The discovery that mammalian cells have the ability to synthesize the free radical nitric oxide (NO) has stimulated an extraordinary impetus for scientific research in all the fields of biology and medicine. Since its early description as an endothelial-derived relaxing factor, NO has emerged as a fundamental signaling device regulating virtually every critical cellular function, as well as a potent mediator of cellular damage in a wide range of conditions. Recent evidence indicates that most of the cytotoxicity attributed to NO is rather due to peroxynitrite, produced from the diffusion-controlled reaction between NO and another free radical, the superoxide anion. Peroxynitrite interacts with lipids, DNA, and proteins via direct oxidative reactions or via indirect, radical-mediated mechanisms. These reactions trigger cellular responses ranging from subtle modulations of cell signaling to overwhelming oxidative injury, committing cells to necrosis or apoptosis. In vivo, peroxynitrite generation represents a crucial pathogenic mechanism in conditions such as stroke, myocardial infarction, chronic heart failure, diabetes, circulatory shock, chronic inflammatory diseases, cancer, and neurodegenerative disorders. Hence, novel pharmacological strategies aimed at removing peroxynitrite might represent powerful therapeutic tools in the future. Evidence supporting these novel roles of NO and peroxynitrite is presented in detail in this review.

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The enzymatic activities of CD39 and CD73 play strategic roles in calibrating the duration, magnitude, and chemical nature of purinergic signals delivered to immune cells through the conversion of ADP/ATP to AMP and AMP to adenosine, respectively. This drives a shift from an ATP-driven proinflammatory environment to an anti-inflammatory milieu induced by adenosine. The CD39/CD73 pathway changes dynamically with the pathophysiological context in which it is embedded. It is becoming increasingly appreciated that altering this catabolic machinery can change the course or dictate the outcome of several pathophysiological events, such as AIDS, autoimmune diseases, infections, atherosclerosis, ischemia-reperfusion injury, and cancer, suggesting these ecto-enzymes are novel therapeutic targets for managing a variety of disorders.

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Endogenous cannabinoids acting at CB 1 receptors stimulate appetite, and CB 1 antagonists show promise in the treatment of obesity. CB 1 -/-mice are resistant to diet-induced obesity even though their caloric intake is similar to that of wild-type mice, suggesting that endocannabinoids also regulate fat metabolism. Here, we investigated the possible role of endocannabinoids in the regulation of hepatic lipogenesis. Activation of CB 1 in mice increases the hepatic gene expression of the lipogenic transcription factor SREBP-1c and its targets acetyl-CoA carboxylase-1 and fatty acid synthase (FAS). Treatment with a CB 1 agonist also increases de novo fatty acid synthesis in the liver or in isolated hepatocytes, which express CB 1 . High-fat diet increases hepatic levels of the endocannabinoid anandamide (arachidonoyl ethanolamide), CB 1 density, and basal rates of fatty acid synthesis, and the latter is reduced by CB 1 blockade. In the hypothalamus, where FAS inhibitors elicit anorexia, SREBP-1c and FAS expression are similarly affected by CB 1 ligands. We conclude that anandamide acting at hepatic CB 1 contributes to diet-induced obesity and that the FAS pathway may be a common molecular target for central appetitive and peripheral metabolic regulation. IntroductionMaintenance of energy homeostasis and body weight involves the coordinated regulation of appetitive behavior and peripheral energy metabolism (1), as illustrated by the ability of the appetitereducing hormone leptin to regulate fat metabolism in the liver (2). Endocannabinoids are novel lipid mediators that modulate appetitive behavior through the activation of CB 1 (3-11). Sites in the hypothalamus (4, 10, 12), limbic forebrain (11-13), and peripheral sensory nerve terminals (7) have been implicated in mediating the orexigenic effect of endocannabinoids, which is potentiated by hunger or in hyperphagia associated with obesity (4-6, 9) and antagonized by CB 1 blockade. Indeed, CB 1 antagonists show promise in the treatment of obesity (14). A number of recent observations suggest that reduction of food intake alone cannot fully account for the antiobesity effects of CB 1 antagonists. In a mouse model of diet-induced obesity, chronic treatment with the CB 1 antagonist SR141716 caused a transient reduction in food intake and a more prolonged reduction in body weight (15). Mice lacking CB 1 are resistant to diet-induced obesity even though their total caloric intake is similar to that of wild-type littermates, which become obese on the same diet (16). CB 1 -/-mice display a moderately lean phenotype throughout adulthood but only a temporary hypophagia in the first few weeks of life (17). These observations suggest that endocannabinoids and CB 1 also regulate peripheral energy metabolism. Indeed, adipocytes have been found to express

Soluble guanylate cyclase (sGC) is a key signal-transduction enzyme activated by nitric oxide (NO). Impaired bioavailability and/or responsiveness to endogenous NO has been implicated in the pathogenesis of cardiovascular and other diseases. Current therapies that involve the use of organic nitrates and other NO donors have limitations, including non-specific interactions of NO with various biomolecules, lack of response and the development of tolerance following prolonged administration. Compounds that activate sGC in an NO-independent manner might therefore provide considerable therapeutic advantages. Here we review the discovery, biochemistry, pharmacology and clinical potential of haem-dependent sGC stimulators (including YC-1, BAY 41-2272, BAY 41-8543, CFM-1571 and A-350619) and haem-independent sGC activators (including BAY 58-2667 and HMR-1766 NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptNitric oxide (NO) is a key signalling molecule that is involved in the regulation of a variety of biological and physiological processes in mammals. Vast experimental and clinical evidence indicates that reduced bioavailability and/or responsiveness to endogenously produced NO contributes to the development of cardiovascular, pulmonary, endothelial, renal and hepatic diseases, as well as erectile dysfunction. Some of these conditions are currently treated with organic nitrates (such as glyceryl trinitrate) and other NO-donor or 'nitrovasodilator' drugs that release NO by spontaneous decomposition 1 or bioconversion 2 to activate soluble guanylate cyclase (sGC). However, the use of such compounds is limited by a potential lack of response due to insufficient biometabolism 3 , development of tolerance following prolonged administration 4 and nonspecific interactions of NO with other biological molecules, including peroxynitrite-mediated tyrosine nitration 5 . The latter reactions are difficult to control owing to the spontaneous release of NO from nitrovasodilators and its free diffusion in biological systems. Moreover, despite the symptomatic improvement in patients with cardiovascular disease treated with organic nitrates, there is no clear evidence that such treatment reduces mortality. Therefore compounds that activate sGC in an NO-independent manner might offer considerable advantages over current therapies (FIG. 1). In this article, we describe the NOsGC-cGMP signalling pathway and explain the therapeutic rationale for sGC stimulators and activators. The discovery, biochemistry and pharmacology of several such compounds are reviewed, and their potential for the treatment of a range of diseases, including arterial and pulmonary hypertension, heart failure, atherosclerosis, thrombosis, erectile dysfunction, renal fibrosis and failure, and liver cirrhosis, is discussed. The NO-sGC-cGMP signal transduction pathwayGuanine nucleotidyl (guanylyl; guanylate) cyclases (GCs) are widely distributed signaltransduction enzymes that, in response to various cellular stimuli, convert GTP into t...

Adenosine is a key endogenous molecule that regulates tissue function by activating four G-protein-coupled adenosine receptors: A1, A2A, A2B and A3. Cells of the immune system express these receptors and are responsive to the modulatory effects of adenosine in an inflammatory environment. Animal models of asthma, ischaemia, arthritis, sepsis, inflammatory bowel disease and wound healing have helped to elucidate the regulatory roles of the various adenosine receptors in dictating the development and progression of disease. This recent heightened awareness of the role of adenosine in the control of immune and inflammatory systems has generated excitement regarding the potential use of adenosine-receptor-based therapies in the treatment of infection, autoimmunity, ischaemia and degenerative diseases.

Ventricular pressure-volume relationships have become well established as the most rigorous and comprehensive ways to assess intact heart function. Thanks to advances in miniature sensor technology, this approach has been successfully translated to small rodents, allowing for detailed characterization of cardiovascular function in genetically engineered mice, testing effects of pharmacotherapies and studying disease conditions. This method is unique for providing measures of left ventricular (LV) performance that are more specific to the heart and less affected by vascular loading conditions. Here we present descriptions and movies for procedures employing this method (anesthesia, intubation and surgical techniques, calibrations). We also provide examples of hemodynamics measurements obtained from normal mice/rats, and from animals with cardiac hypertrophy/heart failure, and describe values for various useful load-dependent and loadindependent indexes of LV function obtained using different types of anesthesia. The completion of the protocol takes 1-4 h (depending on the experimental design/end points).