Filling GAPs in the understanding of cardioprotection induced by GPCR activation: RGS proteins modulate ischaemic injury.

Brief periods of vascular occlusion followed by reperfusion (I/R) have been shown to confer protection against myocardial injury and necrosis induced by subsequent exposure to prolonged I/R, a phenomenon referred to as ischaemic preconditioning. Initially discovered by Murry et al. in canine hearts 25 years ago, subsequent work has shown that preconditioning induces two periods of protection: an initial, early phase that arises with minutes of exposure to the bouts of preconditioning I/R, persists for 1–4 h, and then disappears, followed by the re-emergence of a protected phenotype 24 h later (delayed preconditioning). More recently, the phenomenon of ischaemic post-conditioning has been described, wherein the short bouts of I/R are initiated at the onset of reperfusion to induce cardioprotection. These preconditioning protocols (IPC) activate endogenous cell survival programmes that appear to exist in every species, organ, and tissue tested and may also operate in humans. –4 Moreover, IPC represents the most powerful cardioprotective intervention yet discovered. As a consequence, an intense research effort has ensued in an attempt to elucidate the signalling mechanisms that mediate IPC so that practical therapies can be developed for patients who are predisposed to ischaemic disease. This work has resulted in identification of a number of pharmacological agents that mimic IPC, including adenosine, acetylcholine, opioids, calcitonin gene-related peptide, bradykinin, angiotensin II, and endothelin. 4 Because each of these endogenously produced agonists serves as a ligand for Gai protein-coupled receptors (GPCRs), there is growing interest in developing therapies that potentiate and/or sustain their activity in cardiovascular disease. Since it is difficult to predict when myocardial ischaemia will occur, this therapeutic approach may provide a novel away to target the treatment of the ischaemic heart on a temporal basis that allows for prophylactic management of individuals at risk for myocardial infarction. GPCRs are the largest cell-surface receptor superfamily, with more than 800 of these proteins encoded in the human genome. Of these, more than 100 different GPCRs are expressed in the cardiovascular system. GPCRs respond to a wide variety of stimuli, including hormones, neurotransmitters, peptides, amino acids, nucleotides, lipids and fatty acid derivatives, and calcium ions, as well as light, chemical odorants, and taste molecules. The GPCRs transfer extracellular signals across the plasmalemma to intracellular effectors via heterotrimeric G proteins (a, b, and g). The regulator of G protein signalling (RGS) proteins were discovered almost 15 years ago and are now recognized as important regulators of GPCR activity. They do so by acting as GTPase-activating proteins (GAPs) that enhance GTP hydrolysis, thereby terminating the G protein activation cycle. RGS proteins all share a 120–130 amino acid motif designated as the GAP (or RGS) domain that can increase the rate of Ga-mediated hydrolysis of GTP by 40–2000-fold over basal levels. As a consequence, RGS proteins attenuate G protein signalling by accelerating G protein signal termination kinetics upon removal of the agonist. The GAP domain in RGS proteins can also physically block Ga-binding sites to downstream effectors as another mechanism for inhibiting GPCR signalling. Waterson et al. provide the first evidence that Gai2-mediated cardioprotection is attenuated by RGS proteins. Until this study, the lack of specific inhibitors for specific RGS proteins has made it difficult to address this question, an issue compounded by methodological problems caused by the tandem arrangement of many RGS protein genes on one chromosome, which make it difficult to create knockout mice lacking just one of the RGS proteins. However, Waterson et al. capitalized on the recent development of genetically manipulated mice expressing a mutant Gai2 (G184S) that is RGS insensitive. 8