Brown adipose tissue (BAT) is a highly thermogenic organ that converts lipids and glucose into heat. Many of the metabolic and gene transcriptional hallmarks of BAT activation, namely increased lipolysis, uncoupling protein-1 (UCP1) mRNA, and glucose uptake, are regulated by the adrenergic second messenger, cAMP. Cyclic nucleotide phosphodiesterases (PDEs) catalyze the breakdown of cAMP, thereby regulating the magnitude and duration of this signaling molecule. In the absence of adrenergic stimulus, we found that it required a combination of a PDE3 and a PDE4 inhibitor to fully induce UCP1 mRNA and lipolysis in brown adipocytes, whereas neither PDE inhibitor alone had any substantial effect under basal conditions. Under submaximal b-adrenoceptor stimulation of brown adipocytes, a PDE3 inhibitor alone could potentiate induction of UCP1 mRNA, whereas a PDE4 inhibitor alone could augment lipolysis, indicating differential roles for each of these two PDEs. Neither induction of UCP1 nor lipolysis was altered by inhibition of PDE1, PDE2, or PDE8A. Finally, when injected into mice, the combination of PDE3 and PDE4 inhibitors stimulated glucose uptake in BAT under thermoneutral and fasted conditions, a response that was further potentiated by the global ablation of PDE8A. Taken together, these data reveal that multiple PDEs work in concert to regulate three of the important pathways leading to BAT activation, a finding that may provide an improved conceptual basis for the development of therapies for obesity-related diseases.

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Cardiovascular disease is the leading cause of death in the United States. It is known that heart failure is marked by a decrease in expression of key proteins that regulate cardiac contractility, including the β1‐adrenergic receptor (β1‐AR). Despite a compensatory increase in sympathetic tone, cardiac contractility and inotropic response are substantially impaired, primarily through β1‐AR desensitization and mRNA degradation. Here we aim to elucidate the mechanisms by which the β1‐AR is regulated via mRNA processing at the regulatory 3’‐end (alternative polyadenylation), and identify the role that RNA binding proteins, such as HuR and AUF‐1, play in β1‐AR mRNA stability. We demonstrate a decrease in β1‐AR mRNA in cardiac pathology models of pressure overload induced heart failure (transverse aortic constriction; TAC) and chronic (2 week) β AR stimulation. In addition, we show the effect that cardiac‐specific deletion of HuR has on heart function and β1‐AR mRNA regulation. We also investigate the mechanisms of agonist‐dependent β1‐AR mRNA degradation in murine embryonic fibroblasts treated with β1‐AR agonists and antagonists with overexpression and/or siRNA‐mediated silencing of HuR and AUF1. These findings provide important insight into the regulation of the β1‐AR in heart failure and may suggest new therapeutic targets for the restoration of healthy adrenergic function.

Recent studies indicate that adult humans possess brown adipose tissue (BAT), though the impact of BAT on human energy expenditure is still unclear. The activation of key BAT processes by adrenergic stimulation, namely lipolysis and Ucp1 expression, are mediated through cAMP‐dependent signaling. Phosphodiesterases (PDEs) catalyze the breakdown of cyclic nucleotides, and therein control subsequent downstream effects. This makes PDEs attractive pharmacological targets to activate BAT in obese humans in order to potentially reduce fat stores. We have identified six different PDE subtypes in mouse interscapular BAT that can hydrolyze cAMP. We found that a combination of PDE3 and PDE4 inhibitors can highly synergistically stimulate basal cAMP, CREB‐phosphorylation and Ucp1 mRNA expression, whereas individual inhibitors were ineffective when used alone. However, a PDE3 inhibitor alone could shift the adrenergic agonist dose‐response curves for cAMP and Ucp1 mRNA to the left. Additionally, we found the same synergistic effect of PDE3 and PDE4 inhibitors on lipolysis in primary brown adipocytes, but in this case also identify PDE4 as a major regulator in the basal state. These results identify a functional synergy between PDE3 and PDE4 in the control of BAT function, and suggest that one must inhibit both in order to fully activate the tissue in the basal adrenergic state. Supported by NIH grant GM08392.

Brown adipose tissue (BAT) is the primary center of metabolic heat production in mammals during cold‐induced, adaptive thermogenesis. While white fat is an energy "storing" tissue, BAT is known as an energy "wasting" tissue, where the energy from triglyceride breakdown is lost as heat. Intracellular increases in cAMP are known to regulate a number of key processes in BAT, such as acute lipolysis, precursor proliferation and differentiation, and the induction of thermogenic genes including Ucp1. Phosphodiesterases (PDE) are enzymes that catalyze the breakdown of cyclic nucleotides, thereby modulating the second messenger's downstream effects. Here, we present evidence that Phosphodiesterase 8A (PDE8A) is expressed highly in the interscapular BAT of mice, though not expressed in white fat. Utilizing a PDE8A‐/‐ mouse, which has a LacZ‐Neo cassette inserted into the endogenous PDE8A locus, we observe strong β‐Gal activity in BAT. We confirmed PDE8A expression in wild‐type BAT using mRNA, protein, and activity analyses. Preliminary evidence shows that PDE8A ablation modulates lipolysis in response to isoproterenol in isolated adipocytes, suggesting that PDE8A plays a role in the lypolitic component of the thermogenic response in brown fat. Supported by NIH grants DK21723 and GM08392.