MERCURY UPTAKE BY PRIMARY CULTURES OF RAT RENAL CORTICAL EPITHELIAL-CELLS .1. EFFECTS OF CELL-DENSITY, TEMPERATURE, AND METABOLIC-INHIBITORS

Factors affecting the renal uptake of inorganic mercury were investigated using primary cultures of rat renal cortical epithelial (RCE) cells under protein- and amino acid-free conditions. The cells were isolated from kidneys of adult rats and cultured, Confluence of culture (cell density), monitored morphologically and by total protein content, was achieved on Day 5. The RCE cells were incubated with 1 mu M Hg at 37 degrees C for 30 min, followed by washing with phosphate-buffered saline containing various chelating agents (i.e., EGTA, PEN, DMSA, and BAL) to remove the surface-bound, noninternalized Hg. A substantial portion of Hg was bound to the cell surface. The removal of Hg from these binding sites was dependent on the stability constants of the chelating agents for Hg and lipophilic BAL removed the most Hg. Hg accumulation by the cells was dependent on cell density and decreased as the cell culture became confluent, possibly due to the formation of tight junctions resulting in a majority of the Hg transport occurring through the apical membrane. As measured after BAL washing, metabolic inhibitors, NaF, DNP, and ouabain decreased Hg accumulation by 28% and low temperature (4 degrees C) decreased it by 62%. Dependence of Hg uptake on metabolic energy and temperature suggests that a part of Hg is transported via active transport system, The pronounced decrease of Hg uptake at 4 degrees C indicates that, in addition to active transport, Hg transport also involves simple diffusion, some of which is dependent on membrane fluidity. It is concluded that Hg transport in RCE cells through the apical membrane occurs mainly by diffusion, and to a smaller extent by active transport. (C) 1995 Academic Press, Inc.