Little or no motility is observed when sperm from 5 mammalian species are incubated in vitro in their cauda epididymal fluid (CEF). We examined the effects of pH, lactate, and viscoelastic drag on sperm motility to determine whether these factors are responsible for this inhibition of motility. The pHs of CEF from bull, dog, rat, guinea pig, and hamster were 5.8, 6.2, 6.9, 6.9, and 7.2, respectively. The lactate concentration of epididymal semen collected from anesthetized animals ranged from 0.6 to 0.9, but increased almost 10-fold in samples from rats or dogs when measured 2 h postmortem. Increasing the pH of CEF to 7.0 resulted in the initiation of full motility for bull and dog sperm. Suspensions of sperm in buffer at various pHs (from 4.0 to 7.6) produced a sigmoidal motility curve for all species. All species, including bull and dog, showed almost full motility in buffer at a pH equal to the pH of their own CEF. Motility of bull and dog sperm showed greater inhibition with decreasing pH when suspended in CEF instead of buffer. The addition of 15 mM lactate, which has been shown to lower sperm intracellular pH, shifted the motility versus pH curves of all species toward higher pH. In bull and dog the addition of lactate produced a motility profile that was indistinguishable from that in their own CEF. The viscoelastic drag of the CEF of only two species, rat and hamster, was sufficiently high to inhibit sperm motility. We conclude that the low pH of the CEF from bulls and dogs plus the presence of lactate is sufficient to cause inhibition of motility.(ABSTRACT TRUNCATED AT 250 WORDS)