Agonist-dependent postsynaptic effects of opioids on miniature excitatory postsynaptic currents in cultured hippocampal neurons

Although chronic treatment with morphine is known to alter the function and morphology of excitatory synapses, the effects of other opioids on these synapses are not clear. Here we report distinct effects of several opioids (morphine, [d-ala(2),me-phe(4),gly(5)-ol]enkephalin (DAMGO), and etorphine) on miniature excitatory postsynaptic currents (mEPSCs) in cultured hippocampal neurons: 1) chronic treatment with morphine for >3 days decreased the amplitude, frequency, rise time and decay time of mEPSCs. In contrast, "internalizing" opioids such as etorphine and DAMGO increased the frequency of mEPSCs and had no significant effect on the amplitude and kinetics of mEPSCs. These results demonstrate that different opioids can have distinct effects on the function of excitatory synapses. 2) mu opioid receptor fused with green fluorescence protein (MOR-GFP) is clustered in dendritic spines in most hippocampal neurons but is concentrated in axon-like processes in striatal and corticostriatal nonspiny neurons. It suggests that MORs might mediate pre- or postsynaptic effects depending on cell types. 3) Neurons were cultured from MOR knock-out mice and were exogenously transfected with MOR-GFP. Chronic treatment with morphine suppressed mEPSCs only in neurons that contained postsynaptic MOR-GFP, indicating that opioids can modulate excitatory synaptic transmission postsynaptically. 4) Morphine acutely decreased mEPSC amplitude in neurons expressing exogenous MOR-GFP but had no effect on neurons expressing GFP. It indicates that the low level of endogenous MORs could only allow slow opioid-induced plasticity of excitatory synapses under normal conditions. 5) A theoretical model suggests that morphine might affect the function of spines by decreasing the electrotonic distance from synaptic inputs to the soma.     

Effect of cyclothiazide on spontaneous miniature excitatory postsynaptic currents in rat hippocampal pyramidal cells

 Spontaneous miniature excitatory postsynaptic currents (mEPSCs) were recorded from the CA1 region of slices using the whole-cell patch-clamp technique. Cyclothiazide (0.1 mM), a complete blocker of desensitization of (S)-α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) channels, was applied to determine the changes in amplitude and kinetics of mEPSCs occurring with complete suppression of desensitization. The amplitude of mEPSC (A) was not affected significantly by cyclothiazide, but both the rise (τ_r) and the decay time (τ_d) were consistently increased (from 2.3 to 6.5 ms and from 9.9 to 22.2 ms respectively). The amplitude dependence of both τ_d and τ_r became much greater, but there was no upward shift of the best-fitting lines. The slopes of the control best-fitting lines were (±SD; ms/pA; n=5) 0.39±0.05 for τ_d:A and 0.12±0.07 for τ_r:A, but, in the presence of cyclothiazide, the corresponding slopes were much steeper (2.1±0.60 and 0.68±0.21; holding potential was –50 mV and temperature 32°C). These changes, which were slow to develop, suggest that cyclothiazide blocks AMPA receptor channel desensitization, whilst having no effect on the closing rate of AMPA channels. Judging by the extent of change, the speed of diffusion of glutamate in the synaptic cleft is probably similar to that in water. In conclusion, this study provides evidence that: (1) under control conditions, desensitization of AMPA channels plays a major role in shaping the time course of synaptic currents in CA1; and (2) cyclothiazide prolongs their time course solely by abolishing desensitization. Received: 28 May 1998 / Received after revision: 16 July 1998 / Accepted: 3 September 1998     

Noradrenaline-induced spontaneous inhibitory postsynaptic currents in mouse basolateral nucleus of amygdala pyramidal neurons: Comparison with dopamine-induced currents

The basolateral nucleus of the amygdala (BLA) receives both noradrenergic and dopaminergic projections. These projections are thought to be important for modulation of amygdala neural circuits. In BLA pyramidal neurons, noradrenaline (NA) is known to facilitate gamma-aminobutyric acid (GABA)ergic spontaneous inhibitory postsynaptic currents (sIPSCs) through excitation of interneurons. Dopamine (DA) also is known to facilitate GABAergic sIPSCs in pyramidal neurons of the amygdala region including the BLA. It is unclear which neurotransmitter, NA or DA, is predominant in facilitating sIPSC in the BLA. Whether NA and DA facilitate sIPSC in different or the same pyramidal neurons also remains unknown. Herein, we employed the patch clamp recording technique on BLA pyramidal neurons in mouse brain slices, and compared the facilitating actions of NA and DA on sIPSCs. First NA and then DA, or first DA and then NA, were applied to a slice. NA enhanced sIPSC frequency in the majority (80–90%) of pyramidal neurons tested, whereas DA enhanced sIPSC frequency in relatively few neurons (approximately 30%). Neurons responding to NA alone and DA alone accounted, respectively, for 54.3% and 2.9% of the pyramidal neurons tested (11.4% of neurons responded to neither NA nor DA). Pyramidal neurons in which NA and DA both facilitated sIPSCs accounted for 31.4% of neurons tested. These results suggest that NA facilitates GABAergic sIPSCs in a larger proportion of mouse BLA pyramidal neurons than DA.     

Reduction of glycine receptor-mediated miniature inhibitory postsynaptic currents in rat spinal lamina I neurons after peripheral inflammation

Peripheral inflammation may induce long-lasting sensitization in the central nociceptive system. Neurons in lamina I of the spinal dorsal horn play a pivotal role in the integration and relay of pain-related information. In rats we studied whether changes in passive and active membrane properties and/or alteration of glycine receptor-mediated inhibitory control of spinal lamina I neurons may contribute to central sensitization in a model of peripheral long-lasting inflammation (complete Freund's adjuvant, hindpaw). Spontaneously occurring glycine receptor-mediated miniature inhibitory postsynaptic currents (GlyR-mediated mIPSCs) were recorded in lumbar spinal lamina I neurons. Miniature IPSC rise, decay kinetics and mean GlyR-mediated mIPSC amplitude were not affected by peripheral inflammation. The mean frequency of GlyR-mediated mIPSCs of lamina I neurons ipsilateral to the inflamed hindpaw was, however, significantly reduced by peripheral inflammation when compared with neurons from noninflamed animals. Principal passive and active membrane properties and firing patterns of spinal lamina I neurons were not changed by inflammation. These results indicate that long-lasting peripheral inflammation leads to a reduced glycinergic inhibitory control of spinal lamina I neurons by a presynaptic mechanism.     

Analysis of miniature spontaneous inhibitory postsynaptic currents (sIPSCs) from current noise in crayfish opener muscle

Deviating from the normal situation, some crayfish muscle fibres showed spontaneous inhibitory activity: discharge of large inhibitory postsynaptic currents, IPSCs, alternating with long lasting bursts of current noise. Analysis of the bursts of current noise revealed that they are composed of spontaneous miniature unit currents, sIPSCs. In the burst periods the sIPSCs occurred with an average rate of 3.5–10 kHz and had an amplitude of about ã=90 pA at a driving force E=10 mV. The peak conductance _ã=ã/E of the sIPSCs was _ã=9.2 nS±0.5 (S.D.,n=5) for membrane potentials between E=–60 mV and E=–80 mV. _ã seemed to decrease when the membrane was hyperpolarized. The time constants of decay, , of the sIPSCs were identical with of the IPSCs. Further, and its potential dependence agreed with the mean lifetimes of inhibitory postsynaptic channels operated by -aminobutyric acid (GABA) [Dudel et al. 1977, 1980]. Synchronized opening of about 750 inhibitory synaptic channels generates a sIPSC.Analysis of this anomalous bursting inhibitory activity thus yields the size of the inhibitory quantum of transmission, which could not be obtained from IPSCs.This investigation was supported by the Deutsche Forschungsgemeinschaft     

Prolongation of hippocampal miniature inhibitory postsynaptic currents in mice lacking the GABA(A) receptor alpha1 subunit

GABA(A) receptors (GABA(A)-Rs) are pentameric structures consisting of two alpha, two beta, and one gamma subunit. The alpha subunit influences agonist efficacy, benzodiazepine pharmacology, and kinetics of activation/deactivation. To investigate the contribution of the alpha1 subunit to native GABA(A)-Rs, we analyzed miniature inhibitory postsynaptic currents (mIPSCs) in CA1 hippocampal pyramidal cells and interneurons from wild-type (WT) and alpha1 subunit knock-out (alpha1 KO) mice. mIPSCs recorded from interneurons and pyramidal cells obtained from alpha1 KO mice were detected less frequently, were smaller in amplitude, and decayed more slowly than mIPSCs recorded in neurons from WT mice. The effect of zolpidem was examined in view of its reported selectivity for receptors containing the alpha1 subunit. In interneurons and pyramidal cells from WT mice, zolpidem significantly increased mIPSC frequency, prolonged mIPSC decay, and increased mIPSC amplitude; those effects were diminished or absent in neurons from alpha1 KO mice. Nonstationary fluctuation analysis of mIPSCs indicated that the zolpidem-induced increase in mIPSC amplitude was associated with an increase in the number of open receptors rather than a change in the unitary conductance of individual channels. These data indicate that the alpha1 subunit is present at synapses on WT interneurons and pyramidal cells, although differences in mIPSC decay times and zolpidem sensitivity suggest that the degree to which the alpha1 subunit is functionally expressed at synapses on CA1 interneurons may be greater than that at synapses on CA1 pyramidal cells.     

脑室注射组胺对下丘脑室旁核促皮质素释放激素神经元的作用

目的:观察第三脑室注射组胺对下丘脑室旁核促皮质素释放激素(CRH)神经元活动的影响。方法:Fos癌蛋白免疫组化LSAB法结合双抗原标记法;半定量逆转录聚合酶链反应(RT-PCR)方法。结果:第三脑室注射组胺后,(1)下丘脑室旁核Fos阳性神经元数目明显增加(P＜0.05);(2)室旁核内的Fos阳性神经元中约有31.78%同时呈CRH阳性反应;(3)室旁核CRHmRNA含量明显升高,且有量效关系。结论:中枢组胺可以激活下丘脑室旁核的CRH神经元,并使CRH基因表达增加。     

Cellular mechanisms of orexin actions on paraventricular nucleus neurones in rat hypothalamus

Using whole-cell patch clamp techniques we have examined the cellular mechanisms underlying the effects of orexin A (OX-A) on electrophysiologically identified magnocellular and parvocellular neurones in the rat hypothalamic paraventricular nucleus (PVN). The majority of magnocellular neurones (67 %) showed concentration-dependent, reversible depolarizations in response to OX-A. These effects were abolished in tetrodotoxin (TTX), suggesting them to be indirect effects on this population of neurones. OX-A also caused increases in excitatory postsynaptic current (EPSC) frequency and amplitude in magnocellular neurones. The former effects were again blocked in TTX while increases in mini-EPSC amplitude remained. Depolarizing effects of OX-A on magnocellular neurones were also found to be abolished by kynurenic acid, supporting the conclusion that these effects were the result of activation of a glutamate interneurone. Parvocellular neurones (73 % of those tested) also showed concentration-dependent, reversible depolarizations in response to OX-A. In contrast to magnocellular neurones, these effects were maintained in TTX, indicating direct effects of OX-A on this population of neurones. Voltage clamp analysis using slow voltage ramps demonstrated that OX-A enhanced a non-selective cationic conductance with a reversal potential of -40 mV in parvocellular neurones, effects which probably explain the depolarizing effects of this peptide in this subpopulation of PVN neurones. These studies have identified separate cellular mechanisms through which OX-A influences the excitability of magnocellular and parvocellular PVN neurones.     

Variations in amplitude and time course of inhibitory postsynaptic currents

In order to examine the relative contributions of changes in amplitude and time course to synaptic plasticity, variations in peak amplitude and time constant of decay have been analyzed from inhibitory postsynaptic currents (PSC) recorded in voltage-clamped Aplysia buccal ganglia neurons. In these cells, synaptic currents with single time constant decay can be recorded with low noise under well-controlled space clamp. Over a population of 36 neurons, duration was more narrowly distributed than amplitude, but each varied. The coefficient of variation (CV) was 0.21 for decay time constant (tau) and 0.87 for peak conductance (g peak). Population variances are larger than can be accounted for by such variables as temperature and noise amplitude, suggesting that functional modifications alter each of these determinants of synaptic effectiveness over the long term. Recordings of up to several hundred PSC in each of 16 neurons show that both PSC amplitude and time course recorded in a single cell can vary independently over short time spans. Decay remained single exponential as time course changed. CV for tau averaged 0.11; CV for g peak was 0.19. Variability of tau was not an artifact of amplitude; CV was relatively uncorrelated with current amplitudes or sample size. Smoothing and adding excess noise to each individual PSC of a set produced only small changes to CV, showing that variability was not an artifact of noise. Several specific manipulations of the presynaptic neuron altered both PSC amplitude and time course. Tetanic stimulation of the presynaptic neuron produced short-term potentiation of both amplitude and time course of subsequent PSCs. Peak amplitude was increased by 80%; tau by 12%. Reducing interspike intervals from 10 to 1 s produced habituation of both amplitude and time course, with g peak decreasing by 35 to 40% and tau by 10%. Conditioning DC depolarization of the presynaptic neuron enhanced PSC amplitude with little effect on decay time constant. Although short-term plastic changes affect PSC amplitude more than duration, each is alterable. Parallel changes in both can synergistically alter synaptic charge transfer, and therefore efficacy. Similar mechanisms may produce larger long-term differences seen between neurons.     

Kappa opioid inhibition of somatodendritic dopamine inhibitory postsynaptic currents

In the midbrain, dopamine neurons can release dopamine somatodendritically. This results in an inhibitory postsynaptic current (IPSC) within adjacent dopamine cells that occurs by the activation of inhibitory D(2) autoreceptors. Kappa, but not mu/delta, opioid receptors inhibit this IPSC. The aim of the present study was to determine the mechanism by which kappa-opioid receptors inhibit the dopamine IPSC. In both the ventral tegmental area (VTA) and substantia nigra compacta (SNc) the kappa-receptor agonist U69593 inhibited the IPSC, but not the current induced by the exogenous iontophoretic application of dopamine. The endogenous peptide dynorphin A (1-13) also inhibited IPSCs in the VTA and SNc, but also the dopamine iontophoretic current in the VTA. Although both kappa agonists induced a postsynaptic outward current in the VTA, the current induced by dynorphin was dramatically larger. This suggests that the decrease in iontophoretic dopamine current was the result of occlusion. Occlusion alone, however, could not completely account for suppression of the IPSC. The kappa opioid inhibition of the IPSC was not affected by global increases or decreases in dopamine cell activity within the slice. These findings suggest that, although kappa opioid receptors can hyperpolarize dopamine neurons, they also suppress dopamine release by direct actions at the release site. The results thus demonstrate both pre- and postsynaptic actions of kappa receptor agonists. The actions of dynorphin indicate that VTA dopamine cells are selectively regulated by kappa receptors.     

Activity of neurones in the paraventricular nucleus of the hypothalamus and its control

1. Activity of the paraventricular nucleus neurones was recorded by micro-electrodes during resting conditions and while various osmotic, chemical, direct and indirect neural stimuli were applied. This activity was correlated with evidence of oxytocin release by recording milk ejection responses.2. Seventy-four per cent of all paraventricular nucleus neurones were osmosensitive in that firing was augmented following close arterial injection of hypertonic solutions (1 ml. 1 M-NaCl in 10-15 sec). A very few neurones showed decreased activity and in twenty-two per cent no change at all occurred following injection. Evidence indicated that the observed depressions of cellular firing rate were due to other than osmotic stimulation.3. In post-partum cats an osmotically-induced neurone discharge increase was accompanied by a milk ejection response equivalent to that produced by 2-3 m-u. of oxytocin.4. Paraventricular neurones were also sensitive to acetylcholine. Intracarotid injections of 40-80 mug of acetylcholine greatly increased discharge rates and caused a very definite milk ejection response.5. Stimulation of the nipples by gentle suction, but not by electrical shock, and distension of the uterus in post-partum cats increased unit discharge in the paraventricular nucleus and evoked a milk ejection response.6. Neurones of the paraventricular nucleus, unlike those of the supraoptic nucleus, did not appear to be specifically responsive to electrical stimulation of skin and muscle afferent or of central nervous structures. Such stimuli did cause slight augmentations or depressions in firing rates of cells within and adjacent to the paraventricular nuclei, but many neurones were unaffected. Stimuli such as those applied appeared to have a general central effect involving paraventricular as well as many other neurones.7. Direct electrical excitation of the pituitary stalk produced a milk ejection response in post-partum cats. Electrical pulses applied to the paraventricular nuclei were less effective for reasons discussed.     

Effects of fenamate on inhibitory postsynaptic currents in Purkinje’s cells

The effects of nonsteroid antiinflammatory drugs of the fenamate group (mefenamic and tolfenamic acids) on spontaneous miniature inhibitory postsynaptic currents in Purkinje’s cells were studied in mouse cerebellar slices by the whole cell patch-clamp method. Both drugs in concentrations of 3–30 μM significantly prolonged miniature inhibitory postsynaptic currents and reduced their amplitude. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 145, No. 5, pp. 500–504, May, 2008     

不同高渗刺激对下丘脑视上核和室旁核神经元功能状态的影响

视上核和室旁核是下丘脑中两个与渗透压感受、加压素分泌以及水平衡调节密切相关的核团。为了搞清楚这两个核团在不同刺激条件下的激活状态和反应特性,本文采用慢性和急性渴觉刺激模型,免疫组化和ELISA检测相结合的方法对视上核和室旁核内的Fos表达以及血清加压素水平进行了测定。慢性刺激组动物给予2% NaCl盐水持续2d,而急性刺激组动物皮下直接注射2mol/L的NaCl盐水2.5ml,两组动物的进食保持正常。结果表明,这两种不同的刺激方式引发的Fos表达模式基本相似,视上核、室旁核、下丘脑外侧区以及正中视前区、穹窿下器和终板血管下器等区都检测到大量的Fos阳性胞核。但Fos染色的深浅程度和Fos胞核的数量却在两组之间有明显的差异:急性组胞核浓染,数量多;慢性组胞核淡染,数量少。ELISA检测的结果与此相反,急性组动物血清中加压素的水平很低,与对照组没有明显差异;而慢性组动物血清中的加压素水平很高,几乎是对照组的2倍。以上结果提示,下丘脑神经元的激活和分泌功能与刺激方式密切相关,选择单一刺激模式、单一指标来揭示和衡量其功能状态是缺乏说服力的。     

High-affinity zinc potentiation of inhibitory postsynaptic glycinergic currents in the zebrafish hindbrain

Zinc has been reported to potentiate glycine receptors (GlyR), but the physiological significance of this observation has been put in doubt by the relatively high values of the EC(50), 0.5-1 microM, since such concentrations may not be attained in the synaptic cleft of glycinergic synapses. We have re-evaluated this observation in the frame of the hypothesis that contaminant heavy metals present in usual solutions may have lead to underestimate the affinity of the zinc binding site, and therefore to underestimate the potential physiological role of zinc. Using chelators either to complex heavy metals or to apply zinc at controlled concentrations, we have examined the action of zinc on GlyR kinetics in outside-out patches from 50-h-old zebrafish Mauthner cells. Chelating contaminating heavy metals with tricine or N,N,N',N'-tetrakis-(2-pyridylmethyl)-ethylenediamine (TPEN) decreased the duration of the currents evoked by glycine, confirming that traces of heavy metals alter the GlyR response in control conditions. Using tricine- (10 mM) buffered zinc solution, we then showed that zinc increases the amplitude of outside-out responses evoked by 0.1-0.5 mM glycine with an EC(50) of 15 nM. In contrast zinc had no effect on the amplitude of currents evoked by a saturating concentration (3-10 mM) of glycine. This suggests that zinc enhances GlyR apparent affinity for glycine. The study of the effects of zinc on the kinetics of the response indicates that this increase of apparent affinity is due to a decrease of the glycine dissociation rate constant. We then analyzed the effects of zinc on postsynaptic GlyRs in whole cell recordings of glycinergic miniature inhibitory postsynaptic currents (mIPSCs). Chelation of contaminant heavy metals decreased the amplitude and the duration of the mIPSCs; inverse effects were observed by adding zinc in buffered solutions containing nanomolar free zinc concentrations. Zinc plus tricine or tricine alone did not change the coefficient of variation ( approximately 0.85) of the mIPSC amplitude distributions. These results suggest that postsynaptic GlyRs are not saturated after the release of one vesicle.     

Neurochemistry of the paraventricular nucleus of the hypothalamus: Implications for cardiovascular regulation

The paraventricular nucleus of the hypothalamus (PVN) is an important site for autonomic and endocrine homeostasis. The PVN integrates specific afferent stimuli to produce an appropriate differential sympathetic output. The neural circuitry and some of the neurochemical substrates within this circuitry are discussed. The PVN has at least three neural circuits to alter sympathetic activity and cardiovascular regulation. These pathways innervate the vasculature and organs such as the heart, kidney and adrenal medulla. The basal level of sympathetic tone at any given time is dependent upon excitatory and inhibitory inputs. Under normal circumstances the sympathetic nervous system is tonically inhibited. This inhibition is dependent upon GABA and nitric oxide such that nitric oxide potentiates local GABAergic synaptic inputs onto the neurones in the PVN. Excitatory neurotransmitters such as glutamate and angiotensin II modify the tonic inhibitory activity. The neurotransmitters oxytocin, vasopressin and dopamine have been shown to affect cardiovascular function. These neurotransmitters are found in neurones of the PVN and within the spinal cord. Oxytocin and vasopressin terminal fibres are closely associated with sympathetic preganglionic neurones (SPNs). Sympathetic preganglionic neurones have been shown to express receptors for oxytocin, vasopressin and dopamine. Oxytocin causes cardioacceleratory and pressor effects that are greatest in the upper thoracic cord while vasopressin cause these effects but more significant in the lower thoracic cord. Dopaminergic effects on the cardiovascular system include inhibitory or excitatory actions attributed to a direct PVN influence or via interneuronal connections to sympathetic preganglionic neurones.     

VR1 receptor activation induces glutamate release and postsynaptic firing in the paraventricular nucleus

Neurons in the paraventricular nucleus (PVN) are important in regulating autonomic function through projections to the brain stem and spinal cord. Although the vanilloid receptors (VR(1)) are present in the PVN, their physiological function is scarcely known. In this study, we determined the role of VR(1) receptors in the regulation of synaptic inputs and the excitability of spinally projecting PVN neurons. Whole cell patch-clamp recordings were performed on the PVN neurons labeled by a retrograde fluorescence tracer injected into the thoracic spinal cord of rats. Capsaicin significantly increased the frequency of glutamatergic miniature excitatory postsynaptic currents (mEPSCs) without changing the amplitude and decay time constant of mEPSCs. On the other hand, capsaicin had no effect on GABAergic miniature inhibitory postsynaptic currents (mIPSCs). The effect of capsaicin on mEPSCs was abolished by a specific VR(1) antagonist, iodo-resiniferatoxin (iodo-RTX), or ruthenium red. Importantly, iodo-RTX per se significantly reduced the amplitude of evoked EPSCs and the frequency of mEPSCs. Removal of extracellular Ca(2+), but not Cd(2+) treatment, also eliminated the effect of capsaicin on mEPSCs. Furthermore, capsaicin caused a large increase in the firing rate of PVN neurons, and such an effect was abolished in the presence of ionotropic glutamate receptor antagonists. Additionally, the double-immunofluorescence labeling revealed that all of the VR(1) immunoreactivity was colocalized with a presynaptic marker, synaptophysin, in the PVN. Thus this study provides the first evidence that activation of VR(1) receptors excites preautonomic PVN neurons through selective potentiation of glutamatergic synaptic inputs. Presynaptic VR(1) receptors and endogenous capsaicin-like substances in the PVN may represent a previously unidentified mechanism in hypothalamic regulation of the autonomic nervous system.     

Angiotensin II activates a nitric-oxide-driven inhibitory feedback in the rat paraventricular nucleus

The hypothalamic paraventricular nucleus (PVN) has been shown to play major obligatory roles in autonomic and neuroendocrine regulation. Angiotensin II (ANG) acts as a neurotransmitter regulating the excitability of magnocellular neurons in this nucleus. We report here that ANG also activates a nitric-oxide-mediated negative feedback loop in the PVN that acts to regulate the functional output of magnocellular neurons. Thus in addition to its depolarizing actions on magnocellular neurons, ANG application results in an increase in the frequency of inhibitory postsynaptic potentials in a population of these neurons without effect on the amplitude of these events. ANG was also without significant effect on the mean frequency or amplitude of mini synaptic currents analyzed in voltage-clamp experiments. This increase in inhibitory input after ANG can be abolished by the nitric oxide synthase inhibitor Nomega-nitro-l-arginine methylester, demonstrating a requisite role for nitric oxide in the activation of this pathway. The depolarization of magnocellular neurons that show increased inhibitory postsynaptic potential (IPSP) frequency in response to ANG is significantly smaller than that observed in neurons in which IPSPs frequency was unaffected (3.2 +/- 1.1 vs. 8.0 +/- 0.5 mV, P < 0.05). Correspondingly, after nitric oxide synthase inhibition, the depolarizing effects of ANG on magnocellular neurons are augmented (2.0 +/- 0.7 vs. 6.7 +/- 0.7 mV, P < 0.05). The depolarization was also enhanced in the presence of the GABAergic antagonist bicuculline (1.9 +/- 1.2 vs. 11.9 +/- 2.3, P < 0.001). These data demonstrate that there exists within the PVN an intrinsic negative feedback loop that modulates neuronal excitability in response to peptidergic excitation.