The Effect of General Anaesthesia on the Circadian Clock of the Honey Bee Apis mellifera

Abstract

Circadian clocks are biological timepieces that govern the daily timing of most processes within an organism, thereby providing internal temporal coordination as well as appropriate synchronisation with the external world. Disturbance of this time structure has detrimental effects on well-being, performance and health, undermining immune responses and promoting disease progression. Whereas such disturbance is commonly associated with rapid travel across time zones, resulting in the syndrome of “jet lag”, clinical studies indicate that, after surgery, patients may experience similar circadian disruption. Whether general anaesthesia can affect the circadian clock and may thus underlie postoperative circadian disruption was investigated in this study. The honey bee (Apis mellifera) served as a model system that is not only of great chronobiological interest but also of high clinical relevance owing to the similarity of its molecular circadian clock to that of mammals. In addition, honey bees exhibit a range of circadian clock-based behaviours that are highly amenable to experimentation and provide multiple avenues for investigation. The effect of general anaesthesia on the circadian clock was assessed by analysing a colony activity rhythm as a behavioural output of the clock and by examining the expression of key circadian clock genes as a measure of the circadian clockwork itself. To this end, the circadian rhythm of hive entrance activity was characterised under both natural conditions outdoors and constant conditions in the laboratory using infra-red population recording as well as radiofrequency identification of individual bees. The temporal expression patterns of five putative clock genes in honey bee brains was also verified using an existing RT-qPCR assay. This assay was subsequently expanded and optimised to conform with current guidelines and ensure optimal assay performance for the detection of subtle expression changes after anaesthesia. Using the above assays, it was demonstrated that the common general anaesthetic isoflurane can significantly impact on circadian timing at clinical concentrations (2 %). Six hours of inhaled isoflurane anaesthesia during the subjective day phase delayed free-running rhythms of hive entrance activity by a mean of 4–6 hours. Likewise, the mRNA rhythms of the clock genes Period and Cryptochrome were delayed by 4–5 hours. These results show that anaesthesia not only affects circadian output but also phase shifts the underlying circadian feedback loop. The specificity of this effect was highlighted by its phase-dependency as the same anaesthetic treatment during the subjective night did not markedly alter phase of the activity and clock gene expression rhythms. The finding that the honey bee circadian system is markedly affected by isoflurane anaesthesia suggests that general anaesthesia may be an important contributor or even determinant of postoperative circadian disruption in patients. Therefore, addressing these chronobiotic effects of anaesthesia is likely to improve postoperative recovery.