Role of the calcium/calmodulin-dependent protein kinase II in the regulation of the renal basolateral PAH and dicarboxylate transporters

The aim of the present study was to investigate whether the activities of the renal basolateral organic anion transporter (PAH transporter) and the sodium-dependent dicarboxylate transporter are modulated by the calcium/calmodulin-dependent multifunctional protein kinase II (CaM kinase II). The studies were performed on isolated S2 segments of proximal tubules microdissected from rabbit kidneys without the use of enzymatic agents. 3H-PAH was used as marker substance of the anion transporter, and 14C-glutarate as a marker of the sodium/dicarboxylate cotransporter. Because the tubules were not perfused, and hence were collapsed, the tubular uptake of the marker substances reflects transport across the basolateral cell membrane. To obtain uptake rates most closely related to initial transport rates, 30 s tubular uptake measurements were performed. The results show that a selective inhibitor of CaM kinase II, KN93, inhibited tubular PAH uptake. The smallest effective dose was 10(-7) M. An inactive analogue of KN93, KN92, was without effect, even at the high concentration of 10(-5) M. In contrast to PAH transport, tubular 14C-glutarate uptake was not affected by KN93 (10(-5) M). PAH transport was also inhibited after elevation of intracellular Ca2+ by the Ca(2+)-ionophore A 23187 and by the polycationic antibiotic neomycin, but not by the intracellular Ca2+ modulators thapsigargin and ryanodine. The effect of the Ca(2+)-ionophore could be abolished by KN93, but not by Rp-cAMPs, an inhibitor of protein kinase A, indicating that this event was mediated by CaM kinase II, but not by PKA. The results provide the first evidence that, in addition to the protein kinases A and C (previous studies from this lab), CaM kinase II has a role in the regulation of the renal basolateral PAH transporter, whereas the renal basolateral dicarboxylate transporter does not depend on CaM kinase II activity.