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Glycerotoxin stimulates exocytosis and endocytosis by increasing intracellular Ca²⁺ in N-type calcium channels expressing cells

We recently purified a novel neurotoxin from Glycera convoluta named Glycerotoxin (GLTx), capable of stimulating neurotransmitter release from N-type Ca²⁺ channels expressing neurons for up to 24h (Schenning et al., 2006). Here, we have found that GLTx also stimulates compensatory endocytosis of synaptic vesicles using styryl dyes and electron microscopy. Furthermore, we have adapted a fluorescent-based assay to monitor intracellular Ca²⁺ flux from both rat brain synaptosomes and human embryonic kidney (HEK) cells over-expressing N, L, P/Q and R-type Ca²⁺ channels. GLTx triggers Ca²⁺ influx in HEK cells expressing rat or human N-type Ca²⁺ channels without affecting cells transfected with L, P/Q or R-type Ca²⁺ channels. In addition, GLTx promoted Ca²⁺ influx in rat brain synaptosomes and an increase in endogenous glutamate released with an EC50 of 50 pM. GLTx is therefore a unique tool available to unravel the mechanism controlling Ca²⁺-regulated exocytosis and compensatory endocytosis via the specific activation of N-type Ca²⁺ channels. Importantly, GLTx was found to act on both rat and human clones of N-type Ca²⁺ channels. GLTx or derivatives could therefore be useful in future human therapy strategies aiming at enhancing neurotransmitter release by selectively stimulating N-type Ca²⁺ channel-expressing neurons.

Schenning, M.P., Proctor, D.T., Ragnarsson, L., Barbier, J., Lavidis, N.A., Molgo, J.J., Zamponi, G.W., Schiavo, G. & Meunier, F.A. (2006) Journal of Neurochemistry 98: 894-904.