Contractile activity modulates atrial natriuretic factor gene expression in neonatal rat ventricular myocytes.
[Ca2+]i transients, and the activation of Ca(2+)-sensitive kinases have been considered potential signaling mechanisms regulating ANF gene expression in cultured neonatal rat ventricular myocytes (NRVM). However, it is unclear whether [Ca2+]i is directly involved, or is indirectly involved by generating additional mechanical signals via contractile activity. Primary cultures of spontaneously contracting NRVM (CON), and NRVM treated for 48 h with verapamil (V, 10 microM), KCl (50 mM), or 2,3-butanedione monoxime (BDM, 7.5 mM) were used to delineate the affects of contractile activity v [Ca2+]i. Verapamil, a calcium, channel blocker, inhibits contraction and decreases [Ca2+]i. High [K+]o causes membrane depolarization, loss of contraction, and elevates [Ca2+]i; whereas BDM strongly inhibits contractile activity but only modestly reduces [Ca2+]i transients. ANF production, as assessed by radioimmunoassay, was significantly reduced upon contractile arrest independently of [Ca2+]i levels. Northern blotting analysis demonstrated that contractile arrest also reduced ANF mRNA levels. Transient transfection of a 3003 bp ANF promoter-luciferase expression plasmid in CON, V, KCl, and BDM-treated NRVM demonstrated marked down-regulation of ANF promoter activity in all of the contractile arrested myocytes. These results indicate that the activation of Ca(2+)-sensitive processes alone are insufficient to maintain high levels of ANF gene expression and peptide production in NRVM.
Eble
D M
DM
Cardiovascular Institute, Loyola University Chicago Stritch School of Medicine, Maywood, IL 60153, USA.
Cadre
B M
BM
Qi
M
M
Bers
D M
DM
Samarel
A M
AM
D000818:Animals; D000831:Animals, Newborn; D009320:Atrial Natriuretic Factor / Q000032:analysis / Q000235:genetics*; D002118:Calcium / Q000502:physiology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000502:physiology*; D006352:Heart Ventricles; D009200:Myocardial Contraction / Q000502:physiology*; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics
D002121:Calcium Channel Blockers; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C472610:atrial natriuretic peptide, rat; D009320:Atrial Natriuretic Factor; D002118:Calcium