GABAergic circuits are critical for the synchronization and higher order function of brain networks. Defects in this circuitry are linked to neuropsychiatric diseases, including bipolar disorder, schizophrenia, and autism. Work in cultured neurons has shown that ankyrin-G plays a key role in the regulation of GABAergic synapses on the axon initial segment and somatodendritic domain of pyramidal neurons where it interacts directly with the GABA A receptor associated protein (GABARAP) to stabilize cell surface GABA A receptors. Here, we generated a knock-in mouse model expressing a mutation that abolishes the ankyrin-G/GABARAP interaction ( Ank3 W1989R) to understand how ankyrin-G and GABARAP regulate GABAergic circuitry in vivo. We found that Ank3 W1989R mice exhibit a striking reduction in forebrain GABAergic synapses resulting in pyramidal cell hyperexcitability and disruptions in network synchronization. In addition, we identified changes in pyramidal cell dendritic spines and axon initial segments consistent with compensation for hyperexcitability. Finally, we identified the ANK3 W1989R variant in a family with bipolar disorder, suggesting a potential role of this variant in disease. Our results highlight the importance of ankyrin-G in regulating forebrain circuitry and provide novel insights into how ANK3 loss-of-function variants may contribute to human disease.

Dysfunction in sodium channels and their ankyrin scaffolding partners have both been implicated in neurodevelopmental disorders, including autism spectrum disorder (ASD). In particular, the genes SCN2A, which encodes the sodium channel NaV1.2, and ANK2, which encodes ankyrin-B, have strong ASD association. Recent studies indicate that ASD-associated haploinsufficiency in Scn2a impairs dendritic excitability and synaptic function in neocortical pyramidal cells, but how NaV1.2 is anchored within dendritic regions is unknown. Here, we show that ankyrin-B is essential for scaffolding NaV1.2 to the dendritic membrane of mouse neocortical neurons, and that haploinsufficiency of Ank2 phenocopies intrinsic dendritic excitability and synaptic deficits observed in Scn2a+/- conditions. Thus, these results establish a direct, convergent link between two major ASD risk genes and reinforce an emerging framework suggesting that neocortical pyramidal cell dendritic dysfunction can be etiological to neurodevelopmental disorder pathophysiology.

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The cannabinoid CB1 (CB1R) and dopaminergic D2 (D2R) receptors modify GABAergic transmission in the globus pallidus. Although dopaminergic denervation produces changes in the expression and supersensitization of these receptors, the consequences of these changes on GABAergic neurotransmission are unknown. The aim of this study was to show the effects of CB1R and D2R activation and coactivation on the uptake and release of [(3) H]GABA in the globus pallidus of hemiparkinsonian rats as well as their effects on motor behavior. The activation of CB1R blocked GABA uptake and decreased GABA release in the globus pallidus in the dopamine denervated side, whereas the co-activation of CB1R-D2R increased GABA release and had no effect on GABA uptake. A microinjection of the CB1R agonist ACEA into the globus pallidus ipsilaterally to a 6-OHDA lesion potentiated turning behavior that was induced by methamphetamine. However, a microinjection of the D2R agonist quinpirole did not modify this behavior, and a microinjection of a mixture of CB1R and D2R agonists significantly potentiated turning behavior. The behavioral effects produced after the activation of the CB1R and the co-activation of CB1R and D2R can be explained by increased GABAergic neurotransmission produced by a block of GABA uptake and an increase in the release of GABA in the globus pallidus, respectively.

Down syndrome (DS) is caused by the trisomy of human chromosome 21 (HSA21). A major challenge in DS research is to identify the HSA21 genes that cause specific symptoms. Down syndrome cell adhesion molecule (DSCAM) is encoded by a HSA21 gene. Previous studies have shown that the protein level of the Drosophila homolog of DSCAM determines the size of presynaptic terminals. However, whether the triplication of DSCAM contributes to presynaptic development in DS remains unknown. Here, we show that DSCAM levels regulate GABAergic synapses formed on neocortical pyramidal neurons (PyNs). In the Ts65Dn mouse model for DS, where DSCAM is overexpressed due to DSCAM triplication, GABAergic innervation of PyNs by basket and chandelier interneurons is increased. Genetic normalization of DSCAM expression rescues the excessive GABAergic innervations and the increased inhibition of PyNs. Conversely, loss of DSCAM impairs GABAergic synapse development and function. These findings demonstrate excessive GABAergic innervation and synaptic transmission in the neocortex of DS mouse models and identify DSCAM overexpression as the cause. They also implicate dysregulated DSCAM levels as a potential pathogenic driver in related neurological disorders.

GABAergic circuits are critical for the synchronization and higher order function of brain networks, and defects in this circuitry are linked to neuropsychiatric diseases, including bipolar disorder, schizophrenia, and autism. Work in cultured neurons has shown that ankyrin-G plays a key role in the regulation of GABAergic synapses on the axon initial segment and somatodendritic domain of pyramidal neurons where it interacts directly with the GABAA receptor associated protein (GABARAP) to stabilize cell surface GABAA receptors. Here, we generated a knock-in mouse model expressing a mutation that abolishes the ankyrin-G/GABARAP interaction (Ank3 W1989R) to understand how ankyrin-G and GABARAP regulate GABAergic circuitry in vivo. We found that Ank3 W1989R mice exhibit a striking reduction in forebrain GABAergic synapses resulting in pyramidal cell hyperexcitability and disruptions in network synchronization. In addition, we identified changes in pyramidal cell dendritic spines and axon initial segments consistent with compensation for hyperexcitability. Finally, we identified the ANK3 W1989R variant in a family with bipolar disorder, suggesting a potential role of this variant in disease. Our results highlight the importance of ankyrin-G in regulating forebrain circuitry and provide novel insights into how ANK3 loss-offunction variants may contribute to human disease. molecular mechanisms underlying the subcellular organization of cortical GABAergic synapses remain poorly understood. Abnormalities in GABAergic interneuron circuitry and decreased gamma oscillations have been implicated in many neurodevelopmental and neuropsychiatric disorders 1-8 . Thus, the understanding of the cellular and molecular mechanisms that contribute to the development and function of GABAergic synapses as well as identification of genetic variants that contribute to neuropsychiatric disorders is critical to the discovery of new therapeutic agents for the treatment of diseases involving altered inhibitory circuits.ANK3 encodes ankyrin-G, a fundamental scaffolding protein that organizes critical plasma membrane domains 9, 10 . Alternative splicing of ANK3 in the brain gives rise to three main isoforms of ankyrin-G: the canonical 190 kDa isoform, a 270 kDa isoform, and a giant, 480 kDa isoform. The 190 kDa isoform is expressed in most tissues and cell types throughout the body including brain, heart, skeletal muscle, kidney, and retina. The 270 kDa and 480 kDa isoforms of ankyrin-G are predominantly expressed in the nervous system, and arise from alternative splicing of a 7.8 kb vertebrate-specific exon 9, 11 . The 480 kDa ankyrin-G isoform has been identified as the master organizer of axon initial segments (AIS) and nodes of Ranvier, sites of action potential (AP) initiation and propagation 10 . This splice variant is necessary for the proper clustering of voltage-gated sodium channels, KCNQ2/3 potassium channels, the cell adhesion molecule neurofascin-186, and the cytoskeletal protein βIV-spectrin to excitable domains (reviewed in 12 )....

Two major groups of terminals release GABA within the Globus pallidus; one group is constituted by projections from striatal neurons, while endings of the intranuclear collaterals form the other one. Each neurons' population expresses different subtypes of dopamine D2-like receptors: D 2 R subtype is expressed by encephalin-positive MSNs, while pallidal neurons express the D 4 R subtype. The D 2 R modulates the firing rate of striatal neurons and GABA release at their projection areas, while the D 4 R regulates Globus pallidus neurons excitability and GABA release at their projection areas. However, it is unknown if these receptors control GABA release at pallidopallidal collaterals and regulate motor behavior. Here, we present neurochemical evidence of protein content and binding of D 4 R in pallidal synaptosomes, control of [ 3 H] GABA release in pallidal slices of rat, electrophysiological evidence of the presence of D 4 R on pallidal recurrent collaterals in mouse slices, and turning behavior induced by D 4 R antagonist microinjected in amphetamine challenged rats. As in projection areas of pallidal neurons, GABAergic transmission in pallido-pallidal recurrent synapses is under modulation of D 4 R, while the D 2 R subtype, as known, modulates striato-pallidal projections. Also, as in projection areas, D 4 R contributes to 4564 | CONDE ROJAS Et Al. 2 | EXPERIMENTAL PROCEDURES 2.1 | Animals All procedures were carried out following the National Institutes of Health Guide for Care and Use of Laboratory Animals and approved by the Institutional Animal Care Committee of the CINVESTAV-IPN. Due to conveniences for technics and the amount of tissue required for experiments, Western blot, [ 3 H] YM-09151-2 D 4 R binding, [ 3 H] GABA release, and behavioral observations were done using

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