Longitudinal muscle of myometrial tissues from immature rats were examined by quantitative thin section electron microscopy for the presence of gap junctions after treatment with estradiol with and without tamoxifen, and cycloheximide for 1-6 days. Gap junctions were present between myometrial cells on days 4, 5, and 6 after treatment with estradiol (500 micrograms/day). Tamoxifen administered concomitantly with estradiol over the 6-day period completely prevented induction of the junctions. Gap junctions were not detected in the myometrium after treatment with tamoxifen alone. Administration of cycloheximide together with estradiol on day 0 of the 6-day period had no effect on gap junction frequency but resulted in a reduction in gap junction size in the myometrium after continued treatment with the hormone. Treatment with cycloheximide on day 1, however, significantly suppressed the effect of further estradiol treatment on the induction of gap junctions in the myometrium. Junctions were not visible in the tissues from animals treated with cycloheximide alone or in the control groups treated with sesame oil. These results indicate that estradiol influences the presence of gap junctions in the myometrium by regulating the synthesis of gap junction proteins through the steroid receptor mechanism.