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.
Viggiano
J M
JM
Centro de Estudios Farmacológicos y Botánicos (CEFYBO), Buenos Aires, Argentina.
Herrero
M B
MB
Martínez
S P
SP
De Gimeno
M F
MF
D000177:Acrosome / Q000187:drug effects / Q000201:enzymology*; D000818:Animals; D001120:Arginine / Q000494:pharmacology; D048429:Cell Size; D002470:Cell Survival / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005456:Fluorescent Dyes; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D013094:Spermatozoa / Q000166:cytology / Q000201:enzymology* / Q000648:ultrastructure; D013194:Staining and Labeling / Q000379:methods; D013997:Time Factors
D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D001120:Arginine; D019001:Nitric Oxide Synthase; D019331:NG-Nitroarginine Methyl Ester