Dopamine inhibits voltage-activated calcium channel currents in rat pars intermedia pituitary cells

Several lines of evidence suggest that dopamine acts as a neurotransmitter that inhibits both hormone secretion and electrical activity in pituitary intermediate cells (melanotrophs). In this study we examined the effects of exogenously applied dopamine on voltage activated calcium currents recorded with the whole-cell mode of the patch-clamp technique from short-term primary cultures of melanotrophs. Two types of calcium currents were distinguished by their voltage dependence and kinetics of inactivation similar to the low voltage-activated currents (LVA; or T-type) and high voltage-activated currents (HVA; N&L-types) of calcium currents. Exogenously applied dopamine (2-20 microM) reversibly reduced both LVA and HVA types of calcium currents. Evidence for these results came from experiments in which LVA and HVA calcium currents were separated by stepping to different membrane potentials from a fixed holding potential (Vh) or by changing Vh. These results suggest that dopamine can regulate the entry of calcium into melanotrophs by acting on at least two different populations of calcium channels thereby affecting hormone secretion and electrical activity.     

Presynaptic calcium currents evoking quantal transmission from avian ciliary ganglion neurons

Using whole-cell patch clamp techniques, we simultaneously recorded presynaptic Ca++ current and excitatory postsynaptic currents (EPSCs) from avian neuromuscular junctions in culture. Quantal synaptic transmission was proportional to evoked presynaptic Ca++ current except with large stimuli, which evoked bursts of quanta, reflecting a shift to synchronized release. Synaptic delay, measured from the onset of presynaptic depolarization to the appearance of the first postsynaptic quantal response, was often greater than 100 msec for weak depolarizations but declined as stimulus intensity was increased. Quantal events evoked by Ca++ tail currents had a mean synaptic delay of 1.67 msec. The single type of presynaptic Ca++ current observed displayed an inactivation time constant of greater than 100 msec and tail currents well fit by a single exponential function.     

Muscarinic Inhibition of Glutamatergic Transmission onto Rat Magnocellular Basal Forebrain Neurons in a Thin-slice Preparation

We have examined excitatory and inhibitory transmission in visually identified rat magnocellular basal forebrain neurons using whole-cell patch-clamp recordings in a thin-slice preparation of the rat brain. In most cells, spontaneous excitatory and inhibitory synaptic activities could be recorded from their resting membrane potential. Following focal stimulation within the basal forebrain nucleus or directly onto visualized neighbouring neurons, postsynaptic currents were elicited in magnocellular basal forebrain cells held at -70 mV (a value close to their resting membrane potential). The synaptic responses were complex, consisting either mainly of excitatory postsynaptic currents (EPSCs), or inhibitory postsynaptic currents (IPSCs), or an EPSC-IPSC sequence. The EPSC component was consistent with the activation of AMPA/KA receptors, as it could be selectively blocked by CNQX. The IPSC component resulted in the activation of GABA_A receptors, and could be blocked by bicuculline. Since GABA-mediated transmissions were not frequently recorded, we focused on the glutamate-mediated transmission. Studies using specific calcium channel blockers suggested that both ω-conotoxin GVIA-sensitive and ω-agatoxin VIA-sensitive calcium channels contribute to the glutamatergic transmission onto magnocellular basal forebrain neurons. Carbachol (0.3–30 μM) had no observable effect on holding current, but produced a dose-dependent inhibition of the amplitude of evoked EPSCs. This cholinergic modulation was mediated by muscarinic receptors, as it could be antagonized by atropine. The inhibitory effect of carbachol on the amplitude of EPSCs could be significantly antagonized by 100 nM methoctramine, an M₂-receptor antagonist. In contrast, only a small degree of antagonism could be obtained with pirenzepine, an M₁-muscarinic receptor antagonist, when present at relatively high concentration of 1 μM. Moreover, the action of carbachol was presynaptic, since the frequency of miniature postsynaptic currents was reduced without affecting their amplitude. In conclusion, the present findings indicate that glutamate-mediated transmission onto magnocellular basal forebrain neurons appeared to involve both N- and P/Q-type calcium channels, and that muscarinic modulation of glutamatergic transmission to MBF neurons is mediated by a presynaptic M₂-muscarinic receptor subtypes.     

Substance P and neurokinin A mediate sensory synaptic transmission in young rat dorsal horn neurons

Spinal nociceptive transmission is mediated by glutamate and neuropeptides such as substance P (SP) and neurokinin A (NKA). The neuropeptide-mediated excitatory postsynaptic potentials (EPSPs) had a slow onset and long duration. Here, we demonstrate SP- and NKA-mediated excitatory postsynaptic currents (EPSCs) in dorsal horn neurons of young rats using whole-cell patch-clamp recording techniques. After complete blockade of glutamate receptor-mediated currents, we observed a small residual EPSC. The residual EPSCs exhibited temporal summation in response to a train of stimulation (six pulses delivered at 10-50 Hz). High intensity stimulation (the same or greater than the stimulation threshold for nociceptive fibers in vivo) was required for evoking these summated EPSCs. Summated EPSCs were attenuated or abolished by capsaicin pretreatment, which depletes SP and NKA from presynaptic terminals; SP and NKA pretreatment; NK(1) or NK(2) receptor antagonists; and inhibition of postsynaptic G proteins. EPSCs were neither blocked by a metabotropic glutamate receptor antagonist nor a gamma-aminobutyric acid(B) receptor antagonist. The summated EPSCs were also sensitive to voltage-gated calcium channel antagonists or mu-opioid receptor activation by DAMGO. The present study provides electrophysiological evidence that suggests the possible contribution of SP and NKA to sensory synaptic transmission between primary afferent fibers and dorsal horn neurons.     

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.     

Exogenous tumor necrosis factor-alpha rapidly alters synaptic and sensory transmission in the adult rat spinal cord dorsal horn

The proinflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) is involved in the generation of inflammatory and neuropathic pain. This study investigated if TNF-alpha has any effect on spinal synaptic and/or sensory transmission by using whole-cell recordings of substantia gelatinosa (SG) neurons in transverse lumbar spinal cord slices of adult rats and by using behavioral tests. After intrathecal administration of TNF-alpha in adult rats, spontaneous hind paw withdrawal behavior and thermal hyperalgesia were rapidly induced (approximately 30 min), while mechanical allodynia slowly developed. Bath application of TNF-alpha (0.1-1 nM, 8 min) depressed peak amplitude of monosynaptic Adelta and C fiber-evoked excitatory postsynaptic currents (EPSCs) without changing in holding currents and input resistances, whereas this application generally potentiated polysynaptic Adelta fiber-evoked EPSCs. Moreover, the frequencies, but not the amplitudes, of spontaneous and miniature EPSCs and spontaneous inhibitory postsynaptic currents were significantly increased by bath-applied TNF-alpha in most of the SG neurons. The effects of TNF-alpha on Adelta/C fiber-evoked monosynaptic and polysynaptic or spontaneous EPSCs were significantly blocked by 5 microM TNF-alpha antagonist that inhibits TNF-alpha binding to its type 1 receptor (TNFR1). Because this study also found high protein expression of TNFR1 in the adult dorsal root ganglion and no change of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) induced whole-cell currents by TNF-alpha, we conclude that presynaptic TNFR1 at Adelta/C primary afferent terminals contributes to the rapid alteration of synaptic transmission in the spinal SG, and the development of abnormal pain hypersensitivity by exogenous TNF-alpha.     

Modulation of spontaneous and evoked EPSCs and IPSCs in optic lobe neurons of cuttlefish Sepia officinalis by the neuropeptide FMRF-amide

The effects of the neuropeptide FMRFa on spontaneous excitatory postsynaptic currents (sEPSCs) and inhibitory postsynaptic currents (sIPSCs), as well as on evoked EPSCs and IPSCs, in two types of neurons within the central optic lobe of cuttlefish were examined using the whole-cell voltage-clamp technique. FMRFa (1-10 micro m) did not affect cell membrane resting potentials, but reversibly reduced both the frequency and amplitude of sEPSCs in neurons within the medulla region of the optic lobe while increasing the frequency and amplitude of their sIPSCs. For centrifugal neurons in the inner granule cell layer of the optic lobe, FMRFa (1-10 micro m) decreased both the frequency and amplitude of sEPSCs. In the presence of tetrodotoxin (0.5 micro m), neither the interevent interval, nor amplitude distributions of the miniature EPSCs or the miniature IPSCs, were affected by FMRFa, implying a presynaptic action of FMRFa on the optic lobe neurons. Bath application of the neuropeptide also abolished or reduced in amplitude the evoked EPSCs and increased the amplitude of evoked IPSCs in optic lobe neurons, showing that FMRFa induced similar effects on evoked as on spontaneous postsynaptic currents. These results demonstrate the complex range of modulatory effects FMRFa can have within central nervous system circuits.     

Whole-cell recordings of spontaneous synaptic currents in medial preoptic neurons from rat hypothalamic slices: mediation by amino acid neurotransmitters

Whole-cell recordings in hypothalamic slices from immature rats were used to test the hypothesis that inhibitory and excitatory amino acid neurotransmitters mediate fast synaptic currents in the medial preoptic area (MPOA). Bicuculline methiodide reversibly blocked spontaneous inhibitory postsynaptic currents (IPSCs), and 6-cyano-2,3-dihydroxy-7-nitroquinoxaline (CNOX) blocked spontaneous excitatory postsynaptic currents (EPSCs). These competitive antagonists act at γ-aminobutyric acid (GABA)_A and (NMDA) glutamate receptors, respectively, thus supporting the hypothesis that these amino acid receptors activate most if not all fast synaptic currents in the MPOA.     

Glutamatergic postsynaptic block by Pamphobeteus spider venoms in crayfish.

The effects of toxins from venom glands of two south american spiders (Pamphobeteus platyomma and P. soracabae) on glutamatergic excitatory synaptic transmission were studied in the neuromuscular junction of the opener muscle of crayfish. The toxins selectively and reversibly blocked both excitatory postsynaptic currents and potentials in a dose-dependent manner. They also reversibly abolished glutamate-induced postsynaptic membrane depolarization. They had no effect on resting postsynaptic membrane conductance nor on postsynaptic voltage-gated currents. The synaptic facilitation and the frequency of miniature postsynaptic potentials were unaffected by the toxins, indicating that presynaptic events were not modified. Picrotoxin, a selective antagonist of the gamma-aminobutyric acid (GABA)A receptor, did not modify toxin effects. We conclude that both toxins specifically block the postsynaptic glutamate receptor-channel complex.     

Control of the temporal fidelity of synaptic transmission by a presynaptic high voltage-activated transient K current

The type of K(+) channels controlling the waveform of the presynaptic spike and synaptic transmission were examined in the lamprey spinal cord. Reticulospinal neuron somata displayed a transient K(+) current with a high voltage-activation and inactivation. This current was selectively blocked by catechol at 100 microM. Reticulospinal axons also displayed a high voltage-activated fast K(+) current sensitive to catechol. The function of this presynaptic high voltage-activated fast K(+) current in controlling synaptic transmission was investigated by using paired intracellular recordings from reticulospinal axons and their targets. Blockade of this current by catechol (100 microM) prolonged the presynaptic spike elicited by a single stimulus leading to a potentiation of the postsynaptic EPSP. Calcium imaging of reticulospinal axons showed an increase in presynaptic calcium transients after blockade of the presynaptic K(+) current by catechol. During high frequency firing, catechol revealed an activity-dependent decrease in the spike duration, which resulted in a depression of synaptic transmission. These results suggest that the presynaptic high voltage-activated transient K(+) current acts to optimize the temporal fidelity of synaptic transmission by minimizing activity-dependent changes in the presynaptic spike waveform and calcium dynamics.     

Reduced presynaptic efficiency of excitatory synaptic transmission impairs LTP in the visual cortex of BDNF-heterozygous mice

The neurotrophin brain-derived neurotrophic factor (BDNF) plays an important role in neuronal survival, axonal and dendritic growth and synapse formation. BDNF has also been reported to mediate visual cortex plasticity. Here we studied the cellular mechanisms of BDNF-mediated changes in synaptic plasticity, excitatory synaptic transmission and long-term potentiation (LTP) in the visual cortex of heterozygous BDNF-knockout mice (BDNF(+/-)). Patch-clamp recordings in slices showed an approximately 50% reduction in the frequency of miniature excitatory postsynaptic currents (mEPSCs) compared to wild-type animals, in the absence of changes in mEPSC amplitudes. A presynaptic impairment of excitatory synapses from BDNF(+/-) mice was further indicated by decreased paired-pulse ratio and faster synaptic fatigue upon prolonged repetitive stimulation at 40 Hz. In accordance, presynaptic theta-burst stimulation (TBS) failed to induce LTP at layer IV to layers II-III synapses during extracellular field-potential recordings in BDNF(+/-) animals. Changes in postsynaptic function could not be detected, as no changes were observed in either the amplitudes of evoked EPSCs, the ratios of AMPA : NMDA currents or the kinetics of evoked AMPA and NMDA EPSCs. In line with this observation, an LTP pairing paradigm that relies on direct postsynaptic depolarization under patch-clamp conditions could be induced successfully in BDNF(+/-) animals. These data suggest that a chronic reduction in the expression of BDNF to nearly 50% attenuates the efficiency of presynaptic glutamate release in response to repetitive stimulation, thereby impairing presynaptically evoked LTP in the visual cortex.     

Excitatory amino acid-evoked membrane currents and excitatory synaptic transmission in lamprey reticulospinal neurons

The characteristics of excitatory amino acid-evoked currents and of excitatory synaptic events have been examined in lamprey Müller neurons using voltage clamp and current clamp recording techniques. Application of glutamate evoked depolarizations associated with a decrease in input resistance. The reversal potential of the responses was -35 mV. Under voltage clamp conditions, a series of excitatory amino acid agonists evoked inward currents associated with little or no increase in baseline current noise. The order of potency of the excitatory amino acid agonists was quisqualate greater than kainate greater than glutamate greater than aspartate, while N-methyl-D-aspartic acid (NMDA) was inactive. Inward currents evoked by glutamate, as well as by kainate and quisqualate were attenuated reversibly by 1 mM kynurenic acid (KYN). In contrast, glutamate-evoked currents were not affected by 100 microM D(-)-2-amino-5-phosphonovaleric acid (APV), a selective NMDA antagonist. Spontaneously occurring and stimulus-evoked excitatory postsynaptic events were antagonized reversibly by 1 mM KYN. At this concentration, KYN had no effect on membrane potential, input resistance, or excitability of the cells. In contrast, excitatory postsynaptic currents were unaffected by APV. It is concluded that both glutamate responses and excitatory synaptic transmission in lamprey Müller neurons are mediated by non-NMDA-type receptors and that these receptors are associated with ionic channels with a low elementary conductance. The combined pharmacological and biophysical characteristics of these responses are therefore different from those previously reported in other preparations. Spontaneous (but not stimulus-evoked) inhibitory synaptic events in Müller neurons were blocked reversibly by 1 mM KYN but not by 100 microM APV, suggesting that excitation of interneurons inhibitory to Müller cells was also mediated by non-NMDA receptors.     

Effects of baclofen on mechanical noxious and innocuous transmission in the spinal dorsal horn of the adult rat: in vivo patch‐clamp analysis

The effects of a GABA_B agonist, baclofen, on mechanical noxious and innocuous synaptic transmission in the substantia gelatinosa (SG) were investigated in adult rats with the in vivo patch‐clamp technique. Under current‐clamp conditions, perfusion with baclofen (10 μm ) on the surface of the spinal cord caused hyperpolarisation of SG neurons and a decrease in the number of action potentials elicited by pinch and touch stimuli applied to the receptive field of the ipsilateral hindlimb. The suppression of action potentials was preserved under blockade of postsynaptic G‐proteins, although baclofen‐induced hyperpolarisation was completely blocked. These findings suggest presynaptic effects of baclofen on the induced action potentials. Under voltage‐clamp conditions, application of baclofen reduced the frequency, but not the amplitude, of miniature excitatory postsynaptic currents (mEPSCs), whereas the GABA_B receptor antagonist CGP55845 increased the frequency of mEPSCs without affecting the amplitude. Furthermore, application of a GABA uptake inhibitor, nipecotic acid, decreased the frequency of mEPSCs; this effect was blocked by CGP55845, but not by the GABA_A antagonist bicuculline. Both the frequency and the amplitude of the pinch‐evoked barrage of excitatory postsynaptic currents (EPSCs) were suppressed by baclofen in a dose‐dependent manner. The frequency and amplitude of touch‐evoked EPSCs was also suppressed by baclofen, but the suppression was significantly smaller than that of pinch‐evoked EPSCs. We conclude that mechanical noxious transmission is presynaptically blocked through GABA_B receptors in the SG, and is more effectively suppressed than innocuous transmission, which may account for a part of the mechanism of the efficient analgesic effects of baclofen.     

Glutamatergic postsynaptic block by Pamphobeteus spider venoms in crayfish

The effects of toxins from venom glands of two south american spiders (Pamphobeteus platyomma and P. soracabae) on glutamatergic excitatory synaptic transmission were studied in the neuromuscular junction of the opener muscle of crayfish. The toxins selectively and reversibly blocked both excitatory postsynaptic currents and potentials in a dose-dependent manner. They also reversibly abolished glutamate-induced postsynaptic membrane depolarization. They had no effect on resting postsynaptic membrane conductance nor on postsynaptic voltage-gated currents. The synaptic facilitation and the frequency of miniature postsynaptic potentials were unaffected by the toxins, indicating that presynaptic events were not modified. Picrotoxin, a selective antagonist of the γ-aminobutyric acid (GABA)_A receptor, did not modify toxin effects. We conclude that both toxins specifically block the postsynaptic glutamate receptor-channel complex.     

Cholinergic, noradrenergic, and serotonergic inhibition of fast synaptic transmission in spinal lumbar dorsal horn of rat

It is known that spinal nociceptive sensory transmission receives descending inhibitory and facilitatory modulation from supraspinal structures. Glutamate is the major fast excitatory transmitter between primary afferent fibers and spinal dorsal horn neurons. In whole-cell patch clamp recordings from dorsal horn neurons in spinal slices, we investigated synaptic mechanisms for inhibitory modulation at the lumbar level of the spinal cord. Application of the cholinergic receptor agonist carbachol produced a dose-dependent inhibition of glutamate-mediated excitatory postsynaptic currents (EPSCs) (IC(50) 13 microM). Postsynaptic injection of two different types of G-protein inhibitors, guanosine 5'-O-2-thiophosphate or guanosine 5'-O-3-thiotriphosphate, blocked the inhibition produced by carbachol. Clonidine, a selective alpha-adrenergic receptor agonist, also produced a dose-dependent inhibition of EPSCs (IC(50) 7 microM) that was reduced by postsynaptic inhibition of G-proteins. The inhibitory effect of serotonin was likewise mediated by postsynaptic G-proteins. Our results suggest that activation of postsynaptic neurotransmitter receptors plays a critical role in inhibition of glutamate mediated sensory responses by acetylcholine, norepinephrine, and serotonin. Our results support the hypothesis that descending sensory modulation may be mediated by multiple neurotransmitter receptors in the spinal cord.     

DAMGO suppresses both excitatory and inhibitory synaptic transmission in supraoptic neurones of mouse hypothalamic slice preparations

Opioid effects on synaptic transmission in the mouse supraoptic nucleus (SON) were investigated using whole-cell, patch-clamp techniques. The mu-opioid receptor agonist, [D-Ala(2), N-Me-Phe(4), Gly(5)-ol]-enkephalin (DAMGO) decreased the amplitude of both evoked excitatory postsynaptic currents (eEPSCs) and inhibitory postsynaptic currents (eIPSCs), and also decreased the frequency of both miniature EPSCs and IPSCs without effect on the amplitude. The selective mu-opioid receptor antagonist, D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH(2), and the nonselective antagonist naloxone, antagonized these inhibitory effects. The application of DAMGO suppressed the amplitude of both the first and second evoked postsynaptic currents with a paired-pulse stimulus protocol, but increased the paired-pulse ratios (second ePSC/first ePSC). DAMGO induced neither inward nor outward currents, and had no significant changes in either glutamate- or GABA-induced currents. When compared with the relatively selective kappa- and delta-opioid receptor agonists dynorphin and [D-Pen(2), D-Pen(5)]-enkephalin, DAMGO showed the most potent inhibitory effects on evoked and miniature postsynaptic currents. Taken together, these results imply that DAMGO strongly suppresses the release of glutamate and GABA via mu-opioid receptors in the mouse SON, and support the involvement of presynaptic regulation by opioids in the control of magnocellular neurosecretory neurones.     

P2X receptor-mediated excitatory synaptic currents in somatosensory cortex

Fast P2X receptor-mediated excitatory postsynaptic current (EPSC) was found in pyramidal neurones of layer V of somatosensory cortex in slices acutely isolated from the brain of 17- to 22-day-old rats. The EPSCs were elicited by field electrical stimulation in the layer VI at 0.1 Hz in the presence of picrotoxin. When the glutamatergic EPSC was blocked by glutamate receptors inhibitors NBQX and D-AP5, a residual EPSC (rEPSC) was recorded from 85% of neurones tested. This rEPSC was not affected by blockers of nicotinic (hexamethonium) and serotonin (Y25130) receptors; however, it was reversibly inhibited by P2X receptors antagonists (NF023, NF279, and PPADS). An application of ATP (20 microM), beta,gamma-methylene ATP (25 microM), and alpha,beta-methylene ATP (20 microM) to acutely isolated pyramidal neurones of layer V evoked inward currents (30 to 400 pA) in 75% of cells tested. We concluded that several subtypes of P2X purinoreceptors participate in synaptic transmission in neocortex.     

Properties of inhibitory and excitatory synapses between hippocampal neurons in very low density cultures

The whole cell patch clamp technique was used to examine the electrophysiological properties of embryonic hippocampal neurons maintained in a very low density (VLD) culture prejparation. The goal of these experiments was to establish the viability of the VLD culture as a model system in which to study regulation of neurotransmission at single monosynaptic connections, in the absence of polysynaptic innervation. Depolarization of neurons in the VLD culture revealed voltage-dependent sodium, calcium, and potassium currents which were blocked with, respectively, tetrodotoxin (TTX), cobalt, and tetraethylammonium and 4-aminopyridine. When pairs of neurons were simultaneously recorded, action potentials evoked in presynaptic neurons elicited either excitatory or inhibitory postsynaptic currents (EPSCs or IPSCs, respectively). The dual component “EPSCs” were due to the activation of both types-of postsynaptic, ionotropic glutamate receptors: N-methyl-D-aspartate (NMDA) and non-NMDA receptors. Evoked IPSCs were due to the activation of postsynaptic γ-aminobutyric acid (GABA) receptors. Both excitatory and inhibitory synapses exhibited short term depression in response to high frequency stimulation, although IPSCs were routinely decreased to a much greater degree than EPSCs. Spontaneous miniature EPSCs and IPSCs were found to persist in TTX, were blocked by the same pharmacological antagonists which blocked evoked responses, increased in frequency in response to hyperosmotic solution, and were unaffected by changes in extracellular calcium concentration. mIPSCS were found to occur at a significantly lower frequency than mEPSCs. These experiments indicated that neurotransmission in the VLD cultures occurs in a manner consistent with the quantal hypothesis and, therefore, the VLD culture is a good model for studying excitatory and inhibitory neurotransmission between isolated pairs of neurons. In addition, these experiments, performed under comparable physiological conditions, demonstrated that there are fundamental differences underlying neurotransmitter release between excitatory and inhibitory neurons. © 1994 Wiley-Liss, Inc.     

Electrophysiological analysis of suprachiasmatic nucleus projections to the ventrolateral preoptic area in the rat

The circadian pacemaker housed in the suprachiasmatic nucleus (SCN) synchronizes daily sleep-wake cycles, presumably by modulating the sleep-wake regulatory system, including ventrolateral preoptic area (VLPO) neurons. We used whole-cell patch-clamp recording to study the projections from the SCN to the VLPO in horizontal slices of rat hypothalamus. Single-pulse stimulation of the SCN region elicited postsynaptic currents (PSCs) in 20 of 66 neurons (30%) recorded within the VLPO region as verified by intracellular biocytin labelling. At a holding potential of -60 mV, the evoked PSCs had an amplitude of 17.6 +/- 3.2 pA (SEM) and a latency of 6.3 +/- 0.5 ms (n = 10). There was a trend for simple excitatory postsynaptic currents (EPSCs) to be evoked in the VLPO cluster, simple inhibitory postsynaptic currents (IPSCs) in the extended VLPO, and a combination of EPSCs and IPSCs in both regions. IPSCs were blocked reversibly by bicuculline (10 microm, n = 11). In both the presence and absence of bicuculline, EPSCs had fast and slow components that were blocked by 6,7-dinitroquinoxaline-2,3-dione (DNQX; 10 microm; n = 7), and (+/-)3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP; 10 microm, n = 6), respectively. Reversal potentials for the evoked IPSCs and EPSCs were consistent with mediation via GABAA and ionotropic glutamate receptors, respectively. These results suggest that the SCN region provides both inhibitory and excitatory inputs to single VLPO neurons, which are mediated, respectively, by GABAA receptors and by both non-NMDA and NMDA glutamate receptors. These projections may play important roles in conveying circadian input to systems in the preoptic area that regulate sleep and waking.     

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.