Inhibiting sperm pyruvate dehydrogenase complex and its E3 subunit, dihydrolipoamide dehydrogenase affects fertilization in Syrian hamsters.
OBJECTIVE
The importance of sperm capacitation for mammalian fertilization has been confirmed in the present study via sperm metabolism. Involvement of the metabolic enzymes pyruvate dehydrogenase complex (PDHc) and its E3 subunit, dihydrolipoamide dehydrogenase (DLD) in hamster in vitro fertilization (IVF) via in vitro sperm capacitation is being proposed through regulation of sperm intracellular lactate, pH and calcium.


RESULTS
Capacitated hamster spermatozoa were allowed to fertilize hamster oocytes in vitro which were then assessed for fertilization, microscopically. PDHc/DLD was inhibited by the use of the specific DLD-inhibitor, MICA (5-methoxyindole-2-carboxylic acid). Oocytes fertilized with MICA-treated (MT) [and thus PDHc/DLD-inhibited] spermatozoa showed defective fertilization where 2nd polar body release and pronuclei formation were not observed. Defective fertilization was attributable to capacitation failure owing to high lactate and low intracellular pH and calcium in MT-spermatozoa during capacitation. Moreover, this defect could be overcome by alkalinizing spermatozoa, before fertilization. Increasing intracellular calcium in spermatozoa pre-IVF and in defectively-fertilized oocytes, post-fertilization rescued the arrest seen, suggesting the role of intracellular calcium from either of the gametes in fertilization. Parallel experiments carried out with control spermatozoa capacitated in medium with low extracellular pH or high lactate substantiated the necessity of optimal sperm intracellular lactate levels, intracellular pH and calcium during sperm capacitation, for proper fertilization.


CONCLUSIONS
This study confirms the importance of pyruvate/lactate metabolism in capacitating spermatozoa for successful fertilization, besides revealing for the first time the importance of sperm PDHc/ DLD in fertilization, via the modulation of sperm intracellular lactate, pH and calcium during capacitation. In addition, the observations made in the IVF studies in hamsters suggest that capacitation failures could be a plausible cause of unsuccessful fertilization encountered during human assisted reproductive technologies, like IVF and ICSI. Our studies indicate a role of sperm capacitation in the post-penetration events during fertilization.
Centre for Cellular and Molecular Biology (Council of Scientific and Industrial Research), Hyderabad, India.
Archana B
AB
Siva
Centre for Cellular and Molecular Biology (Council of Scientific and Industrial Research), Hyderabad, India.
Subbarayalu
S
Panneerdoss
Centre for Cellular and Molecular Biology (Council of Scientific and Industrial Research), Hyderabad, India.
Purnima
P
Sailasree
Centre for Cellular and Molecular Biology (Council of Scientific and Industrial Research), Hyderabad, India.
Durgesh K
DK
Singh
Centre for Cellular and Molecular Biology (Council of Scientific and Industrial Research), Hyderabad, India.
Duvurri B
DB
Kameshwari
Centre for Cellular and Molecular Biology (Council of Scientific and Industrial Research), Hyderabad, India.
Sisinthy
S
Shivaji
D000818:Animals; D002118:Calcium / Q000378:metabolism; D006224:Cricetinae; D003470:Culture Media; D008058:Dihydrolipoamide Dehydrogenase / Q000037:antagonists & inhibitors*; D005260:Female; D005306:Fertilization; D008297:Male; D008647:Mesocricetus; D011768:Pyruvate Dehydrogenase Complex / Q000037:antagonists & inhibitors*; D013094:Spermatozoa / Q000201:enzymology*
D003470:Culture Media; D011768:Pyruvate Dehydrogenase Complex; D008058:Dihydrolipoamide Dehydrogenase; D002118:Calcium