Received for publication September 22, 2006.
Revised January 19, 2007.
Accepted for publication January 22, 2007.

Hypoxic Induction of RAMP2 Alters Regulation of Pulmonary Endothelin-1 by Adrenomedullin: Induction under Normoxia versus Inhibition under Hypoxia

Abstract

The vasodilator adrenomedullin (AM) is upregulated in pulmonary hypertension, and inhaled AM is beneficial in patients. We therefore investigated effects of AM on pulmonary endothelin-1 (ET-1). In normoxic isolated rat lungs (IRL) and rat pulmonary artery endothelial cells (RPAEC), the calcitonin gene-related peptide type-1 receptor (CGRP1R) antagonist hCGRP(8-37) decreased ET-1 secretion, the AM receptor antagonist hAM(22-52) had no effect. Exogenous AM (1 and 10 pmol/l) increased ET-1 levels, which was abolished by hCGRP(8-37) and protein kinase A (PKA) inhibition. At 50 and 100 pmol/l, AM decreased ET-1 -- an effect sensitive to hAM(22-52), NO inhibition, and protein kinase G (PKG) inhibition. In RPAEC, these results were attributed to altered ET-1 gene expression; low exogenous AM also promoted activity of endothelin-converting enzyme, high AM increased the number of endothelin type-B (ETB) receptor sites. Hypoxia significantly elevated AM and ET-1 levels in IRL and RPAEC, and hAM(22-52), NO inhibition, or PKG inhibitors caused a further ET-1 rise. These interventions also prevented the hypoxia-related increase in ETB sites in RPAEC. In RPAEC, both high AM and hypoxia downregulated RAMP1 but upregulated RAMP2 protein and AM receptor sites, and RAMP2 silencing by siRNA proved its pivotal role for signal switching. In conclusion, endogenous pulmonary AM upregulates ET-1 and endothelin-converting enzyme activity under physiological conditions, via CGRP1R and PKA. In contrast, hypoxia-induced high AM levels, via AM1 receptor and NO/PKG, downregulate ET-1 gene expression and promote expression of ETB receptors. This hypoxia-related switch of AM signaling can be attributed to upregulation of the RAMP2/AM1 receptor system.

Key words: RAMP, adrenomedullin, endothelin-1, endothelium, lung, receptors