Fig. 5: β3 GABA_A receptor subunit membrane trafficking mediates CRH excitatory effect in VMH.

a Representative traces of mIPSCs in VMH^SF-1 neurons before and after the application of CRH in Sf-1-Cre mice. b Cumulative amplitude probability plots of mIPSCs in typical VMH^SF-1 neurons before and after CRH application. Inset in b: pooled data. n = 17/4, cells/animals; ***p < 0.001, for mIPSC amplitude. c Schematic of evoked IPSCs recording from SF-1 neurons in Sf-1-Cre mice. d Representative traces of pair pulse ratio in the presence of CNQX in VMH^SF-1 neurons. e CRH treatment did not effect on pair pulse ratio, indicating unlikely a pre-synaptic effect, n = 20/5, cells/animals; p = 0.138. f Representative traces of evoked IPSCs (eIPSCs) in the presence of CNQX in VMH^SF-1 neurons. g CRH treatment significantly decreased eIPSC amplitudes, n = 30/6, cells/animals; p = 0.005. h Representative image showed β3 GABA_A receptor subunit membrane trafficking in response to vehicle and CRH treatment, respectively. i Overall, CRH treatment induced a significant decrease of β3 GABA_A receptor subunit membrane expression when compared to vehicle group (repeat measurement, group effect p < 0.001). n = 12/3, cells/repeats. j GFP-immunoprecipitation (IP) shows increased S408/409 phosphorylation of β3 GABA_A receptors after CRH treatment. The samples derive from the same experiment and the gels/blots were processed in parallel. k Statistic for blotting results showed that CRH treatment significantly increased β3 receptor at S408/409. p = 0.012, n = 3 replicates. l Schematic of infusion of GABA_A receptor antagonist bicuculline in the VMH. m When compared with vehicle group, the administration of bicuculline increased blood glucose levels. p = 0.006. Two-sided paired t test in (b, e, g) or unpaired t test in (k, m), two-way repeated-measures ANOVA in (i). All data shown as means ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001; ns, not significant. Source data are provided as a Source Date file.