Water Channels in Human Sperm Volume Regulation.

Abstract

Human spermatozoa have to regulate their volume as they pass from the male to the female tracts where they are exposed to lower osmolality fluids that cause cell swelling. Regulatory volume decrease (RVD) of human spermatozoa involves removal of cellular water following passive osmolyte efflux through K+ channels, Cl- channels and K+-Cl- co-transporters. As water itself cannot pass through K+ channels it must pass through other channels. Several channels are involved in water efflux in somatic cells: aquaporins (AQPs), Cl- channels, facilitative glucose transporters (GLUTs) and ion-dependent solute [glucose, urea] transporters (SGLTs). AQP1 is not found in human spermatozoa but AQP7 is. In this study human spermatozoa were incubated in hypotonic media with water channels inhibitors to see if they prevented water influx. An experimental paradigm of pre-incubation of Percoll-washed ejaculated spermatozoa from liquefied semen with the inhibitors before the experimental, physiological osmotic challenge was applied. The inhibitors used were the AQP inhibitors mercuric chloride (Hg2+) and silver nitrate (Ag+); the GLUT inhibitor phloretin (Pht: 100, 800 μM) and the SGLT inhibitor phloridzin (500 μM). The K+-channel blocker quinine (800 μM) was used to inhibit RVD and induce swelling of control cells. Sperm volume was measured continuously over 5 min by flow cytometric measurements of laser forward scatter from viable (propidium iodide-excluding) cells. Incubation of human spermatozoa in hypotonic medium with 25 μM Hg2+ or 15 μM Ag+ alone did not reduce viability or alter sperm size from that of the controls. Both Hg2+ and Ag+ completely blocked the increase in volume normally induced by quinine, as there was no difference in cell volume in the presence or absence of quinine with either ion. Incubation of human spermatozoa in hypotonic medium with 100 μM and 800 μM Pht alone did not alter viability or sperm volume but in the presence of quinine both concentrations increased sperm size above that induced by quinine alone. Incubation of human spermatozoa in hypotonic medium with 500 μM phloridzin did not alter viability (78%) or sperm volume or affect the swelling induced by of quinine (data not shown). The results indicate that water movement into spermatozoa under physiologically hypotonic insult is regulated by Hg2+- and Ag+-sensitive channels. As AQP8, but not AQP7, is Hg2+- and Ag+-sensitive and AQP8 is present in human spermatozoa (unpublished data), this channel may be involved in water transport of human spermatozoa. The effect of phloretin in increasing cell volume only in the presence of quinine suggests that osmolytes other than K+ participate in RVD when K+ efflux is blocked by quinine. Phloretin may inhibit this other osmolyte efflux or could prevent water efflux through GLUTs. Sodium-coupled solute transporters seem not to be involved since phloridzin had no effect on volume regulation.