Female reproductive organs show remarkable cyclic changes in morphology and function in response to a combination of hormones. Evidence has accumulated suggesting that phosphoinositide turnover and the consequent diacylglycerol (DG) protein kinase C (PKC) pathway are intimately involved in these mechanisms. The present study has been performed to investigate the gene expression, cellular localization, and enzymatic activity of the DG kinase (DGK) isozymes that control the DG-PKC pathway. Gene expression for DGKalpha, -epsilon, -zeta, and -iota was detected in the ovary and placenta. Intense expression signals for DGKzeta and -alpha were observed in the theca cells and moderate signals in the interstitium and corpora lutea of the ovary. On the other hand, signals for DGKepsilon were seen more intensely in granulosa cells. In the placenta, signals for DGKalpha and -iota were observed in the junctional zone, whereas those for DGKzeta were detected in the labyrinthine zone. At higher magnification, the signals for DGKalpha were mainly discerned in giant cytotrophoblasts, and those for DGKiota were found in small cytotrophoblasts of the junctional zone. DGKzeta signals were observed in all cellular components of the labyrinthine zone, including mesenchyme, trabecular trophoblasts, and cytotrophoblasts. DGKepsilon signals were detected in the junctional zone on day 13 and 15 of pregnancy and were diffusely distributed both in the labyrinthine and junctional zones at later stages. The present study reveals distinct patterns of mRNA localization for DGK isozymes in the rat ovary and placenta, suggesting that each isozyme plays a unique role in distinct cell types in these organs.