Dopamine control of social novelty preference is constrained by an interpeduncular-tegmentum circuit.

Animals are inherently motivated to explore social novelty cues over familiar ones, resulting in a novelty preference (NP), although the behavioral and circuit bases underlying NP are unclear. Combining calcium and neurotransmitter sensors with fiber photometry and optogenetics in mice, we find that mesolimbic dopamine (DA) neurotransmission is strongly and predominantly activated by social novelty controlling bout length of interaction during NP, a response significantly reduced by familiarity. In contrast, interpeduncular nucleus (IPN) GABAergic neurons that project to the lateral dorsal tegmentum (LDTg) were inhibited by social novelty but activated during terminations with familiar social stimuli. Inhibition of this pathway during NP increased interaction and bout length with familiar social stimuli, while activation reduced interaction and bout length with novel social stimuli via decreasing DA neurotransmission. These data indicate interest towards novel social stimuli is encoded by mesolimbic DA which is dynamically regulated by an IPN→LDTg circuit to control NP.

Tesofensine, a novel antiobesity drug, silences GABAergic hypothalamic neurons.

Obesity is a major global health epidemic that has adverse effects on both the people affected as well as the cost to society. Several anti-obesity drugs that target GLP-1 receptors have recently come to the market. Here, we describe the effects of tesofensine, a novel anti-obesity drug that acts as a triple monoamine neurotransmitter reuptake inhibitor. Using various techniques, we investigated its effects on weight loss and underlying neuronal mechanisms in mice and rats. These include behavioral tasks, DeepLabCut videotaped analysis, electrophysiological ensemble recordings, optogenetic activation, and chemogenetic silencing of GABAergic neurons in the Lateral Hypothalamus (LH). We found that tesofensine induces a greater weight loss in obese rats than lean rats, while differentially modulating the neuronal ensembles and population activity in LH. In Vgat-ChR2 and Vgat-IRES-cre transgenic mice, we found for the first time that tesofensine inhibited a subset of LH GABAergic neurons, reducing their ability to promote feeding behavior, and chemogenetically silencing them enhanced tesofensine's food-suppressing effects. Unlike phentermine, a dopaminergic appetite suppressant, tesofensine causes few, if any, head-weaving stereotypy at therapeutic doses. Most importantly, we found that tesofensine prolonged the weight loss induced by 5-HTP, a serotonin precursor, and blocked the body weight rebound that often occurs after weight loss. Behavioral studies on rats with the tastant sucrose indicated that tesofensine's appetite suppressant effects are independent of taste aversion and do not directly affect the perception of sweetness or palatability of sucrose. In summary, our data provide new insights into the effects of tesofensine on weight loss and the underlying neuronal mechanisms, suggesting that tesofensine may be an effective treatment for obesity and that it may be a valuable adjunct to other appetite suppressants to prevent body weight rebound.

Clinical and molecular outcomes from the 5-Year natural history study of SSADH Deficiency, a model metabolic neurodevelopmental disorder.

BACKGROUND: Succinic semialdehyde dehydrogenase deficiency (SSADHD) represents a model neurometabolic disease at the fulcrum of translational research within the Boston Children's Hospital Intellectual and Developmental Disabilities Research Centers (IDDRC), including the NIH-sponsored natural history study of clinical, neurophysiological, neuroimaging, and molecular markers, patient-derived induced pluripotent stem cells (iPSC) characterization, and development of a murine model for tightly regulated, cell-specific gene therapy. METHODS: SSADHD subjects underwent clinical evaluations, neuropsychological assessments, biochemical quantification of γ-aminobutyrate (GABA) and related metabolites, electroencephalography (standard and high density), magnetoencephalography, transcranial magnetic stimulation, magnetic resonance imaging and spectroscopy, and genetic tests. This was parallel to laboratory molecular investigations of in vitro GABAergic neurons derived from induced human pluripotent stem cells (hiPSCs) of SSADHD subjects and biochemical analyses performed on a versatile murine model that uses an inducible and reversible rescue strategy allowing on-demand and cell-specific gene therapy. RESULTS: The 62 SSADHD subjects [53% females, median (IQR) age of 9.6 (5.4-14.5) years] included in the study had a reported symptom onset at ∼ 6 months and were diagnosed at a median age of 4 years. Language developmental delays were more prominent than motor. Autism, epilepsy, movement disorders, sleep disturbances, and various psychiatric behaviors constituted the core of the disorder's clinical phenotype. Lower clinical severity scores, indicating worst severity, coincided with older age (R= -0.302, p = 0.03), as well as age-adjusted lower values of plasma γ-aminobutyrate (GABA) (R = 0.337, p = 0.02) and γ-hydroxybutyrate (GHB) (R = 0.360, p = 0.05). While epilepsy and psychiatric behaviors increase in severity with age, communication abilities and motor function tend to improve. iPSCs, which were differentiated into GABAergic neurons, represent the first in vitro neuronal model of SSADHD and express the neuronal marker microtubule-associated protein 2 (MAP2), as well as GABA. GABA-metabolism in induced GABAergic neurons could be reversed using CRISPR correction of the pathogenic variants or mRNA transfection and SSADHD iPSCs were associated with excessive glutamatergic activity and related synaptic excitation. CONCLUSIONS: Findings from the SSADHD Natural History Study converge with iPSC and animal model work focused on a common disorder within our IDDRC, deepening our knowledge of the pathophysiology and longitudinal clinical course of a complex neurodevelopmental disorder. This further enables the identification of biomarkers and changes throughout development that will be essential for upcoming targeted trials of enzyme replacement and gene therapy.

An insular cortical circuit required for itch sensation and aversion.

Itch encompasses both sensory and emotional dimensions, with the two dimensions reciprocally exacerbating each other. However, whether a shared neural circuit mechanism governs both dimensions remains elusive. Here, we report that the anterior insular cortex (AIC) is activated by both histamine-dependent and -independent itch stimuli. The activation of AIC elicits aversive emotion and exacerbates pruritogen-induced itch sensation and aversion. Mechanistically, AIC excitatory neurons project to the GABAergic neurons in the dorsal bed nucleus of the stria terminalis (dBNST). Manipulating the activity of the AIC → dBNST pathway affects both itch sensation and itch-induced aversion. Our study discovers the shared neural circuit (AIC â†’ dBNST pathway) underlying the itch sensation and aversion, highlights the critical role of the AIC as a central hub for the itch processing, and provides a framework to understand the neural mechanisms underlying the sensation and emotion interaction.

Parvalbumin neurons in the nucleus accumbens shell modulate seizure in temporal lobe epilepsy.

A growing number of clinical and animal studies suggest that the nucleus accumbens (NAc), especially the shell, is involved in the pathogenesis of temporal lobe epilepsy (TLE). However, the role of parvalbumin (PV) GABAergic neurons in the NAc shell involved in TLE is still unclear. In this study, we induced a spontaneous TLE model by intrahippocampal administration of kainic acid (KA), which generally induce acute seizures in first 2 h (acute phase) and then lead to spontaneous recurrent seizures after two months (chronic phase). We found that chemogenetic activation of NAc shell PV neurons could alleviate TLE seizures by reducing the number and period of focal seizures (FSs) and secondary generalized seizures (sGSs), while selective inhibition of PV exacerbated seizure activity. Ruby-virus mapping results identified that the hippocampus (ventral and dorsal) is one of the projection targets of NAc shell PV neurons. Chemogenetic activation of the NAc-Hip PV projection fibers can mitigate seizures while inhibition has no effect on seizure ictogenesis. In summary, our findings reveal that PV neurons in the NAc shell could modulate the seizures in TLE via a long-range NAc-Hip circuit. All of these results enriched the investigation between NAc and epilepsy, offering new targets for future epileptogenesis research and precision therapy.

A concerted neuron-astrocyte program declines in ageing and schizophrenia.

Human brains vary across people and over time; such variation is not yet understood in cellular terms. Here we describe a relationship between people's cortical neurons and cortical astrocytes. We used single-nucleus RNA sequencing to analyse the prefrontal cortex of 191 human donors aged 22-97 years, including healthy individuals and people with schizophrenia. Latent-factor analysis of these data revealed that, in people whose cortical neurons more strongly expressed genes encoding synaptic components, cortical astrocytes more strongly expressed distinct genes with synaptic functions and genes for synthesizing cholesterol, an astrocyte-supplied component of synaptic membranes. We call this relationship the synaptic neuron and astrocyte program (SNAP). In schizophrenia and ageing-two conditions that involve declines in cognitive flexibility and plasticity1,2-cells divested from SNAP: astrocytes, glutamatergic (excitatory) neurons and GABAergic (inhibitory) neurons all showed reduced SNAP expression to corresponding degrees. The distinct astrocytic and neuronal components of SNAP both involved genes in which genetic risk factors for schizophrenia were strongly concentrated. SNAP, which varies quantitatively even among healthy people of similar age, may underlie many aspects of normal human interindividual differences and may be an important point of convergence for multiple kinds of pathophysiology.

A brainstem to hypothalamic arcuate nucleus GABAergic circuit drives feeding.

The obesity epidemic is principally driven by the consumption of more calories than the body requires. It is therefore essential that the mechanisms underpinning feeding behavior are defined. Neurons within the brainstem dorsal vagal complex (DVC) receive direct information from the digestive system and project to second-order regions in the brain to regulate food intake. Although γ-aminobutyric acid is expressed in the DVC (GABADVC), its function in this region has not been defined. In order to discover the unique gene expression signature of GABADVC cells, we used single-nucleus RNA sequencing (Nuc-seq), and this revealed 19 separate clusters. We next probed the function of GABADVC cells and discovered that the selective activation of GABADVC neurons significantly controls food intake and body weight. Optogenetic interrogation of GABADVC circuitry identified GABADVC → hypothalamic arcuate nucleus (ARC) projections as appetite suppressive without creating aversion. Electrophysiological analysis revealed that GABADVC → ARC stimulation inhibits hunger-promoting neuropeptide Y (NPY) neurons via GABA release. Adopting an intersectional genetics strategy, we clarify that the GABADVC → ARC circuit curbs food intake. These data identify GABADVC as a new modulator of feeding behavior and body weight and a controller of orexigenic NPY neuron activity, thereby providing insight into the neural underpinnings of obesity.

GABAergic/Glycinergic and Glutamatergic Neurons Mediate Distinct Neurodevelopmental Phenotypes of STXBP1 Encephalopathy.

An increasing number of pathogenic variants in presynaptic proteins involved in the synaptic vesicle cycle are being discovered in neurodevelopmental disorders. The clinical features of these synaptic vesicle cycle disorders are diverse, but the most prevalent phenotypes include intellectual disability, epilepsy, movement disorders, cerebral visual impairment, and psychiatric symptoms ( Verhage and Sørensen, 2020; Bonnycastle et al., 2021; John et al., 2021; Melland et al., 2021). Among this growing list of synaptic vesicle cycle disorders, the most frequent is STXBP1 encephalopathy caused by de novo heterozygous pathogenic variants in syntaxin-binding protein 1 (STXBP1, also known as MUNC18-1; Verhage and Sørensen, 2020; John et al., 2021). STXBP1 is an essential protein for presynaptic neurotransmitter release. Its haploinsufficiency is the main disease mechanism and impairs both excitatory and inhibitory neurotransmitter release. However, the disease pathogenesis and cellular origins of the broad spectrum of neurological phenotypes are poorly understood. Here we generate cell type-specific Stxbp1 haploinsufficient male and female mice and show that Stxbp1 haploinsufficiency in GABAergic/glycinergic neurons causes developmental delay, epilepsy, and motor, cognitive, and psychiatric deficits, recapitulating majority of the phenotypes observed in the constitutive Stxbp1 haploinsufficient mice and STXBP1 encephalopathy. In contrast, Stxbp1 haploinsufficiency in glutamatergic neurons results in a small subset of cognitive and seizure phenotypes distinct from those caused by Stxbp1 haploinsufficiency in GABAergic/glycinergic neurons. Thus, the contrasting roles of excitatory and inhibitory signaling reveal GABAergic/glycinergic dysfunction as a key disease mechanism of STXBP1 encephalopathy and suggest the possibility to selectively modulate disease phenotypes by targeting specific neurotransmitter systems.

Epidermal Growth Factor Suppresses the Development of GABAergic Neurons Via the Modulation of Perineuronal Net Formation in the Neocortex of Developing Rodent Brains.

Previously, we reported that epidermal growth factor (EGF) suppresses GABAergic neuronal development in the rodent cortex. Parvalbumin-positive GABAergic neurons (PV neurons) have a unique extracellular structure, perineuronal nets (PNNs). PNNs are formed during the development of PV neurons and are mainly formed from chondroitin sulfate (CS) proteoglycans (CSPGs). We examined the effect of EGF on CSPG production and PNN formation as a potential molecular mechanism for the inhibition of inhibiting GABAergic neuronal development by EGF. In EGF-overexpressing transgenic (EGF-Tg) mice, the number of PNN-positive PV neurons was decreased in the cortex compared with that in wild-type mice, as in our previous report. The amount of CS and neurocan was also lower in the cortex of EGF-Tg mice, with a similar decrease observed in EGF-treated cultured cortical neurons. PD153035, an EGF receptor (ErbB1) kinase inhibitor, prevented those mentioned above excess EGF-induced reduction in PNN. We explored the molecular mechanism underlying the effect of EGF on PNNs using fluorescent substrates for matrix metalloproteinases (MMPs) and a disintegrin and metalloproteinases (ADAMs). EGF increased the enzyme activity of MMPs and ADAMs in cultured neurons. These enzyme activities were also increased in the EGF-Tg mice cortex. GM6001, a broad inhibitor of MMPs and ADAMs, also blocked EGF-induced PNN reductions. Therefore, EGF/EGF receptor signals may regulate PNN formation in the developing cortex.

Long-term exposure to 6-PPD quinone at environmentally relevant concentrations causes neurotoxicity by affecting dopaminergic, serotonergic, glutamatergic, and GABAergic neuronal systems in Caenorhabditis elegans.

6-PPD quinone (6-PPDQ), an emerging environmental pollutant, is converted based on 6-PPD via ozonation. However, a systematic evaluation on possible neurotoxicity of long-term and low-dose 6-PPDQ exposure and the underlying mechanism remain unknown. In the present work, 0.1-10 µg/L 6-PPDQ was added to treat Caenorhabditis elegans for 4.5 days, with locomotion behavior, neuronal development, sensory perception behavior, neurotransmitter content, and levels of neurotransmission-related genes being the endpoints. 6-PPDQ exposure at 0.1-10 µg/L significantly reduced locomotion behavior, and that at 1-10 µg/L decreased sensory perception behavior in nematodes. Moreover, 6-PPDQ exposure at 10 µg/L notably induced damage to the development of dopaminergic, glutamatergic, serotonergic, and GABAergic neurons. Importantly, nematodes with chronic 6-PPDQ exposure at 10 µg/L were confirmed to suffer obviously decreased dopamine, serotonin, glutamate, dopamine, and GABA contents and altered neurotransmission-related gene expression. Meanwhile, the potential binding sites of 6-PPDQ and neurotransmitter synthesis-related proteins were further shown by molecular docking method. Lastly, Pearson's correlation analysis showed that locomotion behavior and sensory perception behavior were positively correlated with the dopaminergic, serotonergic, glutamatergic, and GABAergic neurotransmission. Consequently, 6-PPDQ exposure disturbed neurotransmitter transmission, while such changed molecular foundation for neurotransmitter transmission was related to 6-PPDQ toxicity induction. The present work sheds new lights on the mechanisms of 6-PPDQ and its possible neurotoxicity to organisms at environmentally relevant concentrations.

Inhibitory medial zona incerta pathway drives exploratory behavior by inhibiting glutamatergic cuneiform neurons.

The cuneiform nucleus (CnF) regulates locomotor activity, which is canonically viewed as being primarily involved in initiating locomotion and regulating speed. Recent research shows greater context dependency in the locomotor functions of this nucleus. Glutamatergic neurons, which contain vesicular glutamate transporter 2 (vGLUT2), regulate context-dependent locomotor speed in the CnF and play a role in defensive behavior. Here, we identify projections from the medial zona incerta (mZI) to CnF vGLUT2 neurons that promote exploratory behavior. Using fiber photometry recordings in male mice, we find that mZI gamma-aminobutyric acid (GABA) neurons increase activity during periods of exploration. Activation of mZI GABAergic neurons is associated with reduced spiking of CnF neurons. Additionally, activating both retrogradely labeled mZI-CnF GABAergic projection neurons and their terminals in the CnF increase exploratory behavior. Inhibiting CnF vGLUT2 neuronal activity also increases exploratory behavior. These findings provide evidence for the context-dependent dynamic regulation of CnF vGLUT2 neurons, with the mZI-CnF circuit shaping exploratory behavior.

Analogs of α-conotoxin PnIC selectively inhibit α7ß2- over α7-only subtype nicotinic acetylcholine receptors via a novel allosteric mechanism.

This study was undertaken to identify and characterize the first ligands capable of selectively identifying nicotinic acetylcholine receptors containing α7 and ß2 subunits (α7ß2-nAChR subtype). Basal forebrain cholinergic neurons express α7ß2-nAChR. Here, they appear to mediate neuronal dysfunction induced by the elevated levels of oligomeric amyloid-ß associated with early Alzheimer's disease. Additional work indicates that α7ß2-nAChR are expressed across several further critically important cholinergic and GABAergic neuronal circuits within the central nervous system. Further studies, however, are significantly hindered by the inability of currently available ligands to distinguish heteromeric α7ß2-nAChR from the closely related and more widespread homomeric α7-only-nAChR subtype. Functional screening using two-electrode voltage-clamp electrophysiology identified a family of α7ß2-nAChR-selective analogs of α-conotoxin PnIC (α-CtxPnIC). A combined electrophysiology, functional kinetics, site-directed mutagenesis, and molecular dynamics approach was used to further characterize the α7ß2-nAChR selectivity and site of action of these α-CtxPnIC analogs. We determined that α7ß2-nAChR selectivity of α-CtxPnIC analogs arises from interactions at a site distinct from the orthosteric agonist-binding site shared between α7ß2- and α7-only-nAChR. As numerous previously identified α-Ctx ligands are competitive antagonists of orthosteric agonist-binding sites, this study profoundly expands the scope of use of α-Ctx ligands (which have already provided important nAChR research and translational breakthroughs). More immediately, analogs of α-CtxPnIC promise to enable, for the first time, both comprehensive mapping of the distribution of α7ß2-nAChR and detailed investigations of their physiological roles.

Electroacupuncture promotes the repair of the damaged spinal cord in mice by mediating neurocan-perineuronal net.

AIMS: This study aimed to investigate the effect of perineuronal net (PNN) and neurocan (NCAN) on spinal inhibitory parvalbumin interneuron (PV-IN), and the mechanism of electroacupuncture (EA) in promoting spinal cord injury (SCI) repair through neurocan in PNN. METHODS: A mouse model of SCI was established. Sham-operated mice or SCI model mice were treated with chondroitin sulfate ABC (ChABC) enzyme or control vehicle for 2 weeks (i.e., sham+veh group, sham+ChABC group, SCI+veh group, and SCI+ChABC group, respectively), and then spinal cord tissues were taken from the T10 lesion epicenter for RNA sequencing (RNA-seq). MSigDB Hallmark and C5 databases for functional analysis, analysis strategies such as differential expression gene analysis (DEG), Kyoto Encyclopedia of Genes and Genomes (KEGG), gene set enrichment analysis (GSEA), and protein-protein interaction (PPI). According to the results of RNA-seq analysis, the expression of NCAN was knocked down or overexpressed by virus intervention, or/and EA intervention. Polymerase chain reaction (PCR), immunofluorescence, western blot, electrophysiological, and behavioral tests were performed. RESULTS: After the successful establishment of SCI model, the motor dysfunction of lower limbs, and the expression of PNN core glycan protein at the epicenter of SCI were reduced. RNA-seq and PCR showed that PNN core proteoglycans except NCAN showed the same expression trend in normal and injured spinal cord treated with ChABC. KEGG and GSEA showed that PNN is mainly associated with inhibitory GABA neuronal function in injured spinal cord tissue, and PPI showed that NCAN in PNN can be associated with inhibitory neuronal function through parvalbumin (PV). Calcium imaging showed that local parvalbumin interneuron (PV-IN) activity decreased after PNN destruction, whether due to ChABC treatment or surgical bruising of the spinal cord. Overexpression of neurocan in injured spinal cord can enhance local PV-IN activity. PCR and western blot suggested that overexpression or knockdown of neurocan could up-regulate or down-regulate the expression of GAD. At the same time, the activity of PV-IN in the primary motor cortex (M1) and the primary sensory cortex of lower (S1HL) extremity changed synchronously. In addition, overexpression of neurocan improved the electrical activity of the lower limb and promoted functional repair of the paralyzed hind limb. EA intervention reversed the down-regulation of neurocan, enhanced the expression of PNN in the lesioned area, M1 and S1HL. CONCLUSION: Neurocan in PNN can regulate the activity of PV-IN, and EA can promote functional recovery of mice with SCI by upregulating neurocan expression in PNN.

The proneural factors Ascl1a and Ascl1b contribute to the terminal differentiation of dopaminergic GABAergic dual transmitter neurons in zebrafish.

The proneural factor Ascl1 is involved in several steps of neurogenesis, from neural progenitor maintenance to initiation of terminal differentiation and neuronal subtype specification. In neural progenitor cells, Ascl1 initiates the cell-cycle exit of progenitors, and contributes to their differentiation into mainly GABAergic neurons. Several catecholaminergic neuron groups in the forebrain of zebrafish use GABA as co-transmitter, but a potential role of the two paralogues Ascl1a and Ascl1b in their neurogenesis is not understood. Here, we show that ascl1a, ascl1b double mutant embryos develop a significantly reduced number of neurons in all GABAergic and catecholaminergic dual transmitter neuron anatomical clusters in the fore- and hindbrain, while glutamatergic catecholaminergic clusters develop normally. However, none of the affected catecholaminergic cell clusters are lost completely, suggesting an impairment in progenitor pools, or a requirement of Ascl1a/b for differentiation of a subset of neurons in each cluster. Early progenitors which are dlx2a+, fezf2 + or emx2 + are not reduced whereas late progenitors and differentiating neurons marked by the expression of dlx5a, isl1 and arxa are severely reduced in ascl1a, ascl1b double mutant embryos. This suggests that Ascl1a and Ascl1b play only a minor or no role in the maintenance of their progenitor pools, but rather contribute to the initiation of terminal differentiation of GABAergic catecholaminergic neurons.

Dissecting neural circuits from rostral ventromedial medulla to spinal trigeminal nucleus bidirectionally modulating craniofacial mechanical sensitivity.

Chronic craniofacial pain is intractable and its mechanisms remain unclarified. The rostral ventromedial medulla (RVM) plays a crucial role in descending pain facilitation and inhibition. It is unclear how the descending circuits from the RVM to spinal trigeminal nucleus (Sp5) are organized to bidirectionally modulate craniofacial nociception. We used viral tracing, in vivo optogenetics, calcium signaling recording, and chemogenetic manipulations to investigate the structure and function of RVM-Sp5 circuits. We found that most RVM neurons projecting to Sp5 were GABAergic or glutamatergic and facilitated or inhibited craniofacial nociception, respectively. Both GABAergic interneurons and glutamatergic projection neurons in Sp5 received RVM inputs: the former were antinociceptive, whereas the latter were pronociceptive. Furthermore, we demonstrated activation of both GABAergic and glutamatergic Sp5 neurons receiving RVM inputs in inflammation- or dysfunction-induced masseter hyperalgesia. Activating GABAergic Sp5 neurons or inhibiting glutamatergic Sp5 neurons that receive RVM projections reversed masseter hyperalgesia. Our study identifies specific cell types and projections of RVM-Sp5 circuits involved in facilitating or inhibiting craniofacial nociception respectively. Selective manipulation of RVM-Sp5 circuits can be used as potential treatment strategy to relieve chronic craniofacial muscle pain.

Sleep restoration by optogenetic targeting of GABAergic neurons reprograms microglia and ameliorates pathological phenotypes in an Alzheimer's disease model.

BACKGROUND: Alzheimer's disease (AD) patients exhibit memory disruptions and profound sleep disturbances, including disruption of deep non-rapid eye movement (NREM) sleep. Slow-wave activity (SWA) is a major restorative feature of NREM sleep and is important for memory consolidation. METHODS: We generated a mouse model where GABAergic interneurons could be targeted in the presence of APPswe/PS1dE9 (APP) amyloidosis, APP-GAD-Cre mice. An electroencephalography (EEG) / electromyography (EMG) telemetry system was used to monitor sleep disruptions in these animals. Optogenetic stimulation of GABAergic interneurons in the anterior cortex targeted with channelrhodopsin-2 (ChR2) allowed us to examine the role GABAergic interneurons play in sleep deficits. We also examined the effect of optogenetic stimulation on amyloid plaques, neuronal calcium as well as sleep-dependent memory consolidation. In addition, microglial morphological features and functions were assessed using confocal microscopy and flow cytometry. Finally, we performed sleep deprivation during optogenetic stimulation to investigate whether sleep restoration was necessary to slow AD progression. RESULTS: APP-GAD-Cre mice exhibited impairments in sleep architecture including decreased time spent in NREM sleep, decreased delta power, and increased sleep fragmentation compared to nontransgenic (NTG) NTG-GAD-Cre mice. Optogenetic stimulation of cortical GABAergic interneurons increased SWA and rescued sleep impairments in APP-GAD-Cre animals. Furthermore, it slowed AD progression by reducing amyloid deposition, normalizing neuronal calcium homeostasis, and improving memory function. These changes were accompanied by increased numbers and a morphological transformation of microglia, elevated phagocytic marker expression, and enhanced amyloid ß (Aß) phagocytic activity of microglia. Sleep was necessary for amelioration of pathophysiological phenotypes in APP-GAD-Cre mice. CONCLUSIONS: In summary, our study shows that optogenetic targeting of GABAergic interneurons rescues sleep, which then ameliorates neuropathological as well as behavioral deficits by increasing clearance of Aß by microglia in an AD mouse model.

CRISPR-Cas9 knockdown of ESR1 in preoptic GABA-kisspeptin neurons suppresses the preovulatory surge and estrous cycles in female mice.

Evidence suggests that estradiol-sensing preoptic area GABA neurons are involved in the preovulatory surge mechanism necessary for ovulation. In vivo CRISPR-Cas9 editing was used to achieve a 60-70% knockdown in estrogen receptor alpha (ESR1) expression by GABA neurons located within the regions of the rostral periventricular area of the third ventricle (RP3V) and medial preoptic nuclei (MPN) in adult female mice. Mice exhibited variable reproductive phenotypes with the only significant finding being mice with bilateral ESR1 deletion in RP3V GABA neurons having reduced cFos expression in gonadotropin-releasing hormone (GnRH) neurons at the time of the surge. One sub-population of RP3V GABA neurons expresses kisspeptin. Re-grouping ESR1-edited mice on the basis of their RP3V kisspeptin expression revealed a highly consistent phenotype; mice with a near-complete loss of kisspeptin immunoreactivity displayed constant estrus and failed to exhibit surge activation but retained pulsatile luteinizing hormone (LH) secretion. These observations demonstrate that ESR1-expressing GABA-kisspeptin neurons in the RP3V are essential for the murine preovulatory LH surge mechanism.

Glucose-dependent insulinotropic polypeptide regulates body weight and food intake via GABAergic neurons in mice.

The development of single-molecule co-agonists for the glucagon-like peptide-1 (GLP-1) receptor (GLP-1R) and glucose-dependent insulinotropic polypeptide (GIP) receptor (GIPR) is considered a breakthrough in the treatment of obesity and type 2 diabetes. But although GIPR-GLP-1R co-agonism decreases body weight with superior efficacy relative to GLP-1R agonism alone in preclinical1-3 and clinical studies4,5, the role of GIP in regulating energy metabolism remains enigmatic. Increasing evidence suggests that long-acting GIPR agonists act in the brain to decrease body weight through the inhibition of food intake3,6-8; however, the mechanisms and neuronal populations through which GIP affects metabolism remain to be identified. Here, we report that long-acting GIPR agonists and GIPR-GLP-1R co-agonists decrease body weight and food intake via inhibitory GABAergic neurons. We show that acyl-GIP decreases body weight and food intake in male diet-induced obese wild-type mice, but not in mice with deletion of Gipr in Vgat(also known as Slc32a1)-expressing GABAergic neurons (Vgat-Gipr knockout). Whereas the GIPR-GLP-1R co-agonist MAR709 leads, in male diet-induced obese wild-type mice, to greater weight loss and further inhibition of food intake relative to a pharmacokinetically matched acyl-GLP-1 control, this superiority over GLP-1 vanishes in Vgat-Gipr knockout mice. Our data demonstrate that long-acting GIPR agonists crucially depend on GIPR signaling in inhibitory GABAergic neurons to decrease body weight and food intake.

The homeodomain transcriptional regulator DVE-1 directs a program for synapse elimination during circuit remodeling.

The elimination of synapses during circuit remodeling is critical for brain maturation; however, the molecular mechanisms directing synapse elimination and its timing remain elusive. We show that the transcriptional regulator DVE-1, which shares homology with special AT-rich sequence-binding (SATB) family members previously implicated in human neurodevelopmental disorders, directs the elimination of juvenile synaptic inputs onto remodeling C. elegans GABAergic neurons. Juvenile acetylcholine receptor clusters and apposing presynaptic sites are eliminated during the maturation of wild-type GABAergic neurons but persist into adulthood in dve-1 mutants, producing heightened motor connectivity. DVE-1 localization to GABAergic nuclei is required for synapse elimination, consistent with DVE-1 regulation of transcription. Pathway analysis of putative DVE-1 target genes, proteasome inhibitor, and genetic experiments implicate the ubiquitin-proteasome system in synapse elimination. Together, our findings define a previously unappreciated role for a SATB family member in directing synapse elimination during circuit remodeling, likely through transcriptional regulation of protein degradation processes.

Sex-specific modulation of the medial prefrontal cortex by glutamatergic median raphe neurons.

A substantial proportion of raphe neurons are glutamatergic. However, little is known about how these glutamatergic neurons modulate the forebrain. We investigated how glutamatergic median raphe nucleus (MRN) input modulates the medial prefrontal cortex (mPFC), a critical component of fear circuitry. We show that vesicular glutamate transporter 3 (VGLUT3)-expressing MRN neurons activate VGLUT3- and somatostatin-expressing neurons in the mPFC. Consistent with this modulation of mPFC GABAergic neurons, activation of MRN (VGLUT3) neurons enhances GABAergic transmission in mPFC pyramidal neurons and attenuates fear memory in female but not male mice. Serotonin plays a key role in MRN (VGLUT3) neuron-mediated GABAergic plasticity in the mPFC. In agreement with these female-specific effects, we observed sex differences in glutamatergic transmission onto MRN (VGLUT3) neurons and in mPFC (VGLUT3) neuron-mediated dual release of glutamate and GABA. Our results demonstrate a cell type-specific modulation of the mPFC by MRN (VGLUT3) neurons and reveal a sex-specific role of this neuromodulation in mPFC synaptic plasticity.