the voltage-dependent K+ conductance changes with development Stem Cell Compound Library chemical structure (Marcotti et al., 2003), its adult value was measured in P18 animals (Figures 8C–8E). The conductance-voltage relationships could be fit with a single Boltzmann (Figure 8E) with GMAX = 470 ± 96 nS, V0.5 = −31 ± 3 mV and VS = 10.5 ± 3.5 mV (n = 5). The K+ conductance is larger than in OHCs and when combined with the smaller standing MT conductance suggests a more hyperpolarized resting potential than in OHCs. The resting potential was determined in two ways as for OHCs. During current clamp recordings in isolated cochleas of P18 animals (Figures 8F–8H), perfusing 0.02 mM Ca2+ depolarized the IHC from −70 ± Akt inhibitor 3 mV (n = 4) to −59 ± 3 mV and reduced the membrane time constant from 1.08 ± 0.05 ms in 1.3 mM Ca2+ to 0.70 ± 0.06 ms in 0.02 mM Ca2+. A second method was to apply Equation 1, using 5.7 nS for the resting MT conductance, and determining which membrane potential, VR, satisfied the equation for each of the measured GK-V relationships; EK was assumed to be −75 mV. This calculation improves on the direct recording by taking into account the endolymphatic potential and thus predicting IHC properties in vivo. The resting potential was calculated as −55 ± 2 mV (n = 5) comparable to that obtained by direct measurement in the isolated
cochlea. With the measured IHC capacitance (12.5 ± 0.5 pF), the membrane time constant was 0.26 ± 0.03 ms (n = 5), equivalent to a corner frequency of 0.61 kHz, which is similar to that found in vivo (Palmer and Russell, 1986). The difficulties of recording from and directly
stimulating OHCs in the in vivo cochlea has motivated work on isolated pieces of the organ of Corti or cochlear slices in which large transduction currents can be obtained from single hair cells (Kros et al., 1992, Kennedy et al., 2003 and He et al., 2004). However, because the organ of Corti is a tight epithelium dividing two fluid compartments with distinct ionic compositions, use of isolated preparations has the drawback that the environmental conditions usually differ from those in vivo: the hair bundles are not exposed to endolymph containing low, 20–40 μM, Ca2+, the 90 mV endolymphatic potential across the epithelium is absent and, to prolong the viability of the preparation, Non-specific serine/threonine protein kinase measurements are mostly made at room temperature. We therefore corrected for these differences with the justification that OHC MT currents obtained in isolated preparations of younger animals (P7–P13) are the best currently achievable (Kennedy et al., 2003). Our results showed that OHCs have a relatively depolarized resting potential (−30 to −40 mV), based both on direct current-clamp measurements near body temperature in animals around the onset of hearing (P11–P13), and from extrapolations to the mature in vivo condition (P16–P19).