Drosophila Ca(V)2 channels harboring human migraine mutations cause synapse hyperexcitability that can be suppressed by inhibition of a Ca(2+) store release pathway

Gain-of-function mutations in the human Ca(V)2.1 gene CACNA1A cause familial hemiplegic migraine type 1 (FHM1). To characterize cellular problems potentially triggered by Ca(V)2.1 gains of function, we engineered mutations encoding FHM1 amino-acid substitutions S218L (SL) and R192Q (RQ) into transgenes of Drosophila melanogaster Ca(V)2/cacophony. We expressed the transgenes pan-neuronally. Phenotypes were mild for RQ-expressing animals. By contrast, single mutant SL- and complex allele RQ,SL-expressing animals showed overt phenotypes, including sharply decreased viability. By electrophysiology, SL- and RQ,SL-expressing neuromuscular junctions (NMJs) exhibited enhanced evoked discharges, supernumerary discharges, and an increase in the amplitudes and frequencies of spontaneous events. Some spontaneous events were gigantic (10–40 mV), multi-quantal events. Gigantic spontaneous events were eliminated by application of TTX–or by lowered or chelated Ca(2+)–suggesting that gigantic events were elicited by spontaneous nerve firing. A follow-up genetic approach revealed that some neuronal hyperexcitability phenotypes were reversed after knockdown or mutation of Drosophila homologs of phospholipase Cβ (PLCβ), IP(3) receptor, or ryanodine receptor (RyR)–all factors known to mediate Ca(2+) release from intracellular stores. Pharmacological inhibitors of intracellular Ca(2+) store release produced similar effects. Interestingly, however, the decreased viability phenotype was not reversed by genetic impairment of intracellular Ca(2+) release factors. On a cellular level, our data suggest inhibition of signaling that triggers intracellular Ca(2+) release could counteract hyperexcitability induced by gains of Ca(V)2.1 function.