Visualization of GM1 with Cholera Toxin B in Live Epididymal Versus Ejaculated Bull, Mouse, and Human Spermatozoa1

Abstract

The organization of membrane subdomains in mammalian sperm has recently generated controversy, with several reports describing widely differing localization patterns for the ganglioside GM1. Using the pentameric B subunit of cholera toxin (CTB), we found GM1 to be restricted to the plasma membrane overlying the acrosome in the heads of live murine sperm. Interestingly, CTB had minimal binding to live bovine and human sperm. To investigate whether this difference in GM1 localization was because of species differences or differences between collection from the epididymis (mouse) or an ejaculate (bull, human), we examined epididymal bovine and human sperm. We found that GM1 localized to the plasma membrane overlying the acrosome in sperm from these species. To determine whether some component of seminal plasma was interfering with the ability of CTB to access GM1, we incubated epididymal mouse sperm with fluid from murine seminal vesicles and epididymal bull sperm with bovine seminal plasma. This treatment largely abolished the ability of the CTB to bind to GM1, producing a fluorescence pattern similar to that reported for the human. The most abundant seminal plasma protein, PDC-109, was not responsible for this loss. As demonstration that the seminal plasma was not removing GM1, sperm exposed to seminal plasma were fixed before CTB addition, and again displayed fluorescence over the acrosome. These observations reconcile inconsistencies reported for the localization of GM1 in sperm of different species, and provide evidence for the segregation of GM1 to a stable subdomain in the plasma membrane overlying the acrosome.

Mouse sperm undergo GPI-anchored protein release associated with lipid raft reorganization and acrosome reaction to acquire fertility

“…GM1 is a characteristic component and reliable biomarker of lipid rafts. Other studies showed that it dynamically moves with methyl--cyclodextrin (M--CD) treatment in the mouse sperm membrane (Buttke et al, 2006;Selvaraj et al, 2007). Here, we found that M-…”

Protective effects of exogenous gangliosides on ROS-induced changes in human spermatozoa

“…The endogenous ganglioside GM1 has been found in the plasma membrane overlying the acrosome of the sperm head in several mammalian species 44 or over the whole sperm surface in human spermatozoa. 45 GM1 is known to influence several signalling pathways in somatic cells and its importance in sperm functional maturation, such as capacitation, has been recently established.…”

“…It has been localized in lipid raft microdomains, which serve as platforms for the assembly of key recognition molecules on the sperm surface during capacitation. [44][45][46][47] In animal sperm, increasing evidence suggests that capacitation is regulated by the redistribution of GM1 on the plasma membrane. 48 For example, when murine sperm migrate to the uterine cavity, GM1 shifts from the postacrosomal region and disappears from the sperm head in the oviduct, at which time, spermatozoa can undergo the acrosome reaction.…”

Remodelling of Lipid Rafts during In vitro Capacitation and Acrosome Reaction of Ram Spermatozoa

“…Conversely, GM1 was reported as nonexistent or patchy and inconsistent throughout the whole sperm cell of human and boar . One likely difference among these studies could be the fixation procedure [60], which might affect viability. In fact, it has been reported that GM1 moves rapidly from its position overlying the acrosome to the post-acrosomal area after cell death in unfixed or lightly fixed samples [59].…”