Lamotrigine increases intracellular Ca2+ levels and Ca2+/calmodulin-dependent kinase II activation in mouse dorsal root ganglion neurones

Abstract

Aim Lamotrigine is a neuroprotective agent that is used clinically for the treatment of seizures and neuropathic pain. A significant volume of literature has reported that lamotrigine exerts analgesic effect by blocking Ca2+ channels. However, little is known regarding the effect of lamotrigine on the intracellular Ca2+ concentration ([Ca2+]i). The aim of this study was to determine whether lamotrigine modulates [Ca2+]i in sensory neurones. Methods Lamotrigine-induced changes in [Ca2+]i were measured in mouse dorsal root ganglion (DRG) neurones using the Ca2+-sensitive fluorescent indicator Fluo 3-AM and a confocal laser scanning microscope. Ca2+/calmodulin-dependent kinase II (CaMKII) activation was assessed by the fluorescence intensity using immunocytochemical procedures. Results Treatment with 1, 10, 30 or 100 mu m lamotrigine transiently increased [Ca2+]i in DRG neurones in a dose-dependent manner. Treatment with 100 mu m lamotrigine induced a significant (threefold) increase in the Ca2+ peak in the presence or absence of extracellular Ca2+. The lamotrigine-induced Ca2+ increase was abolished or decreased by the treatment with a specific PLC inhibitor (U73122), IP3R antagonist (xestospongin C) or RyR antagonist (dantrolene). In some cells, treatment with 100 mu m lamotrigine caused a transient Ca2+ increase, and the Ca2+ levels quickly fell to below the basal Ca2+ level observed prior to lamotrigine application. The decrease in basal Ca2+ levels was blocked by the treatment with a CaMKII inhibitor (KN93). Immunocytochemical analysis indicated that lamotrigine treatment increased the expression of phosphorylated CaMKII in DRG neurones. Conclusion Treatment with lamotrigine increased [Ca2+]i apparently as a result of Ca2+ release from intracellular stores and CaMKII activity.