A quantitative microscopic technique was employed to examine the distribution of mitotic activity in the rat mammary gland. The frequency distribution of mitoses per unit volume of lobuloalveolar mammary gland epithelium in virgin Lewis/Mai rats at each phase of the estrous cycle were determined and compared to the expected Poisson frequency distributions, assuming random mitotic activity. Both pooled data and data from individual rats were compared to expected Poisson distributions. At each phase of the estrous cycle, the pooled observed distributions deviated significantly from Poisson distributions. Sixty-seven percent (72/108) of the observed frequency distributions obtained from individual rats also deviated significantly from expected Poisson distributions. These data indicate a nonrandom distribution of mitoses in rat lobuloalveolar mammary gland epithelium. This observation suggests that local cell products and/or a variation in the extent of replicative synchrony of lobuloalveolar cell populations may determine in part the pattern of mitotic activity in this tissue. A nonrandom distribution of mitoses in mammary epithelium may have significance in relation to the genesis of hyperplastic and neoplastic lesions of the mammary gland.