Analysis of the effect of nitric oxide synthase inhibition on mouse sperm employing a modified staining method for assessment of the acrosome reaction

A commercially available staining kit (Spermac) was combined with swelling in a hypoosmotic medium (HOS) for simultaneous assessment of viability and acrosome reaction in mouse spermatozoa. We compared the results obtained with the combined technique (HOS‐Spermac) with those obtained with currently used techniques: the chlortetracycline fluorescence assay and eosin exclusion. The results obtained with HOS‐Spermac were the same as those obtained with the chlortetracycline fluorescence assay. Viability assessment with HOS‐Spermac showed a good correlation with the percentage of spermatozoa showing eosin dye exclusion. Using this novel technique, we studied the effect of a nitric oxide synthase inhibitor (NG‐nitro‐L‐arginine methyl ester, L‐NAME) on the acrosome reaction. L‐NAME produced a dose‐dependent inhibition of spontaneous acrosome reaction and its inhibitory effect was specifically counteracted by L‐arginine. We conclude that HOS‐Spermac provides a simple and reliable tool for assessment of the acrosome reaction and that nitric oxide synthase participates in this important function of the spermatozoon.