Persistent stimulation of adenylate cyclase and urea transport by an AVP photolabel

Abstract

The effects of a photoaffinity label for arginine vasopressin receptors, [Phe2, Phe(p-N3)3]AVP (N3-AVP), on urea permeability and adenylate cyclase activity have been investigated in the toad urinary bladder. This compound, when activated by ultraviolet light, induced a maximal and persistent increase in the urea permeability of the intact bladder and a persistent increase in the adenylate cyclase activity of toad bladder epithelial cell homogenates. Covalent attachment of the analogue to target tissue during photolysis was equivalent at 4 and 20 degrees C. Bladders exposed to N3-AVP in the presence of AVP during photolysis were substantially less permeable to urea than controls that had been exposed to N3-AVP alone. These findings constitute further evidence in support of our previous suggestion that N3-AVP binds covalently to AVP receptors and, in addition, demonstrates that N3-AVP evokes a persistent increase in adenylate cyclase activity which, in turn, triggers a persistent increase in bladder permeability to urea.

Photoaffinity, biotinyl, and iodo analogs as probes for vasotocin receptors

“…The present studies shows that analogues of vasotocin with major modifications in position 9 are biologically active in increasing toad bladder permeability to water. [2-phenylalanine,9-tyrosine]vasotocin (2) 2.8 ± 0.6 0 l-desamino[9-(p-aminophenylalanine)]vasotocin (3) [2-phenylalanine,9-(iodotyrosine)]vasotocin (4) l-desamino[9-(p-azidophenylalanine)]vasotocin (5) l-desamino[9-(biotinylphenylalanine)] vasotocin (6) 66 ± 13 0.41 ± 0.14 40 ± 7 23 ± 10 0 "Units/mg; mean ± SD. 6Reference 13.…”

“…These observations emphasize the structural difference between the amphibian hydroosmotic receptor and the mammalian antidiuretic (V-2) receptor. Nevertheless, the photoreactive analogue (compound 5) and the biotinyl analogue (compound 6) do have substantial antidiuretic activity and may, therefore, be useful in the isolation and characterization of the mammalian V-2 receptor.…”

“…0.82 (A), 0.75 (B), 0.50 (C); HPLC t (min) 21.87, k'= 2.25 (20% i-PrOH), single peak, 99% purity. d[biotinyl-Phe9]AVT (6). Compound 3 (20 mg) was dissolved in 2 mL of DMF and treated at pH 7.5 (JV-methylmorpholine as base) with 15 mg of biotinyl succinimide ester (Pierce) overnight at room temperature.…”