Golgi‐resident β1‐adrenergic Receptor Signaling to Cardiac Hypertrophy in Cardiac Myocytes in Vitro and in Failing Hearts in Vivo

Pathological cardiac hypertrophy contributes to a higher risk of developing cardiac dysfunction, such as heart failure, one of the major causes of morbidity or mortality in the western world. Chronic stimulation of Gs‐coupled β 1 ‐adrenergic receptor (β 1 ‐AR) signaling by catecholamines during long‐term pathological stress induces cardiac hypertrophy and ultimately heart failure. However, the mechanism of action remains incompletely understood. GPCRs at different intracellular compartments are rapidly emerging as novel mediators of distinct local signaling outputs. We have demonstrated activation of Golgi‐β 1 ‐ARs generates a local pool of cAMP which, in turn, stimulates Golgi‐localized phosphatidylinositol 4‐phosphate (PI4P) hydrolysis via EPAC dependent PLCε activation, leading to cardiac hypertrophy in a cellular model. This novel prohypertrophic, phosphoinositide hydrolysis pathway is not accessed by plasma membrane β 1 ‐AR stimulation. Remarkably, specific inhibition of activation of these preexisting Golgi‐resident β 1 ‐ARs prevents hypertrophy induced by a physiological neurotransmitter norepinephrine (NE) in cardiac myocytes in vitro. These data defined significant and unique roles of Golgi‐β 1 ‐ARs in regulating downstream hypertrophic signaling in a biologically relevant cellular model. To determine if Golgi‐β 1 ‐ARs play an important role in a whole animal model of pathological cardiac hypertrophy, we developed a Golgi targeted construct, eNos‐mApple‐Nb80 to deliver the β 1 ‐AR inhibitor, Nb80, specifically to the Golgi apparatus without interrupting surface β 1 ‐ARs. To deliver our target genes into animal heart tissue, I adopted recombinant adeno‐associated virus 9 (rAAV9) gene delivery system. We confirmed selective localization to the Golgi apparatus of cardiac myocytes and equal expression of eNos‐mApple‐Nb80 and eNos‐mApple (negative control without Nb80) in mouse hearts. Additionally, activation of the prohypertrophic Golgi PI4P hydrolysis pathway by dobutamine was inhibited in cardiac myocytes isolated from animals expressing eNos‐mApple‐Nb80. To evaluate the effect of inhibition of Golgi‐localized β 1 ‐ARs in a mouse model of pressure overload‐induced cardiac hypertrophy, we will perform transaortic constriction (TAC) on eNos‐mApple‐Nb80 expressing mice as well as eNos‐mApple and GFP controls. After 6‐8 weeks animals will be assessed by echocardiography to measure ejection fraction, followed by sacrifice of the mice and evaluation of heart size, heart weight to tibia length, myocyte area, and interstitial fibrosis. Heart tissues will also be analyzed by western blotting and mRNA analysis for activation of key signal transduction pathways and expression of hypertrophic genes. This will help us elucidate the role of Golgi β 1 ‐ARs in cardiac hypertrophy, informing new strategies for the development of β‐blocker therapies for heart failure.