Characterization of Sleep in Zebrafish and Insomnia in Hypocretin Receptor Mutants

Abstract

Sleep is a fundamental biological process conserved across the animal kingdom. The study of how sleep regulatory networks are conserved is needed to better understand sleep across evolution. We present a detailed description of a sleep state in adult zebrafish characterized by reversible periods of immobility, increased arousal threshold, and place preference. Rest deprivation using gentle electrical stimulation is followed by a sleep rebound, indicating homeostatic regulation. In contrast to mammals and similarly to birds, light suppresses sleep in zebrafish, with no evidence for a sleep rebound. We also identify a null mutation in the sole receptor for the wake-promoting neuropeptide hypocretin (orexin) in zebrafish. Fish lacking this receptor demonstrate short and fragmented sleep in the dark, in striking contrast to the excessive sleepiness and cataplexy of narcolepsy in mammals. Consistent with this observation, we find that the hypocretin receptor does not colocalize with known major wake-promoting monoaminergic and cholinergic cell groups in the zebrafish. Instead, it colocalizes with large populations of GABAergic neurons, including a subpopulation of Adra2a-positive GABAergic cells in the anterior hypothalamic area, neurons that could assume a sleep modulatory role. Our study validates the use of zebrafish for the study of sleep and indicates molecular diversity in sleep regulatory networks across vertebrates.

Augmented generation of protein fragments during wakefulness as the molecular cause of sleep: a hypothesis

“…Adult zebrafishes are diurnal, being active during the day and less active at night, with prolonged episodes of sleep, defined behaviorally. 12,256,257 If adult zebrafishes were sleepdeprived, in darkness, through repetitive electric stimulation and thereafter observed during an extended period of darkness, they exhibited prolonged sleep, as would be expected of responses to sleep deprivation in mammals. However, this compensatory prolongation of sleep after sleep deprivation was not observed if sleep-deprived zebrafishes were released into light.…”

“…However, this compensatory prolongation of sleep after sleep deprivation was not observed if sleep-deprived zebrafishes were released into light. 256 Moreover, sleep-deprived zebrafishes that were kept under constant light conditions exhibited little sleep to begin with, despite the preceding sleep deprivation in darkness. 256 A similar and nearly complete suppression of sleep was also observed with zebrafishes kept under constant light for at least 3 days.…”

“…256 Moreover, sleep-deprived zebrafishes that were kept under constant light conditions exhibited little sleep to begin with, despite the preceding sleep deprivation in darkness. 256 A similar and nearly complete suppression of sleep was also observed with zebrafishes kept under constant light for at least 3 days. Sleep reappeared and progressively increased by 8 days in the presence of light.…”

“…No significant rebound sleep was observed afterwards in darkness, despite the (formally defined) sleep deprivation under constant-light conditions. 256 In the framework of the FG hypothesis, these, currently unexplained, sleep patterns in zebrafishes may be accounted for by different rates of reactions (under different conditions of sleep deprivation described above) that mediate the generation or degradation of specific protein fragments and the remodeling of protein complexes that initially contained these fragments. Addressing these issues experimentally with zebrafishes can involve, at first, quantitative comparisons of the levels of proteasegenerated protein fragments, either of a kind described in Figures 3 to 6 or of any kind, using proteome-scale, mass spectrometry-based techniques.…”

Sleepless in Drosophila

“…Although sleep is an essential process conserved from flies to humans, 1 finding a human homolog of the gene may prove to be difficult. However, regulation of sleep remains a poorly understood phenomenon, and knowing the homolog would aid in the design of better insomnia drugs.…”