Effect of amino acids on goat cauda epididymal sperm cryopreservation using a chemically defined model system

Studies were carried out to analyze the cryoprotecting efficacy of at sever al amino acids by use of a chemically defined synthetic medium (modified Ri nger's solution) and goat cauda epididymal sperm as the model system. Motil e goal cauda sperm dispersed in the synthetic medium were subjected to a fr eezing protocol in a computer-controlled bio-freezer. cooling 0.25 degreesC min(-1) to 5 degreesC. 5 degreesC min(-1) to -20 degreesC. and 20 degreesC min(-1) to -100 degreesC. prior to being plunged into liquid nitrogen. In the absence of amino acids. sperm cells completely lost their flagellar mot ility. Of all the amino acids tested. L-alanine showed maximal cryoprotecti on potential. L-Alanine at 135 mM offered optimum cryoprotection potential: recovery of sperm forward motility and total motility were 14 +/- 2% and 1 9 +/- 2%, respectively. L-Glutamine, L-proline. and glycine at optimum conc entration (100-150 mM) cryopreserved approx. 11-17% total motility of the s perm cells. whereas amino acids such as L-arginine. L-lysine. and L-histidi ne offered little cryoprotection (0-5%) to the cells. Increasing the amino acid concentration beyond the optimum level sharply decreased the recovery of the sperm motility. which therefore showed a biphasic cryoprotection pro file. Addition of amino acids enhanced (approx. 7-10%) the cryoprotection e fficacy of the well-known cryoprotectants glycerol and a combination of gly cerol and dimethyl sulfoxide. The presence of glycerol caused a marked redu ction (from 100-150 mM to 20-70 mM levels) in the optimal cryopreservation concentration of the amino acids. The combined cryoprotecting action of gly cerol, dimethyl sulfoxide and amino acids provided motility recovery as hig h as 52%. The observation that amino acids and dimethyl sulfoxide had an ad ditive effect in augmenting the cryoprotecting potential of glycerol sugges ts that the mechanism of their action is different from that of glycerol. T his cocktail of cryoprotectants may be useful for cryopreservation of semen of various species. (C) 2001 Acadmeic Press.