Molecularly distinct subtypes of Lhx6-positive neurons of the zona incerta differentially regulate sleep pressure and recovery sleep

Recent work implicates Lhx6-positive GABAergic neurons of the zona incerta (ZI) in regulating sleep pressure, but their precise role remains unclear. Using sleep deprivation and HiPlex single-molecule fISH, we show that Lhx6-positive ZI neurons are broadly activated by both natural and induced increases in sleep pressure and remain active for more than three hours into recovery sleep.

Anterior Lhx6-positive neurons showed stronger activation. Fos induction differed across molecularly distinct subpopulations, with Nkx2-2-positive cells showing robust responses and Calb2-positive cells showing reduced activation. We also identified distinct sleep pressure-responsive Lhx6-negative Slc32a1-positive GABAergic ZI subpopulations.