SPECIFIC BINDING-SITES FOR RAT PREPRO-TRH-(160-169) ON C6 GLIOMA AND BN1010 CLONAL NEURAL CELLS

A connecting decapeptide corresponding to rat pre-pro-TRH-(160-169) (P s4) displays several biological activities that are related or unrelat ed to TRH. We have previously characterized pituitary binding sites fo r this connecting peptide and elucidated structural determinants for h igh peptide binding affinity, In the current study, a series of cell l ines was screened for the presence of specific binding sites with a hi ghly potent derivative of Ps4, the monoiodinated radioligand [I-125-Ty r(0)]Ps4. Neuroblastoma X glioma hybrid NG108-15, glioma C6 and neurob lastoma BN1010 cell lines were found to have high-affinity [I-125-Tyr( 0)]Ps4 binding sites containing 600, 9700 and 130 000 sites/cell, resp ectively. The specific binding of [I-125-Tyr(0)]Ps4 was rapid, time-de pendent, reversible and proportional to the amount of C6 and BN1010 me mbrane preparation, Furthermore, Scatchard or Hill analysis revealed t hat [I-125-Tyr(0)]Ps4 was bound by a single population of non-interact ing sites with dissociation constants in the subnanomolar range, Compe tition studies made with Ps4 analogues indicated that [I-125-Tyr(0)]Ps 4 binding sites on C6 and BN1010 cells, were similar to those previous ly described on rat pituitary membranes. It is concluded that C6 and B N1010 cells are suited for studies on the intracellular events followi ng binding of the Ps4 and for the molecular characterization of the Ps 4 binding sites. (C) 1997 Federation of European Biochemical Societies .