GABAB receptors induce phasic release from medial habenula terminals through activity-dependent recruitment of release-ready vesicles.

GABAB receptor (GBR) activation inhibits neurotransmitter release in axon terminals in the brain, except in medial habenula (MHb) terminals, which show robust potentiation. However, mechanisms underlying this enigmatic potentiation remain elusive. Here, we report that GBR activation on MHb terminals induces an activity-dependent transition from a facilitating, tonic to a depressing, phasic neurotransmitter release mode. This transition is accompanied by a 4.1-fold increase in readily releasable vesicle pool (RRP) size and a 3.5-fold increase of docked synaptic vesicles (SVs) at the presynaptic active zone (AZ). Strikingly, the depressing phasic release exhibits looser coupling distance than the tonic release. Furthermore, the tonic and phasic release are selectively affected by deletion of synaptoporin (SPO) and Ca2+-dependent activator protein for secretion 2 (CAPS2), respectively. SPO modulates augmentation, the short-term plasticity associated with tonic release, and CAPS2 retains the increased RRP for initial responses in phasic response trains. The cytosolic protein CAPS2 showed a SV-associated distribution similar to the vesicular transmembrane protein SPO, and they were colocalized in the same terminals. We developed the "Flash and Freeze-fracture" method, and revealed the release of SPO-associated vesicles in both tonic and phasic modes and activity-dependent recruitment of CAPS2 to the AZ during phasic release, which lasted several minutes. Overall, these results indicate that GBR activation translocates CAPS2 to the AZ along with the fusion of CAPS2-associated SVs, contributing to persistency of the RRP increase. Thus, we identified structural and molecular mechanisms underlying tonic and phasic neurotransmitter release and their transition by GBR activation in MHb terminals.

Synaptotagmin-11 facilitates assembly of a presynaptic signaling complex in post-Golgi cargo vesicles.

GABAB receptors (GBRs), the G protein-coupled receptors for GABA, regulate synaptic transmission throughout the brain. A main synaptic function of GBRs is the gating of Cav2.2-type Ca2+ channels. However, the cellular compartment where stable GBR/Cav2.2 signaling complexes form remains unknown. In this study, we demonstrate that the vesicular protein synaptotagmin-11 (Syt11) binds to both the auxiliary GBR subunit KCTD16 and Cav2.2 channels. Through these dual interactions, Syt11 recruits GBRs and Cav2.2 channels to post-Golgi vesicles, thus facilitating assembly of GBR/Cav2.2 signaling complexes. In addition, Syt11 stabilizes GBRs and Cav2.2 channels at the neuronal plasma membrane by inhibiting constitutive internalization. Neurons of Syt11 knockout mice exhibit deficits in presynaptic GBRs and Cav2.2 channels, reduced neurotransmitter release, and decreased GBR-mediated presynaptic inhibition, highlighting the critical role of Syt11 in the assembly and stable expression of GBR/Cav2.2 complexes. These findings support that Syt11 acts as a vesicular scaffold protein, aiding in the assembly of signaling complexes from low-abundance components within transport vesicles. This mechanism enables insertion of pre-assembled functional signaling units into the synaptic membrane.

Cell-Type-Specific Effects of Somatostatin on Synaptic Transmission in the Anterior Cingulate Cortex.

Inhibitory modulation of glutamatergic information processing is a prerequisite for proper network function. Among the many groups of interneurons (INs), somatostatin-expressing interneurons (SOM-INs) play an important role in the maintenance of physiological brain activity. We have previously shown that somatostatin (SOM) causes a reduction in pyramidal cell (PC) excitability. However, the mechanisms of action of the peptide on cortical synaptic circuits are still unclear. To understand the effects of the neuropeptide SOM on cortical synaptic circuits, we performed a detailed side-by-side comparison of its postsynaptic effects on PCs, SOM-INs, and layer 1 interneurons (L1-INs) in the anterior cingulate cortex of male and female mice and found that SOM produced pronounced postsynaptic effects in PCs while having little to no effect on either IN type. This comparison allowed us to link the observed postsynaptic effects to SOM-induced modulations of glutamatergic and GABAergic synaptic transmission and to trace the impact of the neuropeptide on the neuronal circuitry between these three cell types. We show here that SOM depresses glutamatergic synaptic transmission via a presynaptic mechanism while exerting a differential impact on GABAA receptor- and GABAB receptor-mediated transmission at the pre- and postsynaptic level resulting in a shift of inhibition in L2/3 PCs from L1-INs to SOM-INs. In summary, this study unravels a novel aspect by which SOM modulates synaptic signaling between PCs, L1-INs, and SOM-INs.

Presynaptic hyperexcitability reversed by positive allosteric modulation of a GABABR epilepsy variant.

GABABRs are key membrane proteins that continually adapt the excitability of the nervous system. These G-protein coupled receptors are activated by the brain's premier inhibitory neurotransmitter GABA. They are obligate heterodimers composed of GABA-binding GABABR1 and G-protein-coupling GABABR2 subunits. Recently, three variants (G693W, S695I, I705N) have been identified in the gene (GABBR2) encoding for GABABR2. Individuals that harbour any of these variants exhibit severe developmental epileptic encephalopathy and intellectual disability, but the underlying pathogenesis that is triggered in neurons, remains unresolved. Using a range of confocal imaging, flow cytometry, structural modelling, biochemistry, live cell Ca2+ imaging of presynaptic terminals, whole-cell electrophysiology of HEK-293T cells and neurons, and two-electrode voltage clamping of Xenopus oocytes we have probed the biophysical and molecular trafficking and functional profiles of G693W, S695I and I705N variants. We report that all three point mutations impair neuronal cell surface expression of GABABRs, reducing signalling efficacy. However, a negative effect evident for one variant perturbed neurotransmission by elevating presynaptic Ca2+ signalling. This is reversed by enhancing GABABR signalling via positive allosteric modulation. Our results highlight the importance of studying neuronal receptors expressed in nervous system tissue and provide new mechanistic insights into how GABABR variants can initiate neurodevelopmental disease whilst highlighting the translational suitability and therapeutic potential of allosteric modulation for correcting these deficits.

Structural and Biophysical Mechanisms of Class C G Protein-Coupled Receptor Function.

Groundbreaking structural and spectroscopic studies of class A G protein-coupled receptors (GPCRs), such as rhodopsin and the ß2 adrenergic receptor, have provided a picture of how structural rearrangements between transmembrane helices control ligand binding, receptor activation, and effector coupling. However, the activation mechanism of other GPCR classes remains more elusive, in large part due to complexity in their domain assembly and quaternary structure. In this review, we focus on the class C GPCRs, which include metabotropic glutamate receptors (mGluRs) and gamma-aminobutyric acid B (GABAB) receptors (GABABRs) most prominently. We discuss the unique biophysical questions raised by the presence of large extracellular ligand-binding domains (LBDs) and constitutive homo/heterodimerization. Furthermore, we discuss how recent studies have begun to unravel how these fundamental class C GPCR features impact the processes of ligand binding, receptor activation, signal transduction, regulation by accessory proteins, and crosstalk with other GPCRs.

Inhibitory Gating of Thalamocortical Inputs onto Rat Gustatory Insular Cortex.

In primary gustatory cortex (GC), a subregion of the insular cortex, neurons show anticipatory activity, encode taste identity and palatability, and their activity is related to decision-making. Inactivation of the gustatory thalamus, the parvicellular region of the ventral posteromedial thalamic nucleus (VPMpc), dramatically reduces GC taste responses, consistent with the hypothesis that VPMpc-GC projections carry taste information. Recordings in awake rodents reported that taste-responsive neurons can be found across GC, without segregated spatial mapping, raising the possibility that projections from the taste thalamus may activate GC broadly. In addition, we have shown that cortical inhibition modulates the integration of thalamic and limbic inputs, revealing a potential role for GABA transmission in gating sensory information to GC. Despite this wealth of information at the system level, the synaptic organization of the VPMpc-GC circuit has not been investigated. Here, we used optogenetic activation of VPMpc afferents to GC in acute slice preparations from rats of both sexes to investigate the synaptic properties and organization of VPMpc afferents in GC and their modulation by cortical inhibition. We hypothesized that VPMpc-GC synapses are distributed across GC, but show laminar- and cell-specific properties, conferring computationally flexibility to how taste information is processed. We also found that VPMpc-GC synaptic responses are strongly modulated by the activity regimen of VPMpc afferents, as well as by cortical inhibition activating GABAA and GABAB receptors onto VPMpc terminals. These results provide a novel insight into the complex features of thalamocortical circuits for taste processing.SIGNIFICANCE STATEMENT We report that the input from the primary taste thalamus to the primary gustatory cortex (GC) shows distinct properties compared with primary thalamocortical synapses onto other sensory areas. Ventral posteromedial thalamic nucleus afferents in GC make synapses with excitatory neurons distributed across all cortical layers and display frequency-dependent short-term plasticity to repetitive stimulation; thus, they do not fit the classic distinction between drivers and modulators typical of other sensory thalamocortical circuits. Thalamocortical activation of GC is gated by cortical inhibition, providing local corticothalamic feedback via presynaptic ionotropic and metabotropic GABA receptors. The connectivity and inhibitory control of thalamocortical synapses in GC highlight unique features of the thalamocortical circuit for taste.

Postsynaptic GABAB-receptor mediated currents in diverse dentate gyrus interneuron types.

The processing of rich synaptic information in the dentate gyrus (DG) relies on a diverse population of inhibitory GABAergic interneurons to regulate cellular and circuit activity, in a layer-specific manner. Metabotropic GABAB-receptors (GABABRs) provide powerful inhibition to the DG circuit, on timescales consistent with behavior and learning, but their role in controlling the activity of interneurons is poorly understood with respect to identified cell types. We hypothesize that GABABRs display cell type-specific heterogeneity in signaling strength, which will have direct ramifications for signal processing in DG networks. To test this, we perform in vitro whole-cell patch-clamp recordings from identified DG principal cells and interneurons, followed by GABABR pharmacology, photolysis of caged GABA, and extracellular stimulation of endogenous GABA release to classify the cell type-specific inhibitory potential. Based on our previous classification of DG interneurons, we show that postsynaptic GABABR-mediated currents are present on all interneuron types albeit at different amplitudes, dependent largely on soma location and synaptic targets. GABABRs were coupled to inwardly-rectifying K+ channels that strongly reduced the excitability of those interneurons where large currents were observed. These data provide a systematic characterization of GABABR signaling in the rat DG to provide greater insight into circuit dynamics.

GABAB receptor-mediated modulation in the developing dorsal nucleus of the lateral lemniscus.

The dorsal nucleus of the lateral lemniscus (DNLL) is a GABAergic, reciprocally connected auditory brainstem structure that continues to develop postnatally in rodents. One key feature of the DNLL is the generation of a strong, prolonged, ionotropic, GABAA receptor-mediated inhibition. Possible GABAB receptor-mediated signalling is unexplored in the DNLL. Here, we used Mongolian gerbils of either sex to describe GABAB receptor-mediated modulation of postsynaptic potassium currents and synaptic inputs in postnatal (P) animals of days 10/11 and 23-28. Throughout development, we observed the presence of a Baclofen-activated GABAB receptor-enhanced potassium outward conductance that is capable of suppressing action potential generation. In P10/11, old gerbils GABAB receptor activation enhances glutamatergic and suppresses ionotropic GABAergic synaptic transmission. During development, this differential modulation becomes less distinct, because in P22-28, old animals Baclofen-activated GABAB receptors rather enhance ionotropic GABAergic synaptic transmission, whereas glutamatergic transmission is both enhanced and suppressed. Blocking GABAB receptors causes an increase in ionotropic GABAergic transmission in P10/11 old gerbils that was independent on stimulation frequency but depended on the type of short-term plasticity. Together with the lack of Baclofen-induced changes in the synaptic paired-pulse ratio of either input type, we suggest that GABAB receptor-mediated modulation is predominantly postsynaptic and activates different signalling cascades. Thus, we argue that in DNLL neurons, the GABAB receptor is a post-synaptically located signalling hub that alters signalling cascades during development for distinct targets.

Acute Cerebellar Manifestations without Limbic Involvement in GABAB Receptor Autoimmune Encephalitis: Case Report and Literature Review.

Autoimmune encephalitis is a rapidly progressive inflammatory brain disease. Gamma-aminobutyric acid type B (GABAB) receptor autoimmune encephalitis is a rare subtype characterized by distinct clinical features. Diagnosis can be especially challenging when typical limbic symptoms and neuroimaging findings are absent. This case report underscores the importance of identifying this condition and starting immunosuppressive treatment promptly. A 59-year-old man presented with gait disturbances, dysarthria, and severe ataxia without cognitive impairment. Initial examinations, including a brain MRI, were unremarkable, except for an elevated cell count and protein in the cerebrospinal fluid. Despite receiving initial empirical antiviral treatment, his symptoms worsened, prompting the administration of intravenous methylprednisolone and immunoglobulin. After these immunosuppressive therapies, the cerebellar symptoms showed gradual improvement. Subsequent testing for antibodies to the GABAB receptor was positive in both the serum and cerebrospinal fluid. Follow-up MRI revealed cerebellar atrophy, consistent with a diagnosis of GABAB receptor-associated acute cerebellitis. This case illustrates that cerebellar symptoms can occur in the absence of more common limbic manifestations in GABAB receptor autoimmune encephalitis. The progression of cerebellar atrophy following an initially normal MRI is a significant finding that offers supporting evidence for the diagnosis of cerebellitis. A review of the literature identified similar cases of acute cerebellitis without limbic symptoms, although neuroimaging abnormalities in the cerebellum were not reported. Our case underscores the importance of increased clinical awareness and consideration of autoimmune causes, even when neuroimaging appears normal. Early and appropriate immunosuppressive therapy may help change the course of the disease and enhance patient outcomes.

Positive allosteric modulators of the GABAB receptor: a new class of ligands with therapeutic potential for alcohol use disorder.

BACKGROUND: Positive allosteric modulators (PAMs) of the GABAB receptor constitute a new class of GABAB-receptor ligands. GABAB PAMs reproduce several pharmacological effects of the orthosteric GABAB receptor agonist, baclofen, although displaying a better safety profile. AIMS: This paper reviews the reducing or, frequently, even suppressing effects of all GABAB PAMs tested to date on multiple alcohol-related behaviours in laboratory rodents exposed to validated experimental models of human alcohol use disorder. RESULTS: Acute or repeated treatment with CGP7930, GS39783, BHF177, rac-BHFF, ADX71441, CMPPE, COR659, ASP8062, KK-92A, and ORM-27669 reduced excessive alcohol drinking, relapse- and binge-like drinking, operant alcohol self-administration, reinstatement of alcohol seeking, and alcohol-induced conditioned place preference in rats and mice. CONCLUSIONS: These effects closely mirrored those of baclofen; notably, they were associated to remarkably lower levels of tolerance and toxicity. The recent transition of ASP8062 to clinical testing will soon prove whether these highly consistent preclinical data translate to AUD patients.

Brain carbon monoxide can suppress the rat micturition reflex through brain γ-aminobutyric acid receptors.

OBJECTIVES: To investigate roles of brain carbon monoxide (CO), an endogenous gasotransmitter, in regulation of the rat micturition reflex. METHODS: In urethane-anesthetized (0.8 g/kg, ip) male rats, evaluation of urodynamic parameters was started 1 h before intracerebroventricular administration of CORM-3 (CO donor) or ZnPP (non-selective inhibitor of heme oxygenase, a CO producing enzyme) and continued for 2 h after the administration. We also investigated effects of centrally pretreated SR95531 (GABAA receptor antagonist) or SCH50911 (GABAB receptor antagonist) on the CORM-3-induced response. RESULTS: CORM-3 significantly prolonged intercontraction intervals (ICIs) without changing maximal voiding pressure (MVP), while ZnPP significantly shortened ICI and reduced single-voided volume and bladder capacity without affecting MVP, post-voided residual volume, or voiding efficiency. The ZnPP-induced ICI shortening was reversed by CORM-3. The CORM-3-induced ICI prolongation was significantly attenuated by centrally pretreated SR95531 or SCH50911, respectively. CONCLUSIONS: Brain CO can suppress the rat micturition reflex through brain γ-aminobutyric acid (GABA) receptors.

KCTD8 and KCTD12 Facilitate Axonal Expression of GABAB Receptors in Habenula Cholinergic Neurons.

GABAB receptors in habenula cholinergic neurons mediate strong presynaptic excitation and control aversive memory expression. K+ channel tetramerization domain (KCTD) proteins are key interacting partners of GABAB receptors; it remains unclear whether and how KCTDs contribute to GABAB excitatory signaling. Here, we show that KCTD8 and KCTD12 in these neurons facilitate the GABAB receptors expression in axonal terminals and contribute to presynaptic excitation by GABAB receptors. Genetically knocking out KCTD8/12/16 or KCTD8/12, but not other combinations of the three KCTD isoforms, substantially reduced GABAB receptors-mediated potentiation of glutamate release and presynaptic Ca2+ entry in response to axonal stimulation, whereas they had no effect on GABAB-mediated inhibition in the somata of cholinergic neurons within the habenulo-interpeduncular pathway in mice of either sex. The physiological phenotypes were associated with a significant decrease in the GABAB expression within the axonal terminals but not the somata. Overexpressing either KCTD8 or KCTD12 in the KCTD8/12/16 triple knock-out mice reversed the changes in axonal GABAB expression and presynaptic excitation. In mice lacking the KCTDs, aversion-predicting cues produced stronger neuronal activation in the interpeduncular nucleus, and the infusion of GABAB agonist in this nucleus produced a weaker effect on fear extinction. Collectively, our results reveal isoform-specific roles of KCTD proteins in enriching the axonal expression of GABAB receptors, facilitating their presynaptic signaling, and modulating aversion-related memory processes.SIGNIFICANCE STATEMENT GABAB receptors represent the principal inhibitory neurotransmitter receptor, but they mediate strong presynaptic excitation in the habenulo-interpeduncular pathway and modulate aversion memory expression. KCTD proteins are integral constituents of GABAB receptors. By analyzing the physiological, neuroanatomical, and behavioral phenotypes of multiple KCTD knock-out mouse lines, we show that KCTD8 and KCTD12 facilitate the axonal expression and hence presynaptic excitation of GABAB receptors in habenula cholinergic neurons and control cued-aversion memory formation and expression in the habenulo-interpeduncular pathway. These results expand the physiological and behavioral functions of KCTDs in modulating the brain neural circuits.

Deletion of GABAB receptors from Kiss1 cells affects glucose homeostasis without altering reproduction in male mice.

Kisspeptin and γ-amino butyric acid (GABA), synthesized in the central nervous system, are critical for reproduction. Both are also expressed in peripheral organs/tissues critical to metabolic control (liver/pancreas/adipose). Many kisspeptin neurons coexpress GABAB receptors (GABABR) and GABA controls kisspeptin expression and secretion. We developed a unique mouse lacking GABABR exclusively from kisspeptin cells/neurons (Kiss1-GABAB1KO) to evaluate the impact on metabolism/reproduction. We confirmed selective deletion of GABABR from Kiss1 cells in the anteroventral periventricular nucleus/periventricular nucleus continuum (AVPV/PeN; immunofluorescence and PCR) and arcuate nucleus (ARC), medial amygdala (MeA), pituitary, liver, and testes (PCR). Young Kiss1-GABAB1KO males were fertile, with normal LH and testosterone. Kiss1 expression was similar between genotypes in AVPV/PeN, ARC, MeA, bed nucleus of the stria terminalis (BNST), and peripheral organs (testis, liver, pituitary). Kiss1-GABAB1KO males presented higher fasted glycemia and insulin levels, an impaired response to a glucose overload, reduced insulin sensitivity, and marked insulin resistance. Interestingly, when Kiss1-GABAB1KO males got older (9 mo old) their body weight (BW) increased, in part due to an increase in white adipose tissue (WAT). Old Kiss1-GABAB1KO males showed higher fasted insulin, increased pancreatic insulin content, insulin resistance, and significantly decreased pancreatic kisspeptin levels. In sum, lack of GABABR specifically in Kiss1 cells severely impacts glucose homeostasis in male mice, reinforcing kisspeptin involvement in metabolic regulation. These alterations in glucose homeostasis worsened with aging. We highlight the impact of GABA through GABABR in the regulation of the pancreas kisspeptin system in contrast to liver kisspeptin that was not affected.NEW & NOTEWORTHY We developed a unique mouse lacking GABAB receptors specifically in Kiss1 cells to evaluate the impact on reproduction and metabolism. Knockout males showed a severe impact on glucose homeostasis, which worsened with aging. These results reinforce the proposed kisspeptin involvement in metabolic regulation and highlight the impact of GABA through GABABR in the regulation of the peripheral pancreas kisspeptin system.

Underlying mechanisms of long-term potentiation during the inhibition of the cannabinoid CB1 and GABAB receptors in the dentate gyrus of hippocampus.

BACKGROUND: The release of various neurotransmitters and thereby the excitability of neuronal circuits are regulated by the endocannabinoid system in an activity-dependent manner. Hippocampal long-term potentiation (LTP) is augmented in cannabinoid type 1 (CB1) receptor-deficient mice. CB1 receptors exist on GABAergic axon terminals in the hippocampus. In our previous work, we showed that CB1 antagonists increased the population spike (PS) amplitude, field excitatory post-synaptic potential (fEPSP), and the LTP induction in the dentate gyrus (DG) of the rat hippocampus while the GABAB antagonist decreased these parameters. Determining the underlying mechanisms of the pre- and/or postsynaptic locus of LTP expression is of great importance. In this study, we investigated whether LTP alteration acutely caused by CB1 and GABAB receptor antagonists (AM251 and CGP55845, respectively) happens at the postsynaptic or presynaptic regions, or at both. Therefore, the paired-pulse ratio (PPR) was assessed prior to and following the LTP induction in the studied groups. METHODS: Male Wistar rats were randomly assigned to the groups of control, AM251, CGP55845, CGP55845 + AM251. A high-frequency stimulation (HFS) of the perforant path (PP) was used to induce LTP in the DG region. RESULTS: Statistical analysis revealed that AM251 produced significant increase in excitatory postsynaptic potential (EPSP) slope and amplitude of PS. Conversely, administration of CGP55845 produced decrease in slope of EPSP. The current results indicated that the PPR was not influenced by LTP induction in the presence of AM251 or CGP55845 either alone or their combination. CONCLUSIONS: It can be concluded that the site causing LTP expression is, at least in part, the postsynaptic site because PPR was not influenced by LTP induction in the presence of AM251 or CGP55845 either alone or their combination.

Effect of ASP8062 on morphine self-administration and morphine-induced respiratory suppression in monkeys.

ASP8062 is an orally available GABAB receptor positive allosteric modulator (PAM). This study assessed the potential of ASP8062 for treating opioid use disorder (OUD). Three rhesus monkeys were pretreated with ASP8062 (0.3, 1 or 3 mg/kg) by oral administration 1 h prior to a 2-h morphine self-administration session (0.03 mg/kg, iv, per injection) under a fixed-ratio 5 schedule. We further examined the potential worsening of morphine-induced respiratory suppression by ASP8062 after coadministration of morphine (10 mg/kg, sc) and ASP8062 (10 mg/kg, po) in cynomolgus monkeys using a custom-made whole-body plethysmograph. Plasma concentrations of ASP8062 (3 or 10 mg/kg, po) were assessed in cynomolgus monkeys using liquid chromatography-tandem mass spectroscopy (LC-MS/MS). ASP8062 at 3 mg/kg, po decreased the morphine self-administrations with significant differences from the vehicle-treated group (IC50 = 0.97 ± 0.36 mg/kg). Exposure levels at 3 mg/kg observed in monkeys were comparable to the clinical exposure levels which positive pharmacodynamic effects were previously shown. Further, ASP8062 did not potentiate morphine-induced respiratory suppression up to exposure levels higher than the clinically relevant dose. ASP8062 may reduce opioid use in OUD patients without affecting respiratory system, providing justification for further ASP8062 development as a potential treatment option for OUD.

GIRK channel activity in prelimbic pyramidal neurons regulates the extinction of cocaine conditioned place preference in male mice.

Drug-induced neuroadaptations in the prefrontal cortex (PFC) have been implicated in drug-associated memories that motivate continued drug use. Chronic cocaine exposure increases pyramidal neuron excitability in the prelimbic subregion of the PFC (PL), an adaptation that has been attributed in part to a suppression of inhibitory signalling mediated by the GABAB receptor (GABAB R) and G protein-gated inwardly rectifying K+ (GIRK/Kir3) channels. Although reduced GIRK channel activity in PL pyramidal neurons enhances the motor-stimulatory effect of cocaine in mice, the impact on cocaine reward and associated memories remains unclear. Here, we employed Cre- and CRISPR/Cas9-based viral manipulation strategies to evaluate the impact of GIRK channel or GABAB R ablation in PL pyramidal neurons on cocaine-induced conditioned place preference (CPP) and extinction. Neither ablation of GIRK channels nor GABAB R impacted the acquisition of cocaine CPP. GIRK channel ablation in PL pyramidal neurons, however, impaired extinction of cocaine CPP in male but not female mice. Since ablation of GIRK channels but not GABAB R increased PL pyramidal neuron excitability, we used a chemogenetic approach to determine if acute excitation of PL pyramidal neurons impaired the expression of extinction in male mice. While acute chemogenetic excitation of PL pyramidal neurons induced locomotor hyperactivity, it did not impair the extinction of cocaine CPP. Lastly, we found that persistent enhancement of GIRK channel activity in PL pyramidal neurons accelerated the extinction of cocaine CPP. Collectively, our findings show that the strength of GIRK channel activity in PL pyramidal neurons bi-directionally regulates cocaine CPP extinction in male mice.

Gamma-hydroxybutyrate (GHB), 1,4-butanediol (1,4BD), and gamma-butyrolactone (GBL) intoxication: A state-of-the-art review.

γ-hydroxybutyrate (GHB) is synthesized endogenously from γ-aminobutyric acid (GABA) or exogenously from 1,4-butanediol (butane-1,4-diol; 1,4-BD) or γ-butyrolactone (GBL). GBL, and 1,4-BD are rapidly converted to GHB. The gastric absorption time, volume of distribution, and half-life of GHB are between 5 and 45 min, 0.49 ± 0.9 L/kg, and between 20 and 60 min, respectively. GHB and its analogues have a dose-dependent effect on the activation of GHB receptor, GABA-B, and GABA localized to the central nervous system. After ingestion, most patients present transient neurological disorders (lethal dose: 60 mg/kg). Chronic GHB consumption is associated with disorders of use and a withdrawal syndrome when the consumption is discontinued. GHB, GBL, and 1,4-BD are classified as narcotics but only the use of GHB is controlled internationally. They are used for drug facilitated (sexual) assault, recreational purposes, slamsex, and chemsex. To confirm an exogenous intake or administration of GHB, GBL, or 1-4-BD, the pre-analytical conservation is crucial. The antemortem cutoff doses for detection are 5 and 5-15 mg/L, with detection windows of 6 and 10 h in the blood and urine, respectively Control of GHB is essential to limit the number of users, abuse, associated risks, and death related to their consumption.

Reasons for use and experiences of using phenibut, a mixed methods analysis of online reports.

Background: Phenibut is a drug similar in structure to gabapentin and pregabalin. It is available online without prescription, often marketed as a dietary supplement or amino acid derivative. Little is known about phenibut use despite its increased popularity in the United States over the last decade.Objective: To clarify reasons for taking phenibut, circumstances, and effects of use.Methods: Reports of phenibut, gabapentin, and pregabalin use were downloaded from a publicly-available database, Erowid.org. A mixed methods approach utilizing qualitative content analysis was used.Results: Of 229 reports, 211 were from male authors. People usually purchased phenibut online and reportedly used it for recreation, to manage a medical or psychiatric problem (primarily insomnia, anxiety), as a substitute for other drugs (especially benzodiazepines), to manage withdrawal from another substance (including benzodiazepines, opioids), and/or for performance enhancement. While it shared many reported effects with pregabalin and gabapentin such as anxiolysis, increased talkativeness, and impaired motor coordination, reports of gastrointestinal distress and sedation were more commonly attributed to phenibut. Several people reported difficulty in restricting their use and managing withdrawal.Conclusions: Phenibut reports suggest that phenibut may have some benefits for some people. Use also, however, carries risks of adverse effects, a potentially dangerous withdrawal syndrome, and addiction. Not dissimilar to unprescribed gabapentin or pregabalin, self-medication is a common motive for phenibut use. Physicians should continue to ask their patients about use of any non-prescribed medications, dietary supplements, or "amino acid derivatives."Abbreviation: PWUPh: people who use phenibut; PWUG: people who use gabapentin; PWUPr: people who use pregabalin.

ERK1/2-Dependent Phosphorylation of GABAB1(S867/T872), Controlled by CaMKIIß, Is Required for GABAB Receptor Degradation under Physiological and Pathological Conditions.

GABAB receptor-mediated inhibition is indispensable for maintaining a healthy neuronal excitation/inhibition balance. Many neurological diseases are associated with a disturbed excitation/inhibition balance and downregulation of GABAB receptors due to enhanced sorting of the receptors to lysosomal degradation. A key event triggering the downregulation of the receptors is the phosphorylation of S867 in the GABAB1 subunit mediated by CaMKIIß. Interestingly, close to S867 in GABAB1 exists another phosphorylation site, T872. Therefore, the question arose as to whether phosphorylation of T872 is involved in downregulating the receptors and whether phosphorylation of this site is also mediated by CaMKIIß or by another protein kinase. Here, we show that mutational inactivation of T872 in GABAB1 prevented the degradation of the receptors in cultured neurons. We found that, in addition to CaMKIIß, also ERK1/2 is involved in the degradation pathway of GABAB receptors under physiological and ischemic conditions. In contrast to our previous view, CaMKIIß does not appear to directly phosphorylate S867. Instead, the data support a mechanism in which CaMKIIß activates ERK1/2, which then phosphorylates S867 and T872 in GABAB1. Blocking ERK activity after subjecting neurons to ischemic stress completely restored downregulated GABAB receptor expression to normal levels. Thus, preventing ERK1/2-mediated phosphorylation of S867/T872 in GABAB1 is an opportunity to inhibit the pathological downregulation of the receptors after ischemic stress and is expected to restore a healthy neuronal excitation/inhibition balance.

Impact of GABAA and GABAB Inhibition on Cortical Dynamics and Perturbational Complexity during Synchronous and Desynchronized States.

Quantitative estimations of spatiotemporal complexity of cortical activity patterns are used in the clinic as a measure of consciousness levels, but the cortical mechanisms involved are not fully understood. We used a version of the perturbational complexity index (PCI) adapted to multisite recordings from the ferret (either sex) cerebral cortex in vitro (sPCI) to investigate the role of GABAergic inhibition in cortical complexity. We studied two dynamical states: slow-wave activity (synchronous state) and desynchronized activity, that express low and high causal complexity respectively. Progressive blockade of GABAergic inhibition during both regimes revealed its impact on the emergent cortical activity and on sPCI. Gradual GABAA receptor blockade resulted in higher synchronization, being able to drive the network from a desynchronized to a synchronous state, with a progressive decrease of complexity (sPCI). Blocking GABAB receptors also resulted in a reduced sPCI, in particular when in a synchronous, slow wave state. Our findings demonstrate that physiological levels of inhibition contribute to the generation of dynamical richness and spatiotemporal complexity. However, if inhibition is diminished or enhanced, cortical complexity decreases. Using a computational model, we explored a larger parameter space in this relationship and demonstrate a link between excitatory/inhibitory balance and the complexity expressed by the cortical network.SIGNIFICANCE STATEMENT The spatiotemporal complexity of the activity expressed by the cerebral cortex is a highly revealing feature of the underlying network's state. Complexity varies with physiological brain states: it is higher during awake than during sleep states. But it also informs about pathologic states: in disorders of consciousness, complexity is lower in an unresponsive wakefulness syndrome than in a minimally conscious state. What are the network parameters that modulate complexity? Here we investigate how inhibition, mediated by either GABAA or GABAA receptors, influences cortical complexity. And we do this departing from two extreme functional states: a highly synchronous, slow-wave state, and a desynchronized one that mimics wakefulness. We find that there is an optimal level of inhibition in which complexity is highest.