Thesis

Cytochemical localization of NA+, K+-ATPase in the sea urchin egg and embryo

The sites of activity of Na+,K+-ATPase in the sea urchin egg and embryo were determined using a cytochemical localization technique. Fertilized and unfertilized eggs, and embryos at stages of development from 2 to 16-cells were lightly fixed with 2% paraformaldehyde for 3 minutes. Following the method of Mayahara, they were incubated for 20 minutes in an enzymatic reaction medium with p-Nitro-phenylphosphate (NPP) as the substrate and lead as the capture agent. Specimens were then prepared for light and electron microscopic examination using standard techniques. Light micrographs showed that the surfaces of both fertilized and unfertilized eggs were characterized by a circular area of activity at one end of the egg, with only slight activity seen in the rest of the egg. At first cleavage, activity was heavily concentrated in the furrow between the two dividing cells. Electron micrographs of 8 to 16-cell stage embryos showed that activity was most often found on apposed cell membranes, with very little activity on the free surfaces. This asymmetrical distribution of the enzyme may be a reflection of its unique function in embryonic cells.

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