of the cation-stimulated ATP-ase systems of liver and brain.
Methods
Animals. Male Sprague-Dawley rats, body weight 250-300 gm were used. Animals were maintained on a pellet diet supplemented by vitamin E (20 mg dlatocopherol acetate dissolved in olive oil per rat, given thrice weekly, as described by McLean ('63).
Tissue slices. These were cut with a Stadie-Riggs slicer and were of about 200 mg wet weight.
Cation stimulated ATP-ase preparations were prepared as described by Ahmed and Judah ('64), using rat liver or brain as a source.
Chemicals and chemical methods. The Ringer's solution used for tissue slice incubations was composed of NaCl, 133 mM; KCl, 6.0 mM (unless otherwise stated); CaCL, 2.8 mM; MgCL, 1.42 mM; KHzP04, 1.42 mM. pH was maintained at pH 7.20 with NaHC03, using a gas-phase of 95% Na and K were estimated by Aame photometry, using a Zeiss PMQ spectrophotometer with flame attachment, burning a hydrogen-oxygen mixture.
Adenosinetriphosphate ( ATP) was obtained from Sigma Chemical Co., St. Louis, Missouri. It was rendered cation-free by passage over Dowex-50 resin. The resin had previously been converted to trihydroxymethylaminomethane (Tris) form. KCl as obtained was found to contain material toxic to the ATP-ase preparation and was recrystallized from hot water. Choline chloride was recrystallized from hot ethanol before use. Sucrose solutions used for preparation of enzymes were made up in de-ionized water. The solutions were then passed through a mixed-oz-5% coz.