To investigate the mechanism of action of spermine, we measured the intracellular calcium ([Ca]i) of guinea pig spermatozoa using a probe of fluorescence, Quin 2. Results showed that spermine (0.25-2.0 mmol.L-1) suppressed the membrane permeability to Ca2+ during capacitation, which was similar to that of verapamil (a Ca2+ channel blocker). Furthermore, the rapid increase of [Ca]i induced by calcimycin (A-23187) was inhibited by spermine and verapamil, whereas trifluoperazine (an inhibitor of calmodulin) had no effect on it. The inhibition of the acrosome reaction caused by verapamil (5-100 mumol.L-1) or trifluoperazine (1-60 mumol.L-1) was reversed by calcimycin and cAMP, respectively. In addition, there was no effect on the initiation of the acrosome reaction when verapamil was added to capacitated spermatozoa. This result was in agreement with that of spermine. When addition of spermine (0.5 mmol.L-1) was combined with trifluoperazine (5 mumol.L-1), the acrosome reaction decline almost to zero, indicating that spermine might inhibit Ca2+ sensitive channel during sperm capacitation.