The alpha 1A-adrenergic receptor subtype mediates biochemical, molecular, and morphologic features of cultured myocardial cell hypertrophy

K.U Knowlton,M.C Michel,M Itani,H.E Shubeita,K Ishihara,J.H Brown,K.R Chien

doi:10.1016/s0021-9258(18)82267-0

K.U Knowlton, M.C Michel + Show 5 more

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https://doi.org/10.1016/s0021-9258(18)82267-0

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Abstract

alpha 1-Adrenergic agonists induce a hypertrophic phenotype in cultured neonatal rat ventricular myocytes. Quantifiable markers of this phenotype include stimulation of phosphoinositide hydrolysis, transcriptional induction of atrial natriuretic factor (ANF) gene expression, and an increase in myocardial cell size. The aim of the present work was to determine which alpha 1-adrenergic receptor subtype mediates the acquisition of these parameters of myocardial cell hypertrophy. Phosphoinositide hydrolysis is inhibited by low concentrations of 5-methylurapidil (log Ki = -8.7) and (+)-niguldipine (log Ki = -10.6). The alpha-adrenergic receptor-induced increase in transcriptional activation of an ANF luciferase reporter gene is inhibited over the same range of concentrations of 5-methylurapidil (log Ki = -8.2) and (+)-niguldipine (log Ki = -11.2) that inhibit phosphoinositide hydrolysis. In addition, the increase in cell size that accompanies alpha-adrenergic receptor stimulation of cultured ventricular myocytes is blocked by similar concentrations of 5-methylurapidil (log Ki = -8.0) and (+)-niguldipine (log Ki = -10.6). In contrast, treatment with the alpha 1B selective antagonist chlorethylclonidine at a concentration of 10 microM had no effect on the adrenergically mediated induction of ANF luciferase reporter gene expression or the adrenergically induced increase in myocardial cell size. These findings demonstrate that pharmacologically identifiable alpha 1A-adrenergic receptors mediate not only the early effects of alpha 1-adrenergic stimulation such as phosphoinositide hydrolysis, but that they activate the signaling pathways that control transcriptional induction of the ANF luciferase reporter gene and an increase in myocardial cell size. Studies using alpha 1-adrenergic receptor cDNAs to delineate and alter the direct interaction of this receptor subtype with proximal signaling molecules, e.g. GTP binding proteins, should provide a powerful means of assessing their role in the induction of the molecular and morphologic parameters of myocardial cell hypertrophy.

Highlights

Diego, School of Medicine, La Jolla, California92093-0613and the )IDepartment of Medicine, University of Essen, Essen, Germany al-Adrenergic agonists induce a hypertrophic phe- should provide a powerful meansof assessing their role notype in cultured neonatal rat ventricular myocytes. in the inductionof the molecular and morphologic pa
Theseinclude induction antagonist chlorethylclonidineat a concentration of 10 of a distinct immediate early gene program, appearance of PM had no effect on the adrenergically mediated inducm-arkers of an embryonic program of gene expression, accution of ANF luciferase reporter gene expressionr the mulation andassembly of constitutively expressed contractile adrenergically induced increaseinmyocardial cell proteins into sarcomeric units, and an increase in cell size size. These findings demonstrate that pharmacologi- without concomitant proliferation[7, 8].Activation of a subcally identifiable alA-adrenergic receptors mediate nostet of constitutively expressed contractile proteins and markonly the early effecotsf al-adrenergic stimulation suchers of an embryonic programof gene expression is largely due as phosphoinositide hydrolysis, but that they activate the signaling pathways that control transcriptional duction of the ANF luciferase reporter gene and an increase in myocardial cell size
Phosphoinositide Hydrolysis-To determine the consequences of activation of specific a-adrenergic receptor subtypes, we examined inhibition of phenylephrine-stimulated

Summary

Introduction

Affinity of adrenergic drugs for neonatal rat heart al-adrenoceptors Data are the means f S.E. of values from three experiments in Radioligand Binding Studies-To determine the relative which 18-25 hearts were pooled. 5-Methylurapidil and (+)-niguldipine decreased neonatal rat ventricular cells [24,29].Immunocytofluorescent luciferase expression in these transfectedcells in a concentra- analysis with antibodies against a TrpE/MLC-2 fusion protion-dependentmanner with a log Ki of -8.2 and -11.2, tein demonstrated that myocytes maintained in serum-free respectively (Fig. 3).

Results

Conclusion