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Adenosine inhibits the transfected Na+-H+ exchanger NHE3 in Xenopus laevis renal epithelial cells (A6/C1).
Di Sole F
,
Casavola V
,
Mastroberardino L
,
Verrey F
,
Moe OW
,
Burckhardt G
,
Murer H
,
Helmle-Kolb C
.
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1. Adenosine influences the vectorial transport of Na+ and HCO3- across kidney epithelial cells. However, its action on effector proteins, such as the Na+-H+ exchanger NHE3, an epithelial brush border isoform of the Na+-H+ exchanger (NHE) gene family, is not yet defined. 2. The present study was conducted in Xenopus laevis distalnephron A6 epithelia which express both an apical adenosine receptor of the A1 type (coupled to protein kinase C (PKC)) and a basolateral receptor of the A2 type (coupled to protein kinase A (PKA)). The untransfected A6 cell line expresses a single NHE type (XNHE) which is restricted to the basolateral membrane and which is activated by PKA. 3. A6 cell lines were generated which express exogenous rat NHE3. Measurements of side-specific pHi recovery from acid loads in the presence of HOE694 (an inhibitor with differential potency towards individual NHE isoforms) detected an apical resistant Na+-H+ exchange only in transfected cell lines. The sensitivity of the basolateral NHE to HOE694 was unchanged, suggesting that exogenous NHE3 was restricted to the apical membrane. 4. Stimulation of the apical A1 receptor with N 6-cyclopentyladenosine (CPA) inhibited both apical NHE3 and basolateral XNHE. These effects were mimicked by the addition of the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA) and partially prevented by the PKC inhibitor calphostin C which also blocked the effect of PMA. 5. Stimulation of the basolateral A2 receptor with CPA inhibited apical NHE3 and stimulated basolateral XNHE. These effects were mimicked by 8-bromo-cAMP and partially prevented by the PKA inhibitor H89 which entirely blocked the effect of 8-bromo-cAMP. 6. In conclusion, CPA inhibits rat NHE3 expressed apically in A6 epithelia via both the apical PKC-coupled A1 and the basolateral PKA-coupled A2 adenosine receptors.
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