Synapsin I-associated Phosphatidylinositol 3-Kinase Mediates Synaptic Vesicle Delivery to the Readily Releasable Pool

Abstract

Maintaining synaptic transmission requires replenishment of docked synaptic vesicles within the readily releasable pool (RRP) from synaptic vesicle clusters in the synapsin-bound reserve pool. We show that synapsin forms a complex with phosphatidylinositol 3-kinase (PI 3-kinase) in intact nerve terminals and that synapsinassociated kinase activity increases on depolarization. Disruption of either PI 3-kinase activity or its interaction with synapsin inhibited replenishment of the RRP, but did not affect exocytosis from the RRP. Thus we conclude that a synapsin-associated PI 3-kinase activity plays a role in synaptic vesicle delivery to the RRP. This also suggests that PI 3-kinase contributes to the maintenance of synaptic transmission during periods of high activity, indicating a possible role in synaptic plasticity.A typical nerve terminal in the central nervous system contains about 200 -250 synaptic vesicles (SVs).1 These can be functionally divided into a small readily releasable pool (RRP) and a large reserve pool. The RRP contains less than 5% of total nerve terminal SVs and is defined morphologically as predocked SVs at the active zone and functionally as SVs that are primed and immediately available for the initial rapid phase of neurotransmitter release (1). The remainder constitutes the reserve pool, which ensures that a continual supply of SVs is available for delivery to the RRP and is drawn upon to maintain neurotransmission during periods of intense or prolonged stimulation (2-4). The reserve pool SVs surround the active zone and are clustered together in an actin-based cytoskeletal matrix via an interaction with synapsins (3, 5).There are five synapsin proteins (Ia, Ib, IIa, IIb, and III) from three genes (I, II, and III) (6), which maintain the reserve pool SVs as a cluster and prevent their free dispersal within the terminal. Synapsins I and II control SV replenishment of the RRP, because conditions that disrupt their function result in dispersal or depletion of SV clusters near the active zone and enhance synaptic depression (3, 5, 7). In contrast, disruption of the much less abundant synapsin III does not affect SV clusters and reduces synaptic depression (8). SV liberation from clusters within the actin cytomatrix is mediated by the multi-site phosphorylation of synapsin I and II, which decreases their affinity for SVs or actin (6). Disruption of synapsin I phosphorylation sites thus reduces the number of SVs undergoing exocytosis (9). It is unknown how liberated SVs translocate to the RRP, and the mechanisms governing entry into the RRP (docking/priming) are incompletely understood (10).Synapsin I has a C-terminal proline-rich domain that interacts with src homology 3 (SH3) domains present in a large number of proteins, including the p85 subunit of phosphatidylinositol 3-kinase (PI 3-kinase) (6, 11). A precise role for PI 3-kinase in nerve terminal function has not been established (12)(13)(14), despite that its lipid products play numerous roles the actin cytoskeleton and in vesicle traffic...

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