Adiponectin mediates metabolic feedback to the mediobasal hypothalamic circadian clocks.

The rotation of the Earth around its axis results in a repetitive succession of day and night. The profound environmental changes associated with the day-night cycle drove most organisms to evolve endogenous timekeepers to reliably anticipate predictable events at particular times of day and adjust their behaviors and physiology accordingly. Such endogenous timekeeping machineries are known as circadian (from Latin circa diem – about a day) clocks. In mammals, the cellular time-keeping machinery is comprised of an interlocked transcriptional-translational feedback loop (TTL) that during the daytime the transcriptional activating BMAL1/CLOCK complexes activate their own repressors PERs and CRYs which will then be degraded during the night. This molecular clockwork regulates local cellular physiology and is shared among the central circadian pacemaker – the suprachiasmatic nucleus (SCN) and other tissues in the brain and in the periphery. Recent studies have highlighted an extensive crosstalk between metabolism and circadian clock. For example, circadian misalignments contribute to metabolic disorders and vice versa. However, the mechanism of this link is still poorly understood. The mediobasal hypothalamus (MBH) is an assembly of hypothalamic nuclei which together play a major role in regulating behavioral rhythms such as feeding/fasting and sleep/wake cycles. It has been documented that the autonomous cellular clockwork exists in multiple MBH nuclei and regulates the local physiology such as electrophysiological properties and appetite-regulating neuropeptides (NP) expression, hinting for the role of molecular clock in appetite regulation. One of the most important features of the MBH is its ability to integrate information carried by circulating metabolic hormones to regulate energy homeostasis of the body. I therefore hypothesize that there are metabolic hormones that can modulate the molecular clock in the MBH and thereby regulating feeding rhythms. To search for metabolic hormones that can reset the MBH clock, I engineered a hypothalamic neuronal cell line to stably express a circadian reporter and used it as a model to screen for metabolic signals that are capable of resetting neuronal clocks. In a small scale screening, I identified an adipokines – adiponectin as a novel mediobasal hypothalamic cellular clock modulator. As it is known that circulating adiponectin levels are regulated by the metabolic status of the body, I further hypothesize that adiponectin is a mediator that can feed back to the MBH clocks according to the metabolic status of the body. I demonstrated that adiponectin possesses a phase-resetting effect in multiple in vitro models of MBH neurons and an induction effects on Bmal1 transcription in MBH neurons both in vitro and in vivo. Further molecular analyses revealed that these circadian effects of adiponectin are, at least in part, mediated by a adiponectin receptor 1 (AdipoR1), peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC1α) and RAR-related orphan receptor alpha (RORα) dependent mechanism. Using adiponectin deficient (Adipoq KO) mice as a model, I investigated the role of adiponectin in circadian behavioral rhythms. Adipoq KO mice have largely normal circadian locomotor activity rhythms and photic entrainment of the circadian clock. However, they show significant dampened 24-hr feeding rhythms associated with altered diurnal profiles of clock and appetite-regulating gene expression in the MBH. Moreover, the mutants also show abnormal food entrainment of the locomotor activity under a time-restricted feeding (RF) regime - known as food anticipatory activity (FAA). Conversely, compared to ad libitum fed animals, the RF regime significantly enhances the circadian oscillation of plasma adiponectin, upregulates the diurnal expression of adiponectin receptors and Pgc1a clock genes associated with a profound reorganization of the diurnal expression patterns of appetite-regulating genes in the MBH of wild-type mice. Furthermore, central delivery of an antagonist of RORα in wild-type mice could recapitulate the impaired FAA phenotypes of Adipoq KO mice. These data thus provide evidence to support the role of central adiponectin signaling in food entrainment of MBH clocks and feeding rhythms. Together, these data reveal a novel metabolic feedback mechanism to the central circadian clocks.