Fig. 7: Responsiveness of PVH Sim1 neurons to acute stress.

a, b A diagram showing delivery of AAV-Flex-GCaMP6m viral vectors to bilateral PVH of Sim1-Cre mice (a) and representative expression of GCaMP6m (green) and tdTomato in the PVH after delivery of AAV-Flex-GCaMP6m to Sim1-Cre::Ai9 mice (b). c, e, f, h Sim1-Cre (or Sim1-Cre::Ai9) male mice received stereotaxic delivery of AAV-FLEX-GCaMP6m (c, h), a mixture of AAV-FLEX-GCaMP6m and AAV-DIO-Kir2.1-P2A-dTomato vectors (e), a mixture of AAV-FLEX-GCaMP6m and AAV-EF1a-FLEX-EGFP-P2A-mNachBac vectors (f). d, g Bmal1^flox/flox mice (d) or γ2^flox/flox::γ2-TB^lox/lox mice (g) receiving a mixture of AAV-FLEX-GCaMP6m and AAV-Cre-GFP (to bilateral PVH at 7–8 weeks of age with optic fiber implantation targeting PVH neurons). There mice were then used to monitor acute activity responses to a single water spray with sprayer positioned at a fixed distance toward mouse head (arrows indicating onset of water spray). c–g A representative trace showing responses of PVH Sim1 neurons to water spray in the animal model indicated. h A representative response of the same mice used in c after 6 weeks on HFD. i Comparison in activation of PVH neurons to water-spray among mouse groups. Data are represented as mean ± SEM. One-way ANOVA, n = 5 or 6, ***p < 0.0001, Control vs Bmal1, vs NachBac, vs Kir2.1, vs γ2, or vs HFD. All mice were fed chow unless otherwise noted. j A simplified diagram showing the proposed mechanism based on the presented results. Diurnal BMAL1 expression pattern elicits diurnal patterns in GABAergic inputs through regulation of γ2 subunit, which maintains a diurnal rhythm of PVH neurons activity and metabolism. HFD diminishes diurnal rhythms of PVH neuron activity, likely through blunting neuron responsiveness and promotes obesity through disruption of diurnal rhythms in metabolism. All data are presented as mean ± SEM. Source data are provided as a Source Data file.