Receptor-mediated Gq signaling promotes hypertrophic growth of cultured neonatal rat cardiac myocytes and is postulated to transduce in vivo cardiac pressure overload hypertrophy. Although initially compensatory, hypertrophy can proceed by unknown mechanisms to cardiac failure. We used adenoviral infection and transgenic overexpression of the alpha subunit of Gq to autonomously activate Gq signaling in cardiomyocytes. In cultured cardiac myocytes, overexpression of wild-type G␣q resulted in hypertrophic growth. Strikingly, expression of a constitutively activated mutant of G␣q, which further increased Gq signaling, produced initial hypertrophy, which rapidly progressed to apoptotic cardiomyocyte death. This paradigm was recapitulated during pregnancy in G␣q overexpressing mice and in transgenic mice expressing high levels of wild-type G␣q. The consequence of cardiomyocyte apoptosis was a transition from compensated hypertrophy to a rapidly progressive and lethal cardiomyopathy. Progression from hypertrophy to apoptosis in vitro and in vivo was coincident with activation of p38 and Jun kinases. These data suggest a mechanism in which moderate levels of Gq signaling stimulate cardiac hypertrophy whereas high level Gq activation results in cardiomyocyte apoptosis. The identification of a single biochemical stimulus regulating cardiomyocyte growth and death suggests a plausible mechanism for the progression of compensated hypertrophy to decompensated heart failure.

We evaluated the role of the G alpha-q (G␣q) subunit of heterotrimeric G proteins in the insulin signaling pathway leading to GLUT4 translocation. We inhibited endogenous G␣q function by single cell microinjection of anti-G␣q/11 antibody or RGS2 protein (a GAP protein for G␣q), followed by immunostaining to assess GLUT4 translocation in 3T3-L1 adipocytes. G␣q/11 antibody and RGS2 inhibited insulin-induced GLUT4 translocation by 60 or 75%, respectively, indicating that activated G␣q is important for insulin-induced glucose transport. We then assessed the effect of overexpressing wild-type G␣q (WT-G␣q) or a constitutively active G␣q mutant (Q209L-G␣q) by using an adenovirus expression vector. In the basal state, Q209L-G␣q expression stimulated 2-deoxy-D-glucose uptake and GLUT4 translocation to 70% of the maximal insulin effect. This effect of Q209L-G␣q was inhibited by wortmannin, suggesting that it is phosphatidylinositol 3-kinase (PI3-kinase) dependent. We further show that Q209L-G␣q stimulates PI3-kinase activity in p110␣ and p110␥ immunoprecipitates by 3-and 8-fold, respectively, whereas insulin stimulates this activity mostly in p110␣ by 10-fold. Nevertheless, only microinjection of anti-p110␣ (and not p110␥) antibody inhibited both insulin-and Q209L-G␣q-induced GLUT4 translocation, suggesting that the metabolic effects induced by Q209L-G␣q are dependent on the p110␣ subunit of PI3-kinase. In summary, (i) G␣q appears to play a necessary role in insulin-stimulated glucose transport, (ii) G␣q action in the insulin signaling pathway is upstream of and dependent upon PI3-kinase, and (iii) G␣q can transmit signals from the insulin receptor to the p110␣ subunit of PI3-kinase, which leads to GLUT4 translocation.

Abstract-Expression of the wild-type ␣ subunit of Gq stimulates phospholipase C and induces hypertrophy in cardiomyocytes. Addition of Gq-coupled receptor agonists additionally activates phospholipase C, as does expression of a constitutively active mutant form of G␣q. Under these conditions, hypertrophy is rapidly succeeded by apoptotic cellular and molecular changes, including myofilament disorganization, loss of mitochondrial membrane potential, alterations in Bcl-2 family protein levels, DNA fragmentation, increased caspase activity (Ϸ4-fold), cytochrome c redistribution, and nuclear chromatin condensation in Ϸ12% of the cells. We used various interventions to define the molecular relationships between these events and identify potential sites at which these features of apoptosis could be rescued. Treatment with caspase inhibitors prevented DNA fragmentation and promoted myocyte survival; however, cytochrome c release and loss of mitochondrial membrane potential still occurred. In contrast, treatment with bongkrekic acid, an inhibitor of the mitochondrial permeability transition pore, not only prevented DNA fragmentation and reduced nuclear chromatin condensation but also preserved mitochondrial membrane potential and limited cytochrome c redistribution to only Ϸ2% of cells. These data demonstrate the central role of mitochondrial membrane potential in initiation of caspase activation and downstream apoptotic events and suggest that preservation of mitochondrial integrity is crucial for prolonging the life and function of cardiomyocytes exposed to pathological levels of stress. (Circ Res.

The prevalence of postpartum HIV-infected women retained in care and maintaining viral suppression is low. Interventions seeking to engage women in care shortly after delivery have the potential to improve clinical outcomes.

The acute contractile function of the heart is controlled by the e ects of released nonepinephrine (NE) on cardiac adrenergic receptors. NE can also act in a more chronic fashion to induce cardiomyocyte growth, characterized by cell enlargement (hypertrophy), increased protein synthesis, alterations in gene expression and addition of sarcomeres. These responses enhance cardiomyocyte contractile function and thus allow the heart to compensate for increased stress. The hypertrophic e ects of NE are mediated through Gq-coupled a 1 -adrenergic receptors and are mimicked by the actions of other neurohormones (endothelin, prostaglandin F 2a angiotensin II) that also act on Gq-coupled receptors. Activation of phospholipase C by Gq is necessary for these responses, and protein kinase C and MAP kinases have also been implicated. Gq stimulated cardiac hypertrophy is also evident in transgenic mouse models. In contrast, stimulation of G s -coupled b-adrenergic receptors or G i -coupled receptors do not directly e ect cardiomyocyte hypertrophy. Apoptosis is also induced by G-protein-coupled receptor stimulation in cardiomyocytes. Sustained or excessive activation of either Gq-or Gs-signaling pathways results in apoptotic loss of cardiomyocytes both in vitro and in vivo. Apoptosis is associated with decreased ventricular function in the failing heart. Cardiomyocytes provide an ideal model system for understanding the basis for G-protein mediated hypertrophy and apoptosis, and the mechanisms responsible for the transition from compensatory to deleterious levels of signaling. This information may prove critical for designing interventions that prevent the pathophysiological consequences of heart failure. Oncogene (2001) 20, 1626 ± 1634.

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We previously reported that constitutively activated G␣ q (Q209L) expression in cardiomyocytes induces apoptosis through opening of the mitochondrial permeability transition pore. We assessed the hypothesis that disturbances in Ca 2؉ handling linked G␣ q activity to apoptosis because resting Ca 2؉ levels were significantly increased prior to development of apoptosis. Treating cells with EGTA lowered Ca 2؉ and blocked both loss of mitochondrial membrane potential (an indicator of permeability transition pore opening) and apoptosis (assessed by DNA fragmentation). When cytosolic Ca 2؉ and mitochondrial membrane potential were simultaneously measured by confocal microscopy, sarcoplasmic reticulum (SR)-driven slow Ca 2؉ oscillations (time-to-peak ϳ4 s) were observed in Q209L-expressing cells. These oscillations were seen to transition into sustained increases in cytosolic Ca 2؉, directly paralleled by loss of mitochondrial membrane potential. Ca 2؉ transients generated by caffeine-induced release of SR Ca 2؉ were greatly prolonged in Q209L-expressing cells, suggesting a decreased ability to extrude Ca 2؉. Indeed, the Na ؉

The Mas receptor is a class I G-protein-coupled receptor that is expressed in brain, testis, heart, and kidney. The intracellular signaling pathways activated downstream of Mas are still largely unknown. In the present study, we examined the expression pattern and signaling of Mas in the heart and assessed the participation of Mas in cardiac ischemia-reperfusion injury. Mas mRNA and protein were present in all chambers of human hearts, with cardiomyocytes and coronary arteries being sites of enriched expression. Expression of Mas in either HEK293 cells or cardiac myocytes resulted in constitutive coupling to the G(q) protein, which in turn activated phospholipase C and caused inositol phosphate accumulation. To generate chemical tools for use in probing the function of Mas, we performed a library screen and chemistry optimization program to identify potent and selective nonpeptide agonists and inverse agonists. Mas agonists activated G(q) signaling in a dose-dependent manner and reduced coronary blood flow in isolated mouse and rat hearts. Conversely, treatment of isolated rat hearts with Mas inverse agonists improved coronary flow, reduced arrhythmias, and provided cardioprotection from ischemia-reperfusion injury, an effect that was due, at least in part, to decreased cardiomyocyte apoptosis. Participation of Mas in ischemia-reperfusion injury was confirmed in Mas knockout mice, which had reduced infarct size relative to mice with normal Mas expression. These results suggest that activation of Mas during myocardial infarction contributes to ischemia-reperfusion injury and further suggest that inhibition of Mas-G(q) signaling may provide a new therapeutic strategy directed at cardioprotection.