Hamster sperm Na+, K+-adenosine triphosphatase: increased activity during capacitation in vitro and its relationship to cyclic nucleotides

Abstract

These in vitro studies of golden hamster sperm were undertaken to determine whether: Na+, K+-ATPase activity is required for capacitation; Na+, K+-ATPase activity is altered during capacitation; and cyclic nucleotides can control this enzyme activity. Hamster sperm were incubated in a medium in which capacitation occurred in an asynchronous manner and in which acrosome reactions began to occur after approximately 3.5 h of incubation. Inhibition of the hamster sperm acrosome reaction by the Na+, K+-ATPase inhibitor ouabain (1 .mu.M) added at Time (T) = 2 or T = 3 h could be fully reversed by the addition of the ionophore nigericin (0.1 .mu.M) at T = 3.5 h. When ouabain was added at T = 0 or T = 1 h, similar nigericin addition could not completely reverse the inhibition. Na+, K+-ATPase activity of hamster sperm increased by 2 h of incubation (compared to that measured initially after 15 min) and this activity remained elevated at 3.5 h. Addition of either monobutyryl (BtcAMP) (12.9 .mu.M) or BtcGMP (10.5 .mu.M), or the phosphodiesterase inhibitor SQ20009 [1-ethyl-4-(isopropylidene-hydrazino)-1H-pyrazolo-[3,4-b] pyridine-5- carboxylic acid ethyl ester hydrochloride] (10 .mu.M) at 2 h produced a stimulation of acrosome reactions at 4 and 5 h. While BtcGMP and SQ 20009 also induced a further increase in Na+, K+-ATPase activity measured at 3.5 h, BtcAMP had no effect. Intracellular cAMP and cGMP levels measured showed cAMP increased by 2 h and remained elevated when measured at 3.5 h, while cGMP could not be consistently detected at 15 min, 2 h or 3.5 h. Assays of high numbers of uncapacitated sperm did detect a low level of cGMP. Na+, K+-ATPase activity evidently increases in and is essential for early capacitation [and thereby eventually for the acrosome reaction (AR)] of hamster sperm. The increase in Na+, K+-ATPase activity occurring during capacitation is probably mediated by intracellular cGMP but not cAMP, although both cyclic nucleotides stimulate the hamster sperm AR.