Initiation of the human sperm acrosome reaction by thapsigargin

Abstract

Previous studies have established that the mammalian sperm acrosome reaction (AR) is dependent upon an influx of extracellular Ca2+, but the involvement of a mobilizable store of intracellular Ca2+ has not been shown. In many other cells, the endoplasmic reticulum is the site of such a Ca(2+)-store. Here, we show that thapsigargin, a highly specific inhibitor of the endoplasmic reticulum Ca(2+)-ATPase Ca(2+)-pump (and thus a mobilizer of intracellular Ca2+) in other cells, can initiate the AR in capacitated human sperm. Thapsigargin at concentrations from 50-500 nM significantly increased the AR to the same extent when incubated with capacitated sperm for 1 min (assayed by indirect immunofluorescence). Transmission electron microscopy confirmed the occurrence of normal morphology in the AR initiated by thapsigargin. Thapsigargin (200 nM) did not initiate the AR in noncapacitated sperm. Initiation of the AR by thapsigargin apparently requires an influx of Ca2+ since 1 min preincubation with the calcium channel blockers La3+ (250 microM) or Ni2+ (250 microM) prior to addition of thapsigargin completely inhibits AR-initiation. Mobilization of an intracellular Ca(2+)-store by thapsigargin in capacitated human sperm may lead to an influx of extracellular Ca2+ and subsequently the AR. Putative sites for thapsigargin-sensitive intracellular Ca(2+)-stores in human sperm include the cytoplasmic droplet, the sperm nucleus and the acrosome.

Spermatozoa Epididymal Maturation in the Mexican Big-Eared Bat (Corynorhinus Mexicanus)

“…), respectively, with time during the 60 min incubation [Meizel and Turner 1993]. No statistical significant differences were found in the percentage of sperm undergoing the acrosomal reaction induced both with A23187 or progesterone.…”

The Intraacrosomal Calcium Pool Plays a Direct Role in Acrosomal Exocytosis

“…However, some contrary effects are observed in intact and permeabilized spermatozoa with inhibitors of the calcium pump. A plausible explanation for this difference is that, in intact cells, these compounds quickly trigger SOC channel opening (5,7,8) and acrosome reaction (3,4,9). Therefore, acrosomal exocytosis may occur long before the acrosomal calcium store has been depleted to a point that precludes membrane fusion.…”

“…Recent results from different laboratories suggest that a first transient cytosolic calcium increase (probably mediated by T-type calcium channels (2)) leads to a second sustained increase of cytosolic calcium that is necessary for the acrosome reaction (3)(4)(5)(6)(7)(8)(9). Although the connection between the two events is not completely clear, the current hypothesis is that the first calcium increase causes the activation of a phospholipase C (PLC).…”

“…PLC␦4, in particular, has been implicated in the early events of the acrosome reaction (12). Active PLC would produce inositol 1,4,5-trisphosphate (IP 3 ) that would open IP 3 -sensitive calcium channels in the membrane of intracellular stores. The emptying of these stores would trigger the opening of store-operated calcium (SOC) channels in the plasma membrane causing a second and sustained calcium increase that would trigger acrosomal exocytosis (1, 7).…”

Secretory pathway Ca2+-ATPase (SPCA1) Ca2+ pumps, not SERCAs, regulate complex [Ca2+]i signals in human spermatozoa

“…Thapsigargin elevates internal Ca 2+ in human sperm (Blackmore, 1993;Bonaccorsi et al, 1995;Williams and Ford, 2003) and stimulates acrosomal exocytosis in both human spermatozoa (Meizel and Turner, 1993;Rossato et al, 2001) and sperm of other mammals (Llanos, 1998;Dragileva et al, 1999). Specific inhibition of SERCAs by thapsigargin is beyond dispute (Thastrup et al, 1990), and has been well documented in many somatic cell types (Treiman et al, 1998), but data from studies on the action of thapsigargin on sperm are inconsistent.…”