Diffusion barriers in ram and boar sperm plasma membranes: Directionality of lipid diffusion across the posterior ring

The plasma membrane of mammalian spermatozoa, like that of other differenti ated cells, is compartmentalized into discrete regions or domains that are biochemically and functionally distinct from one another. Physical structur es within the membrane, such as the posterior ring at the juncture of the s perm head and tail, have long been thought to act as diffusion barriers to help segregate important molecules required for fertilization within specif ic domains and to regulate migration of molecules between domains. In this investigation, we used a quantitative photobleaching technique (video-FRAP) to assess the efficacy of the posterior ring as a barrier to exchange of l ipids between the postacrosomal and midpiece plasma membranes. A lipid repo rter probe (1,1'-diduodecyl-3,3,3',3'-tetramethylindocarbocyanine; DilC(12) ) was incorporated into the plasma membrane of live ram and boar spermatozo a, and the directionality of its diffusion across the posterior ring was me asured by line-profile analysis. Results showed that DilC(12) was able to t raverse the posterior ring from the direction of the postacrosomal plasma m embrane and to diffuse onto the midpiece plasma membrane. These results sug gest that the posterior ring is not an immutable barrier to lipid exchange in mature spermatozoa and that there are other mechanisms for maintaining i n-plane lipid asymmetry, such as differential phase behavior and interactio n with the submembranous cytoskeleton.