Figure 4.

Cleft alkalinization is synchronous with Ca²⁺ extrusion from adjacent synaptic compartments. A, NMJ cross-section, showing the distribution of coincidence detector 1.0 construct expressed using the muscle driver 24B-GAL4. B, The topology and composition of the construct relative to the cleft. C, D, Images of coincidence detector 1.0 at a live NMJ (MN6/7-Ib): (C) SE-pHluorin fluorescence and (D) jRGECO1a fluorescence imaged 0.3 s before, 0.09 s after, and 0.3 s after a nerve stimulus. Each image represents an 8 ms exposure averaged from 50 trials (50 stimuli delivered at 0.6 Hz). The look-up-table (LUT) used in C and D is shown in C. E, Fluorescence image from an HRP antibody conjugated with DyLight405 at the live NMJ. F, Plots represent the fluorescence intensity of SE-pHluorin and jRGECO1a at the terminals in C and D, minus background, at each sampling interval over 10 stimuli (fluorescence from each fluorophore was collected on the EMCCD camera at 56 Hz and interdigitated with the other fluorophore). G, Average response (%ΔF/F_rest) of each fluorophore to a single nerve stimulus (data from C and D, i.e., all 50 stimuli), where F_rest is the average fluorescence value immediately before each nerve stimulus. H, Data from G normalized to a peak value of 1. I, Data from H plotted on an expanded time scale at the time of nerve stimulus. 2 mm Ca²⁺, 20 mm Mg²⁺, and 100 μm ryanodine used throughout.