Localization and developmental expression of BK channels in mammalian cochlear hair cells

The expression of Slo channels (alpha subunits of BK channels) was investigated in the developing mouse cochlea using a polyclonal antibody against the C-terminal part of the protein (residues 1098-1196). The first BK channel immunoreactivity was observed in the cochlea at E18, where it was localized within the cytoplasm of cells lining the area of the organ of Corti and the spiral ganglion. There was an increase of immunoreactivity in all cells bordering the scala media (supporting and hair cells of the organ of Corti, the stria vascularis and the Reissner's membrane) in the following stages (postnatal day [P] 0 and P6). From P12 to adult, a strong membranous labeling, increasing with age, appeared in inner hair cells. The distribution of BK channels was mainly observed as dense elongated plaques localized in the lateral membrane below the cuticular plate. In addition, a more discrete immunolabeling for BK channels, as punctuated dots, was observed in the synaptic area of inner hair cells. This dual localization of BK channels within inner hair cells was confirmed by a different technique using a fluorescently labeled high-affinity ligand of these channels: IbTX-D19C-Alexa488. We demonstrated under patch clamp experiments that this fluorescent toxin conserved its native property, i.e. to reversibly inhibit BK currents in isolated inner hair cells. The fluorescent toxin, both in living or fixed tissues, also showed a preferential binding to mature inner hair cells with a similar subcellular distribution described above using immunocytochemical technique. Overall, our present results confirm the appearance of membranous BK channels around P12 in mouse inner hair cells, an age at which the auditory system becomes functional. The expression of BK channels in mature inner hair cells, near the site of mechanical-transduction, might serve to limit receptor potential attenuation due to the space constant, and thus permitting these sensory cells to function as fast and sensitive transducers.