Guy R. Warman

Disruption of daily patterns of activity, physiology, or social interaction is associated with stress and reduced fitness across species. In social insects such as honey bees (Apis mellifera), circadian organisation extends to the colony level, where rhythms emerge from interactions among individual clocks, hive conditions, and environmental cues. Precision apiculture offers a mature sensor toolkit for measuring colony-level variables, and several data streams carry early-warning information about colony condition. Almost all alarms, however, are triggered against set points rather than against the daily rhythm of the signal. We review each sensor technology to establish whether its data have been, or could be, analysed with a chronobiological lens, describing the measurement principle, the achievable temporal resolution and record continuity, the rhythmic signature expected on biological grounds, and any evidence that such a signature has been reported. We then use a synthetic dataset to illustrate the workflow: in this simulated example, rhythm metrics flagged a gradually developing decline approximately three weeks before a conventional threshold alert on the same data. This lead time is a property of the simulation and awaits validation on real colonies. Wherever continuous behavioural monitoring is deployed, rhythm-based metrics may offer a route to earlier, more interpretable detection of change than set point comparisons.

GABAergic neurotransmission plays a central role in regulating the mammalian circadian clock. However, the precise contribution of GABA type A receptor (GABA_AR) subunits to timekeeping within the suprachiasmatic nucleus (SCN) remains incompletely defined. Here, we investigated the spatiotemporal expression patterns of four GABA_AR subunits (α1, α5, β3, and γ2) in the SCN of male and female C57BL/6 mice. Our aim was to determine whether these subunits exhibit circadian, regional, and sex-specific variation under entrained (LD12:12) and free-running (DD) conditions. Fluorescence immunohistochemistry revealed the expression of all four subunits in the SCN, with several displaying rhythmic profiles. The α1 subunit peaked during the day in both sexes and was preferentially expressed in the shell region. The α5 subunit also varied across time but lacked strong regional specificity. Temporal variation in β3 and γ2 expression was observed in males but not females, indicating potential sex-dependent regulation. Notably, the γ2 subunit showed a dynamic shift in regional distribution over the circadian cycle, with higher shell expression during the day and core enrichment at night. Under DD conditions, rhythmic expression of the α1 and γ2 subunits persisted, suggesting endogenous control independent of external light cues. However, overall amplitude and regional specificity were reduced in the absence of light. These findings demonstrate that GABA_AR subunit expression within the SCN is regulated by circadian phase, SCN subregion, and biological sex. This study provides new insight into the molecular architecture of the SCN and supports the hypothesis that subunit-specific GABAergic signalling contributes to the temporal organisation of the circadian clock. Further investigation into the functional significance of these subunit patterns will help clarify their roles in SCN synchrony, photic entrainment, and sex-specific circadian regulation.

The immune response to bacterial infection demonstrates circadian rhythmicity, with heightened antibacterial immunity often observed during the daytime. This circadian‐gated immune response is believed to have evolved to synchronize peak antibacterial immunity with the increased threat of infection when active. Using a larval zebrafish infection model, we have previously shown that light has a positive effect on antibacterial immunity, with enhanced survival and neutrophil recruitment observed when infections are delivered during the light phase. However, the specific contribution of the light‐detecting pineal gland and its principal hormone, melatonin, to this circadian immune response has not been investigated. Here, we examined whether the pineal gland modulates the host response to bacterial infection in a time‐of‐day–dependent manner by targeted ablation of the pineal gland. To generate pineal gland‐deficient larvae we created the transgenic zebrafish line Tg(exorh:YFP‐NTR2.0) that restricts expression of the highly efficient nitroreductase variant NTR2.0 to pinealocytes, enabling metronidazole‐mediated pinealectomy. We also analyzed larvae lacking the brain‐specific homeobox ( bsx ) gene, that encodes a transcription factor essential for pineal gland development. Both approaches resulted in complete loss of the pineal gland, confirmed by live imaging, the absence of arylalkylamine N‐acetyltransferase 2 ( aanat2 ) expression and diminished melatonin production. Pineal gland‐deficient larvae exhibited normal survival and neutrophil recruitment when infected during the light phase, indicating that daytime immune defenses can be maintained independently of the pineal gland. In contrast, infection of pineal gland‐deficient larvae during the dark phase led to significantly reduced survival, which correlated with impaired neutrophil recruitment to the infection site. These findings demonstrate that the pineal gland plays an important role in supporting nighttime antibacterial immunity in larval zebrafish.

The immune response exhibits strong circadian rhythmicity, with enhanced bacterial clearance often synchronized with an organism’s active phase. Despite providing the bulk of cellular antibacterial defense, the neutrophil clockwork is poorly understood. Here, we used larval zebrafish to explore the role of clock genes in neutrophils during infection. Per2 was required in neutrophils for reactive oxygen species (ROS) production and bacterial killing by enhancing infection-responsive expression of high-mobility group box 1a ( hmgb1a ). The Cry binding domain of Per2 was required for regulation of neutrophil bactericidal activity, and neutrophils lacking Cry1a had elevated bactericidal activity and infection-responsive hmgb1a expression. A conserved cis-regulatory element with BMAL1 and nuclear factor κB binding motifs gated infection-responsive hmgb1a expression to the light phase. Mutagenesis of the BMAL1 motif in neutrophils blunted the priming effect of light on bactericidal activity and hmgb1a expression. These findings identify a light-responsive cell-intrinsic timer that controls time-of-day variations in antibacterial activity.

General anaesthesia (GA) is known to affect the circadian clock. However, the mechanisms that underlie GA-induced shifting of the clock are less well understood. Activation of γ-aminobutyric acid (GABA)_-type A receptors (GABA_AR) in the suprachiasmatic nucleus (SCN) can phase shift the clock and thus GABA and its receptors represent a putative pathway via which GA exerts its effect on the clock. Here, we investigated the concurrent effects of the inhalational anaesthetic, isoflurane, and light, on mouse behavioural locomotor rhythms and on α1, β3, and γ2 GABA_AR subunit expression in the SCN of the mouse brain. Behavioural phase shifts elicited by exposure of mice to four hours of GA (2% isoflurane) and light (400 lux) (n = 60) were determined by recording running wheel activity rhythms in constant conditions (DD). Full phase response curves for the effects of GA + light on behavioural rhythms show that phase shifts persist in anaesthetized mice exposed to light. Daily variation was detected in all three GABA_AR subunits in LD 12:12. The γ2 subunit expression was significantly increased following GA in DD (compared to light alone) at times of large behavioural phase delays. We conclude that the phase shifting effect of light on the mouse clock is not blocked by GA administration, and that γ2 may potentially be involved in the phase shifting effect of GA on the clock. Further analysis of GABA_AR subunit expression in the SCN will be necessary to confirm its role.

Following general anaesthesia (GA), patients frequently experience sleep disruption and fatigue, which has been hypothesized to result at least in part by GA affecting the circadian clock. Here, we provide the first comprehensive time-dependent analysis of the effects of the commonly administered inhalational anaesthetic, isoflurane, on the murine circadian clock, by analysing its effects on (a) behavioural locomotor rhythms and (b) PER2::LUC expression in the suprachiasmatic nuclei (SCN) of the mouse brain. Behavioural phase shifts elicited by exposure of mice (n = 80) to six hours of GA (2% isoflurane) were determined by recording wheel-running rhythms in constant conditions (DD). Phase shifts in PER2::LUC expression were determined by recording bioluminescence in organotypic SCN slices (n = 38) prior to and following GA exposure (2% isoflurane). Full phase response curves for the effects of GA on behaviour and PER2::LUC rhythms were constructed, which show that the effects of GA are highly time-dependent. Shifts in SCN PER2 expression were much larger than those of behaviour (c. 0.7 h behaviour vs. 7.5 h PER2::LUC). We discuss the implications of this work for understanding how GA affects the clock, and how it may inform the development of chronotherapeutic strategies to reduce GA-induced phase-shifting in patients.

General anaesthesia (GA) is implicated as a cause of postoperative sleep disruption and fatigue with part of the disturbance being attributed to a shift of the circadian clock. In this study, Drosophila melanogaster was used as a model to determine how Isoflurane affects the circadian clock at the behavioural and molecular levels. We measured the response of the clock at both of these levels caused by different durations and different concentrations of Isoflurane at circadian time 4 (CT4). Once characterized, we held the duration and concentration constants (at 2% in air for 6 h) and calculated the phase responses over the entire circadian cycle in both activity and period expression. Phase advances in behaviour were observed during the subjective day, whereas phase delays were associated with subjective night time GA interventions. The corresponding pattern of gene expression preceded the behavioural pattern by approximately four hours. We discuss the implications of this effect for clinical and research practice.

The way in which the circadian clock mechanism develops and decays throughout life is interesting for a number of reasons and may give us insight into the process of aging itself. The Drosophila model has been proven invaluable for the study of the circadian clock and development and aging. Here we review the evidence for how the Drosophila clock develops and changes throughout life, and present a new conceptual model based on the results of our recent work. Firefly luciferase lines faithfully report the output of known clock genes at the central clock level in the brain and peripherally throughout the whole body. Our results show that the clock is functioning in embryogenesis far earlier than previously thought. This central clock in the fly remains robust throughout the life of the animal and only degrades immediately prior to death. However, at the peripheral (non-central oscillator level) the clock shows weakened output as the animal ages, suggesting the possibility of the breakdown in the cohesion of the circadian network.