Cell suspensions from rat olfactory neuroepithelium: biochemical and histochemical characterization

Abstract

Cell suspensions were generated from rat olfactory epithelium by digestion with collagenase and hyaluronidase followed by gentle mechanical disruption. These cell suspensions excluded nigrosin dye and synthesized RNA, protein and carnosine from radiolabeled precursors. Sustentacular cells, repiratory epithelial cells and olfactory neurons but not basal cells could be identified by phase-contrast microscopy. Sedimentation of these cell suspensions at unit gravity in discontinuous gradients of buffered bovine serum albumin resulted in partial separation of the various cell types as indicated by the distribution of several biochemical markers. Olfactory marker protein and carnosine synthetase activity were found in the upper gradient fractions, while carnosinase activity was present predominantly in the lower gradient fractions. Cellular localization of olfactory neuron marker protein and non-neuronal S-100 protein by immunoperoxidase staining of gradient-fractionated cells indicated that neuronal cells were only partially separated from non-neuronal cells by our fractionation techniques. Evaluation of gradient fractionated cells by histochemical staining for carbohydrates demonstrated that secretory Bowman's gland cells were quite efficiently separated from neurons. This study demonstrates the ease with which cell suspensions may be produced from the olfactory epithelium, and emphasizes the importance of utilizing both biochemical and histochemical approaches in studies of mixed populations of cells, particularly when the purity of the cell fractions is a consideration.