Previous studies have suggested that insulin resistance develops secondary to diminished fat oxidation and resultant accumulation of cytosolic lipid molecules that impair insulin signaling. Contrary to this model, the present study used targeted metabolomics to find that obesity-related insulin resistance in skeletal muscle is characterized by excessive beta-oxidation, impaired switching to carbohydrate substrate during the fasted-to-fed transition, and coincident depletion of organic acid intermediates of the tricarboxylic acid cycle. In cultured myotubes, lipid-induced insulin resistance was prevented by manipulations that restrict fatty acid uptake into mitochondria. These results were recapitulated in mice lacking malonyl-CoA decarboxylase (MCD), an enzyme that promotes mitochondrial beta-oxidation by relieving malonyl-CoA-mediated inhibition of carnitine palmitoyltransferase 1. Thus, mcd(-/-) mice exhibit reduced rates of fat catabolism and resist diet-induced glucose intolerance despite high intramuscular levels of long-chain acyl-CoAs. These findings reveal a strong connection between skeletal muscle insulin resistance and lipid-induced mitochondrial stress.

The obesity epidemic has led to an increased incidence of non–alcoholic fatty liver disease (NAFLD) and type 2 diabetes. AMP–activated protein kinase (Ampk) regulates energy homeostasis and is activated by cellular stress, hormones and the widely prescribed anti–type 2 diabetic drug metformin1,2. Ampk phosphorylates murine acetyl–CoA carboxylase3,4 (Acc) 1 at Ser79 and Acc2 at Ser212, inhibiting the conversion of acetyl–CoA to malonyl–CoA, a precursor in fatty acid synthesis5 as well as an allosteric inhibitor of fatty acid transport into mitochondria for oxidation6. To test the physiological impact of these phosphorylation events we generated mice with alanine knock–in mutations in both Acc1 (Ser79) and Acc2 (Ser212) (Acc double knock–in, AccDKI). These mice have elevated lipogenesis and lower fatty acid oxidation compared to wild–type (WT) mice, which contribute to the progression of insulin resistance, glucose intolerance and NAFLD, but not obesity. Remarkably, AccDKI mice made obese by high–fat feeding, are refractory to the lipid–lowering and insulin–sensitizing effects of metformin. These findings establish that inhibitory phosphorylation of Acc by Ampk is essential for the control of lipid metabolism, and in the setting of obesity, for metformin–induced improvements in insulin action.

Summary A robust and well-organized rhythm is a key feature of many neuronal networks, including those that regulate essential behaviors such as circadian rhythmogenesis, breathing and locomotion. Here we show that excitatory V3-derived neurons are necessary for a robust and organized locomotor rhythm during walking. When V3-mediated neurotransmission is selectively blocked by the expression of the Tetanus toxin light chain subunit (TeNT), the regularity and robustness of the locomotor rhythm is severely perturbed. A similar degeneration in the locomotor rhythm occurs when the excitability of V3-derived neurons is reduced acutely by ligand-induced activation of the allatostatin receptor. The V3-derived neurons additionally function to balance the locomotor output between both halves of the spinal cord, thereby ensuring a symmetrical pattern of locomotor activity during walking. We propose that the V3 neurons establish a robust and balanced motor rhythm by distributing excitatory drive between both halves of the spinal cord.

Background-Matrix metalloproteinases are best recognized for their ability to degrade the extracellular matrix in both physiological and pathological conditions. However, recent findings indicate that some of them are also involved in mediating acute processes such as platelet aggregation and vascular tone. The acute contractile defect of the heart after ischemia-reperfusion may involve the proteolytic degradation of the thin filament protein troponin I; however, the protease responsible for this remains obscure. Methods and Results-Here we report that matrix metalloproteinase-2 is colocalized with troponin I within the thin myofilaments of cardiomyocytes in ischemic-reperfused hearts and that troponin I is a novel intracellular target for proteolytic cleavage by matrix metalloproteinase-2. Inhibition of matrix metalloproteinase-2 activity prevented ischemia-reperfusion-induced troponin I degradation and improved the recovery of mechanical function of the heart. Conclusions-These data reveal for the first time a novel molecular mechanism by which matrix metalloproteinase-2 causes acute myocardial dysfunction after ischemia-reperfusion injury and that matrix metalloproteinase-2 has a biological action within the cell.

Abstract-The pulmonary arteries (PA) in pulmonary arterial hypertension (PAH) are constricted and remodeled;. They have suppressed apoptosis, partly attributable to suppression of the bone morphogenetic protein axis and selective downregulation of PA smooth muscle cell (PASMC) voltage-gated K ϩ channels, including Kv1.5. The Kv downregulation-induced increase in [K ϩ ] i , tonically inhibits caspases, further suppressing apoptosis. Mitochondria control apoptosis and produce activated oxygen species like H 2 O 2 , which regulate vascular tone by activating K ϩ channels, but their role in PAH is unknown. We show that dichloroacetate (DCA), a metabolic modulator that increases mitochondrial oxidative phosphorylation, prevents and reverses established monocrotaline-induced PAH (MCT-PAH), significantly improving mortality. Compared with MCT-PAH, DCA-treated rats (80 mg/kg per day in drinking water on day 14 after MCT, studied on day 21) have decreased pulmonary, but not systemic, vascular resistance (63% decrease, PϽ0.002), PA medial thickness (28% decrease, PϽ0.0001), and right ventricular hypertrophy (34% decrease, PϽ0.001). DCA is similarly effective when given at day 1 or day 21 after MCT (studied day 28) but has no effect on normal rats. DCA depolarizes MCT-PAH PASMC mitochondria and causes release of H 2 O 2 and cytochrome c, inducing a 10-fold increase in apoptosis within the PA media (TUNEL and caspase 3 activity) and decreasing proliferation (proliferating-cell nuclear antigen and BrdU assays). Immunoblots, immunohistochemistry, laser-captured microdissection-quantitative reverse-transcription polymerase chain reaction and patch-clamping show that DCA reverses the Kv1.5 downregulation in resistance PAs. In summary, DCA reverses PA remodeling by increasing the mitochondria-dependent apoptosis/proliferation ratio and upregulating Kv1.5 in the media. We identify mitochondriadependent apoptosis as a potential target for therapy and DCA as an effective and selective treatment for PAH. Key Words: apoptosis Ⅲ proliferation Ⅲ smooth muscle Ⅲ vascular remodeling P ulmonary arterial hypertension (PAH) is defined by an elevated pulmonary vascular resistance (PVR), which leads to right heart failure and premature death. The cause remains unknown and available treatments are limited, expensive, and often associated with significant side effects. 1,2 The pulmonary arteries (PAs) are affected by varying degrees of vasoconstriction and vascular remodeling, including cellular proliferation in both the intima and media and distal PA muscularization. 1,2 Vascular medial remodeling results from an imbalance between smooth muscle cell (SMC) proliferation and apoptosis, favoring proliferation. Gene microarray studies show that lungs from patients with PAH have a decrease in the proapoptotic/antiapoptotic gene expression ratio. 3 Furthermore, several loss-of-function germline or acquired mutations have been described in receptors of the transforming growth factor-␤) superfamily, such as bone morphogenetic protein receptor-2 (BM...

Virtually every mammalian cell, including cardiomyocytes, possesses an intrinsic circadian clock. The role of this transcriptionally based molecular mechanism in cardiovascular biology is poorly understood. We hypothesized that the circadian clock within the cardiomyocyte influences diurnal variations in myocardial biology. We, therefore, generated a cardiomyocyte-specific circadian clock mutant (CCM) mouse to test this hypothesis. At 12 wk of age, CCM mice exhibit normal myocardial contractile function in vivo, as assessed by echocardiography. Radiotelemetry studies reveal attenuation of heart rate diurnal variations and bradycardia in CCM mice (in the absence of conduction system abnormalities). Reduced heart rate persisted in CCM hearts perfused ex vivo in the working mode, highlighting the intrinsic nature of this phenotype. Wild-type, but not CCM, hearts exhibited a marked diurnal variation in responsiveness to an elevation in workload (80 mmHg plus 1 M epinephrine) ex vivo, with a greater increase in cardiac power and efficiency during the dark (active) phase vs. the light (inactive) phase. Moreover, myocardial oxygen consumption and fatty acid oxidation rates were increased, whereas cardiac efficiency was decreased, in CCM hearts. These observations were associated with no alterations in mitochondrial content or structure and modest mitochondrial dysfunction in CCM hearts. Gene expression microarray analysis identified 548 and 176 genes in atria and ventricles, respectively, whose normal diurnal expression patterns were altered in CCM mice. These studies suggest that the cardiomyocyte circadian clock influences myocardial contractile function, metabolism, and gene expression.

Mitochondrial uncoupling protein 1 (UCP1) is enriched within interscapular brown adipose tissue (iBAT) and beige (also known as brite) adipose tissue 1,2 , but its thermogenic potential is reduced with obesity and type 2 diabetes 3-5 for reasons that are not understood. Serotonin (5-hydroxytryptamine, 5-HT) is a highly conserved biogenic amine that resides in non-neuronal and neuronal tissues that are specifically regulated via tryptophan hydroxylase 1 (Tph1) and Tph2, respectively 6-8 . Recent findings suggest that increased peripheral serotonin 9 and polymorphisms in TPH1 are associated with obesity 10 ; however, whether this is directly related to reduced BAT Reprints and permissions information is available online at

Background-Sildenafil was recently approved for the treatment of pulmonary arterial hypertension. The beneficial effects of phosphodiesterase type 5 (PDE5) inhibitors in pulmonary arterial hypertension are thought to result from relatively selective vasodilatory and antiproliferative effects on the pulmonary vasculature and, on the basis of early data showing lack of significant PDE5 expression in the normal heart, are thought to spare the myocardium. Methods and Results-We studied surgical specimens from 9 patients and show here for the first time that although PDE5is not expressed in the myocardium of the normal human right ventricle (RV), mRNA and protein are markedly upregulated in hypertrophied RV (RVH) myocardium. PDE5 also is upregulated in rat RVH. PDE5 inhibition (with either MY-5445 or sildenafil) significantly increases contractility, measured in the perfused heart (modified Langendorff preparation) and isolated cardiomyocytes, in RVH but not normal RV. PDE5 inhibition leads to increases in both cGMP and cAMP in RVH but not normal RV. Protein kinase G activity is suppressed in RVH, explaining why the PDE5 inhibitor-induced increase in cGMP does not lead to inhibition of contractility. Rather, it leads to inhibition of the cGMP-sensitive PDE3, explaining the increase in cAMP and contractility. This is further supported by our findings that, in RVH protein kinase A, inhibition completely inhibits PDE5-induced inotropy, whereas protein kinase G inhibition does not. Conclusions-The ability of PDE5 inhibitors to increase RV inotropy and to decrease RV afterload without significantly affecting systemic hemodynamics makes them ideal for the treatment of diseases affecting the RV, including pulmonary arterial hypertension. (Circulation. 2007;116:238-248.)