Deciphering dose and time dependent effect of supplementing SNP and L-NAME in extender on progressive motility of Murrah bull spermatozoa

Abstract

Male germ cells have the potential to generate reactive oxygen species (ROS) during different stages of differentiation, laboratory manipulation and during their interaction with dissolved oxygen in extender. These ROS and its subset reactive nitrogen species (RNS) perform a concentration specific bimodal role, favouring oxidative eustress at physiological concentrations. The present study was aimed to investigate the dose and time dependent effect of the most potent RNS, i.e. nitric oxide (NO) compounds, specifically SNP (Sodium nitroprusside) and L-NAME (N-Nitro-L-Arginine Methyl Ester) supplementation in extender on progressive motility. The semen samples were divided into 8 aliquots comprising of various concentrations of SNP (Control, 0.1, 1 and 10 μmol/L) and L-NAME (Control, 1, 10 and 100 μmol/L) and refrigerated for 0, 6, 24 and 48 h. The progressive motility (%) improved significantly particularly with the addition of 1 μM SNP and 10 μM L-NAME as compared to control during all time intervals. Subsequently semen sample was cryopreserved with optimum level of SNP (1μmol/L) and L-NAME (10 μmol/L) from these results by adding it in extender and assessed for progressive motility before cryopreservation and after 24 h, 1 month and 2 months of cryopreservation. It is concluded that the addition of SNP and L NAME in cryopreservation media at 1 μM and 10 μM respectively improved the sperm motility significantly. On further evaluation a significant increase in progressive motility was observed for L-NAME treated extender at 24 h and SNP treated extender at 1 and 2 month interval in comparison to control.

Keywords

• Cryopreservation
• Extender
• NO
• Progressive motility
• RNS
• ROS