The dorsal motor nucleus of the vagus (DMV) contains the preganglionic motor neurons important in the regulation of glucose homeostasis and gastrointestinal function. Despite the role of sex in the regulation of these processes, few studies examine the role of sex and/or ovarian cycle in the regulation of synaptic neurotransmission to the DMV. Since GABAergic neurotransmission is critical to normal DMV function, the present study used in vitro whole cell patch-clamping to investigate whether sex differences exist in GABAergic neurotransmission to DMV neurons. It additionally investigated whether the ovarian cycle plays a role in those sex differences. The frequency of phasic GABAA receptor-mediated inhibitory postsynaptic currents in DMV neurons from females was lower compared with males, and this effect was TTX sensitive and abolished by ovariectomy (OVX). Amplitudes of GABAergic currents (both phasic and tonic) were not different. However, females demonstrated significantly more variability in the amplitude of both phasic and tonic GABAA receptor currents. This difference was eliminated by OVX in females, suggesting that these differences were related to reproductive hormone levels. This was confirmed for GABAergic tonic currents by comparing females in two ovarian stages, estrus versus diestrus. Female mice in diestrus had larger tonic current amplitudes compared with those in estrus, and this increase was abolished after administration of a 5α-reductase inhibitor but not modulation of estrogen. Taken together, these findings demonstrate that DMV neurons undergo GABAA receptor activity plasticity as a function of sex and/or sex steroids. NEW & NOTEWORTHY Results show that GABAergic signaling in dorsal vagal motor neurons (DMV) demonstrates sex differences and fluctuates across the ovarian cycle in females. These findings are the first to demonstrate that female GABAA receptor activity in this brain region is modulated by 5α-reductase-dependent hormones. Since DMV activity is critical to both glucose and gastrointestinal homeostasis, these results suggest that sex hormones, including those synthesized by 5α-reductase, contribute to visceral, autonomic function related to these physiological processes.

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Background: Traumatic brain injury (TBI) remains one of the greatest public health concerns with increasing morbidity and mortality rates worldwide. Our group reported stimulation of astrocyte mitochondrial metabolism by P2Y1 receptor agonists significantly reduced cerebral edema and reactive gliosis in a TBI model. Subsequent data on the pharmacokinetics (PK) and rapid metabolism of these compounds suggested neuroprotection was likely mediated by a metabolite, AST-004, which binding data indicated was an adenosine A3 receptor (A3R) agonist. Methods: The neuroprotective efficacy of AST-004 was tested in a controlled closed cortical injury (CCCI) model of TBI in mice. Results: Twenty-four (24) hours post-injury, mice subjected to CCCI and treated with AST-004 (0.22mg/kg) exhibited significantly less secondary brain injury. These effects were quantified with less cell death (PSVue794 fluorescence) and loss of blood brain barrier breakdown (Evans Blue extravasation assay), compared to vehicle treated TBI mice. TBI treated mice also exhibited significantly reduced neuroinflammatory markers, glial-fibrillary acidic protein (GFAP, astrogliosis) and ionized Ca2+ binding adaptor molecule 1 (Iba1, microgliosis), both at the mRNA (gRT-PCR) and protein (western blot and immunofluorescence) levels, respectively. Four (4) weeks post-injury, AST-004 treated TBI mice presented significantly reduced impairment of long-term memory. Spatial memory was assessed with a contextual fear conditioning behavior assay (freezing behavior after shock). Finally, AST-004 treatments were found to increase in vivo ATP production in astrocytes (gfap-targeted luciferase activity), consistent with the proposed mechanism of action. Conclusions: These data reveal AST-004 as a novel A3R agonist that increases astrocyte energy production and enhances their neuroprotective efficacy after brain injury.