[histochemical Study of the Adrenal Cortex by T.t.c. Reaction].

The changes of the adrenal cortex caused by corticosteroids and ACTH, were studied histochemically using T.T.C. (2-3-5 triphenyl tetrazolium chloride), one of the tetrazolium compounds. T.T.C. is a water soluble and colorless crystalline substance, changes unsoluble in water, soluble in oil, and forms a pink colored precipitation (formazan) by a reducing process. Zweifach et al. (1951) incubated the adrenal slices of rats in T.T.C. solution to produce formazan in these slices, and then made frozen sections. From their results, they reported that the findings of the T.T.C. reaction clearly showed the adrenal function. In Japan, the T.T.C. reaction was studied by F. Akasu, H. Fukushima etc.. H. Fukushima reported that the T.T.C. reaction was a specific stain for corticoids after ascorbic acid, adrenalin, SS group and SH group were excluded. In our experiments, when an equal volume mixture of 2% T.T.C. and 1% sodium hydroxide was dropped into adrenalin chloride, ascorbic acid solution or hydrocortisone suspension, deep red sediment appeared immediately in these solutions. In the mixture of hydrocortisone acetate and the T.T.C. solution, the red color began to appear 5 minutes after the mixing, and the deep red sediment appeared in 20 minutes. No red sediment appeared in the mixture of dexamethasone phosphate and the T.T.C. alkaline solution, but the sediment appeared easily after heating the mixture. From these results, it was confirmed that T.T.C. reduced and changed red sediments with reducing substances in the alkaline range. It is suggested that in corticosteroids, an OH group attached to the 21-carbon atom has a reducing action (Tab. 1). Animal experiments. Male Wister rats of 100-200gr. in body weight, were used. After various dosages of corticosteroids were administrated, these rats were killed by cutting their heads. The autopsy was done immediately, and both adrenals were removed. After the adrenals were weighed, the adrenals were fixed in neutral formalin solution, then T.T.C. reaction was peformed. While T.T.C. reactions of the normal adrenal of rats were clearly observed in the zona glomerulosa as beautiful pink colored fine granules, the reactions were most conspicuously present in the outer layer of zona fasciculata. But the reactions decreased gradually in the inner layer of zona fasciculata, and some gross granules were present near the border between the adrenal cortex and the medulla (Fig. 1). In the adrenal cortex of rats given dexamethasone, the initial change was the flecked finding where the pink color was deep in a section and was light in other sections of the zona fasciculata. Then the reaction of the zona fasciculata decreased