Abstract

In previous studies we found that intraperitoneal injection of nicotinamide (NiAm) to rats resulted in increased NAD+ content in proximal tubules, inhibition of brush border membrane (BBM) transport of phosphate (Pi) and decreased activity of alkaline phosphatase (AP). We now studied the effect of NiAm injection on rabbit kidney BBM prepared either directly by Ca2+ precipitation method, or prepared indirectly from sheets of BBM. In BBM vesicles prepared directly from NiAm-injected rabbits, Na+-dependent Pi uptake was inhibited, but no inhibition was found in BBM vesicles prepared by an indirect method. Incubation of both directly prepared BBM vesicles and of BBM sheets with phosphatidylinositol-specific phospholipase C (PI-PLC) released about 85% of AP from BBM. In BBM vesicles prepared indirectly from BBM sheets, incubation with PI-PLC increased by 100% the capacity for Pi transport, but PI-PLC had no effect on Pi transport if rabbits were injected with NiAm. On the other hand, incubation of directly prepared BBM vesicles with PI-PLC did not alter Pi transport capacity both in controls and in NiAm-treated rabbits, although it released AP. Treatment with NiAm decreases significantly AP activity both in BBM vesicles prepared directly or prepared indirectly from BBM sheets. These results suggest that NiAm-induced inhibition of BBM transport system for Pi is reversed by prolonged washing and incubation in the course of indirect preparation of BBM vesicles. Results also suggest that an increase in tissue NAD+ decreases susceptibility of BBM to treatment with PI-PLC in altering Pi transport. Removal of the majority of AP from BBM does not impair Na+-gradient-dependent Pi transport system.