Integration of circadian systems with the light/dark environment involves a widely distributed network of local tissue clocks within both the brain and periphery (33–36) (Figure 3). Interestingly, a variety of factors including food availability, glucocorticoid level, and temperature, so called “zeitgebers” (“time givers”), are able to reset the phase of peripheral clocks. In contrast, the SCN is entrained only to light but not to feeding; thus shifts in feeding time uncouple the phase of peripheral tissue clocks from that of the SCN (37–40).

Figure 3 Interactions between the molecular clock and downstream metabolic genes. The core molecular clock consists of several transcription/translation feedback loops, including posttranscriptional regulation (yellow), that oscillate with an approximately 24-hour periodicity. CLOCK and BMAL1 heterodimerize to drive rhythmic expression of downstream target genes (shown in red), which in turn regulate diverse metabolic processes, including glucose metabolism, lipid homeostasis, and thermogenesis. Many of these clock target genes in turn reciprocally regulate the clock in response to changes in nutrient status (shown in blue) via cellular nutrient sensors (shown in orange), generating a complex network of interlocking feedback loops that fine-tune the clock and coordinate metabolic processes with the daily cycles of sleep/wakefulness and fasting/feeding. Dashed lines represent metabolic inputs; solid lines depict interactions among core clock genes, clock-controlled genes, and nutrient sensors.

Feedback loops between nutrient sensors and molecular clocks in peripheral tissues (NAD+, NAMPT, SIRT1, AMPK). Recent studies have demonstrated that various nutrient sensors are able to relay information regarding the cellular nutrient status to the circadian clock. For example, early studies indicated that the reduced forms of the redox cofactor NADH increase CLOCK/BMAL1 and NPAS2/BMAL1 heterodimer activity, whereas the oxidized forms (NAD+) inhibit their activity (41). The NAD+-dependent deacetylase sirtuin 1 (SIRT1), which is induced by acute nutrient withdrawal (42) or calorie restriction (43), also directly binds to the CLOCK/BMAL1 complex to regulate expression of clock genes (44–47). AMP kinase (AMPK) serves as another highly conserved cellular nutrient sensor that, when activated by exercise, fasting, or hypoxia (48), leads to phosphorylation and degradation of CRY1 (49), as well as activation of NAMPT, the rate-limiting enzyme in NAD+ salvage biosynthesis (50). Interestingly, the connection between nutrient sensors and molecular clocks is bidirectional. Levels of NAMPT, as well as NAD+ itself, exhibit circadian oscillations, and Nampt gene transcription is directly regulated by CLOCK (46, 47). As such, SIRT1 activity is indirectly regulated by the clock network. Together, these data demonstrate on a molecular level that there is a complex crosstalk between highly conserved nutrient sensors and the molecular clock networks, at least at the level of peripheral tissues (see below for discussion regarding potential roles of CNS nutrient sensors in circadian control of metabolism).

Metabolic transcription factor and transcriptional co-activator feedback on the core clock in peripheral tissues (REV-ERB, ROR, PPAR, DBP, PGC1α). NHRs are ligand-activated transcription factors, a majority of which, including REV-ERBα, RORs and PPARs, demonstrate circadian expression in peripheral tissues (51). REV-ERBα regulates hepatic gluconeogenesis, adipocyte differentiation, and lipid metabolism, and also represses Bmal1 transcription (52, 53). RORα competes with REV-ERBα in binding to the Bmal1 promoter and induces Bmal1 expression (54) in addition to regulating lipid metabolism (55). PPARα, when activated by endogenous fatty acids (FAs), stimulates FA oxidation, regulates genes controlling lipid homeostasis, and prevents atherosclerosis (56). In addition, PPARα positively regulates Bmal1 expression, while BMAL1 likewise activates PPARα, generating a positive feedback loop (57). Another subtype, PPARγ, plays an important role in adipocyte differentiation and triglyceride synthesis (58). PPARγ induces Bmal1 expression in the blood vessels, and vasculature-specific PPARγ knockout leads to marked reduction in the circadian variation in blood pressure and heart rate (59).

In addition to NHRs, Dbp, a known clock target gene, regulates expression of key metabolic genes involved in gluconeogenesis and lipogenesis (60). Because DBP levels change 100-fold in response to CLOCK/BMAL1 activation, it is conceivable that DBP generates circadian oscillation in metabolic processes such as gluconeogenesis. Additionally, the transcriptional co-activator PPARγ co-activator 1α (PGC-1α) (61) also displays circadian oscillations and regulates Bmal1 and Rev-erba expression. Knockout of Pgc1a leads to abnormal diurnal rhythms of activity, body temperature, and metabolic rate, in addition to aberrant expression of clock and metabolic genes (62). Interestingly, SIRT1 suppresses PPARγ (63) but activates PGC-1α (64), and thus affects the clock network through multiple mechanisms.

Possible molecular integrators of circadian and metabolic systems in the CNS. Molecular analyses of interplay between circadian and metabolic pathways have primarily emerged from studies in liver and other peripheral tissues, yet it remains uncertain whether similar mechanisms might also couple these processes within the brain. One question is whether specific metabolites that vary according to time of day and nutrient state (fasting vs. feeding), may also affect circadian function of energy-sensing neurons. For instance, 24-hour oscillation in levels of glucose and FAs may in turn influence expression of circadian genes and rhythmic transcriptional outputs within hypothalamic neurons involved in glucose homeostasis (65), food intake (66), and energy expenditure (67–70). Perturbation of metabolic homeostasis with a high-fat diet is sufficient to alter both period length and amplitude of locomotor activity (71). These observations indicate that changes in FA metabolism per se may either alter clock gene function within SCN pacemaker neurons and/or interrupt communication between SCN and extra-SCN neurons.

Additional nutrient factors that may participate in circadian oscillations of SCN and extra-SCN neurons include AMPK, SIRT1, and the mammalian target of rapamycin (mTOR). Hypothalamic AMPK is regulated by nutrient state and hormones such as leptin and insulin (72), and manipulation of its expression alters food intake and body weight (73); however the impact of AMPK on CNS control of locomotor behavior and physiological rhythms is still not known. NPY and POMC neuron-specific knockout of the α2 subunit of AMPK results in lean and obese phenotypes, respectively, so it will be interesting to learn whether AMPK also participates in synchronizing activity and feeding through actions within these cell groups (74). SIRT1 may also represent an additional mechanism coupling nutrient flux with rhythmic activity of hypothalamic neurons, as the nutrient-sensing deacetylase is present within both ARC and DMH/LHA neurons (75, 76). A final potential mediator involved in both energy sensing and circadian function in the CNS is mTOR, a regulator of protein synthesis present in ARC neurons that has been shown to modulate food intake (77). mTOR is also expressed within pacemaker neurons of the SCN, where its expression is activated by light (78). Moreover, activation of mTOR causes phase resetting of SCN explants (79), while inhibition of mTOR alters light induction of the Period gene within the SCN of the intact animal (78). Thus amino acid metabolism may participate in both entrainment of the master clock and in the temporal organization of feeding.

Nutrient signaling, whether through glucose, FAs, AMPK, SIRT1, or mTOR, may function to entrain or “gate” CNS and peripheral clocks, leading to tissue-specific differences in phase and amplitude of gene expression rhythms. Indeed, different zeitgebers may exert differing effects on circadian entrainment in the brain and peripheral tissues, leading to distinct effects on phase and amplitude of gene oscillation within these locales. It is also possible that entrainment of peripheral clocks by specific hormones may be phase dependent (80); that is, local tissue clocks may only respond to phase-resetting hormones and/or metabolites when these are present within a narrow window of time during the 24 hour light/dark cycle. For instance, glucocorticoid secretion from adrenal glands in response to adrenocorticotropin is dependent on time of day (81). In addition to metabolite and hormonal input into the clock, recent studies by Buhr et al. suggest that temperature and heat shock signaling pathways play a central role in entrainment of peripheral tissues (39). Importantly, since alteration in temperature and/or the heat shock pathway is closely associated with other synchronizing signals including feeding and glucocorticoid secretion, it may serve as a unifying signal in entraining peripheral clocks. A major objective in future research will thus be to delineate the role of nutrient signaling in entrainment of CNS and peripheral clocks, and to determine how these signals interact with synchronizing signals such as feeding and neuroendocrine hormones (e.g., glucocorticoids) to maintain phase alignment of behavioral and metabolic rhythms within the whole organism at the levels of both physiology and pathophysiology.