1. Adenosine influences the vectorial transport. of Na+ and HCO3- across ki
dney epithelial cells. However, its action on effector proteins, such as th
e Na+-H+ exchanger NHE3, an epithelial brush border isoform of the Na+-H+ e
xchanger (NHE) gene family: is not yet defined,
2. The present study was conducted in Xenopus laevis distal nephron A6 epit
helia which express both an apical adenosine receptor of the A(1) type (cou
pled to protein kinase C (PKC)) and a basolateral receptor of the A(2) type
(coupled to protein kinase A (PKA)). The untransfected A6 cell line expres
ses a single NHE type (XNHE) which is restricted to the basolateral membran
e and which is activated by PKA.
3. A6 cell lines were generated which express exogenous rat NHE3. Measureme
nts of side-specific pH(i) recovery from acid loads in the presence of HOE6
94 (an inhibitor with differential potency towards individual NHE isoforms)
detected an apical resistant Na+-H+ exchange only in transfected cell line
s. The sensitivity of the basolateral NHE to HOE694 was unchanged, suggesti
ng that exogenous NHE3 was restricted to the apical membrane.
4. Stimulation of the apical A(1) receptor with N-6-cyclopentyladenosine (C
PA) inhibited both epical NHE3 and basolateral;XNHE. These effects were mim
icked by the addition of the protein kinase C (PKC) activator phorbol 12-my
ristate 13-acetate (PMA) and partially prevented by the PKC inhibitor calph
ostin C which also blocked the effect of PMA.
5. Stimulation of the basolateral A(2) receptor with CPA inhibited apical N
HE3 and stimulated basolateral XNHE. These effects were mimicked by 8-bromo
-cAMP and partially prevented by the PKA inhibitor H89 which entirely block
ed the effect of 8-bromo-cAMP.
6. In conclusion, CPA inhibits rat NHE3 expressed apically in A6 epithelia
via both the apical PKC-coupled A(1) and the basolateral PKA-coupled A(2) a