These studies are intended to identify and characterize the cellular mechanisms involved in the transport and accumulation of cadmium by the kidney. The unifying hypothesis of the proposal is that mechanisms normally involved in renal calcium transport represent the means by which cadmium is accumulated by proximal and distal tubular cells. Studies described in this proposal will address several"""""""" problems related to renal cadmium transport by using either continuously cultured opossum kidney cells. Madin-Darby canine kidney (MDCK) cells or primary cell cultures derived from proximal or distal tubules. The cell lines are sensitive to the same hormones that regulate renal calcium transport. and also express phenotypes associated with proximal or distal tubule cells, respectively. The primary cell cultures have been developed and characterized in our lab and are fully operational. Preliminary data demonstrate that parathyroid hormone (PTH) increases cadmium accumulation by MDCK cells by activating dihydropyridine-sensitive calcium channels. Pilot studies also show that beta-estradiol increases PTH-dependent cadmium accumulation. Experiments described in this proposal will characterize: 1) cadmium transport by cultured renal epithelial cells with particular attention to distinguishing between cadmium accumulation, per se, and transepithelial transport. and differences between proximal and distal tubule cells; 2) basal and hormone-modulated cadmium accumulation in cultured kidney cells. Observations of the PTH effects will be extended to other calcitropic hormones including calcitonin and 1.25(OH)2 vitamin D; 3) the chemical forms of cadmium transported by basal and by hormone-stimulated pathways; 4) the cellular mechanisms responsible for entry and exit of ionized cadmium to determine if mechanisms such as Na+/Ca2+ exchange, which also mediate renal calcium transport, may be involved in cadmium uptake or efflux by tubular epithelial cells; 5) the plasma membrane domains of cellular cadmium uptake: 6) the interactions of cadmium with the uptake mechanisms for ionized calcium and other constitutive divalent cations: and. 7) the effects of cadmium on intracellular signal transduction pathways, including cAMP/protein kinase A. inositol lipid/protein kinase C. and intracellular Ca2+. The results of this work should provide important information regarding the mechanisms of renal cadmium transport at the cellular level and may provide a basis for the application or design of pharmacological agents to promote its elimination.
