Role of adenosine triphosphate (ATP) and NaK ATPase in the inhibition of proximal tubule transport with intracellular cystine loading

Abstract

Cellular cystine loading with cystine dimethyl ester inhibits volume absorption, transepithelial potential difference, glucose transport, and bicarbonate transport in proximal convoluted tubules perfused in vitro. This study examined the roles of ATP and NaK ATPase in this in vitro model of the Fanconi syndrome of cystinosis. Intracellular ATP was measured using the luciferin-luciferase assay. Intracellular ATP was reduced by 60% in proximal convoluted tubules incubated with 0.5 mM cystine dimethyl ester for 15 min at 37°C (P < 0.001). Incubation of cystine loaded tubules with 1 mM exogenous ATP increased intracellular ATP to levels not significantly different than that of controls. On the other hand, Vmax NaK ATPase activity was unchanged even though the incubation times and the concentration of cystine dimethyl ester were doubled to 30 min and 1 mM, respectively. In proximal convoluted tubules perfused in vitro, 0.5 mM cystine dimethyl ester resulted in an 89% inhibition in volume absorption (0.81±0.14 to 0.09±0.09 nl/mm · min), while there was only a 45% inhibition in volume absorption (P < 0.01) due to cellular cystine loading in the presence of 1 mM lumen and bath ATP (0.94±0.05 to 0.52±0.11 nl/mm · min). These data demonstrate that proximal tubule cellular cystine loading decreases cellular ATP concentration, but does not directly inhibit NaK ATPase activity. The inhibition in transport and decrease in intracellular ATP due to cellular cystine loading was ameliorated by exogenous ATP. These data are consistent with cellular ATP depletion playing a major role in the inhibition of proximal tubule transport due to intracellular cystine loading.