Characterising the Effects of General Anaesthesia on Circadian Clock and Clock Gene Expression in Drosophila melanogaster

Abstract

General anaesthesia (GA) is implicated in post-operative sleep disruption and fatigue with part of the disturbance being attributed to a shift of the circadian clock. Isoflurane is a commonly used general anaesthetic during surgery. In this research, Drosophila melanogaster was used to characterise the effects of GA on the circadian clock at behavioural (locomotor activity) and molecular (clock gene expression) levels. Drosophila locomotor activity was recorded using a Drosophila activity monitoring (DAM) system and the clock gene (period and timeless) expression was monitored by bioluminescence using the transgenic luciferase reporter strains. The effects of length and dose of GA were investigated at the same circadian time (CT 4) by increasing the length of GA from one hour to six hours and the concentration of Isoflurane from 0.5% to 3%. The results indicated that six hours of 2% Isoflurane intervention induced the most significant phase shifts in locomotor activity. From this, six hours of 2% Isoflurane administered at two-hour intervals was used in subsequent experiments to investigate the effects of GA on locomotor activity and clock gene expression throughout an entire circadian time cycle. The phase response curve (PRC) summarised from these results indicated that subjective daytime GA interventions induced phase advances while subjective night-time GA interventions induced phase delays in locomotor activity. The study also found that the PRC from GA-induced shifts of period expression occurred two hours ahead of the phase shifts in locomotor activity, suggesting that the shifts of clock gene expression preceded the shifts of behavioural rhythms. Furthermore, the locomotor activity and gene expression experiments were held in simulated light and dark (LD) cycles to investigate the effects of GA on circadian rhythms at both behavioural and molecular levels. The results indicated that GA-induced phase shifts in locomotor activity were masked by light but the peripheral clock gene (period and timeless) expressions were shifted by GA. Lastly, GA with and without one-hour light exposure in constant darkness (DD) conditions were conducted to investigate whether a short period of light exposure can block the GA-induced shifts in the circadian clock. The PRC results showed that GA with light exposure has a synergistic effect on activity and period expression. In conclusion, this research has characterised the effects of GA on circadian clocks in Drosophila from various environmental cues and provided theoretical support for further investigation of possible health care treatment to minimise GA-induced negative effects on circadian rhythms.