A Sleep/Wake Circuit Controls Isoflurane Sensitivity in Drosophila

Abstract

General anesthesia remains a mysterious phenomenon, even though a number of compelling target proteins and processes have been proposed [1]. General anesthetics such as isoflurane abolish behavioral responsiveness in all animals, and in the mammalian brain, these diverse compounds probably achieve this in part by targeting endogenous sleep mechanisms [2, 3]. However, most animals sleep [4], and they are therefore likely to have conserved sleep processes. A decade of neurogenetic studies of arousal in Drosophila melanogaster have identified a number of different neurons and brain structures that modulate sleep duration in the fly brain [5-9], but it has remained unclear until recently whether any neurons might form part of a dedicated circuit that actively controls sleep and wake states in the fly brain, as has been proposed for the mammalian brain [10]. We studied general anesthesia in Drosophila by measuring stimulus-induced locomotion under isoflurane gas exposure. Using a syntaxin1A gain-of-function construct, we found that increasing synaptic activity in different Drosophila neurons could produce hypersensitivity or resistance to isoflurane. We uncover a common pathway in the fly brain controlling both sleep duration and isoflurane sensitivity, centered on monoaminergic modulation of sleep-promoting neurons of the fan-shaped body.

Behavioral and electrophysiological analysis of general anesthesia in 3 background strains ofDrosophila melanogaster

“…Behavioral responsiveness under isoflurane anesthesia uncovers a significant difference between 2 white strains We examined adult fly behavior under isoflurane anesthesia using a stimulus-induced locomotion assay 13,14 (Fig. 1A).…”

“…6,7 Behavioral assays Startle assay The stimulus-induced locomotion assay assesses flies' movement response to a startle-inducing vibration stimulus, and has been described previously. 13,14 Briefly, flies are placed in individual tubes, with their movement constantly filmed via a webcam. Locomotion before and after a mechanical vibration stimulus is quantified using a custom Matlab program (Drosophila Arousal Tracking, DART 10 ) before and during isoflurane anesthesia.…”

“…There are numerous studies reporting Drosophila mutant strains with different anesthetic sensitivities. [12][13][14][15][16][26][27][28][29][30][31][32] Differences in general anesthesia sensitivities of different CS stocks has also been previously noted, 12 and individual background strains have been characterized for their anesthetic sensitivity for specific endpoints, 33 but a direct comparison across a range of endpoints in commonly used background strains has not been reported. In this study, we asked 2 broad questions: do different background strains display different sensitivity to isoflurane, and are any differences systemic across various endpoints in different life stages?…”

“…A previous study has shown that w 1118 is slightly resistant to general anesthetics, 15 and it was suggested that this was due to the mutant white gene which may modulate dopaminergic signaling, and hence general anesthesia and sleep. 13 However, it was unknown if this resistance effect in w 1118 is life-stage dependent, and whether it is also evident across different anesthetic endpoints. By comparing the anesthetic phenotypes of another white mutant (isoCJ1) for all endpoints, we addressed whether these anesthesia phenotypes are likely to stem from the white gene.…”

“…Here, we used newly established general anesthesia assays 10,13,14 to characterize behavioral and electrophysiological endpoints in these common genetic background strains (isoCJ1 and w 1118 ), and compared these throughout to CS. A previous study has shown that w 1118 is slightly resistant to general anesthetics, 15 and it was suggested that this was due to the mutant white gene which may modulate dopaminergic signaling, and hence general anesthesia and sleep.…”