In vitro capacitation and acrosome reaction in fresh and cryopreserved porcine spermatozoa

PARIS DUPRAT ML; BREININGER E; PEREYRA V; SATORRE MM; RODRIGUEZ PC

Congreso; XVIII Jornadas Anuales Multidisciplinarias de la Sociedad Argentina de Biología; 2016

The aim of this study was to determine the activity of lactate dehydrogenase (LDH; 1.1.1.27) and study its participation in theprocesses of the in vitro capacitation and acrosome reaction (AR) in fresh and cryopreserved porcine spermatozoa. Fresh andcryopreserved spermatozoa were incubated in TBM medium with bicarbonate and follicular fluid as capacitation and acrosomereaction inducers, respectively in the presence of different concentrations of sodium oxamate (competitive inhibitor of LDH).Capacitation and AR percentages were determined by the CTC technique and trypan blue combined with DIC, respectively.Sperm viability and motility were evaluated by the eosin/nigrosin technique and optic microscopy in thermal stage, respectively.The LDH activity was determined spectrophotometrically at 600 nm, during 2 minutes, at 37°C and the enzyme unit (U) wasdefined as the amount of LDH that oxidizes 1 μmol of NADH/minute. The results were analysed by ANOVA and Bonferronitest. The addition of the competitive inhibitor of the enzyme significantly diminished capacitation and AR without affecting sperm viability, at different concentrations in fresh (50mM for capacitation and 25mM for AR) or cryopreserved sperm (1 mM for both processes).These concentrations also inhibited the activity of LDH in fresh (76,8 ± 6,1 and 86,9 ± 7,5 % of inhibition for 25mM and 50mM, respectively) and cryopreserved spermatozoa (73,0 ± 16,4 % of inhibition for 1 mM). Our results demonstrated the differential participation of the enzyme lactate dehydrogenase in the processes that are involved in the acquisition of the fertilizing ability in fresh and cryopreserved spermatozoa. This information can be useful for the formulation of incubation media to use in biotechnological reproduction techniques.