Fig. 1: LH OXSLD neurons show REM sleep-related activation.
a, b Schematic drawing (a) and representative CTB-555 (red)/CTB-488 (green) injections (b) for retrograde tracing. LH lateral hypothalamus, scp superior cerebellar peduncle. Mo5 trigeminal motor nucleus, 4V 4th ventricle, DMTg dorsomedial tegmental area, PnC caudal pontine reticular nucleus. Scale bar: 500 μm. c Representative SLD-projecting (OXSLD, arrow heads, CTB-488+/orexin-A+, cyan) and LC-projecting (OXLC, arrows, CTB-555+/orexin-A+, purple) orexin-A+ (blue) neurons in the rat LH. Scale bar: 50 μm. Examples (square boxes) were magnified on the right. Scale bars: 10 μm. d Percentage of orexin (OX) neurons projecting to the SLD, LC, and both (OXSLD/LC), respectively (n = 6 rats; analysis of 3902 orexin neurons). e Expression of OXRs (green) and orexin-A+ varicosities (red) in the rat SLD. Scale bars: left, 50 μm; right, 10 μm. f, g Expression of OXRs (red) with VGLUT1 (green, f) and GAD 67 (green, g) in the rat SLD. All scale bars: 20 μm. h Representative sleep/wakefulness architecture following the 72-h RSD. The rat was sacrificed for c-Fos immunostaining (IHC) after 2.5 h from the first REM sleep episode (gray arrow). Control rats were set with normal sleep/wakefulness cycle and underwent same processes. i Percentage of time spent in REM sleep (R), wakefulness (W), and NREM sleep (NR) during the 2.5 h preceding sacrifice in the control (black, n = 4) and RSD (red, n = 7) rats. j Representative c-Fos (red) expression in orexin-A+ (blue) and OXSLD (cyan) neurons (RSD group). C-Fos-positive OXSLD neurons (arrows) from the medial (n1) and lateral (n2) part of the LH were presented below. All scale bars, 20 μm. k Percentage of c-Fos expression in the OXentirety (blue, two-sided unpaired t-test; t9 = 0.817, P = 0.435) and OXSLD neurons (cyan, two-sided unpaired t-test; t9 = 4.839, P = 9.221 × 10−4) in the control (n = 4, analysis of 2767 OXentirety and 178 OXSLD neurons) and RSD (n = 7, analysis of 6012 OXentirety and 370 OXSLD neurons) rats, respectively. Data represent mean ± SEM. **P < 0.01. Source data are provided as a Source Data file.
