HYPERTROPHIC EFFECTS OF CALCITONIN-GENE-RELATED PEPTIDE (CGRP) AND AMYLIN ON ADULT MAMMALIAN VENTRICULAR CARDIOMYOCYTES

Calcitonin gene-related peptide (CGRP), a neuropeptide localized in th e cardiac autonomic nervous supply, shares 46% similarity in sequence of amino acids with amylin, a peptide synthesized in pancreatic beta-c ells. In the present study, the question was addressed whether these p eptides could exert hypertrophic effects in cardiomyocytes isolated fr om the ventricles of adult rats and maintained in short-term, serum-fr ee primary culture. FCS (10% v/v), employed as a positive control, inc reased the incorporation of l-[C-14] phenylalanine into cellular prote in, total content of cellular RNA and total mass of cellular protein s ignificantly. CGRP and amylin also increased each of these parameters significantly and in a concentration-dependent manner; maximum respons es occurred at 100 pM and 10 nM for CGRP and amylin, respectively. The selective antagonist at CGRP(1)-receptors, CGRP(8-37) (100 nM), inhib ited significantly the incorporation of l-[C-14] phenylalanine into ce llular protein in response to CGRP and amylin. The selective inhibitor of protein kinase C (PKC), bisindolylmalemide (BIM) (5 mu M), reduced significantly the incorporation of l-[C-14] phenylalanine into cellul ar protein in response to phenylephrine (1 mu M), employed as a positi ve control,but did not inhibit the response to insulin (1 unit/ml), em ployed as a negative control. BIM (5 mu M) reduced significantly the r esponses to FCS (10% v/v), amylin (10 nM) and CGRP (10 pM), but did no t inhibit the response to CGRP (100 pM). The activity of protein kinas e C in membranes prepared from intact cardiomyocytes pre-treated for 1 0 min with the phorbol ester, phorbol 12-myristate 13-acetate (PMA) (1 00 nM), employed as a positive control, and CGRP (10 pM) was significa ntly greater than in membranes prepared from cardiomyocytes not subjec ted to agonist stimulation. Phenylephrine (1 mu M) increased significa ntly the specific activity of creatine kinase but not of lactate dehyd rogenase in day 1 cultures of freshly isolated cardiomyocytes. Signifi cant induction of creatine kinase, but not lactate dehydrogenase, was also stimulated by CGRP and amylin; the maximum responses occurred at 100 pM and 100 nM CGRP and amylin, respectively. In conclusion, CGRP a nd amylin exert hypertrophic effects directly on ventricular cardiomyo cytes from the hearts of adult rats in vitro. These effects are: (1) d ue to de novo protein synthesis since total content of cellular RNA an d incorporation of l-[C-14] phenylalanine into cellular protein were a lso increased; (2) mediated by a common population of CGRP(1)-preferri ng receptors at which;amylin binds with lower potency; (3) mediated, a t least partly, by the activation of PKC; (4) may be associated with a fetal shift in gene expression, characterized by selective induction of creatine kinase. (C) 1995 Academic Press Limited