Caffeine, a cyclic nucleotide phosphodiesterase inhibitor, markedly increased and maintained the respiration and motility of ejaculated bovine spermatozoa. Caffeine approximately doubled the oxygen consumption of washed ejaculated spermatozoa in the presence of acetate, pyruvate or fructose; it had a smaller effect with citrate as the substrate and no effect on endogenous respiration. The respiration of unwashed ejaculated sperm was increased by caffeine independent of exogenous substrate addition. Motility data collected on ejaculates from bulls demonstrated that spermatozoa in the presence of caffeine (6 mM) maintained the initial percentage of motility for at least 4 hr at 37C, whereas in the untreated sperm samples, 50% of sperm that were motile initially became immotile during 4 hr storage. Comparison of bulls yielding semen of original low motility with bulls yielding originally high motility spermatozoa indicated that an immediate stimulation of sperm motility by caffeine occurred in the low motility specimens (32.2–49.3% motile) but not in those of high motility. It is postulated that the effect of caffeine on spermatozoan motility and respiration is mediated by one of the cyclic nucleotides. This results in a somewhat unique effect, because sperm activity and sperm life, as judged by motility, are both increased with caffeine addition.