Pharmacological Dissection of the Crosstalk between Na(V) and Ca(V) Channels in GH3b6 Cells

Thanks to the crosstalk between Na(+) and Ca(2+) channels, Na(+) and Ca(2+) homeostasis interplay in so-called excitable cells enables the generation of action potential in response to electrical stimulation. Here, we investigated the impact of persistent activation of voltage-gated Na(+) (Na(V)) channels by neurotoxins, such as veratridine (VTD), on intracellular Ca(2+) concentration ([Ca(2+)](i)) in a model of excitable cells, the rat pituitary GH3b6 cells, in order to identify the molecular actors involved in Na(+)-Ca(2+) homeostasis crosstalk. By combining RT-qPCR, immunoblotting, immunocytochemistry, and patch-clamp techniques, we showed that GH3b6 cells predominantly express the Na(V)1.3 channel subtype, which likely endorses their voltage-activated Na(+) currents. Notably, these Na(+) currents were blocked by ICA-121431 and activated by the β-scorpion toxin Tf2, two selective Na(V)1.3 channel ligands. Using Fura-2, we showed that VTD induced a [Ca(2+)](i) increase. This effect was suppressed by the selective Na(V) channel blocker tetrodotoxin, as well by the selective L-type Ca(V) channel (LTCC) blocker nifedipine. We also evidenced that crobenetine, a Na(V) channel blocker, abolished VTD-induced [Ca(2+)](i) elevation, while it had no effects on LTCC. Altogether, our findings highlight a crosstalk between Na(V) and LTCC in GH3b6 cells, providing a new insight into the mode of action of neurotoxins.