle in the rat

Fos immunostaining was used as a marker of neuronal activity following intracranial self-stimulation (ICSS) of the medial forebrain bundle (MFB) in the rat, and was combined with immunostaining for tyrosine hydroxylase (TH), serotonin (5-HT), gamma-aminobutyric acid (GABA), or NR1 (one of the glutamate N-methyl- D-aspartate receptor subunits) for purposes of neurochemical identification. ICSS induced a significant but different degree of increase in the number of Fos-immunopositive (Fos+) cells in the six brainstem monoaminergic nuclei examined, which included the ventral tegmental area (VTA), substantia nigra pars compacta (SNc), dorsal raphe nucleus (DR), median raphe nucleus (MR), locus coeruleus (LC), and A7 noradrenaline cells. Densely labelled Fos+ cells were observed in the LC following ICSS, and many of these Fos+ cells were colocalized with TH. Similarly, many of Fos+ cells in the A7 and DR/MR were colocalized with TH and 5-HT, respectively. By contrast, a smaller number of Fos+ cells was detected in the VTA and SNc following the ICSS, and in these regions the majority of Fos+ cells were not colocalized with TH. Although results among regions quantitatively differed, the ICSS induced a significant increase in the number of double-labelled cells (GABA+/Fos+ or NR1+/Fos+) in all of the VTA, DR, and LC, in which the ICSS produced an ipsilaterally weighted increase in Fos-like immunoreactivity. These results suggest that ICSS of the MFB induces differential Fos expression within monoaminergic and GABAergic neurons in brainstem monoaminergic nuclei under modulation by glutamatergic afferents.     

Glutamatergic input is selectively increased in dorsal raphe subfield 5-HT neurons: role of morphology, topography and selective innervation

Characterization of glutamatergic input to dorsal raphe (DR) serotonin (5-HT) neurons is crucial for understanding how the glutamate and 5-HT systems interact in psychiatric disorders. Markers of glutamatergic terminals, vGlut1, 2 and 3, reflect inputs from specific forebrain and midbrain regions. Punctate staining of vGlut2 was homogeneous throughout the mouse DR whereas vGlut1 and vGlut3 puncta were less dense in the lateral wing (lwDR) compared with the ventromedial (vmDR) subregion. The distribution of glutamate terminals was consistent with the lower miniature excitatory postsynaptic current frequency found in the lwDR; however, it was not predictive of glutamatergic synaptic input with local activity intact, as spontaneous excitatory postsynaptic current (sEPSC) frequency was higher in the lwDR. We examined the morphology of recorded cells to determine if variations in dendrite structure contributed to differences in synaptic input. Although lwDR neurons had longer, more complex dendrites than vmDR neurons, glutamatergic input was not correlated with dendrite length in the lwDR, suggesting that dendrite length did not contribute to subregional differences in sEPSC frequency. Overall, glutamatergic input in the DR was the result of selective innervation of subpopulations of 5-HT neurons and was rooted in the topography of DR neurons and the activity of glutamate neurons located within the midbrain slice. Increased glutamatergic input to lwDR cells potentially synergizes with previously reported increased intrinsic excitability of lwDR cells to increase 5-HT output in lwDR target regions. Because the vmDR and lwDR are involved in unique circuits, subregional differences in glutamate modulation may result in diverse effects on 5-HT output in stress-related psychopathology.     

Influence of inhibitory and excitatory inputs on serotonin efflux differs in the dorsal and median raphe nuclei

The dorsal (DRN) and median raphe nuclei (MRN) are two major sources of serotonergic projections to forebrain that are involved in regulation of behavioral state and motor activity, and implicated in affective disorders such as depression and schizophrenia. To investigate afferent influences on serotonergic neurons, this study compared the role of endogenous GABA and glutamate in the DRN and MRN using microdialysis and measurement of locomotor activity in freely behaving rats. Local infusion of the GABA(A) receptor antagonist bicuculline increased serotonin (5-HT) efflux in the DRN but not the MRN. In contrast, infusion of glutamate receptor antagonists produced larger decreases in 5-HT efflux in the MRN compared with the DRN. Moreover, glutamate receptor antagonists attenuated the increase in 5-HT efflux produced by GABA receptor blockade in the DRN. Thus, the disinhibitory effect of GABA blockers could be ascribed in part to an enhanced influence of glutamate. Measurements of locomotor activity indicate that changes in 5-HT were not simply correlated with behavioral activity induced by drug infusion. In summary, the role of inhibitory and excitatory afferents was strikingly different in the DRN and MRN. GABA afferents were the predominant tonic influence on serotonergic neurons in the DRN. In contrast, glutamatergic but not GABAergic afferents had a strong tonic influence on serotonergic neurons in the MRN.     

离体成年非洲爪蟾视顶盖区突触后电流的研究

用盲法对成年非洲爪蟾视顶盖区的自发微突触后电流（mRSCs)进行全细胞膜片钳记录,观察至了突触后膜有分别由ＡＭＰＡ受体和ＧＡＢＡ受体介导的微兴奋性突触后电流(mEPSCs)和微抑制性突触后电流（mIPSCs),mIPSＣ的发放频率远高于mEPSC,ＧＡＢＡ受体的阻断剂荷包牡丹碱（bicuculline,BM）却能使mEPSC的振幅增加。可能是由于成年期ＡＭＰＡ受体介导的兴奋性活动受到同处于突触后膜的抑制性ＧＡＢＡ受体的制约。尽管因ＮＭＤＡ受体的效能大大下降而记录不到自发电流中的ＮＭＤＡ成分,但远高于生理浓度的外源性ＮＭＤＡ可以诱发去极化的宏电流,说明突触后膜的确仍有ＮＭＤＡ受体存在。而muscimol则能诱发超极化宏电流,这也表明突触后膜上有ＧＡＢＡ受体存在。估计各种受体的一系列变化,使关键期特有的可塑性随神经系统的成熟逐渐降低。     

Control of serotonergic neurons in the dorsal raphe nucleus by the lateral hypothalamus

Anatomical evidence indicates the presence of projections from the lateral hypothalamus to serotonergic (5-hydroxytryptamine, 5-HT) neurons of the dorsal raphe nucleus (DR). Using dual probe microdialysis and extracellular recordings in the DR, we show that the application of GABAergic agents in the lateral hypothalamus modulates the activity of 5-HT neurons in the DR. GABA and bicuculline or baclofen, applied in the lateral hypothalamus significantly reduced and increased, respectively, the 5-HT output in the DR. Likewise, the intrahypothalamic application of GABA and bicuculline reduced (14/20 neurons) and increased (8/12 neurons), respectively, the firing rate of 5-HT neurons in the DR. A smaller percentage of neurons, however, were excited by GABA (3/20) and inhibited by bicuculline (1/12). Application of tetrodotoxin in the lateral hypothalamus suppressed the local 5-HT output and reduced that in the DR. The 5-HT output in the DR increased transiently soon after darkness. The hypothalamic application of GABA attenuated and that of bicuculline potentiated this spontaneous change with an efficacy similar to that seen in light conditions. These results indicate that the lateral hypothalamus is involved in the control of 5-HT activity in the DR, possibly through excitatory (major) and inhibitory (minor) inputs.     

mu-Opioids disinhibit and kappa-opioids inhibit serotonin efflux in the dorsal raphe nucleus

The relative importance of GABAergic and glutamatergic afferents in mediating the effects of mu- and kappa-opioids on serotonin (5-HT) efflux in vivo has not been firmly established. Thus, we used microdialysis in the dorsal raphe nucleus (DRN) of freely behaving rats to study the effect of GABA and glutamate receptor antagonists on opioid-induced changes in 5-HT efflux. Infusing the mu-opioid agonist DAMGO (300 microM) increased extracellular 5-HT in the DRN by approximately 70%. During infusion of the GABA(A) receptor blocker bicuculline (100 microM), extracellular 5-HT increased by approximately 250%, and subsequent infusion of DAMGO decreased 5-HT to approximately 70% above the pre-bicuculline baseline. These data are consistent with the hypothesis that mu-opioids disinhibit 5-HT neurons, an effect attenuated by direct inhibition of 5-HT efflux or inhibition of excitatory influences on 5-HT efflux. To further test this hypothesis, glutamate receptor blockers, AP-5 (1 mM) and DNQX (300 microM), were co-infused with DAMGO. The glutamate receptor antagonists prevented decreases in 5-HT elicited by DAMGO in the presence of bicuculline. This indicates that DAMGO inhibits glutamatergic afferents, which partly offsets the disinhibitory influence of mu-opioids on 5-HT efflux. In contrast, the kappa-opioid agonist, U-50,488 (300 microM), decreased 5-HT by approximately 30% in the DRN. Glutamate and GABA receptor antagonists did not block this effect. In conclusion, mu-opioids inhibit GABAergic and glutamatergic afferents, thereby indirectly affecting 5-HT efflux in the DRN. In contrast, kappa-opioids inhibit 5-HT efflux independent of effects on glutamatergic and GABAergic afferents.     

Brain-derived neurotrophic factor upregulates and maintains AMPA receptor currents in neocortical GABAergic neurons

The regulation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type glutamate receptors is implicated in synaptic plasticity. Although we have found that brain-derived neurotrophic factor (BDNF) triggers surface translocation of AMPA receptor proteins, the physiological significance of the BDNF effect remained to be determined. The present immunohistochemical studies revealed that cortical GABAergic neurons exhibited the most striking response to BDNF. Accordingly, we monitored AMPA-triggered currents through GABAergic neurons: Chronic BDNF treatment increased the AMPA-triggered currents but not NMDA-triggered currents in culture. In parallel, the amplitude, but not frequency, of spontaneous miniature excitatory postsynaptic currents (mEPSCs) was elevated in GABAergic neurons. In agreement, BDNF enhanced GABA release triggered by AMPA compared to the amount triggered by high potassium. Conversely, there was a significant decrease in the mEPSC amplitude of GABAergic neurons in heterozygous BDNF-knockout mice. These findings indicate that the neurotrophin enhances the input sensitivity of GABAergic neurons to facilitate their inhibitory function in the neocortex.     

Median raphe serotonergic innervation of medial septum/diagonal band of broca (MSDB) parvalbumin-containing neurons: possible involvement of the MSDB in the desynchronization of the hippocampal EEG.

Activation of median raphe serotonergic neurons results in the desynchronization of hippocampal electroencephalographic (EEG) activity. This could be a direct effect, because serotonin (5-HT) fibers terminate on a specific population of hippocampal interneurons. On the other hand, it could be an indirect action through the medial septum/diagonal band of Broca (MSDB) pacemaker cells, because, in addition to previously described inhibitory effects, excitatory actions of 5-HT have been demonstrated on MSDB gamma-aminobutyric acid (GABA)-containing neurons through 5-HT2A receptors. Electron microscopic double immunostaining for Phaseolus vulgaris-leucoagglutinin (PHA-L) injected into the median raphe (MR) and parvalbumin, choline acetyltransferase, or calretinin as well as double immunostaining for 5-HT and parvalbumin, and colocalization for parvalbumin and 5-HT2A receptors were done in rats. The results demonstrated that: 1) MR axons form perisomatic and peridendritic baskets and asymmetric synaptic contacts on MSDB parvalbumin neurons; 2) these fibers do not terminate on septal cholinergic and calretinin neurons; 3) 5-HT fibers form synapses identical to those formed by PHA-L-immunolabeled axons with parvalbumin neurons; and 4) MSDB parvalbumin cells contain 5-HT2A receptors. These observations indicate that 5-HT has a dual action on the activity of hippocampal principal cells: 1) an inhibition of the input sector by activation of hippocampal GABA neurons that terminate exclusively on apical dendrites of pyramidal cells, and 2) a disinhibition of the output sector of principal neurons. MSDB parvalbumin-containing GABAergic neurons specifically innervate hippocampal basket and chandelier cells. Thus, 5-HT-elicited activation of MSDB GABAergic neurons will result in a powerful inhibition of these GABA neurons.     

Quantitative analysis of glutamatergic innervation of the mouse dorsal raphe nucleus using array tomography

Serotonin (5-hydroxytryptamine, 5-HT) containing neurons located in the dorsal raphe nucleus (DR) comprise the main source of forebrain 5-HT and regulate emotional states in normal and pathological conditions including affective disorders. However, there are many features of the local circuit architecture within the DR that remain poorly understood. DR neurons receive glutamatergic innervation from different brain areas that selectively express three different types of the vesicular glutamate transporter (VGLUT). In this study we used a new high-resolution imaging technique, array tomography, to quantitatively analyze the glutamatergic innervation of the mouse DR. In the same volumetric images, we studied the distribution of five antigens: VGLUT1, VGLUT2, VGLUT3, the postsynaptic protein PSD-95, and a marker for 5-HT cells, the enzyme tryptophan hydroxylase (TPOH). We found that all three populations of glutamatergic boutons are present in the DR; however, the density of paired association between VGLUT2 boutons and PSD-95 was ≈2-fold higher than that of either VGLUT1- or VGLUT3-PSD-95 pairs. In addition, VGLUT2-PSD-95 pairs were more commonly found associated with 5-HT cells than the other VGLUT types. These data support a prominent contribution of glutamate axons expressing VGLUT2 to the excitatory drive of DR neurons. The current study also emphasizes the use of array tomography as a quantitative approach to understand the fine molecular architecture of microcircuits in a well-preserved neuroanatomical context.     

NT-3 regulates BDNF-induced modulation of synaptic transmission in cultured hippocampal neurons

BDNF and NT-3 can modulate the development and plasticity of central synaptic transmission. Although the expression of NT-3 and BDNF in the rodent hippocampus coincides during perinatal development, little is known about possible functional interactions between both neurotrophins in synaptic development. Here, we have investigated the effects of combined long-term application of NT-3 and BDNF on excitatory glutamatergic (mEPSC) and inhibitory GABAergic miniature synaptic currents (mIPSC) in cultured embryonic hippocampal neurons. Our results show that the BDNF-induced twofold increase in mEPSC frequency is abolished by pre-treatment with NT-3. In addition, the NT-3-induced twofold downregulation of mIPSC frequency is reversed by BDNF. Finally, the BDNF-induced increase in c-fos expression is reduced by 50% after pre-treatment with NT-3. In summary, these data suggest an NT-3 controlled modulation of BDNF signalling in differentiating hippocampal neurons.     

Social Stress Alters Inhibitory Synaptic Input to Distinct Subpopulations of Raphe Serotonin Neurons

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Control of the serotonergic system by the medial prefrontal cortex: potential role in the etiology of PTSD and depressive disorders

The prefrontal cortex is involved in an array of higher brain functions that are altered in psychiatric disorders. Serotonergic neurons of the midbrain rapbe nuclei innervate the prefrontal cortex and are the cellular target for drugs used to treat mood disorders such as the selective serotonin (5-HT) reuptake inhibitors. Anatomical evidence supports the existence of projections from the medial prefrontal cortex (mPFC) to the dorsal raphe nucleus (DR). We report on a functional control of the activity of DR 5-HT neurons by projection neurons in the mPFC. The stimulation of the mPFC elicits two types of responses in DR 5-HT neurons, orthodromic excitations and inhibitions. Excitations are mediated by AMPA/KA and NMDA receptors whereas inhibitions are mediated by GABA(A) and 5-HT(1A) receptors. The activation of a subgroup of 5-HT neurons increases 5-HT release which subsequently activates 5-HT(1A) autoreceptors on other 5-HT neurons. GABA(A)-mediated inhibitions involve GABAergic elements in the DR or adjacent areas. Pyramidal neurons of the mPFC co-express postsynaptic 5-HT(1A) (inhibitory) and 5-HT(2A) (excitatory) receptors. Consistent with the above observations, the selective activation of both receptors in mPFC reduced and increased, respectively, the firing activity of DR 5-HT neurons and the 5-HT release in mPFC. Overall, these data indicate that the activity of the 5-HT system is strongly controlled by the mPFC. Thus, the abnormal prefrontal function in post-traumatic stress disorder and depressive patients may induce a disregulation of 5-HT neurons projecting to other brain areas that can underlie the existing symptomatology in these psychiatric disorders.     

Lack of serotonin neurotoxicity after intraraphe microinjection of (+)-3,4-methylenedioxymethamphetamine (MDMA).

Systemic administration of 3,4-methylenedioxymethamphetamine (MDMA) produces depletions of serotonin (5-HT) and its primary metabolite, 5-hydroxyindoleacetic acid (5-HIAA), decreases 5-HT reuptake sites and diminishes tryptophan hydroxylase activity in various forebrain regions. MDMA has been shown to be neurotoxic to the fine fibers originating from dorsal raphe (DR) 5-HT neurons but not the beaded fibers from the median raphe (MR) nucleus. In the present experiment, MDMA was microinjected directly into the DR or MR to determine whether differential neurotoxicity developed in the DR versus MR fiber systems as measured by 5-HT levels and immunocytochemistry. Two weeks following stereotaxic injection with either vehicle or (+)MDMA (50 micrograms base in 2 microliters) into the DR or MR, rat brains were assayed for 5-HT and catecholamine content or 5-HT immunocytochemistry. HPLC analysis revealed no significant changes in monoamine or metabolite concentrations in the hippocampus and striatum of rats administered intra-DR or -MR (+)MDMA. Raphe sections stained for 5-HT also did not reveal any apparent neurotoxicity. A single cerebral injection of (+)MDMA does not produce neurotoxicity to 5-HT neuronal systems originating in the raphe, although neurotoxicity of multiple MDMA injections into these raphe nuclei cannot be ruled out.     

Serotonin 5-HT(2) receptors activate local GABA inhibitory inputs to serotonergic neurons of the dorsal raphe nucleus

The purpose of the present study was to characterize the synaptic currents induced by bath-applied serotonin (5-HT) in 5-HT cells of the dorsal raphe nucleus (DRN) and to determine which 5-HT receptor subtypes mediate these effects. In rat brain slices, 5-HT induced a concentration-dependent increase in the frequency of inhibitory postsynaptic currents (IPSCs) in 5-HT neurons recorded intracellularly in the ventral part of the DRN (EC(50): 86 microM); 5-HT also increased IPSC amplitude. These effects were blocked by the GABA(A) receptor antagonist, bicuculline (10 microM) and by the fast sodium channel blocker, TTX, suggesting that 5-HT had increased impulse flow in local GABAergic neurons. DAMGO (300 nM), a selective mu-agonist, markedly suppressed the increase in IPSC frequency induced by 5-HT (100 microM) in the DRN. A near maximal concentration of the selective 5-HT(2A) antagonist, MDL100,907 (30 nM), produced a large reduction ( approximately 70%) in the increase in IPSC frequency induced by 100 microM 5-HT; SB242,084 (30 nM), a selective 5-HT(2C) antagonist, was less effective ( approximately 24% reduction). Combined drug application suppressed the increase in 5-HT-induced IPSC frequency almost completely, suggesting involvement of both 5-HT(2A) and 5-HT(2C) receptors. Unexpectedly, the phenethylamine hallucinogen, DOI, a partial agonist at 5-HT(2A/2C) receptors, caused a greater increase (+334%) in IPSC frequency than did 5-HT 100 microM (+80%). This result may be explained by an opposing 5-HT(1A) inhibitory effect since the selective 5-HT(1A) antagonist, WAY-100635, enhanced the 5-HT-induced increase in IPSCs. These results indicate that within the DRN-PAG area there may be a negative feedback loop in which 5-HT induces an increase in IPSC frequency in 5-HT cells by exciting GABAergic interneurons in the DRN via 5-HT(2A) and, to a lesser extent, 5-HT(2C) receptors. Increased GABA tone may explain the previous observation of an indirect suppression of firing of a subpopulation of 5-HT cells in the DRN induced by phenethylamine hallucinogens in vivo.     

Morphine history sensitizes postsynaptic GABA receptors on dorsal raphe serotonin neurons in a stress-induced relapse model in rats

The serotonin (5-hydroxytryptamine, 5-HT) system plays an important role in stress-related psychiatric disorders and substance abuse. Previous work has shown that the dorsal raphe nucleus (DR)-5-HT system is inhibited by swim stress via stimulation of GABA synaptic activity by the stress neurohormone corticotropin-releasing factor (CRF). Additionally, the DR 5-HT system is regulated by opioids. The present study tests the hypothesis that the DR 5-HT system regulates stress-induced opioid relapse. In the first experiment, electrophysiological recordings of GABA synaptic activity in 5-HT DR neurons were conducted in brain slices from Sprague-Dawley rats that were exposed to swim stress-induced reinstatement of previously extinguished morphine conditioned place preference (CPP). Behavioral data indicate that swim stress triggers reinstatement of morphine CPP. Electrophysiology data indicate that 5-HT neurons in the morphine-conditioned group exposed to stress had increased amplitude of inhibitory postsynaptic currents (IPSCs), which would indicate greater postsynaptic GABA receptor density and/or sensitivity, compared to saline controls exposed to stress. In the second experiment, rats were exposed to either morphine or saline CPP and extinction, and then 5-HT DR neurons from both groups were examined for sensitivity to CRF in vitro. CRF induced a greater inward current in 5-HT neurons from morphine-conditioned subjects compared to saline-conditioned subjects. These data indicate that morphine history sensitizes 5-HT DR neurons to the GABAergic inhibitory effects of stress as well as to some of the effects of CRF. These mechanisms may sensitize subjects with a morphine history to the dysphoric effects of stressors and ultimately confer an enhanced vulnerability to stress-induced opioid relapse.     

Excitatory afferent modulation of complex spike synchrony

Inferior olivary neurons receive extensive glutamatergic and GABAergic innervation. Yet, because of the membrane properties of olivary neurons these neurotransmitters can produce only small changes in the firing rates of these cells. Moreover, olivary neurons can generate spontaneous spike activity in the absence of excitatory glutamatergic input. These facts suggest that glutamate and GABA have additional roles within the olivocerebellar system beyond simply modulating single cell firing probability. Indeed, one of the characteristics of the olivocerebellar system is its ability to generate synchronous complex spike activity across populations of Purkinje cells. The pattern of synchronous activity changes rapidly, and is thought to reflect the momentary distribution of effective electrotonic coupling between olivary neurons as shaped by afferent input to the inferior olive. However, it also possible that synchronous olivocerebellar activity is the result of synchrony inherent in the afferent activity itself. The issue of the origin of complex spike synchrony, and the role of glutamatergic olivary afferents in modulating its distribution were recently studied using multiple electrode recordings from Purkinje cells. The results of these studies, reviewed here, demonstrate that synchronous complex spike activity occurs in the absence of glutamatergic (and GABAergic) input to the inferior olive, and therefore indicate that synchronization of complex spike activity primarily results from the electrotonic coupling of olivary neurons, rather than from synchronization present within their afferents. Instead of triggering synchronous discharges directly, the results suggest that the function of tonic excitatory activity is to modulate the effective coupling of spike activity between olivary neurons. Blocking glutamate within the inferior olive causes an enhancement of the normal banding pattern of complex spike synchrony, with higher synchrony among parasagittally aligned Purkinje cells and less synchrony between non-aligned cells. This is in contrast to the more uniform synchrony distribution that follows block of GABAergic olivary afferents. Thus, GABA and glutamate play critical, and complementary, roles in determining the patterns of synchronous complex spike activity that are likely central to the functioning of the olivocerebellar system.     

Actions of the sodium channel inhibitor 202W92 on rat midbrain dopaminergic neurons

Excessive glutamatergic activity is implicated in Parkinson's disease (PD) and sodium channel blockade, resulting in inhibition of glutamate release, is a potential therapeutic approach to PD therapy. Beneficial effects of riluzole and lamotrigine have been reported in animal models of PD, but these compounds have relatively low potency as sodium channel inhibitors and also inhibit N and P/Q-type calcium channels. 202W92, a structural analog of lamotrigine, is a potent sodium channel inhibitor, with no effect on N, P/Q-type channels. Here we present the effects of 202W92 on single patch-clamped dopaminergic neurons. 202W92 (> or =10 microM) inhibited spontaneous action potential firing and reduced amplitude and frequency of evoked action potentials. It also inhibited the frequency of 4-aminopyridine (4-AP)- and electrically evoked excitatory postsynaptic currents (EPSCs) and GABAergic inhibitory postsynaptic currents (IPSCs), with >80% inhibition at 10 microM (IC(50) 1.5 microM). EPSC and IPSC amplitudes were partially inhibited. 202W92 did not affect postsynaptic responses to locally applied glutamate and GABA, nor spontaneously occurring mini-IPSCs. These actions of 202W92 are compatible with sodium channel inhibition and depression of transmitter release.     

Excitatory and inhibitory postsynaptic currents of the superior salivatory nucleus innervating the salivary glands and tongue in the rat

The excitatory and inhibitory synaptic inputs to parasympathetic preganglionic neurons in the superior salivatory (SS) nucleus were investigated in brain slices of neonatal (4-8 days old) rat using the whole-cell patch-clamp technique. The SS neurons innervating the submandibular and sublingual salivary glands and innervating the lingual artery in the anterior region of the tongue were identified by retrograde transport of a fluorescent tracer. Whole-cell currents were evoked by electrical stimulation of tissue surrounding the cell. These evoked postsynaptic currents were completely abolished by antagonists for N-methyl-D-aspartate (NMDA) glutamate, non-NMDA glutamate, gamma-aminobutyric acid type A (GABAA), and glycine receptors, suggesting that SS neurons receive glutamatergic excitatory, and GABAergic and glycinergic inhibitory synaptic inputs. In SS neurons for the salivary glands, the ratio of the NMDA component to the total excitatory postsynaptic current (EPSC) was larger than that of the non-NMDA component. This profile was reversed in the SS neurons for the tongue. In SS neurons for the salivary glands, the ratio of the GABAA component to the total IPSC was larger than the ratio of the glycine component to total inhibitory postsynaptic current (IPSC). The decay time constants of the GABAA component were slower than those for glycine. These characteristics of the excitatory and inhibitory inputs may be involved in determining the firing properties of the SS neurons innervating the salivary glands and the tongue.     

Modulation of excitatory synaptic transmission by endogenous glutamate acting on presynaptic group II mGluRs in rat substantia nigra compacta

Excitatory synaptic inputs from the subthalamic nucleus (STN) have been proposed to underlie burst firing of substantia nigra pars compacta (SNc) dopamine (DA) neurons in Parkinson's disease. Given the potential importance of the STN-SNc synapse in health and disease, our goal was to study how transmission at this synapse is regulated. We tested the hypothesis that neurotransmission at STN-SNc synapses is tonically inhibited by endogenous glutamate acting on presynaptic group II metabotropic glutamate receptors (mGluRs). By using whole-cell recording techniques in brain slices, we examined the effect of LY341495, a mGluR antagonist that is most potent at group II mGluRs, on excitatory postsynaptic currents (EPSCs) that either were evoked in SNc DA neurons by stimulation of the STN or were spontaneously occurred in the presence of tetrodotoxin (miniature EPSCs; mEPSCs). LY341495 increased the evoked EPSC amplitude and mEPSC frequency without changing mEPSC amplitude. In contrast, the group III mGluR antagonist UBP1112 failed to increase the evoked EPSC amplitude. An elevation of extracellular glutamate concentration by a glutamate transporter inhibitor, TBOA, suppressed the evoked EPSCs. LY341495, but not UBP1112, partially reversed the TBOA action. The modulations of EPSCs by TBOA and LY341495 were associated with changes in paired-pulse facilitation ratio. Furthermore, TBOA decreased mEPSC frequency, which was partially reversed by LY341495, without affecting mEPSC amplitude. The results indicate that presynaptic group II mGluRs at STN-SNc synapses appear to be partially activated by a basal level of extracellular glutamate and able to sense the change in extracellular glutamate concentration, subsequently modulating synaptic glutamate release.