Agonist antibody that induces human malignant cells to kill one another.

An attractive, but as yet generally unrealized, approach to cancer therapy concerns discovering agents that change the state of differentiation of the cancer cells. Recently, we discovered a phenomenon that we call "receptor pleiotropism" in which agonist antibodies against known receptors induce cell fates that are very different from those induced by the natural agonist to the same receptor. Here, we show that one can take advantage of this phenomenon to convert acute myeloblastic leukemic cells into natural killer cells. Upon induction with the antibody, these leukemic cells enter into a differentiation cascade in which as many as 80% of the starting leukemic cells can be differentiated. The antibody-induced killer cells make large amounts of perforin, IFN-γ, and granzyme B and attack and kill other members of the leukemic cell population. Importantly, induction of killer cells is confined to transformed cells, in that normal bone marrow cells are not induced to form killer cells. Thus, it seems possible to use agonist antibodies to change the differentiation state of cancer cells into those that attack and kill other members of the malignant clone from which they originate.

Fmr1 deficiency promotes age-dependent alterations in the cortical synaptic proteome.

Fragile X syndrome (FXS) is an X-linked neurodevelopmental disorder characterized by severe intellectual disability and other symptoms including autism. Although caused by the silencing of a single gene, Fmr1 (fragile X mental retardation 1), the complexity of FXS pathogenesis is amplified because the encoded protein, FMRP, regulates the activity-dependent translation of numerous mRNAs. Although the mRNAs that associate with FMRP have been extensively studied, little is known regarding the proteins whose expression levels are altered, directly or indirectly, by loss of FMRP during brain development. Here we systematically measured protein expression in neocortical synaptic fractions from Fmr1 knockout (KO) and wild-type (WT) mice at both adolescent and adult stages. Although hundreds of proteins are up-regulated in the absence of FMRP in young mice, this up-regulation is largely diminished in adulthood. Up-regulated proteins included previously unidentified as well as known targets involved in synapse formation and function and brain development and others linked to intellectual disability and autism. Comparison with putative FMRP target mRNAs and autism susceptibility genes revealed substantial overlap, consistent with the idea that the autism endophenotype of FXS is due to a "multiple hit" effect of FMRP loss, particularly within the PSD95 interactome. Through studies of de novo protein synthesis in primary cortical neurons from KO and WT mice, we found that neurons lacking FMRP produce nascent proteins at higher rates, many of which are synaptic proteins and encoded by FMRP target mRNAs. Our results provide a greatly expanded view of protein changes in FXS and identify age-dependent effects of FMRP in shaping the neuronal proteome.

The Proinflammatory Cytokine Interleukin 18 Regulates Feeding by Acting on the Bed Nucleus of the Stria Terminalis.

UNLABELLED: The proinflammatory cytokine IL-18 has central anorexigenic effects and was proposed to contribute to loss of appetite observed during sickness. Here we tested in the mouse the hypothesis that IL-18 can decrease food intake by acting on neurons of the bed nucleus of the stria terminalis (BST), a component of extended amygdala recently shown to influence feeding via its projections to the lateral hypothalamus (LH). We found that both subunits of the heterodimeric IL-18 receptor are highly expressed in the BST and that local injection of recombinant IL-18 (50 ng/ml) significantly reduced c-fos activation and food intake for at least 6 h. Electrophysiological experiments performed in BST brain slices demonstrated that IL-18 strongly reduces the excitatory input on BST neurons through a presynaptic mechanism. The effects of IL-18 are cell-specific and were observed in Type III but not in Type I/II neurons. Interestingly, IL-18-sensitve Type III neurons were recorded in the juxtacapsular BST, a region that contains BST-LH projecting neurons. Reducing the excitatory input on Type III GABAergic neurons, IL-18 can increase the firing of glutamatergic LH neurons through a disinhibitory mechanism. Imbalance between excitatory and inhibitory activity in the LH can induce changes in food intake. Effects of IL-18 were mediated by the IL-18R because they were absent in neurons from animals null for IL-18Rα (Il18ra(-/-)), which lack functional IL-18 receptors. In conclusion, our data show that IL-18 may inhibit feeding by inhibiting the activity of BST Type III GABAergic neurons. SIGNIFICANCE STATEMENT: Loss of appetite during sickness is a common and often debilitating phenomenon. Although proinflammatory cytokines are recognized as mediators of these anorexigenic effects, their mechanism and sites of action remain poorly understood. Here we show that interleukin 18, an anorexigenic cytokine, can act on neurons of the bed nucleus of the stria terminalis to reduce food intake via the IL-18 receptor. The findings identify a site and a mode of action that indicate targets for the treatment of cachexia or other eating disorders.

High-resolution and cell-type-specific photostimulation mapping shows weak excitatory vs. strong inhibitory inputs in the bed nucleus of the stria terminalis.

The bed nucleus of the stria terminalis (BNST) is a key component of the extended amygdala and has been implicated in anxiety and addiction. As individual neurons function within neural circuits, it is important to understand local microcircuits and larger network connections of identified neuronal types and understand how maladaptive changes in the BNST neural networks are induced by stress and drug abuse. However, due to limitations of classic anatomical and physiological methods, the local circuit organization of synaptic inputs to specific BNST neuron types is not well understood. In this study, we report on the application of high-resolution and cell-type-specific photostimulation methodology developed in our laboratory to local circuit mapping in the BNST. Under calibrated experimental conditions, laser photostimulation via glutamate uncaging or channelrhodopsin-2 photoactivation evokes spiking of BNST neurons perisomatically, without activating spikes from axons of passage or distal dendrites. Whole cell recordings, combined with spatially restricted photostimulation of presynaptic neurons at many different locations over a large region, allow high-resolution mapping of presynaptic input sources to single recorded neurons in the BNST. We constructed maps of synaptic inputs impinging onto corticotrophin-releasing hormone-expressing (CRH+) BNST neurons in the dorsolateral BNST and found that the CRH+ neurons receive predominant local inhibitory synaptic connections with very weak excitatory connections. Through cell-type-specific optogenetic stimulation mapping, we generated maps of somatostatin-expressing neuron-specific inhibitory inputs to BNST neurons. Taken together, the photostimulation-based techniques offer us powerful tools for determining the functional organization of local circuits of specific BNST neuron types.

Oxytocin excites BNST interneurons and inhibits BNST output neurons to the central amygdala.

The dorsolateral bed nucleus of the stria terminalis (BNSTDL) has high expression of oxytocin (OT) receptors (OTR), which were shown to facilitate cued fear. However, the role of OTR in the modulation of BNSTDL activity remains elusive. BNSTDL contains GABA-ergic neurons classified based on intrinsic membrane properties into three types. Using in vitro patch-clamp recordings in male rats, we demonstrate that OT selectively excites and increases spontaneous firing rate of Type I BNSTDL neurons. As a consequence, OT increases the frequency, but not amplitude, of spontaneous inhibitory post-synaptic currents (sIPSCs) selectively in Type II neurons, an effect abolished by OTR antagonist or tetrodotoxin, and reduces spontaneous firing rate in these neurons. These results suggest an indirect effect of OT in Type II neurons, which is mediated via OT-induced increase in firing of Type I interneurons. As Type II BNSTDL neurons were shown projecting to the central amygdala (CeA), we also recorded from retrogradely labeled BNST→CeA neurons and we show that OT increases the frequency of sIPSC in these Type II BNST→CeA output neurons. In contrast, in Type III neurons, OT reduces the amplitude, but not frequency, of both sIPSCs and evoked IPSCs via a postsynaptic mechanism without changing their intrinsic excitability. We present a model of fine-tuned modulation of BNSTDL activity by OT, which selectively excites BNSTDL interneurons and inhibits Type II BNST→CeA output neurons. These results suggest that OTR in the BNST might facilitate cued fear by inhibiting the BNST→CeA neurons.

Protracted withdrawal from alcohol and drugs of abuse impairs long-term potentiation of intrinsic excitability in the juxtacapsular bed nucleus of the stria terminalis.

The juxtacapsular bed nucleus of the stria terminalis (jcBNST) is activated in response to basolateral amygdala (BLA) inputs through the stria terminalis and projects back to the anterior BLA and to the central nucleus of the amygdala. Here we show a form of long-term potentiation of the intrinsic excitability (LTP-IE) of jcBNST neurons in response to high-frequency stimulation of the stria terminalis. This LTP-IE, which was characterized by a decrease in the firing threshold and increased temporal fidelity of firing, was impaired during protracted withdrawal from self-administration of alcohol, cocaine, and heroin. Such impairment was graded and was more pronounced in rats that self-administered amounts of the drugs sufficient to maintain dependence. Dysregulation of the corticotropin-releasing factor (CRF) system has been implicated in manifestation of protracted withdrawal from dependent drug use. Administration of the selective corticotropin-releasing factor receptor 1 (CRF(1)) antagonist R121919 [2,5-dimethyl-3-(6-dimethyl-4-methylpyridin-3-yl)-7-dipropylamino-pyrazolo[1,5-a]pyrimidine)], but not of the CRF(2) antagonist astressin(2)-B, normalized jcBNST LTP-IE in animals with a history of alcohol dependence; repeated, but not acute, administration of CRF itself produced a decreased jcBNST LTP-IE. Thus, changes in the intrinsic properties of jcBNST neurons mediated by chronic activation of the CRF system may contribute to the persistent emotional dysregulation associated with protracted withdrawal.

Developmental NMDA receptor dysregulation in the infantile neuronal ceroid lipofuscinosis mouse model.

Protein palmitoylation and depalmitoylation alter protein function. This post-translational modification is critical for synaptic transmission and plasticity. Mutation of the depalmitoylating enzyme palmitoyl-protein thioesterase 1 (PPT1) causes infantile neuronal ceroid lipofuscinosis (CLN1), a pediatric neurodegenerative disease. However, the role of protein depalmitoylation in synaptic maturation is unknown. Therefore, we studied synapse development in Ppt1-/- mouse visual cortex. We demonstrate that the developmental N-methyl-D-aspartate receptor (NMDAR) subunit switch from GluN2B to GluN2A is stagnated in Ppt1-/- mice. Correspondingly, Ppt1-/- neurons exhibit immature evoked NMDAR currents and dendritic spine morphology in vivo. Further, dissociated Ppt1-/- cultured neurons show extrasynaptic, diffuse calcium influxes and enhanced vulnerability to NMDA-induced excitotoxicity, reflecting the predominance of GluN2B-containing receptors. Remarkably, Ppt1-/- neurons demonstrate hyperpalmitoylation of GluN2B as well as Fyn kinase, which regulates surface retention of GluN2B. Thus, PPT1 plays a critical role in postsynapse maturation by facilitating the GluN2 subunit switch and proteostasis of palmitoylated proteins.

HIV-1 Tat alters neuronal intrinsic excitability.

OBJECTIVE: In HIV+ individuals, the virus enters the central nervous system and invades innate immune cells, producing important changes that result in neurological deficits. We aimed to determine whether HIV plays a direct role in neuronal excitability. Of the HIV peptides, Tat is secreted and acts in other cells. In order to examine whether the HIV Tat can modify neuronal excitability, we exposed primary murine hippocampal neurons to that peptide, and tested its effects on the intrinsic membrane properties, 4 and 24 h after exposure. RESULTS: The exposure of hippocampal pyramidal neurons to Tat for 4 h did not alter intrinsic membrane properties. However, we found a strong increase in intrinsic excitability, characterized by increase of the slope (Gain) of the input-output function, in cells treated with Tat for 24 h. Nevertheless, Tat treatment for 24 h did not alter the resting membrane potential, input resistance, rheobase and action potential threshold. Thus, neuronal adaptability to Tat exposure for 24 h is not applicable to basic neuronal properties. A restricted but significant effect on coupling the inputs to the outputs may have implications to our knowledge of Tat biophysical firing capability, and its involvement in neuronal hyperexcitability in neuroHIV.

c-Fos mapping of brain regions activated by multi-modal and electric foot shock stress.

Real-world stressors are complex and multimodal, involving physical, psychological, and social dimensions. However, the brain networks that mediate stress responses to these stimuli need to be further studied. We used c-Fos mapping in mice to characterize brain circuits activated by exposure to a single episode of multimodal stress (MMS), and compared these to circuits activated by electric foot shocks (EFS). We focused on characterizing c-Fos activity in stress-relevant brain regions including the paraventricular nucleus (PVN) of the hypothalamus and the bed nucleus of the stria terminalis (BNST). We also assessed stress-induced activation of CRH-positive neurons in each of these structures. MMS and EFS activated an overlapping network of brain regions with a similar time course. c-Fos expression within the PVN and the BNST peaked 30-60 min after exposure to both MMS and EFS, and returned to baseline levels within 24 h. Quantification of c-Fos expression within BNST subregions revealed that while c-Fos expression peaked in all subregions 30-60 min after MMS and EFS exposure, the neuronal density of c-Fos expression was significantly higher in the dorsomedial and ventral BNST relative to the dorsolateral BNST. Our preliminary assessment indicated that a great majority of MMS or EFS-activated neurons in the PVN were CRH-positive (>87%); in contrast, about 6-35% of activated neurons in the BNST were CRH-positive. Our findings indicate that both MMS and EFS are effective at activating stress-relevant brain areas and support the use of MMS as an effective approach for studying multidimensional stress in animal models. The results also reveal that the PVN and BNST are part of a common neural circuit substrate involved in neural processing related to stress.

Opiate dependence induces cell type-specific plasticity of intrinsic membrane properties in the rat juxtacapsular bed nucleus of stria terminalis (jcBNST).

RATIONALE: Drugs of abuse can alter circuit dynamics by modifying synaptic efficacy and/or the intrinsic membrane properties of neurons. The juxtacapsular subdivision of the bed nucleus of stria terminalis (jcBNST) has unique connectivity that positions it to integrate cortical and amygdala inputs and provide feed-forward inhibition to the central nucleus of the amygdala (CeA), among other regions. In this study, we investigated changes in the synaptic and intrinsic properties of neurons in the rat jcBNST during protracted withdrawal from morphine dependence using a combination of conventional electrophysiological methods and the dynamic clamp technique. RESULTS: A history of opiate dependence induced a form of cell type-specific plasticity characterized by reduced inward rectification associated with more depolarized resting membrane potentials and increased membrane resistance. This cell type also showed a lower rheobase when stimulated with direct current (DC) pulses as well as a decreased firing threshold under simulated synaptic bombardment with the dynamic clamp. Morphine dependence also decreased excitatory postsynaptic potential amplification, suggesting the downregulation of the persistent Na+ current (I NaP). CONCLUSION: These findings show that a history of morphine dependence leads to persistent cell type-specific plasticity of the passive membrane properties of a jcBNST neuronal population, leading to an overall increased excitability of such neurons. By altering the activity of extended amygdala circuits where they are embedded, changes in the integration properties of jcBNST neurons may contribute to emotional dysregulation associated with drug dependence and withdrawal.

Phosphatidylinositol 3-kinase is required for the expression but not for the induction or the maintenance of long-term potentiation in the hippocampal CA1 region.

Several signal transduction pathways have been implicated in the induction of long-term potentiation (LTP), yet the signal transduction mechanisms behind the maintenance-expression phase of LTP are still poorly understood. We investigated the role of phosphatidylinositol 3-kinase (PI3-kinase) in LTP at Schaffer collateral/commissural fiber-CA1 synapses in rat hippocampal slices using biochemical approaches and extracellular electrophysiological recordings. We observed that PI3-kinase activity was induced in the CA1 region during LTP of field EPSPs (fEPSPs) and that two structurally unrelated PI3-kinase inhibitors, LY294002 and wortmannin, abated established LTP, suggesting that PI3-kinase is involved in the maintenance-expression phase of LTP. However, LTP recovered after washout of the reversible PI3-kinase inhibitor LY294002, confirming that LTP maintenance and expression are distinct events and indicating that PI3-kinase activity is required for LTP expression rather than for its maintenance. Interestingly, preincubation with LY294002 did not prevent LTP induction. In fact, if LY294002 was withdrawn 5 min after high-frequency stimulation, an LTP of fEPSP was seen. Last, a voltage-dependent calcium channel-dependent form of LTP in the CA1 could also be reversibly abated by LY294002, raising the possibility that PI3-kinase could be required for the expression of multiple forms of synaptic potentiation.

Presynaptic modulation of spontaneous inhibitory postsynaptic currents by gamma-hydroxybutyrate in the substantia nigra pars compacta.

The regulation of GABA release from the inhibitory input to dopamine cells in the substantia nigra pars compacta (SNc) plays a key role in different reward-related behaviors. Gamma-hydroxybutyrate (GHB) has therapeutical properties in various psychiatric disorders, especially in alcohol abuse. GHB is also used as a drug of abuse, which induces sedation and euphoria. Using whole-cell patch-clamp recordings, we studied the effects of GHB on GABA release in the SNc by recording spontaneous inhibitory postsynaptic currents (sIPSCs) in brain slices of 21- to 25-day-old rats. We found that GHB depressed the frequency and amplitude of sIPSCs, while the frequency and the amplitude of miniature inhibitory postsynaptic currents (mIPSCs), recorded in the presence of TTX, were not affected. However, in the presence of high extracellular potassium (15 mM), which increases the contribution of voltage-dependent calcium channels, GHB induced a reduction in the frequency of the mIPSCs without any effect on their amplitude. All of these effects were GABA(B)-independent and they were blocked by the GHB receptor antagonist NCS-382. The present results indicate that GHB inhibits spontaneous inhibitory synaptic transmission recorded from dopaminergic neurons in the SNc likely by reducing voltage-dependent calcium influx involved in presynaptic GABA release.

Gamma-hydroxybutyrate reduces GABA(A)-mediated inhibitory postsynaptic potentials in the CA1 region of hippocampus.

Gamma-hydroxybutyric acid (GHB) is a psychoactive drug and a putative neurotransmitter, derived from gamma-aminobutyric acid (GABA). At micromolar concentrations GHB binds to specific high and low affinity binding sites present in discrete areas of the brain, while at millimolar concentrations GHB also binds to GABA(B) receptors. Previous studies indicated that GHB inhibits both NMDA and AMPA receptor mediated excitatory postsynaptic potentials in hippocampal CA1 pyramidal neurons. This action of GHB occurs in the presence of GABA(B) blockade and is antagonized by NCS-382, a specific GHB receptor antagonist, suggesting that it is mediated by GHB receptors. In the present study, we have investigated the effect of GHB on GABA(A) mediated inhibitory postsynaptic potentials (GABA(A)-IPSP) elicited in CA1 hippocampal pyramidal neurons by stimulation of Schaffer collateral-commissural fibers. We observed that GHB inhibited GABA(A)-IPSPs by about 40% at concentrations of 300-600 microM. GHB inhibition was blocked by NCS-382 (500 microM), which per se failed to modify GABA(A)-IPSPs. Moreover, GHB failed to modify cell membrane depolarization induced by the brief pressure application of GABA in the presence of tetrodotoxin (TTX), indicating that GHB does not inhibit postsynaptic GABA responses. However, GHB reduced the amplitude of GABA(A)-IPSPs elicited in pyramidal neurons by paired pulse stimulation and enhanced paired pulse facilitation with respect to control condition, suggesting that GHB reduces GABA release from nerve terminals. Finally, GHB failed to reduce the amplitude of GABA(A)-IPSPs in the presence of BaCl(2), suggesting that the effect of GHB is due to GHB receptor-mediated presynaptic inhibition of Ca(2)+ influx.

HIV secreted protein Tat prevents long-term potentiation in the hippocampal CA1 region.

HIV-associated dementia (HAD) is a complication of advanced HIV disease. Both viral products and host cytokines are believed to be involved in the pathogenesis of HIV-associated neurological manifestations. Among the viral products released by HIV-infected cells is the soluble protein Tat. We investigated the effect of exposure of organotypic hippocampal slices to 100 nM recombinant Tat 1-86 on long-term potentiation (LTP) of field excitatory postsynaptic potential (fEPSP) at Schaffer collateral/commissural fiber-CA1 synapses. Exposure to Tat 1-86 prevented the induction of LTP without affecting post-tetanic potentiation. Tat 1-72delta31-61, which lacks the neurotoxic domain of Tat, had no significant effect on LTP. Tat's ability to disrupt synaptic plasticity may be relevant to the pathogenesis of the cognitive impairments seen in patients with HIV disease.

The metabotropic glutamate receptor 5 is necessary for late-phase long-term potentiation in the hippocampal CA1 region.

Selective antagonists of the metabotropic receptors 1 (mGluR1), +/-2-methyl-4-carboxyphenylglycine (LY367385), and mGluR5, 2-methyl-6-(phenylethynyl)-pyridine (MPEP), were used to investigate the role of group I metabotropic receptors in late-phase long-term potentiation (L-LTP) at Schaffer collateral/commissural fiber-CA1 synapses in rat hippocampal slices. L-LTP was induced with three trains of tetanization of 1 s duration at 100 Hz separated by 10-min intervals. Neither LY367385 nor MPEP affected basal synaptic responses at the doses used (200 and 10 microM, respectively) and only the mGluR5 inhibitor MPEP blocked L-LTP. However, in agreement with previous mouse mutant studies, we found that both LY367385 and MPEP inhibited the induction of an LTP obtained with a single train of tetanization of 1 s duration at 100 Hz. MPEP's ability to disrupt L-LTP was not due to an effect on NMDA responses since it did not affect pharmacologically isolated N-methyl-D-aspartate (NMDA) excitatory postsynaptic potentials (EPSPs). However, MPEP prevented the increased phosphorylation in dendrites of p70 S6 kinase (p70(S6K)) at Thr3889, a major regulator of translation required for the induction of protein synthesis-dependent forms of LTP.

Gamma-hydroxybutyrate and ethanol depress spontaneous excitatory postsynaptic currents in dopaminergic neurons of the substantia nigra.

Gamma-hydroxybutyrate (GHB) has been shown to have therapeutical properties in various psychiatric disorders, especially in alcohol abuse, and to mimic different actions of ethanol at the cellular and system level. Using whole-cell patch-clamp recordings on brain slices of 21- to 25-day-old rats, the present study investigated the effects of GHB and ethanol on spontaneous excitatory postsynaptic currents (sEPSCs) in dopaminergic neurons of the substantia nigra pars compacta (SNc). sEPSCs are an index of glutamate release from the excitatory input to dopamine cells, which play a key role in different reward-related behaviors. We found that GHB and ethanol depressed both the frequency and the amplitude of sEPSCs. These effects were GABA(B)-independent and the GHB-induced depression was blocked by the GHB receptor antagonist 6,7,8,9-tetrahydro-5[H]benzocyclohepte-5-ol-4-ylidene acetic acid (NCS-382), pointing to a specific effect of this drug. The effects of ethanol were not affected by NCS-382. This study indicates that GHB and ethanol share the effect of reducing the efficacy of excitatory glutamatergic neurotransmission in the SNc by acting through different mechanisms.

Excitability of jcBNST neurons is reduced in alcohol-dependent animals during protracted alcohol withdrawal.

Alcohol dependence and withdrawal has been shown to cause neuroadaptive changes at multiple levels of the nervous system. At the neuron level, adaptations of synaptic connections have been extensively studied in a number of brain areas and accumulating evidence also shows the importance of alcohol dependence-related changes in the intrinsic cellular properties of neurons. At the same time, it is still largely unknown how such neural adaptations impact the firing and integrative properties of neurons. To address these problems, here, we analyze physiological properties of neurons in the bed nucleus of stria terminalis (jcBNST) in animals with a history of alcohol dependence. As a comprehensive approach, first we measure passive and active membrane properties of neurons using conventional current clamp protocols and then analyze their firing responses under the action of simulated synaptic bombardment via dynamic clamp. We find that most physiological properties as measured by DC current injection are barely affected during protracted withdrawal. However, neuronal excitability as measured from firing responses under simulated synaptic inputs with the dynamic clamp is markedly reduced in all 3 types of jcBNST neurons. These results support the importance of studying the effects of alcohol and drugs of abuse on the firing properties of neurons with dynamic clamp protocols designed to bring the neurons into a high conductance state. Since the jcBNST integrates excitatory inputs from the basolateral amygdala (BLA) and cortical inputs from the infralimbic and the insular cortices and in turn is believed to contribute to the inhibitory input to the central nucleus of the amygdala (CeA) the reduced excitability of the jcBNST during protracted withdrawal in alcohol-dependent animals will likely affect ability of the jcBNST to shape the activity and output of the CeA.

Consistency and diversity of spike dynamics in the neurons of bed nucleus of stria terminalis of the rat: a dynamic clamp study.

Neurons display a high degree of variability and diversity in the expression and regulation of their voltage-dependent ionic channels. Under low level of synaptic background a number of physiologically distinct cell types can be identified in most brain areas that display different responses to standard forms of intracellular current stimulation. Nevertheless, it is not well understood how biophysically different neurons process synaptic inputs in natural conditions, i.e., when experiencing intense synaptic bombardment in vivo. While distinct cell types might process synaptic inputs into different patterns of action potentials representing specific "motifs" of network activity, standard methods of electrophysiology are not well suited to resolve such questions. In the current paper we performed dynamic clamp experiments with simulated synaptic inputs that were presented to three types of neurons in the juxtacapsular bed nucleus of stria terminalis (jcBNST) of the rat. Our analysis on the temporal structure of firing showed that the three types of jcBNST neurons did not produce qualitatively different spike responses under identical patterns of input. However, we observed consistent, cell type dependent variations in the fine structure of firing, at the level of single spikes. At the millisecond resolution structure of firing we found high degree of diversity across the entire spectrum of neurons irrespective of their type. Additionally, we identified a new cell type with intrinsic oscillatory properties that produced a rhythmic and regular firing under synaptic stimulation that distinguishes it from the previously described jcBNST cell types. Our findings suggest a sophisticated, cell type dependent regulation of spike dynamics of neurons when experiencing a complex synaptic background. The high degree of their dynamical diversity has implications to their cooperative dynamics and synchronization.

A potential role for adiponectin receptor 2 (AdipoR2) in the regulation of alcohol intake.

The anterior cingulate cortex (ACC) has been implicated in alcohol and drug addiction. We recently identified the small G protein K-ras as an alcohol-regulated gene in the ACC by gene expression analysis. We show here that the adiponectin receptor 2 (AdipoR2) was differentially regulated by alcohol in the ACC in a K-ras-dependent manner. Additionally, withdrawal-associated increased drinking was attenuated in AdipoR2 null mice. Intracellular recordings revealed that adiponectin increased the excitability of ACC neurons and that this effect was more pronounced during alcohol withdrawal, suggesting that AdipoR2 signaling may contribute to increased ACC activity. Altogether, the data implicate K-ras-regulated pathways involving AdipoR2 in the cellular and behavioral actions of alcohol that may contribute to overactivity of the ACC during withdrawal and excessive alcohol drinking.

Intrinsic neuronal plasticity in the juxtacapsular nucleus of the bed nuclei of the stria terminalis (jcBNST).

The juxtacapsular nucleus of the anterior division of the BNST (jcBNST) receives robust glutamatergic projections from the basolateral nucleus of the amygdala (BLA), the postpiriform transition area, and the insular cortex as well as dopamine (DA) inputs from the midbrain. In turn the jcBNST sends GABAergic projections to the medial division of the central nucleus of the amygdala (CEAm) as well as other brain regions. We recently described a form of long-term potentiation of the intrinsic excitability (LTP-IE) of neurons of the juxtacapsular nucleus of BNST (jcBNST) in response to high-frequency stimulation (HFS) of the stria terminalis that was impaired during protracted withdrawal from alcohol, cocaine, and heroin and in rats chronically treated with corticotropin-releasing factor (CRF) intracerebroventricularly. Here we show that DAergic neurotransmission is required for the induction of LTP-IE of jcBNST neurons through dopamine (DA) D1 receptors. Thus, activation of the central CRF stress system and altered DAergic neurotransmission during protracted withdrawal from alcohol and drugs of abuse may contribute to the disruption of LTP-IE in the jcBNST. Impairment of this form of intrinsic neuronal plasticity in the jcBNST could result in inadequate neuronal integration and reduced inhibition of the CEA, contributing to the negative affective state that characterizes protracted abstinence in post-dependent individuals. These results provide a novel neurobiological target for vulnerability to alcohol and drug dependence.