| The overall goal of this study was to examine the effect of streptozotocin (STZ)-induced diabetes on the function of the active renal organic cation transport system. Previous work completed in our laboratory demonstrated decreased tetraethylammonium (TEA) uptake in freshly isolated proximal tubule (PT) cells from diabetic rats. Because TEA uptake in cells represents both influx and efflux processes, conclusions are limited by nature of the method. Additional experiments were therefore necessary to better define the mechanisms mediating the observed transport impairment. This study was designed to test the hypothesis that impaired TEA uptake in PT cells is due, in part, to a decrease in uptake across the basolateral membrane (BLM) caused by a reduction in the number of functional transporters.; Kinetic analysis of TEA uptake in isolated cells from 21-day diabetic rats revealed a significant 46% decrease in Vmax and a non-significant higher affinity constant, Km. Results from slice experiments, a model that predominately reflects BLM uptake, document a progressive decline in TEA accumulation with increasing duration of diabetes. A maximal decline of 40% occurred after 21 days of diabetes. The accumulation in slices was energy dependent, saturable, and quinine sensitive, indicating that the uptake was primarily mediated by the organic cation transport system. Normal oxygen consumption was found in diabetic slices, suggesting that the transport impairment at the BLM is not due to altered aerobic metabolism.; Instead, the observed transport impairment in slices was attributable to molecular adaptations of the BLM organic cation transporters (OCTs). OCT1 and OCT2 protein expression was found to be decreased 50% and 70% respectively, in 21-day diabetic slices. Examination of OCT mRNA revealed OCT1 was unaltered, while OCT2 was decreased by 50% after 21 days of diabetes. These results suggest differential regulation exists for the cloned transporters. As expected, studies using insulin prevented all diabetes-associated impairments, arguing against the likelihood of either an acute toxic action of STZ or an artifact of the methods. This study implies diabetes-associated changes in the organic cation transport system may alter renal handling of drugs, toxins, and endogenous compounds. |
