Adenosine modulates Mg2+ uptake in distal convoluted tubule cells via A(1)and A(2) purinoceptors

Adenosine plays a role in the control of water and electrolyte reabsorption in the distal tubule. As the distal convoluted tubule is important in the regulation of renal Mg2+ balance, we determined the effects of adenosine on cellular Mg2+ uptake in this segment. The effect of adenosine was studied on immortalized mouse distal convoluted tubule (MDCT) cells, a model of the intact distal convoluted tubule. The rate of Mg2+ uptake was measured with fluorescence techniques using mag-fura 2. To assess Mg2+ uptake, MDCT cell s were first Mg2+ depleted to 0.22 +/- 0.01 mM by being cultured in Mg2+-fr ee media for 16 h and then placed in 1.5 mM MgCl2; next, changes in intrace llular Mg2+ concentration ([Mg2+](i)) were determined. [Mg2+](i) returned t o basal levels, 0.53 +/- 0.02 mM, with a mean refill rate, d([Mg2+](i))/dt, of 137 +/- 16 nM/s. Adenosine stimulates basal Mg2+ uptake by 41 +/- 10%. The selective A(1) purinoceptor agonist N-6-cyclopentyladenosine (CPA) incr eased intracellular Ca2+ and decreased parathyroid hormone (PTH)-stimulated cAMP formation and PTH-mediated Mg2+ uptake. On the other hand, the select ive A(2) receptor agonist 2-[p-(2-carbonyl-ethyl)-phenylethylamino]-5'-N-et hyl-carboxamidoadenosine (CGS) stimulated Mg2+ entry in a concentration-dep endent fashion. CGS increased cAMP formation and the protein kinase A inhib itor RpcAMPS inhibited CGS-stimulated Mg2+ uptake. Selective inhibition of phospholipase C, protein kinase C, or mitogen-activated protein kinase enzy me cascades with U-73122, Ro-31-8220, and PD-98059, respectively, diminishe d A(2) agonist-mediated Mg2+ entry. Aldosterone potentiated CGS-mediated Mg 2+ entry, and elevation of extracellular Ca2+ diminished CGS-responsive cAM P formation and Mg2+ uptake. Accordingly, MDCT cells possess both A(1) and A(2) purinoceptor subtypes with intracellular signaling typical of these re spective receptors. We conclude that adenosine has dual effects on Mg2+ upt ake in MDCT cells through separate A(1) and A(2) purinoceptor pathways.