Circadian clocks are intracellular molecular mechanisms that allow the cell to anticipate the time of day. We have previously reported that the intact rat heart expresses the major components of the circadian clock, of which its rhythmic expression in vivo is consistent with the operation of a fully functional clock mechanism. The present study exposes oscillations of circadian clock genes [brain and arylhydrocarbon receptor nuclear translocator-like protein 1 (bmal1), reverse strand of the c-erba␣ gene (rev-erba␣), period 2 (per2), albumin D-element binding protein (dbp)] for isolated adult rat cardiomyocytes in culture. Acute (2 h) and/or chronic (continuous) treatment of cardiomyocytes with FCS (50% and 2.5%, respectively) results in rhythmic expression of circadian clock genes with periodicities of 20 -24 h. In contrast, cardiomyocytes cultured in the absence of serum exhibit dramatically dampened oscillations in bmal1 and dbp only. Zeitgebers (timekeepers) are factors that influence the timing of the circadian clock. Glucose, which has been previously shown to reactivate circadian clock gene oscillations in fibroblasts, has no effect on the expression of circadian clock genes in adult rat cardiomyocytes, either in the absence or presence of serum. Exposure of adult rat cardiomyocytes to the sympathetic neurotransmitter norephinephrine (10 M) for 2 h reinitiates rhythmic expression of circadian clock genes in a serumindependent manner. Oscillations in circadian clock genes were associated with 24-h oscillations in the metabolic genes pyruvate dehydrogenase kinase 4 (pdk4) and uncoupling protein 3 (ucp3). In conclusion, these data suggest that the circadian clock operates within the myocytes of the heart and that this molecular mechanism persists under standard cell culture conditions (i.e., 2.5% serum). Furthermore, our data suggest that norepinephrine, unlike glucose, influences the timing of the circadian clock within the heart and that the circadian clock may be a novel mechanism regulating myocardial metabolism.heart; metabolism; neurohumoral; rat; zeitgebers CIRCADIAN CLOCKS are controlled by a set of genes that generate self-sustained positive and negative transcriptional feedback loops with a free-running period of 24 h (7,11,40). This molecular mechanism is intrinsic to the cell, persisting in cultured cells such as fibroblasts, vascular smooth muscle cells, and various cell lines (3,13,20,23). Circadian clocks confer the selective advantage of anticipation, conditioning the cell to changes in its environment, such that it can respond rapidly to a specific extracellular signal at an appropriate time of the day. For example, our laboratory (30) has recently hypothesized that the circadian clock within cardiac and skeletal muscle allows the myocytes to anticipate diurnal variations in circulating fatty acids through modulation of fatty acid oxidative capacity at a transcriptional level. However, the existence of an intracellular circadian clock mechanism that is intrinsic to either the cardiomyocyte or...

BackgroundExcess caloric intake is strongly associated with the development of increased adiposity, glucose intolerance, insulin resistance, dyslipidemia, and hyperleptinemia (i.e., the cardiometabolic syndrome). Research efforts have focused attention primarily on the quality (i.e., nutritional content) and/or quantity of ingested calories as potential causes for diet-induced pathology. Despite growing acceptance that biological rhythms profoundly influence energy homeostasis, little is known regarding how the timing of nutrient ingestion influences development of common metabolic diseases.ObjectiveTo test the hypothesis that the time of day at which dietary fat is consumed significantly influences multiple cardiometabolic syndrome parameters.ResultsWe report that mice fed either low or high fat diets in a contiguous fashion during the 12 hour awake/active period adjust both food intake and energy expenditure appropriately, such that metabolic parameters are maintained within a normal physiologic range. In contrast, fluctuation in dietary composition during the active period (as occurs in humans) markedly influences whole body metabolic homeostasis. Mice fed a high fat meal at the beginning of the active period retain metabolic flexibility in response to dietary challenges later in the active period (as revealed by indirect calorimetry). Conversely, consumption of high fat meal at the end of the active phase leads to increased weight gain, adiposity, glucose intolerance, hyperinsulinemia, hypertriglyceridemia, and hyperleptinemia (i.e., cardiometabolic syndrome) in mice. The latter perturbations in energy/metabolic homeostasis are independent of daily total or fat-derived calories.ConclusionsThe time-of-day at which carbohydrate versus fat is consumed markedly influences multiple cardiometabolic syndrome parameters.

Cells/organs must respond both rapidly and appropriately to increased fatty acid availability; failure to do so is associated with the development of skeletal muscle and hepatic insulin resistance, pancreatic ␤-cell dysfunction, and myocardial contractile dysfunction. Here we tested the hypothesis that the intrinsic circadian clock within the cardiomyocytes of the heart allows rapid and appropriate adaptation of this organ to fatty acids by investigating the following: 1) whether circadian rhythms in fatty acid responsiveness persist in isolated adult rat cardiomyocytes, and 2) whether manipulation of the circadian clock within the heart, either through light/dark (L/D) cycle or genetic disruptions, impairs responsiveness of the heart to fasting in vivo. We report that both the intramyocellular circadian clock and diurnal variations in fatty acid responsiveness observed in the intact rat heart in vivo persist in adult rat cardiomyocytes. Reversal of the 12-h/12-h L/D cycle was associated with a re-entrainment of the circadian clock within the rat heart, which required 5-8 days for completion. Fasting rats resulted in the induction of fatty acid-responsive genes, an effect that was dramatically attenuated 2 days after L/D cycle reversal. Similarly, a targeted disruption of the circadian clock within the heart, through overexpression of a dominant negative CLOCK mutant, severely attenuated induction of myocardial fatty acid-responsive genes during fasting. These studies expose a causal relationship between the circadian clock within the cardiomyocyte with responsiveness of the heart to fatty acids and myocardial triglyceride metabolism.