Adrenomedullin upregulates M2-muscarinic receptors in cardiomyocytes from P19 cell line

Br J Pharmacol. 2003 Jul;139(6):1219-27. doi: 10.1038/sj.bjp.0705350.

Abstract

1. The effects of AM on expression of muscarinic (M) receptors from P19-derived cardiomyocytes were examined. 2. RT-PCR experiments revealed expression of M(1)-M(4) receptor genes. Immuno-histochemistry indicated that M(2) expression is restricted to contractile cells. Carbachol inhibition of isoprenaline-induced increase in beating rate was prevented by atropine and methoctramine (pA(2): 8.1). Inhibition of [(3)H]-NMS binding by atropine (pK(i): -8.4+/-0.2) and methoctramine (pK(i): -8.3+/-0.2) suggests that M(2) is the functional expressed isoform. 3. [(3)H]-NMS binding and semiquantitative RT-PCR studies showed a dome shaped time course of M(2) expression with a maximum at 7 days of differentiation followed by a progressive decline. 4. AM concentration-dependently upregulated M(2) receptor mRNA during late differentiation stages in P19 cells but also in rat atrial cardiomyocytes. This effect was potentiated by factor H. AM (100 nM) plus factor H (50 nM) treatment of P19 cells for 24 h significantly increased [(3)H]-NMS-specific binding (B(max): 81+/-7 vs 31+/-6 fmol mg(-1) prot). The effect of AM on mRNA levels was prevented by AM receptor antagonist AM(22-52) (1 micro M) but not by CGRP antagonist, CGRP(8-37) (1 micro M). 5. The mRNA levels encoding CRLR receptor declined with culture duration, whereas those encoding L1/G10D receptor remained stable. 6. Our findings demonstrate that AM regulates M(2) receptors expression in cardiomyocytes probably through a mechanism involving L1/G10D receptors. The 'in vivo' significance of this phenomenon remains to be demonstrated.

MeSH terms

  • Adrenomedullin
  • Animals
  • Cell Differentiation / physiology
  • Cell Line, Tumor
  • Mice
  • Myocytes, Cardiac / cytology
  • Myocytes, Cardiac / metabolism
  • Myocytes, Cardiac / physiology*
  • Peptides / physiology*
  • Protein Binding / physiology
  • RNA, Messenger / biosynthesis
  • RNA, Messenger / genetics
  • Receptor, Muscarinic M2 / biosynthesis*
  • Receptor, Muscarinic M2 / genetics
  • Up-Regulation / physiology*

Substances

  • Peptides
  • RNA, Messenger
  • Receptor, Muscarinic M2
  • Adrenomedullin