CHARACTERIZATION AND MOLECULAR-IDENTIFICATION OF ADRENOMEDULLIN BINDING-SITES IN THE RAT SPINAL-CORD - A COMPARISON WITH CALCITONIN-GENE-RELATED PEPTIDE RECEPTORS

Calcitonin gene-related peptide (CGRP) and its receptors are found in mammalian spinal cord. We show, for the first time, binding sites for the novel related peptide adrenomedullin in rat spinal cord microsomes . I-125-Adrenomedullin binding showed high affinity (KD = 0.45 +/- 0.0 6 nM) and sites were abundant (B-max = 723 +/- 71 fmol/mg of protein). CGRP, amylin, and calcitonin did not compete at these sites (K-i > 10 mu M). High-affinity CGRP binding sites (K-D = 0.18 +/- 0.01 nM) were much less numerous (B-max = 17.7 +/- 2.4 fmol/mg of protein) and show ed competition by unlabeled adrenomedullin (K-i = 34.6 +/- 2.4 nM). Ch emical cross-linking revealed a major band for I-125-adrenomedullin of M(r) = 84,400 +/- 1,200 and a minor band of M(r) = 122,000 +/- 8,700. I-125-CGRP cross-linking showed bands of lower molecular weight (M(r) = 74,500 +/- 5,000 and 61,000 +/- 2,200). Enzymic deglycosylation of the adrenomedullin binding site showed a considerable carbohydrate con tent. Neither adrenomedullin nor CGRP was able to increase cyclic AMP in spinal cord, Adrenomedullin mRNA was present in spinal cord, at one -third of its level in lung, and adrenomedullin immunoreactivity was p resent, at a low concentration (40 fmol/g of tissue). Thus, the presen ce of abundant binding sites acid adrenomedullin mRNA and immunoreacti vity anticipate an as yet undefined function for this peptide in spina l cord.