Bicarbonate stimulated phospholipid scrambling induces cholesterol redistribution and enables cholesterol depletion in the sperm plasma membrane

Mammalian sperm cells are activated prior to fertilization by high bicarbon ate levels, which facilitate lipoprotein-mediated cholesterol efflux. The r ole of bicarbonate and cholesterol acceptors on the cholesterol organizatio n in the sperm plasma membrane was tested. Bicarbonate induced an albumin-i ndependent change in lipid architecture that was detectable by an increase in merocyanine staining (due to protein kinase A-mediated phospholipid scra mbling). The response was limited to a subpopulation of viable sperm cells that were sorted from the non-responding subpopulation by flow cytometry. T he responding cells had reduced cholesterol levels (30% reduction) compared with non-responding cells. The subpopulation differences were caused by va riable efficiencies in epididymal maturation as judged by cell morphology. Membrane cholesterol organization was observed with filipin, which labeled the entire sperm surface of non-stimulated and nonresponding cells, but lab eled only the apical surface area of bicarbonate-responding cells. Addition of albumin caused cholesterol efflux, but only in bicarbonate-responding c ells that exhibited virtually no filipin labeling in the sperm head area. A lbumin had no effect on other lipid components, and no affinity for cholest erol in the absence of bicarbonate. Therefore, bicarbonate induces first a lateral redistribution in the low cholesterol containing spermatozoa, which in turn facilitates cholesterol extraction by albumin. A model is proposed in which phospholipid scrambling induces the formation of an apical membra ne raft in the sperm head surface that enables albumin mediated efflux of c holesterol.