This study was undertaken to investigate the influence of prostaglandin E₁(PGE₁) upon the activity of carbonic anhydrase and upon the inhibitory action of acetazolamide on carbonic anhydrase. The heparinized blood was sampled by cardiac puncture from Sprague-Dawley strain rats under ether anesthesia and was hemolysed by adding distilled water 1,000 times the amount of the blood. The activity of carbonic anhydrase of 0.1 ml of the hemolysate was measured by Maren s simplified micro-method. In the first experiment, the 7 rats were used, and the activity was measured by adding 0.1 ml of various concentrations of PGE₁(0.5, 1.25, 2.5, 5.0, 10 and 20 μg/ml). In the second experiment, the 6 rats were used and the activity was measured by adding 0.1 ml of PGE₁(5 μg/ml) and 0.1 ml of acetazolamide(6 X 10^-7M/l) respectively or simultaneously. Obtained results were as follows: 1) The activity of carbonic anhydrase was significantly inhibited by PGE₁ at doses of 0.5 ~ 10 μg/ml, maximally at a dose of 2.5 μg/ml, but inhibition was no more observed at a dose of 20 μg/ml. 2) The activity of the acetazolamide group was significantly less than that of the control group. 3) The activity of the PGE₁+acetazolamide group was significantly less than those of the PGE₁ group and the acetazolamide group. It is inferred from the above results that the PGE₁ inhibits the activity of carbonic anhydrase dose-dependently and strengthens the inhibitory effect of acetazolamide on carbonic anhydrase.