Recent studies by Ramsey et al.
] and Nakahata et al
] reveal a direct link between metabolism and the core transcription-translation feedback loop of the mammalian circadian clock in mice. At the center of this link are nicotinamide phosphoribosyltransferase (Nampt), the rate limiting enzyme in the major mammalian NAD+
salvage pathway [26
], and silent mating type information regulation 2 - homolog 1 (SIRT1), an NAD+
-activated deacetylase [28
]. These studies demonstrate that cellular nutrient status reflected by intracellular [NAD+
] has a strong influence on transcription of genes at the core of circadian rhythms and metabolism (reviewed in [24
]). These discoveries suggest a new and exciting avenue of investigation that is likely to reveal mechanisms that integrate nutrient status, biological timing, and metabolic substrate use during torpor. In fact, the concentration of NAD in liver of thirteen-lined ground squirrels (Spermophilus tridecemlineatus
) entering torpor is on average two to three times higher than that found in summer active ground squirrels, or ground squirrels that are about to exit torpor and return to activity [32
], suggesting that Nampt and SIRT1 are likely to be activated during torpor induction.
A hypothetical role for Nampt, SIRT1 and the core circadian clock genes in the induction of torpor can be easily envisioned (). First, as a result of reduced food supply due to a changing environment, an animal enters a period of fasting. During a fast, cellular [NAD+
] increases due to nutrient depletion [16
], causing SIRT1 deacetylase activity to increase. SIRT1 activation of Nampt activity upregulates the NAD+
salvage pathway, maintaining elevated [NAD+
] and reinforcing SIRT1 activity. Activated SIRT1 also deacetylates the circadian clock protein BMAL1, inactivating the heterodimeric transcription activation complex that BMAL1 forms with CLOCK. Inactivation of the BMAL1:CLOCK complex inhibits expression of the circadian clock genes Per
], disrupting the normal circadian feedback loop and causing the clock to either slow or stand still. This would prolong a transcription profile typical of an animal’s inactive phase (e.g. during sleep) for genes regulated by the circadian clock.
Figure 3 A model for torpor induction in mammals that links reduced availability of food in the environment with the energy status of the cell, the mammalian circadian clock, and response to oxidative stress. An animal begins to fast as a result of reduced availability (more ...)
The molecular details of gene regulation by the mammalian circadian clock have been reviewed recently [24
]. In mice, genes encoding key, rate-limiting enzymes of glycolysis, lipolysis, and cholesterol metabolism are regulated by the circadian clock in liver [19
], and there is evidence that lipid metabolism in adipocytes is regulated by BMAL1 [35
]. Additionally, SIRT1 activation of lipolytic pathways would shift metabolism away from carbohydrates and toward catabolism of lipid [36
]. The combined effects of prolonged fasting, SIRT1 activity, inactive/night-time mode circadian clock, and a switch to lipolysis would ultimately poise the animal for entry into torpor ().
The nuclear receptors REV-ERBα, a negative regulator of BMAL1:CLOCK, and RORα, a positive regulator of BMAL1:CLOCK, might also play a role in regulating mammalian torpor [38
]. Supporting the model in , the expression of REV-ERBα in the heart of Siberian hamsters, an animal that exhibits daily torpor, was higher in torpid than in normothermic animals [42
]. Carey and coworkers also observed that the liver concentration of cholesterol sulfate, a ligand of RORα [43
], was low during torpor and rose slightly during interbout arousals (IBAs; brief, periodic arousals from hibernation torpor) and in summer active animals [44
]. Activation of BMAL1:CLOCK by RORα might be necessary for IBAs to occur. These results suggest that REV-ERBα and RORα function to integrate cellular nutrient status, the circadian clock and metabolism (reviewed in [41
]). Further investigation of these nuclear receptors in active and torpid animals will possibly reveal more about the regulation and maintenance of torpor.
In addition to SIRT1 activation in response to fasting (reviewed in [29
]), SIRT1 activity might also blunt oxidative stress encountered during torpor or arousal from torpor by its direct activation of hypoxia-inducible factor-2 alpha (HIF-2α) and the resulting expression of HIF-2α target genes [46
] (). It is not currently understood how animals enter and exit from torpor with no apparent damage due to reduced oxygen utilization during torpor, or due to rapid reperfusion of tissues during arousal. A recent study of Arctic ground squirrels found no evidence of oxidative stress in brain cortex or liver tissues of torpid or cold-adapted squirrels [47
]. In the same study, brown adipose tissue showed limited oxidative stress in association with arousal from torpor. It has been hypothesized that antioxidant defense mechanisms are set in place prior to torpor induction [47
]. SIRT1 activation of hypoxia inducible factors might provide a mechanism for preparation of these defenses.