Iodinated neurohypophyseal hormones as potential ligands for receptor binding and intermediates in synthesis of tritiated hormones.

[3-Iodo-Tyr2]oxytocin (MIOT), [3,5-diiodo-Tyr2]oxytocin (DIOT), [3-iodo-Tyr2,Lys8]vasopressin (MILVP), [3,5-diiodo-Tyr2,Lys8]vasopressin (DILVP), [3-iodo-Tyr2,Arg8]vasopressin (MIAVP), and [3,5-diiodo-Tyr2,Arg8]vasopressin (DIAVP) were synthesized by iodination of the respective hormones, pruified, and characterized. All the monoiodo hormones had to be freshly prepared prior to bioassays, since on storage they gave rise to hormonal-like biological activity. The biological activities of these iodo analogues were measured in an adenylate cyclase assay employing neurohypophyseal hormone (NHH) sensitive bovine renal medullary membranes, and/or the rat oxytocic assay. In the cyclase assay, DIOT, DILVP, and DIAVP were inactive as agonists or antagonists. MIOT shows no agonistic activity in the renal cyclase system and uterus, but is a weak reversible inhibitor of oxytocin (OT) in both systems. When MIOT (10(-4) M) was preincubated with renal membranes for 10 min at 37 degrees C before addition of OT, it behaved as a noncompetitive inhibitor of NHH-stimulated adenylate cyclase. MILVP and MIAVP appear to be partial agonists with Km (half maximal response) 3 X 10(-6) and 3 X 10(-7) M, respectively, as determined in the cyclase assay. Upon preincubation with renal medullary membranes, MILVP (10(-6) M) behaves as a more potent noncompetitive inhibitor of OT than MIOT. Accordingly, iodo derivatives of NHH do not exhibit sufficient affinity to serve an specific ligands to measure OT, LVP, or AVP receptors in the uterus and kidney. Study of the specificity of inhibition produced by MIOT revealed that this analogue does not act selectively upon NHH receptors. Thus, MIOT modified adenylate cyclase systems which do not have NHH receptors, e.g., the PTH-sensitive adenylate cyclase in bovine renal cortex and the glucagon-sensitive adenylate cyclase in rat liver. DIOT, DILVP, and DIAVP were subjected to catalytic tritiation (employing carrier free tritium) and were converted to [3H]OT (25, 31, and 25 Ci/mmol), [3H]LVP (26 and 23 Ci/mmol), and [3H]AVP (17 Ci/mmol), respectively. These tritiated ligands have been successfully used to measure NHH receptor sites both in kidney and uterine membranes as described in other studies.