The intracellular localization of carbonic anhydrase and a carbonic anhydrase inhibitor in the brains of mice

Homogenates prepared from the perfused brains of mice were differentially centrifuged in 0.25 M sucrose. On the basis of activity per mg of nitrogen, only the supernatant showed a concentration of carbonic anhydrase activity greater than that of the original homogenate (about two- to three-fold). No concentration of activity was found in the particulate fraction, in any of its components studied, or in the washes of the various residues. Partition of per cent activity varied with the procedure used, but approximately two-thirds of the total activity of the original homogenate were in the supernatant fraction; the total particulate matter accounted for about one-third. No differences of any consequence were found amongst whole brain, cortex, and brain with the cortex removed. The data are believed to demonstrate the intracellular localization of carbonic anhydrase in the soluble fraction of cells; the activity of particulate matter is considered to be the result of contamination. The inhibitor of carbonic anhydrase found in the brains of mice following the intravenous administration of methazolamide was almost entirely localized in the supernatant fractions; an intercellular localization in the soluble fraction is also postulated. The localization of both carbonic anhydrase and carbonic anhydrase inhibitor in the soluble fraction of homogenates of the brains of mice does not constitute evidence of the inhibition of the enzyme in the brain in vivo. However, the concentrations of inhibitor found in the whole homogenates 15 min and 2 hr after the administration of doses of methazolamide at the level of the ED 95 for anticonvulsant effect were theoretically capable of causing maximum inhibition of carbonic anhydrase in the brains of mice. Anticonvulsant action and inhibition of carbonic anhydrase are probably related.