Bimodal Acute Effects of A1 Adenosine Receptor Activation on Na+/H+ Exchanger 3 in Opossum Kidney Cells

Abstract

Abstract. Regulation of renal apical Na ϩ /H ϩ exchanger 3 (NHE3) activity by adenosine has been suggested to contribute to acute control of mammalian Na ϩ homeostasis. The mechanism by which adenosine controls NHE3 activity in a renal cell line was examined. The adenosine analog, N 6 -cyclopentyladenosine (CPA) exerts a bimodal effect on NHE3: CPA concentrations Ͼ10 Ϫ8 M inactivate NHE3, whereas concentrations Ͻ10 Ϫ8 M stimulate NHE3 activity. Acute CPA-induced control of NHE3 was blocked by antagonists of A 1 adenosine receptors and inhibition of phospholipase C, pretreatment with BAPTA-AM (chelator of cellular calcium), and exposure to pertussis toxin. Stimulatory and to some extent also inhibitory CPA concentrations attenuated 8-bromo-cAMP and dopaminemediated inhibition of NHE3. BAPTA eliminated the ability of a stimulatory dose of CPA to attenuate 8-bromo-cAMP-induced suppression of NHE3 activity. Upon inhibition of protein kinase C, CPA at an inhibitory dose provoked activation of NHE3, which is partially reverted by 8-bromo-cAMP and suppressed by pre-incubation with BAPTA-AM. Cytochalasin B, an actin-modifying agent, selectively prevented downregulation but did not affect upregulation of NHE3 activity by CPA. In conclusion, these observations demonstrate that (1) CPA modulates NHE3 activity by elevation of cellular Ca 2ϩ exerting a negative control on adenylate cyclase activity, (2) protein kinase C is the determining factor leading to CPAinduced downregulation of NHE3 activity, and (3) alterations of surface NHE3 abundance may contribute to A 1 adenosine receptor-dependent inhibition of NHE3 activity.

Regulation of apical NHE3 trafficking by ouabain-induced activation of the basolateral Na+-K+-ATPase receptor complex

“…Interestingly, the binding of ouabain to Na ϩ -K ϩ -ATPase not only tethers PLC-␥ 1 and the inositol 1,4,5-trisphosphate receptor (IP 3 R) together to form a Ca 2ϩ -regulatory complex (54) but also stimulates the interaction of the NH 2 terminus of the endoplasmic reticulum-localized IP 3 R with the NH 2 terminus of the Na ϩ -K ϩ -ATPase ␣-subunit (55). These observations support the notion that ouabain-activated Na ϩ -K ϩ -ATPase signaling may also regulate NHE3 in the renal proximal tubule, since intracellular Ca 2ϩ has been shown to regulate NHE3 activity and trafficking (16,29).…”

Adenosine and Kidney Function

“…Notably, studies in opossum kidney cells indicated that activation of adenosine A 1 receptors exerts a bimodal effect on the Na ϩ -H ϩ exchanger NHE3, which is known to play a major role for Na ϩ reabsorption in proximal tubule (349): low concentrations (Ͻ10 Ϫ8 M) of the adenosine analog N 6 -cyclopentyladenosine (CPA) stimulated NHE3 and high concentrations (Ͼ10 Ϫ8 M) inactivated NHE3. Moreover, CPA-induced control of NHE3 was blocked by adenosine A 1 receptor antagonists (68). The inactivation of NHE3 by adenosine was suggested to involve calcineurin homologous protein-mediated regulation of NHE3 (67).…”

“…The intracellular signaling pathways that contribute to adenosine A 1 receptor-mediated increases in proximal tubular reabsorption are still under investigation but may include increases of intracellular Ca 2ϩ (68) and/or reductions of intracellular cAMP levels (174). Notably, studies in opossum kidney cells indicated that activation of adenosine A 1 receptors exerts a bimodal effect on the Na ϩ -H ϩ exchanger NHE3, which is known to play a major role for Na ϩ reabsorption in proximal tubule (349): low concentrations (Ͻ10 Ϫ8 M) of the adenosine analog N 6 -cyclopentyladenosine (CPA) stimulated NHE3 and high concentrations (Ͼ10 Ϫ8 M) inactivated NHE3.…”

Requirement of Intact Adenosine A1Receptors for the Diuretic and Natriuretic Action of the Methylxanthines Theophylline and Caffeine

“…In accordance, in vitro studies in the proximal tubule showed that activation of A 1 receptor can stimulate transport of fluid, HCO 3 Ϫ as well as Na ϩ -glucose and Na ϩ -phosphate symport (Coulson et al, 1991;Takeda et al, 1993;Cai et al, 1994Cai et al, , 1995. The intracellular signaling pathways that contribute to these changes in tubular transport are still under investigation but may include reductions of intracellular cAMP levels (Kost et al, 2000), increases of intracellular Ca 2ϩ (Di Sole et al, 2003), and calcineurin homologous proArticle, publication date, and citation information can be found at http://jpet.aspetjournals.org. doi:10.1124/jpet.104.080432.…”