Figure 2.

Preconditioning induces presynaptic muting but no detectable postsynaptic changes. A. Images of vGluT-1 positive puncta from representative fields from cultures preconditioned by 4 hr KCl (30 mM) depolarization or by 30 mM NaCl control preconditioning. Middle panels show uptake of FM1-43fx during brief depolarization in the same field. The merged images reveal more inactive (FM1-43fx negative) vGluT-1 positive puncta after depolarizing preconditioning. Green = FM1-43fx. Red = vGluT-1. Red puncta with no green overlap are mute synapses, while yellow indicates overlap and active synapses. White arrowheads indicate examples of co-labeled, active glutamatergic synapses. B. The percentage of FM1-43fx positive (FM (+)) terminals is summarized from 5 experiments like that depicted in A. C. Representative fluorescence images from somatic Ca²⁺ signals measured in control (n = 18 cells in 6 fields) and depolarization-conditioned (n = 15 cells in 5 fields) cells. Cells were loaded with a 30 min bath application of 2 μM Fluo3- FF AM. Representative regions of interest (labeled with “1”) show typical locations of measurement in the soma, chosen based on a phase contrast image (not shown). Dendrites and axons are below the plane of focus. Images of baseline (prior to glutamate perfusion), peak fluorescence (5 s following 10 μM glutamate onset), and return to baseline (30 s wash) fluorescence are shown. Region 2 indicates a region devoid of cells and typical of regions from which background fluorescence was measured. D. Summary of glutamate-evoked somatic intracellular Ca²⁺ signals measured over the entire experiment (67 images over 40 seconds) with control-conditioned and depolarization-conditioned cells overlaid.