Proceedings of The Physiological Society

Sleep Sleep and Circadian Rhythms (London, UK) (2018) Proc Physiol Soc 42, C03

Poster Communications

Sleep-wake regulation in mice: insights from a synaptobrevin-2 mutant line and computational modelling

M. Guillaumin1, P. Achermann2, P. Nolan4, S. Peirson1, V. Vyazovskiy3

1. Nuffield Department of Clinical Neurosciences, University of Oxford, Oxford, United Kingdom. 2. Institute of Pharmacology and Toxicology, University of Zurich, Zurich, Switzerland. 3. Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom. 4. MRC Harwell Institute, Harwell, United Kingdom.

The alternation between waking and sleep is regulated by the internal circadian clock and sleep-wake history, and is also influenced by the external environment. Although our understanding of the circadian aspect of sleep regulation has increased, the mechanisms underlying sleep homeostasis are still largely unknown. Independent of the circadian clock, only a limited number of genes have been associated with specific sleep-wake properties. Forward genetics provides an unbiased approach, which seeks to identify genes involved in specific biological processes1. This project has focused on the Sleepy6 mouse line, obtained via a forward genetics sleep screen, with a mutation in synaptobrevin 2, giving rise to decreased sleep duration. We aimed to further characterise the sleep phenotype of this line, at a molecular and behavioural level, to gain novel insights into the regulation of sleep. Using molecular techniques (high-performance liquid chromatography, quantitative PCR) to evaluate neurotransmitter levels and gene expression, we found no significant difference in the neurotransmitter pathways investigated (n(wild-type-WT)= 8, n(mutant)=10, measured compounds and levels of expression of genes involved in serotonergic and dopaminergic pathways, mixed-design ANOVA followed by t-tests, p-values all > 0.05). Behavioural assays highlighted hyperactivity, with a mild learning impairment (n(WT)=28, n(mutant)=21, mixed-design ANOVA revealed a significant main effect of genotype: F(1,45)=8.93, p=0.005). Electrophysiology recordings revealed striking differences at global (electroencephalography - EEG) and neuronal levels, with Sleepy6 homozygous mice showing a decreased ability to switch between vigilance states, and notable alterations in neuronal firing patterns during slow-wave sleep (surgeries were performed under isoflurane anaesthesia, inhalation, 1.5-2.5%; n(WT)=5, n(mutant)=5, mixed-design ANOVA, p-values ranging from 0.001 to 0.041 when analysing time spent in vigilance states and episode durations). Finally, the successful adaption of an elaborated version of the "two-process" model2 of sleep regulation furthered our understanding of sleep/wake control in both wild-types and Sleepy6 homozygotes (n(WT)=5, n(mutant)=5, non-parametric Mann-Whitney tests, p<0.01 for the rate of decrease (WT: 60±8 *10-5 (4s)-1; mutant: 23±3 *10-5 (4s)-1, mean±SEM) and upper asymptote (WT: 447±33 %; mutant: 172±15 %, mean±SEM) of the simulated homeostatic process in the frontal EEG derivation). This combination of in vivo and computational work provides new insights into the mechanisms that underlie the homeostatic regulation of sleep, and more particularly, the alternation between vigilance states. It furthers our comprehension of a putative "sleep-switch", which allows animals to transfer between sleep and wakefulness in a biologically relevant manner.

Where applicable, experiments conform with Society ethical requirements