FIG. 3.

NKAα3 is expressed in both calyx and bouton afferent endings. A–C High magnification micrographs of the central region of the horizontal crista triple-immunolabeled with a mouse monoclonal (IgG1) anti-NKAα3 (green, A), mouse monoclonal (IgG2A) anti-TuJ (blue, B), and a rabbit polyclonal anti-Myo6 (red, C). NKAα3 immunoreactivity was associated with TuJ-identified bouton terminals (labeled B) on type II hair cells as well as simple (labeled S) and complex (labeled C) calyx terminals around Myo6-positive type I hair cells (merge, C). D Higher magnification views of bouton afferents also show NKAα3 immunoreactivity (green) in the membranes of TuJ-positive (blue) boutons and identify the Myo6-positive (red) hair cells as type II. E–F Higher magnification views of simple (E) and complex (F) calyx terminals show NKAα3 immunoreactivity (green) that completely surrounds TuJ-positive (blue) calyces and identifies the Myo6-positive (red) hair cells as type I. Where clearly resolved, the inner-face (open arrowhead) and outer-face (closed arrowhead) calyx membranes are indicated (D, E). Across samples, NKAα3 immunoreactivity appeared more intense in complex calyces compared with either simple or bouton calyces (as appreciated in Fig. 3A). Micrographs represent projections through the minimal number of optical sections required to completely visualize the hair cells and their afferent terminals. Similar patterns of immunoreactivity were observed in high magnification views of the anterior crista and utricle (data not shown). Scale bars measure 10 μm.