Behavioral evaluation of mice deficient in GABAB(1) receptor isoforms in tests of unconditioned anxiety

Rationale: Emerging data support a role for GABAB receptors in anxiety. GABAB receptors are comprised of a heterodimeric complex of GABAB1 and GABAB2 receptor subunits. The predominant neuronal GABAB1 receptor isoforms are GABAB(1a) and GABAB(1b). Recent findings indicate specific roles for these isoforms in conditioned fear responses, although their influence on behavior in tests of unconditioned anxiety is unknown. Objective: The aim of this study was to examine the role of the GABAB(1) isoforms in unconditioned anxiety. Materials and methods: Mice deficient in the GABAB(1a) or GABAB(1b) receptor isoforms were examined in a battery of anxiety tests. Results: In most tests, genotype did not significantly affect anxious behavior, including the elevated plus maze, marble burying, and stress-induced hypothermia tests. Corticosterone and adrenocorticotropic hormone levels were similarly unaffected by genotype. Female, but not male, {\text{GABA}}^{{ - \mathord{\left/ {\vphantom { - - }} \right. \kern-\nulldelimiterspace} - }}_{{{\text{B}}{\left( {1{\text{a}}} \right)}}} and {\text{GABA}}^{{ - \mathord{\left/ {\vphantom { - - }} \right. \kern-\nulldelimiterspace} - }}_{{{\text{B}}{\left( {1{\text{b}}} \right)}}} mice showed increased anxiety relative to wild-type controls in the elevated zero maze. In the staircase test, male {\text{GABA}}^{{ - \mathord{\left/ {\vphantom { - - }} \right. \kern-\nulldelimiterspace} - }}_{{{\text{B}}{\left( {1{\text{b}}} \right)}}} mice defecated more than male {\text{GABA}}^{{ - \mathord{\left/ {\vphantom { - - }} \right. \kern-\nulldelimiterspace} - }}_{{{\text{B}}{\left( {1{\text{a}}} \right)}}} mice, although no other test parameter was influenced by genotype. In the light-dark box, female {\text{GABA}}^{{ - \mathord{\left/ {\vphantom { - - }} \right. \kern-\nulldelimiterspace} - }}_{{{\text{B}}{\left( {1{\text{a}}} \right)}}} mice spent less time in the light compartment compared to the {\text{GABA}}^{{ - \mathord{\left/ {\vphantom { - - }} \right. \kern-\nulldelimiterspace} - }}_{{{\text{B}}{\left( {1{\text{b}}} \right)}}} females, whereas male {\text{GABA}}^{{ - \mathord{\left/ {\vphantom { - - }} \right. \kern-\nulldelimiterspace} - }}_{{{\text{B}}{\left( {1{\text{b}}} \right)}}} mice made fewer light-dark transitions than {\text{GABA}}^{{ - \mathord{\left/ {\vphantom { - - }} \right. \kern-\nulldelimiterspace} - }}_{{{\text{B}}{\left( {1{\text{a}}} \right)}}} males. Conclusions: Specific roles for either GABAB(1) isoform in unconditioned anxiety were not explicit. This differs from their contribution in conditioned anxiety and from the anxious phenotype of GABAB1 and GABAB2 subunit knockout mice. The findings suggest that the GABAB(1) isoforms have specific relevance for anxiety with a cognitive component, rather than for innate anxiety per s

Conditional gene deletion reveals functional redundancy of GABAB receptors in peripheral nociceptors in vivo

Background Gamma-aminobutyric acid (GABA) is an important inhibitory neurotransmitter which mainly mediates its effects on neurons via ionotropic (GABAA) and metabotropic (GABAB) receptors. GABAB receptors are widely expressed in the central and the peripheral nervous system. Although there is evidence for a key function of GABAB receptors in the modulation of pain, the relative contribution of peripherally- versus centrally-expressed GABAB receptors is unclear. Results In order to elucidate the functional relevance of GABAB receptors expressed in peripheral nociceptive neurons in pain modulation we generated and analyzed conditional mouse mutants lacking functional GABAB(1) subunit specifically in nociceptors, preserving expression in the spinal cord and brain (SNS-GABAB(1)-/- mice). Lack of the GABAB(1) subunit precludes the assembly of functional GABAB receptor. We analyzed SNS-GABAB(1)-/- mice and their control littermates in several models of acute and neuropathic pain. Electrophysiological studies on peripheral afferents revealed higher firing frequencies in SNS-GABAB(1)-/- mice compared to corresponding control littermates. However no differences were seen in basal nociceptive sensitivity between these groups. The development of neuropathic and chronic inflammatory pain was similar across the two genotypes. The duration of nocifensive responses evoked by intraplantar formalin injection was prolonged in the SNS-GABAB(1)-/- animals as compared to their control littermates. Pharmacological experiments revealed that systemic baclofen-induced inhibition of formalin-induced nociceptive behaviors was not dependent upon GABAB(1) expression in nociceptors. Conclusion This study addressed contribution of GABAB receptors expressed on primary afferent nociceptive fibers to the modulation of pain. We observed that neither the development of acute and chronic pain nor the analgesic effects of a systematically-delivered GABAB agonist was significantly changed upon a specific deletion of GABAB receptors from peripheral nociceptive neurons in vivo. This lets us conclude that GABAB receptors in the peripheral nervous system play a less important role than those in the central nervous system in the regulation of pain

Behavioural endophenotypes in mice lacking the auxiliary GABAB receptor subunit KCTD16

Gamma-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the brain and is implicated in the pathophysiology of a number of neuropsychiatric disorders. The GABAB receptors are G-protein coupled receptors consisting of principle subunits and auxiliary potassium channel tetramerization domain (KCTD) subunits. The KCTD subunits 8, 12, 12b and 16 are cytosolic proteins that determine the kinetics of the GABAB receptor response. Previously, we demonstrated that Kctd12 null mutant mice (Kctd12(-/-)) exhibit increased auditory fear learning and that Kctd12(+/-) mice show altered circadian activity, as well as increased intrinsic excitability in hippocampal pyramidal neurons. KCTD16 has been demonstrated to influence neuronal excitability by regulating GABAB receptor-mediated gating of postsynaptic ion channels. In the present study we investigated for behavioural endophenotypes in Kctd16(-/-) and Kctd16(+/-) mice. Compared with wild-type (WT) littermates, auditory and contextual fear conditioning were normal in both Kctd16(-/-) and Kctd16(+/-) mice. When fear memory was tested on the following day, Kctd16(-/-) mice exhibited less extinction of auditory fear memory relative to WT and Kctd16(+/-) mice, as well as more contextual fear memory relative to WT and, in particular, Kctd16(+/-) mice. Relative to WT, both Kctd16(+/-) and Kctd16(-/-) mice exhibited normal circadian activity. This study adds to the evidence that auxillary KCTD subunits of GABAB receptors contribute to the regulation of behaviours that could constitute endophenotypes for hyper-reactivity to aversive stimuli in neuropsychiatric disorders

GABAB receptors in GtoPdb v.2021.2

Functional GABAB receptors (nomenclature as agreed by the NC-IUPHAR Subcommittee on GABAB receptors [11, 71]) are formed from the heterodimerization of two similar 7TM subunits termed GABAB1 and GABAB2 [11, 70, 28, 71, 87]. GABAB receptors are widespread in the CNS and regulate both pre- and postsynaptic activity. The GABAB1 subunit, when expressed alone, binds both antagonists and agonists, but the affinity of the latter is generally 10-100-fold less than for the native receptor. Co-expression of GABAB1 and GABAB2 subunits allows transport of GABAB1 to the cell surface and generates a functional receptor that can couple to signal transduction pathways such as high-voltage-activated Ca2+ channels (Cav2.1, Cav2.2), or inwardly rectifying potassium channels (Kir3) [12, 11, 5]. The GABAB1 subunit harbours the GABA (orthosteric)-binding site within an extracellular domain (ECD) venus flytrap module (VTM), whereas the GABAB2 subunit mediates G protein-coupled signalling [11, 70, 40, 39]. The cryo-electron microscopy structures of the human full-length GABAB1-GABAB2 heterodimer have been solved in the inactive apo state, two intermediate agonist-bound forms and an active state in which the heterodimer is bound to an agonist and a positive allosteric modulator [81]. The positive allosteric modulator binds to the transmembrane dimerization interface and stabilizes the active state. Recent evidence indicates that higher order assemblies of GABAB receptor comprising dimers of heterodimers occur in recombinant expression systems and in vivo and that such complexes exhibit negative functional cooperativity between heterodimers [69, 22]. Adding further complexity, KCTD (potassium channel tetramerization proteins) 8, 12, 12b and 16 associate as tetramers with the carboxy terminus of the GABAB2 subunit to impart altered signalling kinetics and agonist potency to the receptor complex [86, 3, 79] and are reviewed by [72]. The molecular complexity of GABAB receptors is further increased through association with trafficking and effector proteins [80] and reviewed by [68]. The predominant GABAB1a and GABAB1b isoforms, which are most prevalent in neonatal and adult brain tissue respectively, differ in their ECD sequences as a result of the use of alternative transcription initiation sites. GABAB1a-containing heterodimers localise to distal axons and mediate inhibition of glutamate release in the CA3-CA1 terminals, and GABA release onto the layer 5 pyramidal neurons, whereas GABAB1b-containing receptors occur within dendritic spines and mediate slow postsynaptic inhibition [74, 91]. Amyloid precursor protein (APP) and soluble APP (sAPP) bind to the N- terminal sushi domain of the GABAB1a isoform to regulate axonal trafficking of GABAB receptors and release of neurotransmitters [76]

GABAB receptors (version 2019.4) in the IUPHAR/BPS Guide to Pharmacology Database

Functional GABAB receptors (nomenclature as agreed by the NC-IUPHAR Subcommittee on GABAB receptors [11, 72]) are formed from the heterodimerization of two similar 7TM subunits termed GABAB1 and GABAB2 [11, 71, 28, 72, 85]. GABAB receptors are widespread in the CNS and regulate both pre- and postsynaptic activity. The GABAB1 subunit, when expressed alone, binds both antagonists and agonists, but the affinity of the latter is generally 10-100-fold less than for the native receptor. Co-expression of GABAB1 and GABAB2 subunits allows transport of GABAB1 to the cell surface and generates a functional receptor that can couple to signal transduction pathways such as high-voltage-activated Ca2+ channels (Cav2.1, Cav2.2), or inwardly rectifying potassium channels (Kir3) [12, 11, 5]. The GABAB1 subunit harbours the GABA (orthosteric)-binding site within an extracellular domain (ECD) venus flytrap module (VTM), whereas the GABAB2 subunit mediates G protein-coupled signalling [11, 71, 40, 39]. The two subunits interact by direct allosteric coupling [63], such that GABAB2 increases the affinity of GABAB1 for agonists and reciprocally GABAB1 facilitates the coupling of GABAB2 to G proteins [71, 54, 39]. GABAB1 and GABAB2 subunits assemble in a 1:1 stoichiometry by means of a coiled-coil interaction between α-helices within their carboxy-termini that masks an endoplasmic reticulum retention motif (RXRR) within the GABAB1 subunit but other domains of the proteins also contribute to their heteromerization [5, 71, 15]. Recent evidence indicates that higher order assemblies of GABAB receptor comprising dimers of heterodimers occur in recombinant expression systems and in vivo and that such complexes exhibit negative functional cooperativity between heterodimers [70, 22]. Adding further complexity, KCTD (potassium channel tetramerization proteins) 8, 12, 12b and 16 associate as tetramers with the carboxy terminus of the GABAB2 subunit to impart altered signalling kinetics and agonist potency to the receptor complex [84, 3, 79] and are reviewed by [73]. The molecular complexity of GABAB receptors is further increased through association with trafficking and effector proteins [Schwenk et al., 2016, Nature Neuroscience 19(2): 233-42] and reviewed by [69]. Four isoforms of the human GABAB1 subunit have been cloned. The predominant GABAB1a and GABAB1b isoforms, which are most prevalent in neonatal and adult brain tissue respectively, differ in their ECD sequences as a result of the use of alternative transcription initiation sites. GABAB1a-containing heterodimers localise to distal axons and mediate inhibition of glutamate release in the CA3-CA1 terminals, and GABA release onto the layer 5 pyramidal neurons, whereas GABAB1b-containing receptors occur within dendritic spines and mediate slow postsynaptic inhibition [75, 89]. Only the 1a and 1b variants are identified as components of native receptors [11]. Additional GABAB1 subunit isoforms have been described in rodents and humans [55] and reviewed by [5]

Inhibition of Notch2 by Numb/Numblike controls myocardial compaction in the heart

Aims The ventricular wall of the heart is composed of trabeculated and compact layers, which are separated by yet unknown processes during embryonic development. Here, we wanted to explore the role of Notch2 and Numb/Numblike for myocardial trabeculation and compaction. Methods and results We found that Notch2 activity is specifically down-regulated in the compact layer during cardiac development in the mouse. The biological role of Notch2 down-regulation was investigated by the expression of constitutively active Notch2 in the myocardium of transgenic mice, resulting in hypertrabeculation, reduced compaction, and ventricular septum defects. To disclose the mechanism that inhibited Notch2 activity during the formation of myocardial layers, we analysed potential suppressors of Notch signalling. We unveiled that concomitant but not separate ablation of Numb and Numblike in the developing heart leads to increased Notch2 activity along with hypertrabeculation, reduced compaction, and ventricular septum defects, phenocopying effects gained by overexpression of constitutively active Notch2. Expression profiling revealed a strong up-regulation of Bmp10 in Numb/Numblike mutant hearts, which might also interfere with trabeculation and compaction. Conclusion This study identified potential novel roles of Numb/Numblike in regulating trabeculation and compaction by inhibiting Notch2 and Bmp10 signallin

Differential association of GABAB receptors with their effector ion channels in Purkinje cells

Metabotropic GABAB receptors mediate slow inhibitory effects presynaptically and postsynaptically through the modulation of different effector signalling pathways. Here, we analysed the distribution of GABAB receptors using highly sensitive SDS-digested freeze-fracture replica labelling in mouse cerebellar Purkinje cells. Immunoreactivity for GABAB1 was observed on presynaptic and, more abundantly, on postsynaptic compartments, showing both scattered and clustered distribution patterns. Quantitative analysis of immunoparticles revealed a somato-dendritic gradient, with the density of immunoparticles increasing 26-fold from somata to dendritic spines. To understand the spatial relationship of GABAB receptors with two key effector ion channels, the G protein-gated inwardly rectifying K+ (GIRK/Kir3) channel and the voltage-dependent Ca2+ channel, biochemical and immunohistochemical approaches were performed. Co-immunoprecipitation analysis demonstrated that GABAB receptors co-assembled with GIRK and CaV2.1 channels in the cerebellum. Using double-labelling immunoelectron microscopic techniques, co-clustering between GABAB1 and GIRK2 was detected in dendritic spines, whereas they were mainly segregated in the dendritic shafts. In contrast, co-clustering of GABAB1 and CaV2.1 was detected in dendritic shafts but not spines. Presynaptically, although no significant co-clustering of GABAB1 and GIRK2 or CaV2.1 channels was detected, inter-cluster distance for GABAB1 and GIRK2 was significantly smaller in the active zone than in the dendritic shafts, and that for GABAB1 and CaV2.1 was significantly smaller in the active zone than in the dendritic shafts and spines. Thus, GABAB receptors are associated with GIRK and CaV2.1 channels in different subcellular compartments. These data provide a better framework for understanding the different roles played by GABAB receptors and their effector ion channels in the cerebellar network

Differential association of GABAB receptors with their effector ion channels in Purkinje cells

Metabotropic GABAB receptors mediate slow inhibitory effects presynaptically and postsynaptically through the modulation of different effector signalling pathways. Here, we analysed the distribution of GABAB receptors using highly sensitive SDS-digested freeze-fracture replica labelling in mouse cerebellar Purkinje cells. Immunoreactivity for GABAB1 was observed on presynaptic and, more abundantly, on postsynaptic compartments, showing both scattered and clustered distribution patterns. Quantitative analysis of immunoparticles revealed a somato-dendritic gradient, with the density of immunoparticles increasing 26-fold from somata to dendritic spines. To understand the spatial relationship of GABAB receptors with two key effector ion channels, the G protein-gated inwardly rectifying K+ (GIRK/Kir3) channel and the voltage-dependent Ca2+ channel, biochemical and immunohistochemical approaches were performed. Co-immunoprecipitation analysis demonstrated that GABAB receptors co-assembled with GIRK and CaV2.1 channels in the cerebellum. Using double-labelling immunoelectron microscopic techniques, co-clustering between GABAB1 and GIRK2 was detected in dendritic spines, whereas they were mainly segregated in the dendritic shafts. In contrast, co-clustering of GABAB1 and CaV2.1 was detected in dendritic shafts but not spines. Presynaptically, although no significant co-clustering of GABAB1 and GIRK2 or CaV2.1 channels was detected, inter-cluster distance for GABAB1 and GIRK2 was significantly smaller in the active zone than in the dendritic shafts, and that for GABAB1 and CaV2.1 was significantly smaller in the active zone than in the dendritic shafts and spines. Thus, GABAB receptors are associated with GIRK and CaV2.1 channels in different subcellular compartments. These data provide a better framework for understanding the different roles played by GABAB receptors and their effector ion channels in the cerebellar network

Impaired bidirectional communication between interneurons and oligodendrocyte precursor cells affects social cognitive behavior

Cortical neural circuits are complex but very precise networks of balanced excitation and inhibition. Yet, the molecular and cellular mechanisms that form the balance are just beginning to emerge. Here, using conditional γ-aminobutyric acid receptor B1- deficient mice we identify a γ-aminobutyric acid/tumor necrosis factor superfamily member 12-mediated bidirectional communication pathway between parvalbumin-positive fast spiking interneurons and oligodendrocyte precursor cells that determines the density and function of interneurons in the developing medial prefrontal cortex. Interruption of the GABAergic signaling to oligodendrocyte precursor cells results in reduced myelination and hypoactivity of interneurons, strong changes of cortical network activities and impaired social cognitive behavior. In conclusion, glial transmitter receptors are pivotal elements in finetuning distinct brain functions