Ca²⁺ dynamics along identified synaptic terminals in Drosophila larvae

Changes in intracellular Ca²⁺ concentration ([Ca ²⁺]_i) play an important role in the function and plasticity of synapses. We characterized the changes in [Ca²⁺] _i produced by action potentials (APs) along two identified motor terminals found on separate muscle fibers in Drosophila larvae and examined factors that influence the amplitude and duration of the residual Ca ²⁺ signal. We were able to measure Ca²⁺ transients produced along terminals by both single APs and AP trains using Oregon Green 488 BAPTA-1 and streaming images at 20-50 Hz. The decay of [Ca²⁺] _i after single APs or AP trains was well fit by a single exponential. For single APs, the Ca²⁺ transient amplitude and decay rate were similar at boutons and bottleneck regions and much smaller at the axon. Also, the amplitude of single-AP Ca²⁺ transients was inversely correlated with bouton width. During AP trains, the increase in [Ca²⁺] _i became more uniform: the difference in boutons and axons was reduced, and the increase in [Ca²⁺]_i was not correlated with bouton width. The [Ca²⁺]_i decay π was directly correlated with bouton width for both single APs and AP trains. For one terminal, distal boutons had larger single-AP Ca²⁺ transients than proximal ones, probably attributable to greater Ca²⁺ influx for distal boutons. Pharmacological studies showed that Ca²⁺ clearance from these synaptic terminals after single APs and AP trains was primarily attributable to Ca²⁺ extrusion by the plasma membrane Ca²⁺ ATPase (PMCA). Immunostaining of larval muscle fibers showed high levels of the PMCA at the neuromuscular junction.