Vol. 295, Issue 2, 531-545, November 2000

Site of Action of Lubeluzole on Voltage-Sensitive Ca²⁺ Channels in Isolated Dorsal Root Ganglion Cells of the Rat: Influence of pH

Roger Marrannes and Erik De Prins

Central Nervous System Discovery Research, Janssen Research Foundation, Beerse, Belgium

Besides other pharmacological effects, the neuroprotective compound lubeluzole blocks low-voltage-activated (iLVA) and high-voltage-activated (iHVA) calcium channel currents. We investigated the site of action of lubeluzole on Ca²⁺ channels in isolated dorsal root ganglion cells of the rat, using whole-cell voltage clamp. Experiments with extracellular application of 3 µM lubeluzole (pK_a = 7.6) at different values of extracellular pH suggest that the protonated form of lubeluzole contributes to the block of iLVA and iHVA from the extracellular side. The partial block of iLVA and iHVA by 3 µM lubeluzole at extracellular pH 9 and intracellular pH (pH_i) 9 indicates that the uncharged form of lubeluzole (L) may contribute to the block as well. The voltage-dependent acceleration of the apparent inactivation of iHVA by lubeluzole was much more pronounced at lower pH_i, which is consistent with membrane penetration of L and an open channel block of iHVA by the prononated form of lubeluzole acting from the intracellular side. Decreasing pH_i induced a negative shift of the half-inactivation potential of iLVA and increased the lubeluzole-induced block of iLVA. Experiments with extracellular or intracellular application of a quaternary ammonium derivative of lubeluzole (R133121), which was less potent than lubeluzole, support the above conclusions on the side of action of lubeluzole. Application of lubeluzole via the patch pipette affected iLVA and iHVA only minimally compared with extracellular application, probably partly due to efflux of L through the cell membrane. These experiments suggest that lubeluzole blocks Ca²⁺ channels from both the extracellular and the intracellular side.