Acrosomal alkalization triggers Ca²⁺ release and acrosome reaction in mammalian spermatozoa

Abstract

The sperm acrosome reaction (AR), an essential event for mammalian fertilization, involves Ca permeability changes leading to exocytosis of the acrosomal vesicle. The acrosome, an intracellular Ca store whose luminal pH is acidic, contains hydrolytic enzymes. It is known that acrosomal pH (pH ) increases during capacitation and this correlates with spontaneous AR. Some AR inducers increase intracellular Ca concentration ([Ca ] ) through Ca release from internal stores, mainly the acrosome. Catsper, a sperm specific Ca channel, has been suggested to participate in the AR. Curiously, Mibefradil and NNC55-0396, two CatSper blockers, themselves elevate [Ca ] by unknown mechanisms. Here we show that these compounds, as other weak bases, can elevate pH , trigger Ca release from the acrosome, and induce the AR in both mouse and human sperm. To our surprise, μM concentrations of NNC55-0396 induced AR even in nominally Ca free media. Our findings suggest that alkalization of the acrosome is critical step for Ca release from the acrosome that leads to the acrosome reaction.

Cellular and molecular aspects of oocyte maturation and fertilization: a perspective from the actin cytoskeleton

“…The role of Ca 2+ in the regulation of the AR in sea urchin sperm was first suggested by Dan [98], and further evidence was provided by use of the Ca 2+ ionophore A23187 or by simply increasing the external pH in the presence of Ca 2+ . Both treatments initiated the internal polymerization of actin in the acrosomal process [99][100][101][102]. The sperm of starfish has been a particularly useful model to study the egg-jelly signaling molecules for triggering AR as well as for the role played by actin filaments in sperm-egg interaction and penetration (see below).…”

Quantitative Intracellular pH Determinations in Single Live Mammalian Spermatozoa Using the Ratiometric Dye SNARF-5F

“…To understand how sperm pH i is regulated, it is indispensable to determine where and when it changes in individual cells. Although sperm pH i measurements in suspension have been performed using fluorescence indicators for more than three decades (Schackmann and Boon Chock, 1986;Darszon et al, 2004;Hamzeh et al, 2019), there are few reports of imaging single sperm pH i (Zeng et al, 1996;Chávez et al, 2014Chávez et al, , 2018González-Cota et al, 2015). All these experiments were performed with BCECF (Rink et al, 1982), the most popular fluorescent pH i indicator in cell physiology.…”

pH-dependent effects of procaine on equine gamete activation†

“…Percoll-separated sperm was washed twice using NCM and stained with 5 μM of the cytoplasmic pH-sensitive dye BCECF-AM [10, 11] or with 5 μM of the cell organelle pH-sensitive marker Lysosensor green DND-189 [28] in NCM, by incubation at 38.5 °C for 30 min. The extracellular dye was then removed by washing the sperm twice in NCM (400 g, 5 min).…”

“…Moreover, weak bases are able to partition into acidic cell organelles, alkalinizing them and releasing Ca 2+ from them [26, 27]. Recently, Chávez et al [28] reported a similar action of weak bases resulting in the acrosome reaction in mouse and human sperm. In contrast, Loux et al [11] suggested that the relationship between hyperactivated motility and Ca 2+ influx is likely to be weak in stallion sperm cells because analysis of the equine CatSper1 protein revealed species-specific differences in the structure of the pH sensor region.…”