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Characterization of binding sites for N-Tyr-MIF-1 (Tyr-Pro-Leu-Gly-NH2) in rat brain

1982; Elsevier BV; Volume: 17; Issue: 6 Linguagem: Inglês

10.1016/0091-3057(82)90119-8

1873-5177

James E. Zadina, Abba J. Kastin, Edward F. Krieg, David H. Coy,

Cardiovascular, Neuropeptides, and Oxidative Stress Research

Binding sites for N-Tyr-Pro-Leu-Gly-NH2 (Tyr-MIF-1), a novel peptide structurally related but immunoreactively different from Pro-Leu-Gly-NH2 (MIF-1), were investigated. The presence of Tyr-MIF-1-like material in brain tissue has previously been demonstrated by RIA and its levels were shown to vary with the diurnal cycle and after pinealectomy. We now demonstrate a high affinity binding site in rat brain that is saturable and specific for Tyr-MIF-1. Crude P2 synaptosomal fractions from rat brains were incubated with 125I-Tyr-MIF-1 in the presence or absence of μM unlabelled Tyr-MIF-1 (nonspecific binding). Binding reached equilibrium at 30–40 min at 23°C and at about 4 hr on ice, after which it was relatively stable for at least 18 hr. None of the other peptides (including MIF-1) or amino acid residues tested were found to effectively compete for 125I-Tyr-MIF-1 binding. Binding was linear with protein from 280 μg to at least 1.1 mg protein per tube. Scatchard analysis of the striatum-thalamus revealed the presence of binding sites with an apparent KD of 91 nM and maximum number of sites in the range of 45 fmol/mg tissue. Analysis of several brain areas revealed a differential distribution of the binding sites with relatively high concentrations in cortex, striatum, and amygdala and low concentrations in pons-medulla. Together with the previously published RIA results, the demonstration of a receptor for Tyr-Pro-Leu-Gly-NH2 supports the concept of the presence of this novel peptide and its receptor in the brain.