The objective of this study was to determine whether the cumulus oophorus cells surrounding the human oocyte release a factor(s) that stimulates sperm velocity. Medium from in vitro fertilization (IVF) that was exposed to cumulus/oocyte complexes was tested for its ability to stimulate sperm velocity by use of a computerized sperm motion analyzer. In subsequent experiments, primary cultures of cumulus cells, granulosa cells, and ovarian cancer cells were established, and conditioned medium from these cells was assessed for sperm velocity-stimulating activity. In the initial series of assays using four sperm donors, cumulus/oocyte complex-conditioned medium from IVF increased sperm curvilinear velocity and amplitude of lateral head displacement by an average of 12.3% and 19.3% (p < 0.0001), respectively, compared to medium from IVF that was not exposed to cumulus/oocyte complexes. These results were confirmed in a subsequent, more extensive series of assays using a single sperm donor. Testing of conditioned media after 24 h of primary cell culture showed that cumulus-conditioned medium consistently stimulated curvilinear velocity (8.8% increase) and lateral head amplitude (18.7% increase) compared to unconditioned medium (p < 0.0001) in a manner similar to cumulus/oocyte complex-conditioned medium. In contrast, granulosa-conditioned medium and ovarian cancer cell-conditioned medium did not stimulate curvilinear velocity and lateral head amplitude compared to unconditioned medium. The ability of cumulus-conditioned medium to stimulate curvilinear velocity and lateral head amplitude declined gradually during the 5-day culture period. Nevertheless, the results indicate that the cumulus released velocity-stimulating activity throughout the culture period.(ABSTRACT TRUNCATED AT 250 WORDS)