V(2)-LIKE VASOPRESSIN RECEPTOR MOBILIZES INTRACELLULAR CA2+ IN RAT MEDULLARY COLLECTING TUBULES

Cytosolic free calcium concentration ([Ca2+]i) was measured in single microdissected rat medullary collecting tubules [outer (OMCD) and inne r (IMCD)] to identify receptors involved in vasopressin (AVP)-induced [Ca2+]i increases. In both segments, [Phe2, Orn8]vasotocin ([Phe2,Orn8 ]VT), a specific V1 agonist, as well as the V2 agonist 1-desamino-8-D- AVP (dDAVP) triggered [Ca2+]i variations. In OMCD, the mean response t o 10 nM AVP roughly corresponded to the sum of V1 and V2 agonists effe cts. In IMCD, dDAVP (10 nM) alone reproduced the calcium response to A VP (DELTA[Ca2+]i = 243 +/- 34 nM, n = 6, and 248 +/- 27 nM, n = 8, wit h dDAVP and AVP, respectively). Furthermore, in the same experiments V 1 and V2 maximal effects were not additive ([Phe2 Orn8]VT = 154 +/- 21 nM, n = 6; dDAVP + [Phe2 Orn8]VT = 233 +/- 23 nM, n = 9). As AVP, dDA VP released intracellular calcium (DELTA[Ca2+]i in calcium-free medium = 182 +/- 24 nM, n = 8, vs. 182 +/- 14 nM, n = 6 with 10 nM dDAVP and AVP, respectively). Neither 8-(4-chlorophenylthio)adenosine 3',5'-cyc lic monophosphate nor forskolin modified [ Ca2+]i. A cross-reaction of dDAVP with an oxytocin (OT) receptor can be excluded since 1) the spe cific OT agonist [Thr4,Gly7] OT (10 nM) increased only slightly [Ca2+] i (DELTA-[Ca2+]i = 20 +/- 5 nM, n = 11) 2) the dDAVP response was not altered by the specific OT antagonist [1-(O-mercapto-O,O-cyclopentamet hylene propionic acid), 2-(O-methyl)tyrosine,4-threonine, 8-ornithine, 9-tyrosylamide]vasotocin [d(CH2)51,O-Me-Tyr2, Thr4, Tyr-NH29] OVT; 3) it was insensitive to V1 antagonists but was totally blocked by the V1 /V2 antagonist [d(CH2)5,O-Et-Tyr2,Val4]AVP ([DELTA[Ca2+]i = 18 +/- 4 n M, n = 6). These results indicate that in IMCD AVP increases [Ca2+]i v ia both V1 and V2 receptors. [Ca2+]i, variations due to V2 receptors i nvolve a mechanism independent of adenylate cyclase and coupled to the same intracellular calcium pool as V1 and V2 receptors.