Susceptibility of cerebellar granule neurons from GM2/GD2 synthase-null mice to apoptosis induced by glutamate excitotoxicity and elevated KCl: Rescue by GM1 and LIGA20

Our previous study showed an impaired regulation of Ca2+ homeostasis in cultured cerebellar granule neurons (CGN) from neonatal mice lacking GM2, GD2 and all gangliotetraose gangliosides, due to disruption of the GM2/GD2 synthase (GalNAc-T) gene. In the presence of depolarizing concentration (55 mM) K+, these cells showed persistent elevation of intracellular Ca2+ ([Ca2+](i)) leading to apoptosis and cell destruction. This was in contrast to CGN from normal littermates whose survival was enhanced by high K+. In this study we demonstrate that glutamate has the same effect as K+ on CGN from these ganglioside-deficient knockout (KO) mice and that apoptosis in both cases is averted by exogenous GM1. Even more effective rescue was obtained with LIGA20, a semi-synthetic derivative of GM1. LC50 of glutamate in the KO cells was 3.1 muM, compared to 46 muM in normal CGN. [Ca2+](i) measurement with fura-2 revealed no difference in glutamate-stimulated Ca2+ influx between the 2 cell types. However, reduction of [Ca2+](i) following application of Mg2+ was significantly impaired in the mutant CGN. The rescuing effects of exogenous GM1 and LIGA20 corresponded to their ability to restore Ca2+ homeostasis. The greater potency of LIGA20 is attributed to its greater membrane permeability with resultant ability to insert into both plasma and nuclear membranes at low concentration (less than or equal to1 muM); GM1 at the same concentration was incorporated only into the plasma membrane and required much higher concentration to influence Ca2+ homeostasis and CGN viability.