The impact of circadian entrainment and melatonin in kidney damage associated with ischaemia-reperfusion injury

Circadian rhythms regulate cellular, behavioural and physiological processes through central and peripheral clocks over a 24-hour period, with the suprachiasmatic nucleus (SCN) acting as the main enforcer of synchronising peripheral oscillators. Increasing evidence links circadian signalling to immune responses and susceptibility to pathological conditions such as ischaemia-reperfusion (IR) injury, an inevitable condition in organ transplantation and major cause of graft dysfunction. The kidneys are particularly sensitive to ischaemic injury, as renal microvasculature is vulnerable to hypoxia. Renal microvasculature, particularly pericytes that regulate capillary blood flow stability and immune cell infiltration, may represent a key site where circadian messengers influence IR onset and severity. This study aimed to investigate (i) whether circadian messengers; GABA, arginine vasopressin, corticosterone and melatonin directly modulate renal pericyte activity, and (ii) whether GABA can reset circadian responses in the kidney live slice model. Live kidney slices from C57BL/6J mice were exposed to individual circadian mediators (GABA 50 µM, arginine vasopressin 300 nM, corticosterone 50 µM and melatonin 50 µM) over a period of 500s, then observed to a total 1200s and 1800s and real-time vascular responses of vasa recta pericytes were observed and assessed under microscope. To assess GABA resetting potential and if it impacts the studies on pericyte regulation of medullary blood flow in the kidney slice model, slices were pre-treated with GABA (50 µM) for 30 minutes prior to corticosterone or melatonin exposure. All tested circadian messengers induced pericyte-mediated constriction, supporting their role in circadian regulation of renal microvasculature and a possible role in IR injury. Corticosterone evoked robust constriction, consistent with glucocorticoid-mediated stress responses, and could play a role in irreversible constriction observed in IR injury and acute kidney injury. Melatonin also constricted pericytes, potentially contributing to nocturnal suppression of micturition activity in mice. GABA induced significant constriction at low concentrations, though effects were inconsistent at 50 µM. Importantly, GABA pre-treatment did not alter pericyte response to corticosterone or melatonin, suggesting there is a capacity to reset peripheral renal clocks in this model. In conclusion, renal pericytes respond directly to circadian messengers, highlighting a mechanism by which circadian signals may influence renal vascular tone during IR injury. Corticosterone and melatonin emerge as key regulators, while the role of GABA in clock resetting requires further investigation. These findings support the potential for circadian-based therapeutic strategies, particularly targeting melatonin, to mitigate and possibly ameliorate renal IR injury.