Changes in 2-Deoxyglucose Transport during Epididymal Maturation of Ram Sperm1

The initial rate of sugar uptake can be measured using 2-deoxy-D-glucose as a model substrate. Uptake of 2-deoxy-D-glucose involves both transport and phosphorylation and can be monitored accurately by measuring the intracellular content of 2-deoxyglucose-6-phosphate. Uptake of 2-deoxy-D-glucose was linear for at least 10 seconds at 22°C. Testicular, cauda epididymal and ejaculated ram sperm all had a similar Vmax for 2-deoxy-D-glucose uptake (1.5 μmole/h/108 cells). However, testicular sperm had a higher Km for 2-deoxy-D-glucose (310 vs 140 and 160 μM), a higher Ki for phloretin (10 vs 3 and 4 μM) and a lower Ki for cytochalasin B (0.7 vs 1.6 and 1.0μ M) than cauda epididymal or ejaculated sperm for which the kinetic constants did not differ significantly. In all sperm types, phloretin was a competitive inhibitor and cytochalasin B a noncompetitive inhibitor of 2-deoxy-D-glucose uptake. We conclude that transport rather than phosphorylation is the rate determining step in sugar uptake. Sugar transport is probably not the rate determining step in spermatozoal glycolysis under steady state conditions since the Vmax for transport is very much greater than the measured glucose consumption rate. The reduction in Km associated with sperm maturation may not reflect an important physiological modification of sugar metabolism since glucose concentrations in the female reproductive tract are >1 mM. However, measurement of 2-deoxyglucose transport provides a sensitive assay for changes in membrane function and our data document that membrane function is altered during sperm maturation.