Effects of Methylmercury on Human Neuronal L-Type Calcium Channels Transiently Expressed in Human Embryonic Kidney Cells (HEK-293)

摘要

Methylmercury (MeHg) disrupts the function of native, high voltage-activated neuronal Ca~(2+) channels in several types of cells. However, the effects of MeHg on isolated Ca~(2+) channel phenotypes have not been examined. The aim of the present study was to examine the action of MeHg on recombinant, neuronal L-type voltage-sensitive Ca~(2+) channels. Human embryonic kidney cells (HEK-293) were transfected with human neuronal cDNA clones of the alpha_(1C-1) subunit in combination with alpha_2b and beta_3b Ca~(2+) channel subunits and the reporter jellyfish expressed channels (I_Ba) and their response to MeHg applied acutely were measured using whole-cell voltage-clamp recording techniques and Ba~(2+) (5 mM) as charge carrier. Amplitude of I_Ba in these cells was reduced by the dihydropyridine (DHP), nimodipine, and enhanced by Bay K8644 [S-(-)-1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-[trifluoromethyl]phenyl)-3 pyridine carboxylic acid methyl ester]. MeHg (0.125-5.0 muM) caused a time-and concentration-dependent reduction in amplitude of the peak and sustained current through these channels. However, even at the highest concentration of MeHg tested, reduction of current amplitude by MeHg was incomplete. Washing with MeHg-free solution could not reverse its effects. The steady-state inactivation curve was unaltered by MeHg. Increasing the stimulation frequency or the extracellular Ba~(2+) concentration each attenuated slightly the reduction in amplitude of I_Ba by MeHg. In the presence of MeHg (5.0 muM), Bay K8644 still increased the remaining current, and nimodipine (10 muM) reduced residual current that was resistant to MeHg. Thus, although MeHg reduces the amplitude of recombinant, heterologously expressed L-type channel current, a portion of current is resistant to reduction by MeHg. Furthermore, DHP agonists and antagonists retain their ability to affect L-type Ca~(2+) channel current even in the presence of MeHg.