Unaltered Tonic Inhibition in the Arcuate Nucleus of Diet-induced Obese Mice

Abstract

The principal inhibitory transmitter, γ-aminobutyric acid (GABA), is critical for maintaining hypothalamic homeostasis and released from neurons phasically, as well as from astrocytes tonically. Although astrocytes in the arcuate nucleus (ARC) of the hypothalamus are shown to transform into reactive astrocytes, the tonic inhibition by astrocytic GABA has not been adequately investigated in diet-induced obesity (DIO). Here, we investigated the expression of monoamine oxidase-B (MAOB), a GABA-synthesizing enzyme, in reactive astrocytes in obese mice. We observed that a chronic high-fat diet (HFD) significantly increased astrocytic MAOB and cellular GABA content, along with enhanced hypertrophy of astrocytes in the ARC. Unexpectedly, we found that the level of tonic GABA was unaltered in chronic HFD mice using whole-cell patch-clamp recordings in the ARC. Furthermore, the GABA-induced current was increased with elevated GABA A receptor α5 (GABRA5) expression. Surprisingly, we found that a nonselective GABA transporter (GAT) inhibitor, nipecotic acid (NPA)-induced current was significantly increased in chronic HFD mice. We observed that GAT1 inhibitor, NO711-induced current was significantly increased, whereas GAT3 inhibitor, SNAP5114-induced current was not altered. The unexpected unaltered tonic inhibition was due to an increase of GABA clearance in the ARC by neuronal GAT1 rather than astrocytic GAT3. These results imply that increased astrocytic GABA synthesis and neuronal GABA A receptor were compensated by GABA clearance, resulting in unaltered tonic GABA inhibition in the ARC of the hypothalamus in obese mice. Taken together, GABA-related molecular pathways in the ARC dynamically regulate the tonic inhibition to maintain hypothalamic homeostasis against the HFD challenge.

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“…To establish the DIO mouse model, male C57BL/6J mice, starting at six weeks of age, were fed either a high-fat diet (HFD; consisting of 60% of calories from fat, sourced from D12492, Research Diets Inc.) or a standard chow diet (Teklad, 2018S, Envigo) for a period of 6 to 18 weeks, following protocols previously established ( Fig. 1 , 2a ) [ 20 - 23 ]. All experimental procedures were carried out using age-matched control groups to ensure accuracy and reliability.…”