Expression and distribution of the calcitonin receptor-like receptor in the developing rat heart

During ontogenesis the 52 amino acid peptide adrenomedullin is first expressed in the heart and it is essential for normal cardiovascular development. Recent work suggests that most adrenomedullin effects are conveyed via the calcitonin receptor-like receptor (CRLR) in combination with appropriate receptor activity-modifying proteins (RAMPs). Here, we investigated the expression of these components during the development of the rat heart, focusing on the period of coronary vascular development. Using RT-PCR, transcripts for CRLR, RAMP1 and RAMP2 were detected at all stages from E 14 to adulthood. The distribution of CRLR was investigated by immunohistochemistry, and endothelial cells and their precursors identified with monoclonal antibodies against RECA-1 and flk-1. On E 14, intense CRLR immunoreactivity was observed in endothelial cells of the large vessels and the endocardial cushions at the AV-junction. Small CRLR immunoreactive cell clusters were located in the wall of the outflow tract and subepicardially in the ventricular wall. On E 16, tubes of CRLR immunoreactive cells formed a subepicardial plexus, from which they penetrated radially towards the trabecular network and entered at E 18. Smooth muscle cells of coronary arteries gained a moderate CRLR immunoreactivity at E 20 which persisted at this intensity up to P 8 and then decreased. At the same time, CRLR immunoreactivity of endothelial cells in coronary arteries vanished while those of coronary veins still exhibited intense CRLR immunoreactivity. These data suggest multiple functions of the adrenomedullin/CRLR signaling pathway in cardiac development, among which the most prominent appears to be the early outgrowth and proliferation of the immature endothelial cells of the coronary vasculature.