VALPROIC ACID IMPAIRS CARNITINE UPTAKE IN CULTURED HUMAN SKIN FIBROBLASTS - AN IN-VITRO MODEL FOR THE PATHOGENESIS OF VALPROIC ACID ASSOCIATED CARNITINE DEFICIENCY

The mechanisms of valproate-associated carnitine deficiency are controversial. The urinary excretion of valproylcarnitine is insufficient to account for tissue carnitine depletion. To explore this mechanism, we studied the effects of valproic acid (VPA) on carnitine uptake in cultured human skin fibroblasts by the method of Tein et al. (Pediatr Res 28:247-255, 1990). Fibroblasts were preincubated with varying concentrations (0-2000 muM) of VPA for 1, 3, 5, 7, 10, 14, 21, and 28 d and then incubated with fixed carnitine concentrations of 50 muM (normal physiologic concentration), 20 muM (as seen in secondary carnitine deficiency disorders), or 5 muM (as seen in the plasma membrane carnitine transport defect). There was an exponential dose-dependent decrease in carnitine uptake with increasing VPA concentrations, and the relative inhibitory effect was the same for all three carnitine concentrations. The mean percentages +/- SD (n - 1) of residual carnitine uptake for all combined preincubation periods (1-28 d) and combined carnitine concentrations (5, 20, and 50 mumol/L) with increasing concentrations of VPA varied from 83.4 +/- 2.6% (10 muM VPA) to 56.7 +/- 0.1% (500 muM) to 19.8 +/-1.3% (2000 muM). The degree of inhibition was directly proportional to the time of VPA preincubation and parallel for all three carnitine concentrations; the longer the preincubation period, the lower the toxic dose of VPA (to a minimum of 450 muM), resulting in a 50% suppression of carnitine uptake (TD50). The mean TD50 of the combined carnitine concentrations for increasing preincubation periods of VPA varied from 1898 +/- 214 muM (1 d) to 447 +/-9 muM (28 d), tapering toward an asymptote of 450 muM when the preincubation period exceeded 14 d. This in vitro TD50 value may be comparable to the in vivo therapeutic range of serum VPA concentrations (350-700 mumol/L) for anticonvulsant therapy. We conclude that one mechanism by which long-term VPA therapy induces serum and tissue carnitine depletion is through inhibition of plasmalemmal carnitine uptake, including decreased renal reabsorption of free carnitine. This effect is directly proportional to the duration of exposure and concentration of VPA.