, 2009), indicative of strong Ca2+ clearance followed by an integrating increase in signal. Sites distal to the synapse show little change initially, consistent with the source of Ca2+ being at a distance, followed by a slow increase. The later signals (Figures 7I–7K), corresponding to the onset of the superlinear response, present
a different picture. Ca2+ at the synapse shows an abrupt VE-821 cost increase in signal, followed by a plateau and decrease in signal even in the face of constant Ca2+ entry (Figures 7D and 7J). Ca2+ signals away from the synapse show a slower integrating signal followed by a sudden increase in signal whose rate is faster and peak greater than that observed at the synaptic region (Figure 7J). The change in kinetics at these distant sites suggests a secondary source of Ca2+. Similar results
were obtained in five cells where the superlinear release component was observed. Smaller depolarizations revealed simple integrating responses that diminished away from the synapse while larger depolarizations yielded similar complex responses (Figure S7). Together these data suggest learn more that Ca2+ dynamics are complex, clearance near the synapse is strong, and a second source of Ca2+ may play a role in vesicle trafficking. Also, although the second component of release appears to be superlinear when compared to the Ca2+ integral, indicating more release per Ca2+
for the second component, when compared to the Ca2+ fluorescent signal the opposite is true. By using fluorescence changes at the synapse, the ratio (Cap/Fluor) of the first component divided by the second component provided an indicator of relative efficiency of release and was 1.5 ± 0.4 (n = 3), indicating that release was more efficient at lower values of Ca2+. Utilization MRIP of a two-sine technique for real-time tracking of vesicle fusion has allowed for more detailed investigation of presynaptic release components at the auditory hair cell-afferent fiber synapse. By using stimuli that did not elicit maximal ICa, saturable pools were clearly identified, whereas variability between and within cells made this impossible (in turtle) with the single-sine technique (Schnee et al., 2005). A superlinear release component whose onset varied with Ca2+ load and correlated with release of an additional source of Ca2+ was also revealed. The superlinear component of release is postulated to reflect the ability of hair cells to rapidly recruit vesicles from regions distant from the synapse, which may underlie the inability to deplete vesicle pools and the ability of hair cell synapses to operate at high rates for sustained periods of time.
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