PGE2-INDUCED INHIBITION OF AVP-DEPENDENT CAMP ACCUMULATION IN THE OMCD OF THE RAT-KIDNEY IS CUMULATIVE WITH RESPECT TO THE EFFECTS OF ALPHA-2-ADRENERGIC AND ADENOSINE-A1 AGONISTS, INSENSITIVE TO PERTUSSIS TOXIN AND DEPENDENT ON EXTRACELLULAR CALCIUM

The accumulation of cyclic adenosine 3',5'-phosphate (cAMP) elicited by antidiuretic hormone (arginine vasopressin, AVP) in the medullary collecting tubule (OMCD) microdissected from the rat kidney is inhibited by different factors: the A1 agonist of adenosine (-)-N6-(R-phenylisopropyl) adenosine (PIA), an alpha-2-adrenergic agonist clonidine (CLO), and prostaglandin E2 (PGE2). The negative regulation elicited by PGE2 was further characterized by measuring summation of inhibition with other inhibitors, by testing the effect of pertussis toxin and by studying the part played by extracellular calcium. Inhibitors were used at concentrations inducing maximum effects. The simultaneous addition of 0.3 muM PGE2 with either 0.1 muM PIA or 1 muM CLO led to an inhibition of the response to AVP (80.0 +/- 3.5 %, SEM, N = 7 and 92.6 +/- 0.8 %, N = 5, respectively) greater than those elicited by each agent alone. In contrast, PIA and CLO added together induced an inhibition similar to that due to CLO alone. The action of PGE2 in combination with either PIA or CLO corresponded to a partial summation fitting with the values calculated by assuming a cumulative inhibition. Preincubation of OMCD samples with pertussis toxin (100 ng/ml or 1 mug/ml) relieved the inhibitory effects of CLO and PIA but did not affect the action of PGE2. PGE2-induced inhibition was prevented, in a calcium-free medium [0 Ca2+ + 0.1 mM [ethylene-bis (oxyethylene-nitrilo)] tetraacetate (EGTA)]: values were 67.0 +/- 2.1 % and 5.8 +/- 8.7 % (+/- SEM) in 2 mM Ca2+ and 0 Ca2+ medium, respectively, N = 7. When applied to Fura-2-loaded OMCD, 0.3 muM PGE, increased intracellular calcium concentration ([Ca2+]i) with a peak phase (in 2 mM or 0 Ca2+ medium) followed by a plateau phase (observed only in 2 mM Ca2+ medium). It is concluded that: (1) in the rat OMCD, PGE2, PIA and CLO act on the same AVP-sensitive cell, (2) PGE2 induces a cumulative inhibition on the cAMP level when combined with other inhibitors by a mechanism insensitive to pertussis toxin, (3) the presence of extracellular calcium is a prerequisite condition to observe PGE2-induced inhibition, and (4) the inhibition by PGE2 might be linked to its capacity of increasing [Ca2+]i.