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Proleta Datta, Leigh Latham, Roger Janz, Ruth Heidelberger; Functional Organization of Synaptic Vesicle Pools in a Retinal Bipolar Cell. Invest. Ophthalmol. Vis. Sci. 2011;52(14):4805.
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Exocytosis of neurotransmitter occurs via a calcium-dependent, SNARE (Soluble N-ethylmaleimide sensitive factor Attachment Protein Receptor) complex -mediated process. In Mb1 bipolar cells, three kinetic components of exocytosis have been reported. These have been attributed to: i) fusion of synaptic vesicles that are docked at the plasma membrane and tethered to the synaptic ribbon (the rapidly-releasing pool, RRP), ii) fusion of the entire ribbon-tethered vesicle pool (the releasable pool, RP) and iii) the recruitment of cytoplasmic reserve vesicles. As a test of this hypothesis, we asked: 1) is the RRP a subset of the RP, such that these pools fuse in series rather than in parallel, and 2) which vesicle pools are associated with SNARE complexes.
Membrane capacitance and calcium currents were monitored in isolated terminals of goldfish Mb1 bipolar cells. To block refilling of the RP, terminals were voltage clamped and the whole-terminal pipette solution contained ATPγS in place of ATP. To probe the association with SNARE complexes, the pipette solution contained a SNARE complex inhibiting peptide (SCIP) or a scrambled control peptide.
First, 1s pulses were given to deplete and probe the state of the RP. Then a 20ms pulse was given to probe the RRP. In ATPγS terminals, but not controls, depletion of the RP resulted in the complete loss of the RRP. In a second set of experiments, 20ms pulses were given to assess the RRP, followed by a 1s pulse to assess the RP. In ATPγS terminals, but not controls, the RRP failed to refill after 4-5 rounds of release, and the RP was significantly depressed. In a third set of experiments, terminals were stimulated with a pulse train that revealed all three components of release. In ATPγS terminals, only the first two components of release were observed; the component attributed to the recruitment of reserve vesicles was blocked. Similar results were seen in SCIP terminals. In the presence of ATPγS or SCIP, significant reduction in the RRP and RP were observed with subsequent trains.
1) The RRP is a subset of the RP and draws from the RP 2) Both the RRP and the RP are initially protected from the SCIP, possibly because both pools are associated with preformed SNARE complexes. These data support a serial organization of vesicle pools that undergo SNARE complex-mediated exocytosis.
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