Characterization and distribution of prolactin releasing peptide (PrRP) binding sites in the rat - evidence for a novel binding site subtype in cardiac and skeletal muscle

1 Prolactin releasing peptide (PrRP) was recently purified from bovine hypo thalamus and binds to the orphan receptor, UHR-1. We examined the distribut ion and kinetics of I-125-PrRP binding in rat tissues together with molecul ar characterization by chemical cross-linking and Northern blotting. 2 In this study I-125-PrRP binding showed specificity and rapid association and dissociation. 3 Specific binding was found in membranes from rat tissues including brain (hypothalamus, medulla oblongata and cerebellum) pituitary, heart, soleus m uscle, adipose tissue, kidney, adrenal gland, testis and small intestine. I n hypothalamus, pituitary, heart and soleus competition analysis indicated only one class of binding site in each tissue. Binding affinity for PrRP (I C50) and binding site density (B-max) respectively were 5.2+/-0.9 nM and 67 4+/-97 fmol mg protein(-1) in hypothalamus (n=5), 1.4+/-0.6 nM and 541+/-12 6 fmol mg protein(-1) in pituitary (n=3), 6.6+/-0.7 nM and 628+/-74 fmol mg protein(-1) in heart (n=4) and 9.8+/-0.9 nM and 677+/-121 fmol mg protein( -1) in soleus muscle (n=4). 4 Analysis of I-125-PrRP-binding site complexes by chemical cross-linking s howed a binding site M-r of 69,000 in hypothalamus and 41,000 in heart and soleus. 5 Northern analysis of polyA(+) RNA from hypothalamus showed a 4.2 kb band as expected for UHR-1, but heart and soleus showed a 4.8 kb band. 6 Taken together these results indicate that there may be different subtype s of PrRP binding sites in rat tissues which may differ from UHR-1.