1. The binding of [3H]-thyrotropin releasing hormone ([3H]-TRH) and [3H]-RX77368 (di-methyl proline TRH) and the ability of these peptides to stimulate phosphoinositide hydrolysis were investigated in the GH3 pituitary cell line. 2. For both peptides binding was found to be saturable with a single component (Hill slopes were, for TRH, 0.98 and for RX77368, 1.13). TRH bound with greater affinity than RX77368 Kd values were 16 nm and 144 nm respectively. Bmax values were 227 fmol mg−1 protein for TRH and 123 fmol mg−1 protein for RX77368. 3. The rank order of potency of a series of TRH analogues to inhibit binding was the same versus each peptide. However, unlike with saturation analysis, Hill slopes of all displacing ligands were less than 1.0 against both TRH and RX77368 suggeting either multiple binding sites, alteration of affinity state, negative co-operativity or some allosteric interaction. 4. Both peptides stimulated phosphoinositide hydrolysis in a dose-dependent fashion. TRH was more potent than RX77368, EC50 values were 7.9 ± 1 nm and 96.3 ± 3 nm respectively. 5. These in vitro data suggest that the greater in vivo potency of RX77368 is not the result of enhanced receptor affinity but is more probably due to its greater metabolic stability.
|Journal||British Journal of Pharmacology|
|Publication status||Published - Nov 1990|