VITAL STAINING OF THE HEARING ORGAN - VISUALIZATION OF CELLULAR STRUCTURE WITH CONFOCAL MICROSCOPY

Cells inside the intact organ of Corti were labelled with fluorescent probes reflecting various aspects of structure and function. The dyes were introduced into the perilymphatic space by perfusion of the scala tympani of the temporal bone from the guinea-pig maintained in isolat ion. The dyes were able to diffuse through the basilar membrane and in to the organ of Corti where they were spontaneously absorbed by the se nsory and supporting cells. Confocal microscopic observation was made through an opening in the apex of the cochlea. A number of different d yes were used; a carbocyanine dye which stains mitochondria; two styry l dyes which are absorbed by the cell membranes and calcein, a cytopla smic marker that fluoresces in vital cells. Extracellular space was st ained by a cell-impermeant Dextran-fluorescein. The most striking find ing was that the membrane dyes preferentially stained the sensory cell s and neural elements whereas the staining of the supporting cells was faint. The cytoplasmic dye in general stained sensory and supporting cells to the same extent. By tilting the organ, a view could be obtain ed from the side like a radial section through the organ. Outer and in ner hair cells with their sensory hairs, nerve fibres and nerve ending s, especially under the inner hair cells, could be seen in profile. In troduction of a high molecular weight Dextran into the endolymphatic s pace outlined the tectorial membrane which was seen in negative contra st. The simultaneous perfusion with a membrane dye stained the hair ce lls and their sensory hairs. Merging of the two images gave the possib ility to examine, in the living tissue, the cilia to tectorial membran e relationship. Of general interest is the finding that the membrane d yes preferentially stained the sensory and neural elements of the nerv ous system, represented here by the hair cells and nerve fibres of the inner ear. (C) 1997 IBRO. Published by Elsevier Science Ltd.