GABA Synapses Research Articles

A synaptocentric view of the neuroendocrine response to stress

The essential role of parvocellular neuroendocrine cells (PNCs) in the paraventricular nucleus of the hypothalamus (PVN) is to translate real or perceived challenges into a comprehensive glucocorticoid (GC) hormone response. Synaptic inputs encoding physical and psychological stress engage the hypothalamic-pituitary-adrenal axis (HPA) by increasing PNC activity, and corticotropin-releasing hormone production and release. Following robust recruitment in response to stress, GCs feedback to dampen PNC responses. Here we review the contributions of glutamate and GABA synapses in PVN to the initiation and termination of the stress response. The reliability of HPA responses to a given stress can vary as a function of prior experience. Within this context, we examine possible synaptic correlates that allow this neuroendocrine system to learn and adapt following stress challenges.

GABA-mediated membrane oscillations as coincidence detectors for enhancing synaptic efficacy in the developing hippocampus

Spontaneously occurring neuronal oscillations constitute a hallmark of developmental networks. They have been observed in the retina, neocortex, hippocampus, thalamus and spinal cord. In the immature hippocampus the so-called ‘giant depolarizing potentials’ (GDPs) are network-driven membrane oscillations characterized by recurrent membrane depolarization with superimposed fast action potentials. Usually, they last hundreds of ms and are separated by intervals of several seconds. GDPs depend on the synergistic action of GABA and glutamate acting on GABAA and (RS)-a-amino-3-hydroxy-5-methyl-4-isoxadepropionate (AMPA) receptors, respectively. Early in postnatal life, GABA depolarizes and excites the postsynaptic cells due to high intracellular [Cl-] which results mainly from the unbalance of two Cl- co-transporter systems, the NKCCl and KCC2 that enhance and lower intracellular [Cl-], respectively. In accord with the Hebb postulate on activity-dependent synaptic strengthening, here the hypothesis has been tested that GDPs may act as coincident detector signals between pre and postsynaptic activity. This assumption has been verified at poorly developed mossy fiber (MF)-CA3 connections. These synapses are particularly interesting because they can release GABA in addition to glutamate. We found that during the first postnatal week correlated pre (MF) and postsynaptic (GDPs) activity persistently enhances synaptic strength at MF-CA3 connections. This effect was usually restricted to the activated synapse. When the interval between GDPs and MF stimulation was increased, the potentiating effect progressively declined and disappeared. The potentiation depended on the activation of voltage-dependent calcium channels and calcium flux. Moreover, it was clear from our experiments that at this developmental stage GABA is the main neurotransmitter released from the MF. This is consistent with the sequential expression of functional GABA and glutamatergic synapses found in the hippocampus at early developmental stages. Synchronous membrane oscillations may therefore contribute to the refinement of neuronal connectivity before the establishment of the adult neuronal circuit. Later in development when the degree of functional connections is sufficiently high, GDPs would be replaced by more subtle types of signal synchronization such as theta or gamma activity characteristic of the adult network.

Single-Synapse Analysis of a Diverse Synapse Population: Proteomic Imaging Methods and Markers

A lack of methods for measuring the protein compositions of individual synapses insitu has so far hindered the exploration and exploitation of synapse molecular diversity. Here, we describe the use of array tomography, a new high-resolution proteomic imaging method, to determine the composition of glutamate and GABA synapses in somatosensory cortex of Line-H-YFP Thy-1 transgenic mice. We find that virtually all synapses are recognized by antibodies to the presynaptic phosphoprotein synapsin I, while antibodies to 16 other synaptic proteins discriminate among 4 subtypes of glutamatergic synapses and GABAergic synapses. Cell-specific YFP expression in the YFP-H mouse line allows synapses to be assigned to specific presynaptic and postsynaptic partners and reveals that a subpopulation of spines on layer 5 pyramidal cells receives both VGluT1-subtype glutamatergic and GABAergic synaptic inputs. These results establish a means for the high-throughput acquisition of proteomic data from individual cortical synapses insitu.

Impaired striatal GABA transmission in experimental autoimmune encephalomyelitis

Synaptic dysfunction triggers neuronal damage in experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis (MS). While excessive glutamate signaling has been reported in the striatum of EAE, it is still uncertain whether GABA synapses are altered. Electrophysiological recordings showed a reduction of spontaneous GABAergic synaptic currents (sIPSCs) recorded from striatal projection neurons of mice with MOG (35−55)-induced EAE. GABAergic sIPSC deficits started in the acute phase of the disease (20–25 days post immunization, dpi), and were exacerbated at later time-points (35, 50, 70 and 90 dpi). Of note, in slices they were independent of microglial activation and of release of TNF-α. Indeed, sIPSC inhibition likely involved synaptic inputs arising from GABAergic interneurons, because EAE preferentially reduced sIPSCs of high amplitude, and was associated with a selective loss of striatal parvalbumin (PV)-positive GABAergic interneurons, which contact striatal projection neurons in their somatic region, giving rise to more efficient synaptic inhibition. Furthermore, we found also that the chronic persistence of pro-inflammatory cytokines were able, per se, to produce profound alterations of electrophysiological network properties, that were reverted by GABA administration. The results of the present investigation indicate defective GABA transmission in MS models depending from alteration of PV cells number and, in part, deriving from the effects of a chronic inflammation, and suggest that pharmacological agents potentiating GABA signaling might be considered to limit neuronal damage in MS patients.

Phase response analysis during in vivo-like high conductance states; dendritic SK determines the mean and variance of responses to dendritic excitation

A neuron’s phase response curve (PRC) describes how synaptic inputs at different times during the spike cycle affect the timing of subsequent spikes, and PRC analysis is a powerful technique for predicting and interpreting the emergence of synchronous modes in synaptically coupled networks and neuronal populations receiving common input. However, neuronal PRCs are typically measured during intrinsic pacemaking which may not reflect neuronal excitability and dynamics during high conductance states generated by complex network activity in vivo. Using a full morphological model of a globus pallidus (GP) neuron we have recently demonstrated that during intrinsic pacemaking, somatic PRCs for GP neurons are type I, i.e. excitatory inputs at all phases of the spike cycle advance the spontaneous spiking rhythm1. We also demonstrated that synaptic excitation of the distal dendrite can paradoxically delay subsequent spiking when delivered at some phases of the spike cycle (yielding a type II PRC) as a consequence of dendritic activation of the small conductance calcium-activated potassium current, SK1. Since during high conductance states spike timing is determined by a balance between intrinsic mechanisms and synaptic input fluctuations, in this study we investigated how somatic and dendritic phase response properties of the GP model are affected by ongoing stochastic synaptic background activity. We generated high conductance states in the model by applying synaptic backgrounds composed of randomly-timed excitatory and inhibitory synaptic inputs at 1022 GABA synapses and 100 AMPA synapses distributed throughout the dendrite. By varying the synaptic gain and input frequency parameters across the physiological range for inputs to GP, we achieved a diverse set of high conductance states characterized by sub-threshold voltage fluctuations, irregular spiking, and elevated membrane conductance (Figure ​(Figure1A1A &1B) and spanning the range of spike frequencies observed in vivo (15-45 Hz). For each synaptic background parameter set, we generated 100 single-trial PRCs by delivering a single 2.5 nS AMPA-synaptic input to either the soma or distal dendrite at each of 72 time-points (in separate simulations) within the first spike cycle of each of 100 control spike trains (Figure ​(Figure1C1C &1D). We then averaged the single-trial PRCs for each synaptic background to evaluate the dependence of PRC shape on stimulus location, spike frequency, and dendritic SK conductance, in addition to the gain and input frequencies of synaptic backgrounds. Next, we analyzed on a trial-by-trial basis the interactions of PRC stimuli with transient fluctuations in the synaptic background leading to added or skipped spikes. We determined that dendritic SK underlies both the incidence of skipped spikes and the dependence of skipped spike events on stimulus phase. Interestingly, the input-phase dependence of skipped spike events mirrored the phase-dependent variance in the PRC itself (which we plotted as phase response-variance curves, PRVCs). This indicates that dendritic SK determines not only the mean response, but also the variance of responses, to excitatory dendritic inputs. Average somatic and dendritic PRCs were type I and type II, respectively, and are likely to represent the average behavior of populations of GP neurons in response to shared excitation. Furthermore, in-as-much as the GP model and simulated synaptic backgrounds are physiologically realistic, the variance in the model’s responses across trials directly reflects a major source of spike time variance across populations of GP neurons in vivo. Figure 1 A&B. Total synaptic conductance (top) and current (bottom) for synaptic backgrounds composed of 1 nS (A) or 2 nS (B) unitary inputs. C&D. Perturbations of control spike trains (black) by somatic (C) or dendritic (D2D)(D) stimuli. (Trace ...

Temporal dysregulation of cortical gene expression in the isolation reared Wistar rat

The critical sequence of molecular, neurotransmission and synaptic disruptions that underpin the emergence of psychiatric disorders like schizophrenia remain to be established with progress only likely using animal models that capture key features of such disorders. We have related the emergence of behavioural, neurochemical and synapse ultrastructure deficits to transcriptional dysregulation in the medial prefrontal cortex of Wistar rats reared in isolation. Isolation reared animals developed sensorimotor deficits at postnatal day 60 which persisted into adulthood. Analysis of gene expression prior to the emergence of the sensorimotor deficits revealed a significant disruption in transcriptional control, notably of immediate early and interferon-associated genes. At postnatal day 60 many gene transcripts relating particularly to GABA transmission and synapse structure, for example Gabra4, Nsf, Syn2 and Dlgh1, transiently increased expression. A subsequent decrease in genes such as Gria2 and Dlgh2 at postnatal day 80 suggested deficits in glutamatergic transmission and synapse integrity, respectively. Microdialysis studies revealed decreased extracellular glutamate suggesting a state of hypofrontality while ultrastructural analysis showed total and perforated synapse complement in layer III to be significantly reduced in the prefrontal cortex of postnatal day 80 isolated animals. These studies provide a molecular framework to understand the developmental emergence of the structural and behavioural characteristics that may in part define psychiatric illness.

GABAergic synapse properties may explain genetic variation in hippocampal network oscillations in mice

Cognitive ability and the properties of brain oscillation are highly heritable in humans. Genetic variation underlying oscillatory activity might give rise to differences in cognition and behavior. How genetic diversity translates into altered properties of oscillations and synchronization of neuronal activity is unknown. To address this issue, we investigated cellular and synaptic mechanisms of hippocampal fast network oscillations in eight genetically distinct inbred mouse strains. The frequency of carbachol-induced oscillations differed substantially between mouse strains. Since GABAergic inhibition sets oscillation frequency, we studied the properties of inhibitory synaptic inputs (IPSCs) received by CA3 and CA1 pyramidal cells of three mouse strains that showed the highest, lowest and intermediate frequencies of oscillations. In CA3 pyramidal cells, the frequency of rhythmic IPSC input showed the same strain differences as the frequency of field oscillations. Furthermore, IPSC decay times in both CA1 and CA3 pyramidal cells were faster in mouse strains with higher oscillation frequencies than in mouse strains with lower oscillation frequency, suggesting that differences in GABAA-receptor subunit composition exist between these strains. Indeed, gene expression of GABAA-receptor β2 (Gabrb2) and β3 (Gabrb2) subunits was higher in mouse strains with faster decay kinetics compared with mouse strains with slower decay kinetics. Hippocampal pyramidal neurons in mouse strains with higher oscillation frequencies and faster decay kinetics fired action potential at higher frequencies. These data indicate that differences in genetic background may result in different GABAA-receptor subunit expression, which affects the rhythm of pyramidal neuron firing and fast network activity through GABA synapse kinetics.

Nitric oxide alters GABAergic synaptic transmission in cultured hippocampal neurons

Nitric oxide (NO) production increases during hypoxia/ischemia–reperfusion in the immature brain and is associated with neurotoxicity. NO at physiologic concentrations has been shown to modulate GABAergic (gamma-aminobutyric acid) synaptic transmission in the adult brain. However, the effects of neurotoxic concentrations of NO (relevant to hypoxia–ischemia) on GABAergic synaptic transmission remain unknown. The present study tests the hypothesis that nNOS is expressed at GABAergic synapses and that exposure to neurotoxic concentrations of NO results in enhanced GABAergic synaptic transmission in cultured hippocampal neurons (days- in-vitro 10–14) prepared from fetal rats. Using double immunocytochemistry techniques, we were able to demonstrate that nNOS is co-localized to both presynaptic and postsynaptic markers of GABAergic synapses. The effects of NO on GABAergic synaptic transmission were then studied using whole cell patch-clamp electrophysiology. Spontaneous and miniature inhibitory postsynaptic currents (sIPSCS and mIPSCs) were recorded prior to and after exposure to 250 μM of the NO donor diethyleneamine/nitric oxide adduct (DETA-NO). Exposure to DETA-NO resulted in increased sIPSCs and mIPSCs frequency, indicating that neurotoxic concentrations of NO enhance GABAergic synaptic transmission in cultured hippocampal neurons. Because GABA synapses appear to be excitatory in the immature brain, this effect may contribute to overall enhanced synaptic transmission and hyperexcitability. We speculate that NO represents one of the mechanisms by which hypoxia–ischemia increases seizure susceptibility in the immature brain.

Simulating attentional blink with a neocortical attractor model

This study simulates attentional or blink of themind, a phenomenon where a human subject missesperception of a later expected visual pattern as twoexpected visual patterns are presented less than 500 milli-seconds apart. With an abstract computational model ofthe neocortex, a cortical patch is stimulated with asequence of patterns at varying time points. Patterns thatbecome active attractors are considered recognized.A neocortical patch is represented as a square matrix ofhypercolumns, each containing a set of minicolumns withsynaptic connections both within and across both mini-columns and hypercolumns [1]. Each minicolumn con-sists of locally connected layer 2/3 pyramidal cells withinteracting basket and bipolar/double bouquet interneu-rons. All neurons are implemented using the Hodgkin-Huxley multi-compartmental cell formalism and includecalcium dynamics, synaptic depression and saturatingAMPA, NMDA and GABA synapses. Active patterns areencoded with global connectivity of minicolumns acrosshypercolumns and they inhibit each other.Stored patterns were stimulated at sliding differential timepoints to create neural attractor interference measurablewith synthetic electrode traces. The effect correspondedwith human visual attentional studies. Additionally,studies on the inhibitory effect of the drug class benzodi-azopine on attentional in human subjects werecompared with neocortical simulations where an inhibi-tory GABA receptor conductance was increased. Simula-tions showed increases in the attentional duration,agreeing with observations in human studies. Figures 1and 2.

Influence of external input on waxing and waning of neuronal network oscillations

Introduction Experimental observations have reported modulation of cortical oscillations as phases of high synchronization (waxing) followed by periods of reduced synchronization (waning) [1-3]. Although the phenomenon is present in almost all frequency bands, it is still not understood how this is driven. Here we study whether this phenomenon can occur in a network of inhibitory (I) and excitatory (E) cells and what effect external inputs have. Methods Using NEURON, we model a network of Ne excitatory and Ni inhibitory cells such that Ne/Ni = 4. The cells have a single compartment, and include passive channels and voltage dependent Na+, K+ channels. Synaptic connections are random, projecting GABA synapses from I to I and I to E cells and AMPA synapses from E to E and E to I cells. To stimulate the network, each cell receives a baseline of current and a stream of spikes delivered at random intervals across the simulated period. from Eighteenth Annual Computational Neuroscience Meeting: CNS*2009 Berlin, Germany. 18–23 July 2009

Abused Inhalants Enhance GABA-Mediated Synaptic Inhibition

Abused inhalants are widely used, especially among school age children and teenagers, and are ‘gateway’ drugs leading to the abuse of alcohol and other addictive substances. In spite of this widespread use, little is known about the effects produced by inhalants on the central nervous system. The similarity in behavioral effects produced by inhalants and inhaled anesthetics, together with their common chemical features, prompted the present study of inhalant actions on a well characterized anesthetic target, GABA synapses. Whole cell patch clamp recordings were conducted on CA1 pyramidal neurons in rat hippocampal brain slices to measure effects on resting membrane properties, action potential discharge and GABA-mediated inhibitory responses. Toluene, 1,1,1-trichloroethane and trichloroethylene depressed CA1 excitability in a concentration-dependent and reversible manner. This depression appeared to involve enhanced GABA-mediated inhibition, evident in its reversal by a GABA receptor antagonist. Consistent with this, the abused inhalants increased inhibitory postsynaptic potentials produced using minimal stimulation of stratum radiatum inputs to CA1 neurons, in the presence of CNQX and APV to block excitatory synaptic responses and GGP to block GABAB responses. The enhanced inhibition appeared to come about by a presynaptic action on GABA nerve terminals, since spontaneous inhibitory postsynaptic current (IPSC) frequency was increased with no change in the amplitude of postsynaptic currents, both in the presence and absence of tetrodotoxin used to block interneuron action potentials and cadmium used to block calcium influx into nerve terminals. The toluene-induced increase in mIPSC frequency was blocked by dantrolene or ryanodine, indicating that the abused inhalant acted to increase the release of calcium from intracellular nerve terminal stores. This presynaptic action produced by abused inhalants is shared by inhaled anesthetics and would contribute to the altered behavorial effects produced by both classes of drugs, and could be especially important in the context of a disruption of learning and memory by abused inhalants.

Ethanol-induced delta-opioid receptor modulation of glutamate synaptic transmission and conditioned place preference in central amygdala

Alcoholism involves compulsive behaviors of alcohol drinking, which is thought to be related at least initially to the rewarding effect of alcohol. It has been shown that mu-opioid receptors play an essential role in drug reward and dependence for many drugs of abuse including alcohol, but the function of delta-opioid receptors (DOR) in drug reward remains largely unknown at present. Previous animal studies using systemic approaches with DOR antagonists or DOR knockout animals have yielded inconsistent results, showing a decrease, an increase or no change in alcohol consumption and behaviors of alcohol reward after DOR inhibition or deletion. In the present study, we used ethanol-conditioned rats to investigate adaptive DOR function in neurons of the central nucleus of the amygdala (CeA), a key brain site for alcohol reward and addiction. We found that functional DOR was absent in glutamate synapses of CeA neurons from control rats, but it emerged and inhibited glutamate synaptic currents in CeA neurons from rats displaying ethanol-induced behavior of conditioned place preference (CPP). Analysis of paired-pulse ratios and miniature glutamate synaptic currents revealed that the recruited DOR was present on glutamatergic presynaptic terminals. Similar induction of functional DOR was also found on GABA synapses. Furthermore, microinjection of a DOR antagonist into the CeA reversed ethanol-induced CPP behavior in rats in vivo. These results suggest that repeated alcohol exposure recruits new functional DOR on CeA glutamate and GABA synapses, which may be involved in the expression or maintenance of ethanol-induced CPP behavior.

GABA Synapses Mediate Neuroprotection after Ischemic and εPKC Preconditioning in Rat Hippocampal Slice Cultures

Delayed neuroprotection against ischemic challenges is conferred by both ischemic preconditioning (IPC) and preconditioning by activation of the epsilon-isoform of protein kinase C (epsilonPKC-PC). In vivo, ischemic preconditioning enhances GABA release and ameliorates glutamate release during lethal cerebral ischemia. We tested the hypothesis that IPC and epsilonPKC-PC confer neuroprotection by GABA synapses in rat organotypic hippocampal slices. Ischemic preconditioning or epsilonPKC-PC was induced with 15 mins oxygen-glucose deprivation (OGD) or psiepsilonRACK, a selective epsilonPKC activator; and test ischemia consisted of 40 mins OGD. At the time of peak neuroprotection (48 h after preconditioning), we recorded GABA(A) receptor-mediated miniature postsynaptic currents (GABA mPSCs) in vulnerable CA1 pyramidal neurons using whole-cell voltage clamp techniques. The frequency and amplitude of GABA mPSCs significantly increased 48 h after IPC. In contrast, epsilonPKC-PC enhanced only the amplitude of GABA mPSCs with no effect on frequency. We next asked if neuroprotection depended on these changes in GABA synapses. Weak antagonism of the GABA(A) receptor with bicuculline (100 nmol/L) decreased the amplitude of GABA mPSCs by 20.9+/-6.1%. When applied during test ischemia, 100 nmol/L bicuculline abolished neuroprotection conferred by either IPC or epsilonPKC-PC. We conclude that neuroprotection conferred by preconditioning depends on functional modifications of GABA synapses.

New directions in synaptic and network plasticity – a move away from NMDA receptor mediated plasticity

New directions in synaptic and network plasticity – a move away from NMDA receptor mediated plasticity

Vesicle Pool Heterogeneity at Hippocampal Glutamate and GABA Synapses

Glutamate and GABA are the major fast excitatory and inhibitory neurotransmitters, respectively, in the CNS. Although glutamate and GABA have clearly distinct postsynaptic actions, we are just beginning to appreciate that presynaptic differences between glutamatergic and GABAergic neurons may contribute to distinct functions of these transmitter systems. We therefore probed possible differences between the functional synaptic vesicle populations of glutamatergic and GABAergic neurons. We examined superecliptic synaptopHluorin (SpH) fluorescence during 20 Hz electrical stimulation in transfected hippocampal neurons and identified the phenotype of SpH-fluorescent synapses with post hoc immunostaining. With 200 stimuli (10 s), individual glutamate synapses displayed considerably more variability in peak SpH fluorescence than GABA synapses, without a strong difference in the mean SpH fluorescence increase. This spatial heterogeneity could not be accounted for by differences in endocytosis, which was nearly constant over these short time periods across glutamate and GABA synapses. Instead, variability in vesicle exocytosis correlated with variability in total vesicle staining and in measures of the total recycling pool size. Differences were also evident using FM1-43 [N-(3-triethylammoniumpropyl)-4-(4-(dibutylamino)styryl) pyridinium dibromide] uptake. These data support the idea that the population of glutamate synapses exhibits more heterogeneity in release properties than the population of GABA synapses, possibly correlated with glutamatergic synaptic malleability.

Reticular nucleus-specific changes in α3 subunit protein at GABA synapses in genetically epilepsy-prone rats

Differential composition of GABA(A) receptor (GABA(A)R) subunits underlies the variability of fast inhibitory synaptic transmission; alteration of specific GABA(A)R subunits in localized brain regions may contribute to abnormal brain states such as absence epilepsy. We combined immunocytochemistry and high-resolution ImmunoGold electron microscopy to study cellular and subcellular localization of GABA(A)R alpha1, alpha3, and beta2/beta3 subunits in ventral posterior nucleus (VP) and reticular nucleus (RTN) of control rats and WAG/Rij rats, a genetic model of absence epilepsy. In control rats, alpha1 subunits were prominent at inhibitory synapses in VP and much less prominent in RTN; in contrast, the alpha3 subunit was highly evident at inhibitory synapses in RTN. beta2/beta3 subunits were evenly distributed at inhibitory synapses in both VP and RTN. ImmunoGold particles representing all subunits were concentrated at postsynaptic densities with no extrasynaptic localization. Calculated mean number of particles for alpha1 subunit per postsynaptic density in nonepileptic VP was 6.1 +/- 3.7, for alpha3 subunit in RTN it was 6.6 +/- 3.4, and for beta2/beta3 subunits in VP and RTN the mean numbers were 3.7 +/- 1.3 and 3.5 +/- 1.2, respectively. In WAG/Rij rats, there was a specific loss of alpha3 subunit immunoreactivity at inhibitory synapses in RTN, without reduction in alpha3 subunit mRNA or significant change in immunostaining for other markers of RTN cell identity such as GABA or parvalbumin. alpha3 immunostaining in cortex was unchanged. Subtle, localized changes in GABA(A)R expression acting at highly specific points in the interconnected thalamocortical network lie at the heart of idiopathic generalized epilepsy.

NRG1 and Synaptic Function in the CNS

Recent genetic evidence indicates that neuregulin 1 (NRG1) and its receptor erbB4 may be susceptibility genes in schizophrenia, but their function in CNS synaptic transmission and circuitry is not well understood. In this issue of Neuron, studies from Li et al. and Woo et al. show that NRG1 and erbB4 regulate transmission at brain glutamate and GABA synapses. These findings raise the possibility of synaptic defects in schizophrenia.

Phosphoinositide turnover associated with synaptic transmission.

Although pharmacological stimulation of a wide variety of transmitter receptors triggers phosphoinositide (PI) turnover, little is known about the type of synaptic activity required to activate this system. To investigate this question, we have used primary cultures of embryonic cortical neurons, which develop functional glutamate and GABA synapses during maturation in vitro. Mature cultures display spontaneous synaptic activity that is totally suppressed by tetrodotoxin (TTX). PI turnover, assayed by the lithium-sensitive accumulation of [3H]CDP-diacylglycerol, was readily detected under basal conditions and was abolished by TTX. Increased excitatory synaptic activity induced by picrotoxin, an antagonist of GABAA receptor-mediated inhibition, further stimulated PI turnover. Similar results were obtained when PI turnover was assayed using [3H]inositol labeling. With either assay, the magnitude of synaptically induced PI turnover was comparable to maximal responses produced by muscarinic receptor stimulation. Although a component of the spontaneous synaptic currents is sensitive to N-methyl-D-aspartate (NMDA)-preferring glutamate receptor antagonists, blockade of NMDA receptors did not affect PI turnover associated with synaptic transmission. To assess the time course of synaptically mediated PI turnover, the amplitude and duration of spontaneous synaptic currents were reduced by lowering the extracellular Ca2+ concentration from 2.25 to 0.5 mM, a maneuver that suppresses basal PI turnover. Increases in PI turnover were detected as early as 5 min following restoration of the extracellular Ca2+ concentration to 2.25 mM. Taken together, these findings indicate that activation of the PI system is associated with physiological levels of glutamatergic synaptic transmission.

Nociceptin/orphanin FQ presynaptically decreases GABAergic transmission and blocks the ethanol-induced increase of GABA release in central amygdala

Behavioral studies show that the GABAergic system in the central amygdala (CeA) nucleus has a complex role in the reinforcing effects effects of ethanol and the anxiogenic response to ethanol withdrawal. Opioid peptides and nociceptin/orphanin FQ (nociceptin) within the CeA are implicated also in regulating voluntary ethanol consumption and ethanol relapse. Recently, we reported that basal GABAergic transmission was increased in ethanol-dependent rats, and that acute ethanol increases GABA(A) receptor-mediated inhibitory postsynaptic currents (IPSCs) in CeA neurons from both naïve and ethanol-dependent rats to the same extent, suggesting lack of tolerance for the acute effect of ethanol. Here, we investigated the effect of nociceptin on IPSCs in CeA neurons and its interaction with ethanol effects on these GABA synapses. We found that nociceptin moderately decreased IPSC amplitudes, acting mostly presynaptically as it increased paired-pulse facilitation ratio of IPSCs and decreased miniature IPSC frequencies (but not amplitudes). Nociceptin also prevented the ethanol-induced augmentation of IPSCs in CeA of naïve rats. Interestingly, in CeA of ethanol-dependent rats, the nociceptin-induced inhibition of IPSCs was increased, indicating an enhanced sensitivity to nociceptin. Nociceptin also blocked the ethanol-induced augmentation of IPSCs in ethanol-dependent rats. Our data suggest that nociceptin has a role in regulating the GABAergic system and opposing the effect elicited by ethanol. Thus, nociceptin may represent a therapeutic target for alleviating alcohol dependence.

Transcriptional Signatures of Cellular Plasticity in Mice Lacking the α1 Subunit of GABAAReceptors

GABAA receptors mediate the majority of inhibitory neurotransmission in the CNS. Genetic deletion of the alpha1 subunit of GABAA receptors results in a loss of alpha1-mediated fast inhibitory currents and a marked reduction in density of GABAA receptors. A grossly normal phenotype of alpha1-deficient mice suggests the presence of neuronal adaptation to these drastic changes at the GABA synapse. We used cDNA microarrays to identify transcriptional fingerprints of cellular plasticity in response to altered GABAergic inhibition in the cerebral cortex and cerebellum of alpha1 mutants. In silico analysis of 982 mutation-regulated transcripts highlighted genes and functional groups involved in regulation of neuronal excitability and synaptic transmission, suggesting an adaptive response of the brain to an altered inhibitory tone. Public gene expression databases permitted identification of subsets of transcripts enriched in excitatory and inhibitory neurons as well as some glial cells, providing evidence for cellular plasticity in individual cell types. Additional analysis linked some transcriptional changes to cellular phenotypes observed in the knock-out mice and suggested several genes, such as the early growth response 1 (Egr1), small GTP binding protein Rac1 (Rac1), neurogranin (Nrgn), sodium channel beta4 subunit (Scn4b), and potassium voltage-gated Kv4.2 channel (Kcnd2) as cell type-specific markers of neuronal plasticity. Furthermore, transcriptional activation of genes enriched in Bergman glia suggests an active role of these astrocytes in synaptic plasticity. Overall, our results suggest that the loss of alpha1-mediated fast inhibition produces diverse transcriptional responses that act to regulate neuronal excitability of individual neurons and stabilize neuronal networks, which may account for the lack of severe abnormalities in alpha1 null mutants.