Article Figures & Data
Figures
- Fig. 1.
Ca2+-dependent exocytosis can be evoked by depolarizations long before the onset of hearing.A shows [Ca2+]i(top panel) and Cm(bottom panel) of an IHC from a P2 mouse at low time resolution. In response to long depolarizations (200 and 500 msec), sizable increases in Cm (exocytosis) and in [Ca2+]i were observed. Shorter depolarizations resulted in much smaller responses (arrow, 50 msec). The subsequent decline inCm most likely reflects endocytic membrane retrieval. B, Cm (top panel) and membrane current (bottom panel) of an IHC from a P6 mouse in response to 100 msec long depolarizations to −5 mV in the presence or absence of extracellular Ca2+ (Ca2+-free Ringer containing 2 mm Ca2+-free EGTA and 3 mm MgCl2).Cm increments were inhibited by abolition of the Ca2+ influx and restored on readdition of Ca2+. C, Representative Ca2+ currents (bottom panel) and Cm increments (top panel) of IHCs from different developmental stages to a 50 msec long depolarization to −5 mV. Baseline capacitance values are indicated below each trace.
- Fig. 2.
Developmental changes in Ca2+current and exocytosis. A demonstrates the increase in basal cell capacitance during the course of maturation (P0,n = 13; P2, n = 2; P4,n = 10; P6, n = 10; P10,n = 9; and P14–P25, n = 28).B displays normalized ΔCm-responses (top panel) to 20 or 100 msec long depolarizations to −5 mV (number of cells as in A, normalized to the average responses of P6 cells to either depolarization). The bottom panel shows the Ca2+ current densities for 20 and 100 msec depolarizations (peak Ca2+ currents normalized to the cell capacitance). The arrow indicates the onset of hearing. The diagrams depict the preferential abundance of multiple spherical bodies at the active zones of immature IHCs and the typical presence of only one plate-like ribbon at the mature afferent synapse. C displays representative current voltage relationships for three different developmental stages after subtracting the leak current (estimated by a linear fit to the hyperpolarized portion of the data).
- Fig. 3.
Changes in secretion kinetics during maturation. Secretion kinetics was investigated by measuring ΔCm to depolarizations (to −5 mV) of different duration applied in random order. A plots the mean Cm responses recorded in the perforated-patch configuration in IHCs from the different stages (n = 13 cells for P0; n = 10 for P6; n = 9 for P10; and n = 28 for IHCs from hearing mice). B displays the first 30 msec of stimulation with higher resolution. C compares the ΔCm of whole-cell experiments on IHCs from P6 and hearing mice in which we added a mixture of 5 mm Ca2+-free EGTA and 5 mmCa2+-loaded EGTA to the pipette (n = 9 cells for P6; n = 27 for IHCs from P14–P25). The solid lines represent double exponential fits to the data to estimate size and release time constant of the readily releasable pool (see text).
- Fig. 4.
Ca2+ dependence of exocytosis during and after maturation. The plot relates secretory responses of 5 mm EGTA-buffered IHCs from P6 and P14–P25 mice to their Ca2+ current integrals (restricted to small integrals; n = 9 cells for P6 andn = 27 for P14–P25 IHCs). The dashed lines represent linear fits to the first three points of each data set.
- Fig. 5.
Spontaneous action potentials trigger exocytosis and increase [Ca2+]i in immature IHCs.A displays an AP train recorded in the whole-cell configuration in the absence of any holding current from a representative P6 IHC. In this particular cell, spontaneous activity was observed during the first 260 sec of the whole-cell recording.B displays two spontaneous APs from the same cell at higher time resolution. C shows a low-time resolution plot of [Ca2+]i and membrane potential (Vm) of an IHC in which we applied 10 pA of depolarizing current for short intervals (indicated bybars) after spontaneous activity had ceased.D shows an average of three secretory responses of two IHCs from a P6 mouse to a single action potential. We used the voltage-clamp mode to measure capacitance before and after an AP-like voltage-waveform to −10 mV. The time periods ofCm measurements can be recognized from the band-like (sinusoidal) signals in the voltage and currenttraces.
In this issue
