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Pyridoxine Uptake by Rat Renal Proximal Tubular Cells^1,2,

Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322

Uptake of [³H]pyridoxine by freshly isolated rat renal proximal tubular cells was temperature dependent and exhibited saturation kinetics, with estimated K_t and V_max of 1.3 µM and 14 pmol/(10⁶ cells·0.5 min), respectively. Pyridoxamine, 4'-deoxypyridoxine and 5'-deoxypyridoxal were as effective as unlabeled pyridoxine in inhibiting tracer uptake, whereas pyridoxamine-5'-phosphate, pyridoxal and 4'-pyridoxic acid had no effect, and other vitameric forms and structural analogs caused intermediate inhibition of cellular vitamin uptake. After 30 min, 88% of accumulated radioactive vitamin B-6 in cell extracts had been metabolized to phosphorylated forms and pyridoxal, which supports metabolic trapping as a mechanism responsible for the cellular accumulation observed. Initial uptake was significantly decreased in the presence of ethionine and ouabain; however, carbonylcyanide-p-trifluoromethoxyphenylhydrazone had no effect. Partial to complete substitution of NH₄Cl and LiCl for NaCl had no effect, while partial substitution with RbCl caused significant dose-related inhibition of initial pyridoxine uptake. Amiloride also significantly decreased initial uptake. These data are consistent with a facilitated uptake process for pyridoxine that has substrate specificity and may be modulated by sodium-hydrogen exchange and/or pH gradient effects. Cellular uptake is followed by intracellular metabolic trapping catalyzed by pyridoxal kinase.

KEY WORDS: • pyridoxine • vitamin B-6 • renal cells • cellular uptake • rat • radioactive isotope

¹ Presented in part at the International Congress on Chemical and Biological Aspects of Vitamin B-6 Catalysis, June, 1987 in Turku, Finland; and at the 1988 Annual Meeting of the Federation of American Societies for Experimental Biology, May, 1988, Las Vegas, NV [Bowman, B. B. & McCormick, D. B. (1988) Uptake and metabolism of pyridoxine by rat renal proximal tubular cells. FASEB J. 2: A1086 (abs.)].

² Supported in part by funds from The Coca-Cola Foundation and E. I. duPont de Nemours & Company.

³ B. B. Bowman was the recipient of a National Research Service Award (DK 07977).

Manuscript received 26 September 1988. Revision accepted 10 February 1989.

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