Activation of heterologously expressed Drosophila TRPL channels:: Ca2+ is not required and InsP3 is not sufficient

Light-sensitive channels encoded by the Drosophila transient receptor potential-like gene (trpl) are activated in situ by an unknown mechanism requiring activation of G(q) and phospholipase C (PLC). Recent studies have variously concluded that heterologously expressed TRPL channels are activated by direct G(q)-protein interaction, InsP(3) or Ca2+. In an attempt to resolve this confusion we have explored the mechanism of activation of TRPL channels coexpressed with a PLC-specific muscarinic receptor in a Drosophila cell line (S2 cells). Simultaneous whole-cell recordings and ratiometric Indo-1 Ca2+ measurements indicated that agonist (CCh)-induced activation of TRPL channels was not always associated with a rise in Ca2+. Internal perfusion with BAPTA (10 mM) reduced, but did not block, the response to agonist. In most cases, releasing caged Ca2+ facilitated the level of spontaneous channel activity, but similar concentrations (200-500 nM) could also inhibit TRPL activity. Releasing caged InsP(3) invariably released Ca2+ from internal stores but had only a minor influence on TRPL activity and none at all when Ca2+ release was buffered with BAPTA. Caged InsP(3) also failed to activate any light-sensitive channels in situ in Drosophila photoreceptors. Two phospholipase C inhibitors (U-73122 4 muM and bromo-phenacyl bromide 50 muM) reduced both spontaneous and agonist-induced TRPL activity in S2 cells. The results suggest that, as in situ, TRPL activation involves G-protein and PLC; that Ca2+ can both facilitate and in some cases inhibit TRPL channels, but that neither Ca2+ nor InsP(3) is the primary activator of the channel.