Ultrastructural localization of Na+/K(+)-ATPase in rodent olfactory epithelium.

The olfactory epithelium is comprised of bipolar sensory neurons, sustentacular cells, and basal cells. The sensory neurons have apical knobs and cilia, which project into the olfactory mucus toward the nasal lumen, and represent presumptive sites of odorant binding. Ionic currents, measured across this epithelium in both the resting and odorant-stimulated states, are known to be sustained, at least in part, by active transport of sodium. Information identifying the cellular sites of ion transport in olfactory sensory epithelium will therefore aid in elucidating the ionic mechanisms associated with olfactory transduction. The membrane-bound enzyme Na+/K(+)-ATPase mediates active ion transport in many other cells and tissues. We have consequently employed the cytochemical technique reported by Ernst (J. Histochem. Cytochem., 20 (1972) 23-38, 1322) to identify possible sites of elevated Na+/K(+)-ATPase activity in olfactory epithelium. This procedure detects inorganic phosphate (Pi) released from an artificial substrate (nitrophenyl phosphate) by enzyme catalytic activity. In the presence of strontium ion. Pi is precipitated near regions of high enzymatic activity, then converted to a product visible in the electron microscope. Parallel control preparations were incubated in media (1) supplemented with the specific Na+/K(+)-ATPase inhibitor ouabain (to abolish formation of specific reaction product); (2) with substrate deleted (to demonstrate possible non-specific binding of Sr2+ and/or Pb2+); or (3) with the necessary cofactor K+ deleted. In tissues incubated for demonstration of Na+/K(+)-ATPase activity, reaction product was associated with apical knobs, cilia and dendrites of olfactory receptor neurons at the apical surface. In the more proximal region of the epithelium, reaction product was associated with cell bodies and axons of the sensory neurons, and with the lateral membranes of sustentacular cells. Reaction product was deposited intracellularly, compatible with the known mechanism of the Na+/K(+)-ATPase enzymatic reaction. In control specimens incubated with ouabain, with substrate deleted, or with K+ deleted, only a small quantity of non-specific precipitate was observed. These results are discussed with reference to the various sodium currents implicated in olfactory transduction and transepithelial transport.