Fig. 2: Optogenetic activation of RMTg GABAergic neurons terminated REM sleep and inhibited REM sleep induction.

a Schematic illustration of virus injection. Adeno-associated virus (AAV) encoding channelrhodopsin 2 (ChR2) fused to an enhanced red fluorescent protein (mCherry) reporter was introduced into the bilateral RMTg in VGAT-Cre mice. b The coronal section shows the superimposed virus-injected area in five mice. c A ChR2-mCherry expressing RMTg GABAergic neuron showed spiking following 30 Hz blue light illumination in current–clamp mode (top). Fidelity analysis of the action potential evoked by stimulus at frequencies ranging from 1 Hz to 100 Hz (bottom). d Representative EEG and EMG traces and corresponding heat map of EEG power spectrum in different sleep–wake states from a mouse in the mCherry (top) and ChR2 (bottom) group, respectively. Blue line, laser stimulation (30 Hz, 120 s,10 ms per pulse). e, h Sleep–wake state changes (top) and probability of brain state transitions (bottom) after photostimulation was applied during REM sleep lasting no less than 16 s before laser on (e) and NREM sleep lasting no less than 60 s before laser on (h) in all trials from mice in the mCherry (left) and ChR2 (right) groups, respectively. Shading indicates 95% confidence intervals. f Mean latency of REM sleep transitions to wakefulness or NREM sleep after blue laser stimulation during REM sleep in the first eight trials per mouse. p = 0.0159, Mann–Whitney U test. g, i During 120 s laser stimulation, the cumulative probability for REM sleep termination, wake, and NREM sleep initiation when laser illuminated during REM sleep (g) and REM sleep and wake initiation when laser illuminated during NREM sleep (i). Colorful stairs, ChR2 group. Black stairs, mCherry group. Data represent mean ± SEM. *p < 0.05 vs. mCherry group, Kolmogorov–Smirnov test. ChR2: n = 5, mCherry: n = 4. Source data are provided as a Source Data file.