Abstract
Vesicles within presynaptic terminals are thought to be segregated into a variety of readily releasable and reserve pools. The nature of the pools and trafficking between them is not well understood, but pools that are slow to mobilize when synapses are active are often assumed to feed pools that are mobilized more quickly, in a series. However, electrophysiological studies of synaptic transmission have suggested instead a parallel organization where vesicles within slowly and quickly mobilized reserve pools would separately feed independent reluctant- and fast-releasing subdivisions of the readily releasable pool. Here we use FM-dyes to confirm the existence of multiple reserve pools at hippocampal synapses and a parallel organization that prevents intermixing between the pools, even when stimulation is intense enough to drive exocytosis at the maximum rate. The experiments additionally demonstrate extensive heterogeneity among synapses in the relative sizes of the slowly and quickly mobilized reserve pools, which suggests equivalent heterogeneity in the numbers of reluctant and fast-releasing readily releasable vesicles that may be relevant for understanding information processing and storage.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
The most significant addition is Figure 3, which is an analysis of FM4-64 destaining during 1 Hz stimulation at synapses that are clearly separated spatially from neighbors. We have additionally moved results at body temperature from supplementary information to Figure 12. We did this because Figure 12 shows that the relative sizes of quickly and slowly mobilized reserve pools are not altered by increasing the temperature. This is relevant because so called asynchronous release is eliminated in these cultures near body temperature, so the continued presence of a slowly mobilized reserve counters the concern raised in peer review that slowly releasable reserve vesicles might be primarily released asynchronously.
