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Matthew Van Hook, Wallace Thoreson; Calmodulin regulates vesicle replenishment and shapes kinetics of synaptic transfer from cones. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1757.
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© ARVO (1962-2015); The Authors (2016-present)
Dynamics of synaptic depression and vesicle replenishment at ribbon synapses enable photoreceptors and bipolar cells to signal changes in contrast and luminance to other retinal neurons. Like many other synapses, the rate of vesicle replenishment in cones is regulated by Ca2+ but the mechanisms are unclear. Additionally, photoreceptor signaling to second-order neurons is bandpass with a peak at ~3 Hz. Replenishment has been proposed as a major determinant of this feature of synaptic transfer. Here, we test a role for calmodulin (CaM) in vesicle replenishment and synaptic transfer in cones.
Whole-cell recordings from cones, horizontal cells (HCs), and bipolar cells (BCs) were performed with retinal slices from tiger salamanders. Using paired recordings, replenishment was measured with a paired pulse protocol or with trains of depolarizing pulses (25 ms, 13.3 Hz) to deplete the releasable pool and reach an equilibrium state where release is rate-limited by vesicle replenishment. In this protocol, the cumulative release rate provides a measure of the replenishment rate. Electroretinograms (ERGs) were recorded intraretinally from superfused eyecups with L-AP4 (10 μM) to isolate the d-wave (Off BC responses).
In paired recordings from cones and HCs, inhibition of CaM by introduction of the peptide MLCK (20 μM) through the cone patch pipette slowed recovery from paired-pulse depression, especially with shorter inter-stimulus intervals. MLCK also inhibited Ca2+-dependent acceleration of replenishment measured with trains of depolarizing pulses. Bath-application of the CaM inhibitor calmidazolium (20 μM) inhibited post-synaptic currents at light offset after short but not long flashes in HCs and Off BCs, consistent with a role for CaM in fast vesicle replenishment. Similarly, ERG d-waves were inhibited by calmidazolium following short, but not long flashes. HC and BC responses to sinusoidal light stimuli were bandpass, with a peak at ~2 Hz. In the presence of calmidazolium, the response peak was shifted to 0.5 Hz.
These results suggest that Ca2+-dependent acceleration of replenishment involves CaM and shapes the synaptic transfer kinetics at cone synapses. Thus, in addition to roles in signaling luminance and contrast, replenishment at the photoreceptor synapse is important for signaling the timing of visual stimuli to second-order neurons.
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