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 inner (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 to 10 nM AVP roughly corresponded to the sum of V1 and V2 agonists effects. In IMCD, dDAVP (10 nM) alone reproduced the calcium response to AVP (DELTA[Ca2+]i = 243 +/- 34 nM, n = 6, and 248 +/- 27 nM, n = 8, with dDAVP and AVP, respectively). Furthermore, in the same experiments V1 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, dDAVP 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'-cyclic monophosphate nor forskolin modified [ Ca2+]i. A cross-reaction of dDAVP with an oxytocin (OT) receptor can be excluded since 1) the specific 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-cyclopentamethylene 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 nM, n = 6). These results indicate that in IMCD AVP increases [Ca2+]i via both V1 and V2 receptors. [Ca2+]i, variations due to V2 receptors involve a mechanism independent of adenylate cyclase and coupled to the same intracellular calcium pool as V1 and V2 receptors.