| Summary: | <p>Sleep is a universal and highly conserved phenomenon across species; yet its operational mechanisms remain one of the greatest biological mysteries. In Drosophila, sleep homeostasis is largely mediated by the activity of neurons projecting to the dorsal fan-shaped body (dFB). Dopamine promotes wakefulness by inhibiting the activity of dFB neurons, switching them to a silent state. This wake-promoting effect is hypothesised to require the exocytosis of the potassium “leak” channel, Sandman, to the plasma membrane, where it exerts its effects by blunting the firing of the neurons. In the chronically sleep-deprived flies bearing a mutation for the crossveinless-c (cv-c) gene, the dFB neurons were previously found in electrical quiescence as well. It is therefore possible that in these mutants, Sandman resides at the plasma membrane of the neurons, reducing their excitability and causing insomnia in the flies. However, the signals responsible for Sandman’s recycling to and from the plasma membrane remain elusive.</p>
<p>To visualise the exo-endocytic pathway of Sandman, I attempted to localise the channel with different fluorescent probes and identify proteins acting in the same pathway using genetic interaction experiments. Depletion of Sandman from the dFB neurons rescues the sleep loss of cv-c mutants, supporting the notion that insomnia in these flies results from the channel’s presence at the plasma membrane. As cv-c encodes for a Rho GTPase-activating protein (Rho-GAP) and small GTPases are often regulated by neurotrophin-related pathways, sleep induction involving the Cv-c-Sandman pathway may further couple to the upstream action of neurotrophic molecules.</p>
<p>dFB-restricted interference with Toll-6, a receptor with neurotrophic function, reduces sleep. Toll-6/cv-c mutants mirror the sleepless phenotype of cv-c mutants, suggesting that Toll-6 may signal through Cv-c, conveying sleep pressure to the neurons. A mutation for Drosophila neurotrophin 1 (DNT1), a ligand of Toll-6, also diminishes sleep. RNAi-mediated knockdown of DNT1 in the R5 neurons, a defined input to the dFB, decreases sleep and abolishes sleep drive encoded by the circuit.</p>
<p>It is proposed here that activity-dependent release of DNT1 from the R5 neurons signals sleep pressure that is transduced by the Toll-6 receptor in the dFB neurons. Activation of the receptor likely feeds into a small GTPase cascade that involves Cv-c and Sandman.</p>
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