Fig. 6. Exemplary EPSC amplitude distribution used for estimation of quantal size. Amplitude distribution for evoked EPSCs occurring in the AF response during a sustained depolarization of the IHC from -89 to -29 mV in the presence of CTZ (Recording 6). All events in the second half of a 1-s pulse, n = 280, were selected to construct the distribution. The distribution had a modal peak at 39 pA and was skewed toward larger amplitudes of several hundreds of pA. The average amplitude was 117.1 ± 4.2 pA (coefficient of variation of 59.5%). This distribution in CTZ is very reminiscent of the results by Glowatzki and Fuchs (1), who recorded without CTZ, in several aspects: shape of the distribution, modal peak value (30-36 pA), and average amplitude.

1. Glowatzki E, Fuchs PA (2002) Nat Neurosci 5:147-154.

Fig. 7. Estimation of channel-vesicle distance by local nonequilibrium approximation calculation of [Ca²⁺] spread. An estimate of the Ca²⁺ signal spread in the cell can be obtained by the local nonequilibrium approximation (1) as a function of the distance from the Ca²⁺ source with the following equation:

where D[Ca²⁺] is the change in Ca²⁺ concentration, f is the Ca²⁺ flux through a channel in units of mol/s, D_Ca is the diffusion of free Ca²⁺, k_B is the rate constant of Ca²⁺ binding to buffer, [B] is the concentration of free buffer,_{and^{r is the distance from the Ca2+ source. This provides an estimate of the Ca2+ concentration at different locations around the channel and of how the buffering affects the spread. Additionally, the amount of release (as a function of the fourth power of [Ca2+]) can be evaluated from this calculation, again, at different distances from the channel. Specifically, we are interested in the relative release as a ratio between our standard buffer condition 1 mM EGTA and test conditions 5 mM EGTA and 5 mM BAPTA. This theoretical calculation tells us what fraction of the release process is expected to be affected by a certain buffer condition (relative to the standard) when a fixed distance, r, is assumed between Ca2+ source and sensor (no assumption are required upon the number of channels necessary to open to produce the release event). Or taking the opposite approach, we can estimate the distance by using the experimental result that we found for the effect of 5 mM EGTA or 5 mM BAPTA relative to 1 mM EGTA (at steady state, Fig. 4B). Considering a 56% reduction in 5 mM EGTA, the calculated distance was ~ 23 nm. When the same procedure is applied for the effect of 5 mM BAPTA, the number is even smaller.}}

1. Neher E (1998) Cell Calcium 24:345-357.