RESUMO
When human sperm was incubated in medium deprived of glucose, glucose restoration caused a transient hyperpolarization of the plasma membrane. This hyperpolarization was also induced by fructose but not by 2-deoxyglucose, a substrate that cannot be metabolized. The hyperpolarization was inhibited by NaF, a glycolysis inhibitor, but not by mitochondrial inhibitors (cyanide, rotenone and antimycin), suggesting that it depended on glycolysis. Furthermore, the hyperpolarization was still induced in medium containing a high concentration of KCl and was insensitive to the K(+) channel blocker TEA and the Cl(-) channel blocker niflumic acid, but it was blocked by ouabain. This suggested that upon glucose addition, there was an increase in the concentration of ATP, that in turns increased the Na(+),K(+)-ATPase activity. Since this pump is electrogenic (2K(+)/3Na(+)) the plasma membrane hyperpolarized. On the other hand, CCCP, a proton ionophore, inhibited the hyperpolarization induced by glucose. When CCCP was added to glucose-treated hyperpolarized sperm, it caused a depolarization that triggered a Ca(2+) influx sensitive to nickel, an inhibitor of voltage-dependent calcium channels. Moreover, CCCP caused hyperpolarization in sperm incubated in medium without calcium, a known condition that depolarizes sperm. This indicated that CCCP induced proton permeability in the plasma membrane that was able to change the membrane potential to a value corresponding to the E(H) and that was also able to clamp it, so that it prevented the hyperpolarization induced by glucose.
