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Mature female mice, grouped in the absence of a male stimulus, exhibit a suppressed estrous cycle (the so-called Lee-Boot effect). We have designed a series of experiments to elucidate the involvement of the adrenal gland in this phenomenon. Our initial results indicate that adrenalectomized mice exhibit a regular estrous cycle in either isolated or grouped conditions. A single, intact mouse caged with five adrenalectomized females showed repeated normal cycles. When the urine samples from group-caged intact mice or group-caged adrenalectomized mice were applied to the external nares of singly caged females, estrous cycles were inhibited in the animals receiving urine from the intact mice but not from the adrenalectomized mice. In addition, corticosterone therapy restored the function of estrus suppression in grouped, adrenalectomized mice. We had previously shown that the urinary excretion of several volatile compounds (2-heptanone, trans-5-hepten-2-one, trans-4-hepten-2-one, pentyl acetate, cis-2-penten-1-yl acetate, and 2,5-dimethylpyrazine) was adrenal mediated (Science 1986; 231:722-725). A further testing of these compounds in relation to estrus suppression has now revealed that a mixture of these compounds is effective, but removing 2, 5-dimethylpyrazine from the mixture abolished the biological response. The overall results of this study show conclusively an important role of the adrenal gland and adrenal-mediated urinary metabolites in estrus suppression.