Is replenishment of the readily releasable pool associated with vesicular movement?

At the excitatory synapse of rat hippocampus the short-term synaptic depression observed during long high-frequency stimulation is associated with slower replenishment of the readily-releasable pool. Given that the replenishment rate is also not [Ca⁺⁺]_o sensitive this puts into question a widely held notion that the vesicles—constrained by the cytoskeleton and rendered free from such constraints by Ca⁺⁺ entry that renders them more mobile—are important in the replenishment of the readily-releasable pool. This raises a question—Is vesicular replenishment of the readily releasable pool associated with significant movement? To answer this question we evaluated how okadaic acid and staurosporine (compounds known to affect vesicular mobility) influence the replenishment rate. We used patterned stimulation on the Schaffer collateral fiber pathway and recorded the excitatory post-synaptic currents (EPSCs) from rat CA1 neurons, in the absence and presence of these drugs. The parameters of a circuit model with two vesicular pools were estimated by minimizing the squared difference between the ESPC amplitudes and simulated model output. [Ca²⁺]_o did not influence the progressive decrease of the replenishment rate during long, high frequency stimulation. Okadaic acid did not significantly affect any parameters of the vesicular storage and release system, including the replenishment rate. Staurosporine reduced the replenishment coupling, but not the replenishment rate, and this is owing to the fact that it also reduces the ability of the readily releasable pool to contain quanta. Moreover, these compounds were ineffective in influencing how the replenishment rate decreases during long, high frequency stimulation. In conclusion at the excitatory synapses of rat hippocampus the replenishment of the readily releasable pool does not appear to be associated with a significant vesicular movement, and during long high frequency stimulation [Ca⁺⁺]_o does not influence the progressive decrease of vesicular replenishment.