Sub-cellular Ca2+ dynamics affected by voltage- and Ca2+-gated K+ channels: Regulation of the soma-growth cone disparity and the quiescent state in Drosophila neurons

Using Drosophila mutants and pharmacological blockers, we provide the first evidence that distinct types of K+ channels differentially influence sub-cellular Ca2+ regulation and growth cone morphology during neuronal development. Fura-2-based imaging revealed in cultured embryonic neurons that the loss of either voltage-gated, inactivating Shaker channels or Ca2+-gated Slowpoke BK channels led to robust spontaneous Ca2+ transients that preferentially occurred within the growth cone. In contrast, loss of voltage-gated, non-inactivating Shab channels did not show such a disparity and sometimes produced soma-specific Ca2+ transients. The fast spontaneous transients in both the soma and growth cone were suppressed by the Na+ channel blocker tetrodotoxin, indicating that these Ca2+ fluctuations stemmed from increases in membrane excitability. Similar differences in regional Ca2+ regulation were observed upon membrane depolarization by high K+-containing saline. In particular, Shaker and slowpoke mutations enhanced the size and dynamics of the depolarization-induced Ca2+ increase in the growth cone. In contrast, Shab mutations greatly prolonged the Ca2+ increase in the soma. Differential effects of these excitability mutations on neuronal development were indicated by their distinct alterations in growth cone morphology. Loss of Shaker currents increased the size of lamellipodia and the number of filopodia, structures associated with the actin cytoskeleton. Interestingly, loss of Slowpoke currents strongly influenced tubulin regulation, enhancing the number of microtubule loop structures per growth cone. Together, our findings support the idea that individual K+ channel subunits differentially regulate spontaneous sub-cellular Ca2+ fluctuations in growing neurons that may influence activity-dependent growth cone formation.