IONIC AND OSMOTIC CONDITIONS 



81 



Salisbury, 1960). The data show for other semen samples that, while 

 the free sullhydryl radical of cysteine will inhibit the release by sul- 

 fite, the bound sulfhydryl radical of methionine will not do so (Lodge 

 and Salisbury, 1960). 



Evidence for a Diffusible Metabolic Regulator 



The second condition resulted from our need to obtain spermato- 

 zoa free from absorbed substrate which had not been stimulated to 

 maximum metabolism and motility. This we have done by collect- 

 ing semen directly into 100 ml of freshly made C0 2 -saturated diluent 

 prepared by mixing citric acid with a potassium and sodium bicar- 

 bonate solution (Graves and Salisbury, 1959). With the proper con- 

 tent of K+ the cells are completely immobilized and do not take up 

 substrate when left in the mixture for 24 hours at 5°C (Graves and 

 Salisbury, 1959, 1960). In Fig. 11 is shown the endogenous respira- 



INCUBATED IN 09% NaCI 



SAME AS ABOVE PLUS FRUCTOSE 



2 3 



TIME-HOURS 



Fig. 11. Typical oxygen uptake by washed epididymal-like spermato- 

 zoa soon after collection with and -without substrate, and respiratory stim- 

 ulation after 24 hours storage at 5°C in the inhibitory diluent. I. Collected 

 in lNa:3K, citrate-bicarbonate C0 2 -saturated diluent and stored at 5°C 

 for 25 hours before washing and incubation at 37°C: (a) in saline-fructose 

 (500 mg %); (b) in saline. Number of semen samples, n, equals 12. II. Col- 

 lected in 0.9% NaCI, and INarIK or lNa:3K citrate-bicarbonate C0 2 - 

 saturated diluent, immediately washed and incubated at 37°C: (a) in sa- 

 line-fructose (500 mg %); (b) in saline. Number of semen samples, n, 

 equals 78 (Graves and Salisbury, 1960). 



