Adolescent alcohol exposure increases orexin-A/hypocretin-1 in the anterior hypothalamus

Abstract

Adolescence is a time of marked changes in sleep, neuromaturation, and alcohol use. While there is substantial evidence that alcohol disrupts sleep and that disrupted sleep may play a role in the development of alcohol use disorders (AUD), there is very little known about the brain mechanisms underlying this phenomenon. The orexin (also known as hypocretin) system is fundamental for a number of homeostatic mechanisms, including the initiation and maintenance of wakefulness that may be impacted by adolescent alcohol exposure. The current study investigated the impact of adolescent ethanol exposure on adult orexin-A/hypocretin-1 immunoreactive (orexin-A + IR) cells in hypothalamic nuclei in two models of adolescent intermittent ethanol (AIE) exposure. Both models assess adult hypothalamic orexin following either an AIE vapor exposure paradigm, or an AIE intragastric gavage paradigm during adolescence. Both AIE exposure models found that binge levels of ethanol intoxication during adolescence significantly increased adult orexin-A + IR expression in the anterior hypothalamic nucleus (AHN). Further, both AIE models found no change in orexin-A + IR in the posterior hypothalamic area (PH), perifornical nucleus (PeF), dorsomedial hypothalamic nucleus dorsal part (DMD) or lateral hypothalamic area (LH). However, AIE vapor exposure reduced orexin-A + IR in the paraventricular nucleus (PVN), but AIE gavage exposure did not. These findings suggest that the AHN orexinergic system is increased in adults following binge-like patterns of intoxication during adolescence. Altered adult AHN orexin could contribute to long-lasting changes in sleep.

Keywords: adolescence; hypocretin/orexin; hypothalamus.

Figure 1.. Schematic drawings of the hypothalamic regions

Schematic drawings of the hypothalamic regions analyzed for orexin-A+IR expression in adolescent intermittent ethanol (AIE) and control (CONT) rats. The regions of interest included (A) anterior hypothalamic nucleus (AHN), periventricular hypothalamic nucleus (Pe) and paraventricular hypothalamic nucleus (PVN) at −1.80 mm; (B) lateral hypothalamic area (LH), dorsomedial hypothalamic nucleus, dorsal part (DMD) and perifornical nucleus (PeF) at −2.80 mm; and (C) posterior hypothalamic area (PH) at −4.18 mm. Figures have been adapted from the Paxino and Watson (1998) rat atlas.

Figure 2.. Orexin-A+IR Expression in AHN after AIE Exposure

(A) The density of hypothalamic orexin-A+IR expression (x1000, pixels/mm²) was analyzed in young adult rats after intermittent ethanol vapor (top) or intragastric gavage (bottom) during adolescence. Orexin-A+IR in the AHN was significantly increased in AIE rats compared to controls (CONT) for both methods of exposure. (B) Example orexin-A+IR staining in AHN for AIE and CONT rats after vapor (top) or intragastric (bottom) exposure models. Scale bar = 50μm. *p < 0.05 vs. controls.

Figure 3.. Orexin-A in various hypothalamic nuclei after AIE Exposure

The density of hypothalamic orexin-A+IR expression (x1000, pixels/mm²) was analyzed after adolescent rats were exposed to either adolescent intermittent vapor (top) or gavage (bottom) models with ethanol (AIE) or non-ethanol controls (CONT). (A) Orexin-A+IR expression was analyzed in the Pe and PVN at Bregma −1.8 mm. AIE vapor exposure significantly decreased PVN orexin-A+IR when compared to air-matched controls, however this result was not replicated in the gavage model. No difference in orexin-A+IR were seen in the Pe of the hypothalamus for either model. (B) There was also no significant effect of AIE vapor or gavage model on orexin-A+IR density at Bregma −2.8 mm in the LH, DMD, or PeF of the hypothalamus. (C) Orexin-A+IR (pixel/mm²) in the PH at a depth of −4.16mm was no different for AIE compared to CONT for either vapor (top) or gavage (bottom) exposure model. *p < 0.05 vs. controls