The lateral mobility of membrane integral receptors has been implicated as playing a significant role in signal transduction. The adenylate cyclase-coupled vasopressin V2 receptor has been shown to be highly laterally mobile in membranes of LLC-PK1 renal epithelial cells at physiological temperature using a fluorescent vasopressin agonist, with lateral mobility of the V2 receptor proposed to play a role in both adenylate cyclase activation and ligand induced receptor internalization and down-regulation. This study reports the synthesis and characterization of two new fluorescent antagonists [(β-mercapto-β,β-cyclopentamethylene propionic acid)1,d-Tyr2,Ile4,Lys9(N6-fluoresceinylaminothiocarbonyl)]AVP (FL-AVP-data) and [(β-mercapto-β,β-cyclopentamethylene propionic acid)1,d-Tyr2,Ile4,Lys9(N6-tetramethylrhodamylaminothiocarbonyl)]AVP (TR-AVP-anta) for the V2 receptor. The latter was used to determine the parameters of lateral mobility of the V2 receptor in the non-activated antagonist-occupied form. Using fluorescence photobleaching techniques, results were largely comparable to those for agonist-occupied receptor, indicating high mobility at 37°C. Antagonistic properties of the V2 receptor ligands are apparently not related to decreased receptor lateral mobility. Photobleaching measurements, however, did show that in contrast to V2 agonist, V2 antagonist did not induce receptor immobilization due to aggregation with time at 37°C, indicating that this could be of mechanistic importance in the internalization process.
- Receptor antagonist
- Receptor mobility