Examination of the cupula and stereocilia of the horizontal semicircular canal in the toadfish Opsanus tau

We imaged the horizontal semicircular canal (HSCC) crista and cupula of toadfish, Opsanus tau by using a) confocal light microscopy of isolated vital HSCC; b) serial sections of fixed, trichrome-stained HSCC; and c) scanning electron microscopy of fixed HSCCs. HSCC were dissections which included an ampulla and an attached canal tube (long and slender canal portion), and, in some cases, a small portion of the utricular wall. Cupulae were seen as multipartite mucus connective tissue shells rising from the crista and extending toward the ampullary roof. They were composed of several refractile bands traversing the cupulae perpendicular to longitudinal fibers extending from the cupular base to its apex. Alcian green-stained cupulae showed an asymmetric alcianphilic, dark, X-shaped structure, indicating that the pillar is rich in mucin and carbohydrate, an interpretation supported by images of trichrome-stained sections. The cupular antrum is devoid of prominent refractile fibers. No tubes or channels were observed in the cupula or antrum of vital preparations. Cupular shell fibers cover the surface of the crista, are roughly parallel, and are associated with a translucent material having a refractive index greater than the surrounding endolymph. Stereocilia were thin, 100-μm-long structures, with little longitudinal curvature, which end with no end bulb. No strands extend from stereocilia to the roof or other portions of the cupular antrum. Gross movements of stereocilia were not seen in mechanically quiescent preparations. Within the cupular antrum, stereocilia were parallel to connective tissue fibers, all embedded in an isotropic gel. This fiber-reinforced gel and cupular matrix are sensitive to N-acetlyneuraminidase and β-N-acetyl glucosaminidase, and minimally sensitive to β-N-acetyl hexosaminidase. Connective tissue fibers may serve to stiffen the gel, whose matrix would restrict lateral motion of embedded fibers and stereocilia thereby providing mechanical support for stereocilia.