Masetto, Sergio and Manning J. Correia. Electrophysiological properties of vestibular sensory and supporting cells in the labyrinth slice before and during regeneration. J. Neurophysiol. 78: 1913–1927, 1997. The whole cell patch-clamp technique in combination with the slice preparation was used to investigate the electrophysiological properties of pigeon semicircular canal sensory and supporting cells. These properties were also characterized in regenerating neuroepithelia of pigeons preinjected with streptomycin to kill the hair cells. Type II hair cells from each of the three semicircular canals showed similar, topographically related patterns of passive and active membrane properties. Hair cells located in the peripheral regions (zone I, near the planum semilunatum) had less negative resting potentials [0-current voltage in current-clamp mode ( V z) = −62.8 ± 8.7 mV, mean ± SD; n = 13] and smaller membrane capacitances ( C m = 5.0 ± 0.9 pF, n = 14) than cells of the intermediate (zone II; V z = −79.3 ± 7.5 mV, n = 3; C m = 5.9 ± 1.2 pF, n = 4) and central (zone III; V z = −68.0 ± 9.6 mV, n = 17; C m = 7.1 ± 1.5 pF, n = 18) regions. In peripheral hair cells, ionic currents were dominated by a rapidly activating/inactivating outward K+ current, presumably an A-type K+ current ( I KA). Little or no inwardly rectifying current was present in these cells. Conversely, ionic currents of central hair cells were dominated by a slowly activating/inactivating outward K+ current resembling a delayed rectifier K+ current ( I KD). Moreover, an inward rectifying current at voltages negative to −80 mV was present in all central cells. This current was composed of two components: a slowly activating, noninactivating component ( I h), described in photoreceptors and saccular hair cells, and a faster-activating, partially inactivating component ( I K1) also described in saccular hair cells in some species. I h and I K1 were sometimes independently expressed by hair cells. Hair cells located in the intermediate region (zone II) had ionic currents more similar to those of central hair cells than peripheral hair cells. Outward currents in intermediate hair cells activated only slightly more quickly than those of the cells of the central region, but much more slowly than those of the peripheral cells. Additionally, intermediate hair cells, like central hair cells, always expressed an inward rectifying current. The regional distribution of outward rectifying potassium conductances resulted in macroscopic currents differing in peak–to–steady state ratio. We quantified this by measuring the peak ( G p) and steady-state ( G s) slope conductance in the linear region of the current-voltage relationship (−40 to 0 mV) for the hair cells located in the different zones. G p/ G s average values (4.1 ± 2.1, n = 15) from currents in peripheral hair cells were higher than those from intermediate hair cells (2.3 ± 0.8, n = 4) and central hair cells(1.9 ± 0.8, n = 21). The statistically significant differences ( P < 0.001) in G p/ G s ratios could be accounted for by KA channels being preferentially expressed in peripheral hair cells. Hair cell electrophysiological properties in animals pretreated with streptomycin were investigated at ∼3 wk and ∼9–10 wk post injection sequence (PIS). At 3 wk PIS, hair cells (all zones combined) had a statistically significantly ( P < 0.001) lower C m (4.6 ± 1.1 pF, n = 24) and a statistically significantly ( P < 0.01) lower G p(48.4 ± 20.8 nS, n = 26) than control animals ( C m = 6.2 ± 1.6 pF, n = 36; G p = 66 ± 38.9 nS, n = 40). Regional differences in values of V z, as well as the distribution of outward and inward rectifying currents, seen in control animals, were still obvious. But, differences in the relative contribution of the expression of the different ionic current components changed. This result could be explained by a relative decrease in I KA compared with I KD during that interval of regeneration, which was particularly evident in peripheral hair cells. At 9–10 wk PIS, hair cells of all zones had membrane properties not statistically different ( P > 0.5) from those in untreated normal animals. C m was 6.1 ± 1.3 pF ( n = 30) and G p was 75.9 ± 36.6 nS ( n = 30). Thus it appears that during regeneration, avian semicircular canal type II hair cells are likely to recover all their functional properties. At 9–10 wk PIS, regenerated hair cells expressed the same macroscopic ionic currents and had the same topographic distribution as normal hair cells. Measurements obtained at 3 wk PIS suggest that regenerated hair cells come from smaller cells (smaller mean values of C m) endowed with fewer potassium channels (smaller mean values of G p). In addition, differences observed in peripheral hair cells' kinetics and G p/ G s ratios at 3 wk PIS suggest that different ionic channels follow different schedules of expression during hair cell regeneration. We recorded from nine supporting cells both in normal ( n = 5) and regenerating ( n = 4) epithelia. These cells had an average negative resting potential of V z = −49.5 ± 14.1 mV ( n = 9), but no obvious sign of voltage- and time-dependent ionic currents, except for a very weak inward rectification at very negative potentials, both in normal and streptomycin-recovering animals. Therefore, if all semicircular canal supporting cells are like the small sample we tested and if supporting cells are actually the progenitors of regenerating hair cells, then they must change shape, develop hair bundles, become reinnervated, and also acquire a complete set of ionic channels ex novo.
Expression and Localization of Ryanodine Receptors in the Frog Semicircular Canal
