Ouabain-sensitive, K-dependent p-nitrophenyl phosphatase (p-NPPase) activity was localized ultra-Ocytochemically in the lateral plasma membranes of secretory ameloblasts and the stratum intermedium and principally in the papillary layer cells of aldehyde-fixed rat incisor enamel organs by the one-step lead method. Daily intraperitoneal injection of ouabain (250 μg, 500 μg, and 1 mg/100 g body weight) for two weeks reduced p-NPPase activity in the enamel organ cells. However, the degree to which this activity was reduced appeared to vary among the experimental animals. Addition of ouabain to the cytochemical incubation medium completely inhibited p-NPPase activity in the tissues. Although long-term ouabain injection did not result in any morphological alterations of the enamel organ cells, it caused, in part, an appearance of electron-dense, homogeneous matrix-like substances (MS) in the extracellular spaces of the ameloblast layers at both the secretion and maturation stages. In addition, long-term ouabain injection appeared to have resulted in delayed maturation of enamel as measured by energy-dispersive x-ray analysis of Ca and P in surface enamel. These results suggest that Na-K-ATPase of enamel organ cells may participate in the net flow (removal) of organic matrix components and water from the enamel during the maturation stage of enamel formation. It is suggested that this flow is maintained by local osmotic gradients generated by Na-K-ATPase within the papillary layer.
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