Vital Staining by Trypan Blue; Its Selectivity for Olfactory Receptor Cells of the Brown Bullhead, Ictalurus Natalis

Abstract

The affinity of olfactory receptors in fish for trypan blue as a vital stain is similar to that in amphibia and mammals. But, so far, only Ictalurus has given satisfactory staining. After anesthetizing the animals with MS 222 (tricaine methane sulfonate), the olfactory folds were laid bare by excising the flap of skin between anterior and posterior nares. Next, filter paper was employed to absorb the water and mucus between the folds which were then covered with staining solution. The variations in the treatment were: (a) exposure to 0.5, 1.0 and 2.0% stain concentration, (b) use of distilled water or of 0.7% NaCl solution, (c) staining times of 20, 40, 60 and 80 min. The treated tissues were fixed in Heidenhain's SUSA for 12 hr, dehydrated in isopropyl alcohol, and embedded by passage through methyl benzoate and benzene into paraffin. The sections were cut at 7 μ and mounted in Canada balsam. Vital staining in a 1% distilled water solution for 60 min gave the best results. The receptors were deep blue and contrasted well with the supporting cells. Also, the central receptor-cell processes had taken the stain. Preparations exposed longer than 60 min showed a loss of stain from the receptors. In Ictalurus all receptors are spindle-shaped with peripheral processes of little varying length and thickness depending on the depth of their nuclei within the epithelium. Most nuclei lie near the center of the epithelium. The central processes are thinner (0.25-0. 28 μ) and more curved than the peripheral ones (0.7-0. 9 μ) and may contain small pale blue apparently vesicular swellings. Trypan blue delimits receptor cells more sharply than silver impregnation and seems to be well suited for studies of comparative histology and cytology of vertebrate olfactory epithelium. Methylene blue vital staining of the same tissue shows selectivity for the intraepithelial bundles of central receptor processes rather than for the receptor nuclei.