Diacylglycerol (DAG) is a derivative of the membrane phospholipids and the first identified second messenger to activate protein kinase C (PKC), a multifunctional serine/threonine kinase with multiple isozymes, involved in wide varieties of functions. DAG is phosphorylated by DAG kinase (DGK) to generate  phosphatidic acid (PA), another signal molecule to activate some other proteins such as atypical PKC. Therefore, DGK is regarded as an important regulator of PKC activities through attenuation of DAG and augmentation of PA. With a hypothesis that DGK is involved in the steroidogenesis through regulation of PKCs, the present study was attempted to examine the localization of the three DGK isozymes in immunohistochemistry. Distinct immunoreactivity for DGKb was detected in almost all the adrenocortical cells and testicular Leydig cells of adult mice. The immunoreactive materials appeared in forms of fine granules and they were distributed throughout the cytoplasm. The immuno-intensity was variable among the positive Leydig cells and cortical cells. Although the immunoreactivity for DGK was found in the cytoplasm of almost all the cortical cells, it was negative in all the Leydig cells. No immunoreactivity for DGKz was found in the adrenal cortex and testicular Leydig cells. Judging from the occurrence of distinct immunoreactivity for DGKb common to the adrenocortical cells and Leydig cells, it is highly possible that this DGK isozyme is involved in the steroidogenesis through the regulation of PKC.