Matrix metalloproteinases (MMPs) have been implicated in embryonal implantation processes such as trophoblast invasion and decidualization. The temporal and spatial distributions of MMP bioactivities were analyzed by in situ zymography, which indicated these activities to be markedly increased in the postcoital mouse uterus compared with those in the later implantation stage. Activity was ascribed to proMMP9, which moved from the uterine serosa to the endometrium but was not associated with mRNA up-regulation. The activity was colocalized with infiltrating neutrophils, and neutropenic mice did not exhibit MMP9 expression. Removing the seminal vesicles from male mice abolished the postcoital increase in MMP9 in the female. These results indicate the major MMP activity in the preimplantation uterus to originate in proMMP9-bearing neutrophils attracted by seminal plasma. Considering our results together with those of previous reports of reduced fertility in Mmp9-deficient female mice, we speculate that neutrophil infiltration participates in the extracellular matrix degradation needed to support pregnancy.