Deciphering the function of synaptic release sites is central to understanding neuronal communication. Here, we review studies of the lamprey giant reticulospinal synapse, a model that can be used to dissect synaptic vesicle trafficking at single release sites. The presynaptic axon is large and contains active zones that are spatially separated from each other. During activity, synaptic vesicle membrane is shuttled between the active zone and the periactive zone at which endocytosis occurs. Recent studies have shown that the periactive zone contains an actin-rich cytomatrix that expands during synaptic activity. This cytomatrix has been implicated in multiple functions that include (1) activity-dependent trafficking of proteins between the synaptic vesicle cluster and the periactive zone, (2) synaptic vesicle endocytosis, and (3) the movement of newly formed synaptic vesicles to the vesicle cluster. The actin cytomatrix thus provides a link between the active zone and the periactive zone; this link appears to be critical for sustained cycling of synaptic vesicles.