Enhanced calcium cycling and contractile function in transgenic hearts expressing constitutively active Gα_o* protein

Abstract

In contrast to the other heterotrimeric GTP-binding proteins (G proteins) G s and Gi, the functional role of Go is still poorly defined. To investigate the role of G␣o in the heart, we generated transgenic mice with cardiac-specific expression of a constitutively active form of G␣ o1 ‫ء‬ (G␣ o ‫,)ء‬ the predominant G␣ o isoform in the heart. G␣ o expression was increased 3-to 15-fold in mice from 5 independent lines, all of which had a normal life span and no gross cardiac morphological abnormalities. We demonstrate enhanced contractile function in G␣ o‫ء‬ transgenic mice in vivo, along with increased L-type Ca 2ϩ channel current density, calcium transients, and cell shortening in ventricular G␣ o‫-ء‬expressing myocytes compared with wild-type controls. These changes were evident at baseline and maintained after isoproterenol stimulation. Expression levels of all major Ca 2ϩ handling proteins were largely unchanged, except for a modest reduction in Na ϩ /Ca 2ϩ exchanger in transgenic ventricles. In contrast, phosphorylation of the ryanodine receptor and phospholamban at known PKA sites was increased 1.6-and 1.9-fold, respectively, in G␣ o‫ء‬ ventricles. Density and affinity of ␤-adrenoceptors, cAMP levels, and PKA activity were comparable in G␣ o‫ء‬ and wild-type myocytes, but protein phosphatase 1 activity was reduced upon G␣ o‫ء‬ expression, particularly in the vicinity of the ryanodine receptor. We conclude that G␣ o‫ء‬ exerts a positive effect on Ca 2ϩ cycling and contractile function. Alterations in protein phosphatase 1 activity rather than PKA-mediated phosphorylation might be involved in hyperphosphorylation of key Ca 2ϩ handling proteins in hearts with constitutive G␣ o activation. G proteins; signal transduction; calcium; contraction; transgenic mice CARDIAC CONTRACTILE FUNCTION is determined by the intrinsic contractile properties of the heart and is subject to neurohumoral regulation. The main receptors involved in regulating contraction are prototypical G protein-coupled receptors (42). They activate heterotrimeric G proteins that are comprised of ␣-, ␤-, and ␥-subunits (40). G proteins are classified according to their ␣-subunits, because they primarily determine downstream signaling specificity. Several different G proteins are expressed in the heart (53). Members of the G s and G i/o subfamilies play a key role in transmitting extracellular signals that regulate myocyte cell shortening (42): 1) activation of G s protein-coupled receptors (such as ␤-adrenergic receptors) leads to increased adenylyate cyclase activity and cAMP and PKA activation. PKA then phosphorylates several Ca 2ϩ handling and contractile proteins, resulting in increased contraction and relaxation. 2) Activation of cardiac receptors that are coupled to members of the pertussis toxin-sensitive G i/o subfamily (such as A 1 adenosine and M 2 muscarinic receptors) negatively regulate contractile function in mammalian ventricles in the presence of elevated cAMP ("accentuated antagonism"). While G␣ i has the capacity to directly inhib...

Myocardial expression, signaling, and function of GPR22: a protective role for an orphan G protein-coupled receptor

“…Cardiomyocyte G i /G o signaling in response to adenosine and opioid receptor activation has also been shown to activate prosurvival pathways in the setting of ischemia-reperfusion injury, implicating a role for G i /G o -coupled receptors in cardioprotection (26,32). Furthermore, we cannot rule out a operative role for G o signaling by GPR22, especially in light of recent findings suggesting modulation of cardiac calcium signaling and contractile function by G o in transgenically altered mice (40).…”

Deletion of Ptpn11 (Shp2) in Cardiomyocytes Causes Dilated Cardiomyopathy via Effects on the Extracellular Signal–Regulated Kinase/Mitogen-Activated Protein Kinase and RhoA Signaling Pathways

“…30 Briefly, adult cardiomyocytes were obtained by heart Langendorff perfusion with Ca Taurine (Sigma) for 3 to 5 minutes. Perfusion was continued for 7 to 10 minutes with recirculating Tyrode solution containing collagenase D (0.3 mg/g body weight; Roche, Indianapolis, Ind), collagenase B (0.4 mg/g body weight; Roche), and proteinase XIV (0.05 mg/g body weight; Sigma).…”

“…30 Briefly, adult cardiomyocytes were obtained by heart Langendorff perfusion with Ca 2ϩ -free Tyrode buffer (135 mmol/L NaC1, 4 mmol/L KC1, 1 mmol/L MgC12, 0.33 mmol/L NaH2PO4, 10 mmol/L HEPES) and 10 mmol/L glucose (pH 7.4), 10 mmol/L 2,3-butanedione monoxime (Sigma, St Louis, Mo), and 5 mmol/L Taurine (Sigma) for 3 to 5 minutes. Perfusion was continued for 7 to 10 minutes with recirculating Tyrode solution containing collagenase D (0.3 mg/g body weight; Roche, Indianapolis, Ind), collagenase B (0.4 mg/g body weight; Roche), and proteinase XIV (0.05 mg/g body weight; Sigma).…”

Is overall blockade superior to selective blockade of adrenergic receptor subtypes in suppressing left ventricular remodeling in spontaneously hypertensive rats?

“…Studies have found that although aortic AR-a 1A , AR-a 1B and AR-a 1D are associated with Gqa, AR-a 1B is also linked to Goa and the couple AR-a 1B -Goa mediates the contractile response of rat aorta. 41,42 Although AR-b binding to Gs and Gi-coupled receptors stimulate adenylyl cyclase and induce accumulation of cyclic adenosine monophosphate and activation of PKA, the a-subunit of Gs (Gsa) and Gi (Gia) has the main role. In the vasculature, these pathways have opposite effects: the Gq cascade results in the constriction of blood vessels, whereas the Gs and Gi signals mediate vasodilation.…”