Production of superoxide anion and hydrogen peroxide by capacitating buffalo (Bubalus bubalis) spermatozoa.
In the present study attempts were made to detect and quantify the generation of superoxide anion (O(2)(*-)) and hydrogen peroxide (H(2)O(2)) by capacitating buffalo spermatozoa. Ejaculated buffalo spermatozoa were suspended in sp-TALP medium at 50x10(6)mL(-1) and incubated at 38.5 degrees C with 5% CO(2) in air in the absence or presence of heparin (a capacitation inducer) or reduced nicotinamide adenine dinucleotide phosphate (NADPH) or diphenyleneiodonium (DPI, a flavoprotein inhibitor) for 6h. Production rate of O(2)(*-) and H(2)O(2) by spermatozoa at different hours of capacitation were measured by cytochrome c reduction and phenol red oxidation assays, respectively. Spermatozoa generated both O(2)(*-) and H(2)O(2) spontaneously and following stimulation with heparin and a significant increase of O(2)(*-) production was observed in the presence of NADPH. However, DPI inhibited this NADPH-induced O(2)(*-) production and suggested for existence of putative NADPH-oxidase that constitute a specific O(2)(*-) generating systems in buffalo spermatozoa. Results of our study indicated that buffalo spermatozoa generate O(2)(*-) and H(2)O(2) and production of these free radicals is induced during capacitation.
Roy
S C
SC
Division of Animal Biochemistry, National Dairy Research Institute, Karnal 132001, India. sudhircroy@rediffmail.com
Atreja
S K
SK
D000818:Animals; D002020:Buffaloes / Q000502:physiology*; D045304:Cytochromes c / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005343:Fibrinolytic Agents / Q000494:pharmacology; D006493:Heparin / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000378:metabolism*; D008297:Male; D009249:NADP / Q000494:pharmacology; D009861:Onium Compounds / Q000494:pharmacology; D010637:Phenolsulfonphthalein / Q000378:metabolism; D013075:Sperm Capacitation / Q000187:drug effects / Q000502:physiology*; D013094:Spermatozoa / Q000187:drug effects / Q000502:physiology*; D013481:Superoxides / Q000378:metabolism*; D013997:Time Factors
D004791:Enzyme Inhibitors; D005343:Fibrinolytic Agents; D009861:Onium Compounds; D013481:Superoxides; D009249:NADP; C007517:diphenyleneiodonium; D006493:Heparin; D045304:Cytochromes c; D006861:Hydrogen Peroxide; D010637:Phenolsulfonphthalein