G_M1 Dynamics as a Marker for Membrane Changes Associated With the Process of Capacitation in Murine and Bovine Spermatozoa

Abstract

ABSTRACT:We previously showed that in live murine and bovine sperm heads, the ganglioside G M1 localizes to the sterol-rich plasma membrane overlying the acrosome (APM). Labeling G M1 using the pentameric cholera toxin subunit B (CTB) induced a dramatic redistribution of signal from the APM to the sterol-poor postacrosomal plasma membrane (PAPM) upon sperm death. We now show a similar phenomenon in the flagellum where CTB induces G M1 redistribution to sterol-poor membrane subdomains of the annulus and flagellar zipper. Because sterol efflux from the plasma membrane is required for capacitation, we examined whether G M1 localization might be useful to detect membrane changes associated with capacitation and/or acrosomal exocytosis. First, incubation of murine and bovine sperm with their respective stimuli for capacitation did not change G M1 distribution in live cells. However, incubation of sperm of both species with specific stimuli for capacitation, followed by the use of specific fixation conditions, induced reproducible, stimulus-specific patterns of G M1 distribution. By assessing changes in G M1 distribution in response to progesterone-induced AE, we show that these patterns reflect the response of murine sperm populations to capacitating stimuli. These data suggest that G M1 localization can be used as a diagnostic tool for evaluating sperm response to stimuli for capacitation and/or AE. Such information could be useful when deciding between technologies of assisted reproduction or when screening for male fertility. Furthermore, stimulus-specific changes in G M1 distribution showed that sperm could respond to NaHCO 3 or mediators of sterol efflux independently, thereby refining existing models of capacitation.

Sperm head membrane reorganisation during capacitation

“…Incubation of Percoll-washed sperm in an IVF medium caused the redistribution of both lipid raft marker proteins into the more confined region of the apical ridge and the labelling appears as a clustering of the punctuated structures that were observed prior to the incubation (van Gestel et al, 2005a). Similar dynamics were described by other groups (Shadan et al, 2004;Cross, 2004;Selvaraj et al, 2007) Interestingly, it is possible to isolate a detergent resistant membrane (DRM) fraction at 4°C using 0.1% Triton X-100 or similar detergent strength buffers. The non-solubilized membranes floated over a discontinuous density gradient and this floating fraction containing the DRM.…”

Tssk6 is required for Izumo relocalization and gamete fusion in the mouse

“…Deficient actin polymerization in Tssk6-null sperm Changes in the immunofluorescence pattern of sperm surface proteins during capacitation and acrosome reaction have been described previously (Miranda et al, 2009;Saxena et al, 1986;Primakoff and Myles, 2007;Selvaraj et al, 2007). It has been shown that changes in protein localization are at least in part due to actincytoskeleton dynamics (Saxena et al, 1986;Hernandez-Gonzalez et al, 2000).…”

Calpain modulates capacitation and acrosome reaction through cleavage of the spectrin cytoskeleton

“…In the same way as the principal piece, the cytoskeleton associated with PM in the acrosomal region is mainly composed of spectrin/F-actin, but with the difference that it is remodeled during capacitation. During capacitation and AR, proteins associated with PM as well as with membrane lipids compartmentalize in the acrosome region; they are redistributed inside the acrosome or distribute towards the equatorial segment and postacrosomal region as well (Rochwerger & Cuasnicu 1992, Da Ros et al 2004, Selvaraj et al 2007, Tsai et al 2007, Pasten-Hidalgo et al 2008. These molecules in some way are maintained as compartmentalized in the acrosome before capacitation.…”