A Transgenic Mouse Model of Heart Failure Using Inducible Gαq

Gaofeng Fan,Ya-Ping Jiang,Zhongju Lu,Dwight W Martin,Damon J Kelly,Joan M Zuckerman,Lisa M Ballou,Ira S Cohen,Richard Z Lin

doi:10.1074/jbc.m506810200

Abstract

Receptors coupled to Galpha q play a key role in the development of heart failure. Studies using genetically modified mice suggest that Galpha q mediates a hypertrophic response in cardiac myocytes. Galpha q signaling in these models is modified during early growth and development, whereas most heart failure in humans occurs after cardiac damage sustained during adulthood. To determine the phenotype of animals that express increased Galpha q signaling only as adults, we generated transgenic mice that express a silent Galpha q protein (Galpha qQ209L-hbER) in cardiac myocytes that can be activated by tamoxifen. Following drug treatment to activate Galpha q Q209L-hbER, these mice rapidly develop a dilated cardiomyopathy and heart failure. This phenotype does not appear to involve myocyte hypertrophy but is associated with dephosphorylation of phospholamban (PLB), decreased sarcoplasmic reticulum Ca2+-ATPase activity, and a decrease in L-type Ca2+ current density. Changes in Ca2+ handling and decreased cardiac contractility are apparent 1 week after Galpha qQ209L-hbER activation. In contrast, transgenic mice that express an inducible Galpha q mutant that cannot activate phospholipase Cbeta (PLCbeta) do not develop heart failure or changes in PLB phosphorylation, but do show decreased L-type Ca2+ current density. These results demonstrate that activation of Galpha q in cardiac myocytes of adult mice causes a dilated cardiomyopathy that requires the activation of PLCbeta. However, increased PLCbeta signaling is not required for all of the Galpha q-induced cardiac abnormalities.

Highlights

Receptors coupled to G␣q play a key role in the development of heart failure
Transgenic mice that express an inducible G␣q mutant that cannot activate phospholipase C␤ (PLC␤) do not develop heart failure or changes in PLB phosphorylation, but do show decreased L-type Ca2؉ current density. These results demonstrate that activation of G␣q in cardiac myocytes of adult mice causes a dilated cardiomyopathy that requires the activation of PLC␤
Active G␣qQ209L-hormonebinding domain of the murine estrogen receptor (hbER) Constructs—The G␣q proteins used in this study are G␣qQ209L, which signals constitutively to all of its downstream effectors, and a G␣qQ209L mutant in which Arg256 and Thr257 are changed to Ala

Summary

EXPERIMENTAL PROCEDURES

Materials—Antibodies against the carboxyl terminus of the estrogen receptor and G␣q were from Santa Cruz Biotechnology (Santa Cruz, CA). The supernatant was centrifuged at 72,000 ϫ g for 15 min at 4 °C, and the resulting pellet was suspended in 50 mM Tris-HCl, pH 7.4. SERCA Activity—Crude ventricular membranes were prepared by homogenizing the tissue as described above followed by centrifugation at 14,000 ϫ g for 20 min at 4 °C. Ca2ϩ-dependent ATPase activity was calculated as TAA-CIA and is expressed as nmol ATP hydrolyzed per mg of protein per min. The heart was removed and rinsed in three changes of PS solution (112 mM NaCl, 5.4 mM KCl, 1.7 mM NaH2PO4, 1.63 mM MgCl2, 4.2 mM NaHCO3, 20 mM HEPES, 5.4 mM glucose, 4.1 mM L-glutamine, 10 mM taurine, minimal essential medium vitamins, and minimal essential medium amino acids, pH 7.4) containing 20 units/ml sodium heparin. Experimental groups were compared using Student’s t test

RESULTS

TABLE ONE

TABLE TWO

Ejection fraction

DISCUSSION