Role of Cholesterol in Sperm Capacitation

TC # 216 BIOLOGY OF REPRODUCTION 59, 7–11 (1998) M i n i r ev i ew Role of Cholesterol in Sperm Capacitation1 Nicholas L. Cross2 Department of Anatomy, Pathology, and Pharmacology, Oklahoma State University, Stillwater, Oklahoma 74078 Forty years ago, M.C. Chang found that exposing capacitated rabbit sperm to seminal plasma made them incapable of fertilizing eggs in vivo [1]. The results of this rather unphysiological experiment had a strong impact on the study of sperm capacitation, which Chang [2] and Austin [3] had discovered six years earlier. It was widely believed that understanding how seminal plasma reverses capacitation would shed light on how sperm become capacitated, and considerable effort was devoted to identifying the inhibitory agents in seminal plasma. The candidates included proteins, peptides, and lipids [4, 5]. Beginning with his work on the inhibitory activity of seminal plasma, Brian Davis and his coworkers built a strong case for lipids playing an important role in capacitation. Davis showed that sperm could be ‘‘decapacitated’’ by a vesicle fraction prepared from seminal plasma (reviewed in [6]). The activity of the fraction was diminished by partially extracting the lipids and could be mimicked by synthetic phospholipid vesicles containing cholesterol. Davis suggested that the vesicles changed the lipid composition of the sperm plasma membrane and, secondly, that the changes were the reverse of alterations that normally occur during capacitation. In Davis’s model the cholesterol/phospholipid (C/PL) ratio of the sperm plasma membrane determines the capacitation state of the sperm. A freshly ejaculated sperm has a high C/PL ratio; and during capacitation, cholesterol moves from the sperm membrane to soluble protein acceptors, and/ or phospholipid moves into the sperm membrane. At the time this model was formulated, it was generally believed that capacitation caused sperm to acrosome-react spontaneously (that is, without exposure to a specific inducer). In Davis’s model, the falling C/PL ratio triggered an acrosome reaction. Explanations for how this might happen were based on information obtained in other systems; and it was suggested that a lower C/PL ratio decreased the membrane microviscosity, relaxed the packing of phospholipids in the membrane, and perhaps permitted greater calcium influx, all leading through unspecified intermediate steps to fusion of the plasma and outer acrosomal membranes. The early work that supported a role for cholesterol in the control of sperm function has been extensively reviewed [6–9]. The present review will emphasize more recent work and will summarize current information regarding how cholesterol might act. To conform to the format of CONTENT AND DISTRIBUTION OF STEROLS IN MAMMALIAN SPERM In most species that have been studied, cholesterol is the major sterol of ejaculated sperm. Other sterols are often present, including desmosterol (the immediate precursor of cholesterol), cholesta-7,24-dien-3b-ol, desmosterol sulfate, cholesterol sulfate, and cholesterol esters [10–14]. Some of these molecules have been suggested to be important regulators of sperm function [15], but this review will focus on cholesterol because of its abundance. The extent to which sperm can interconvert these forms needs to be better understood. Most of the work that will be discussed below assumes that cholesterol is the active agent, but in fact a molecule whose concentration mirrors the concentration of cholesterol could be just as important or even more so. In somatic cells, most unesterified cholesterol is in membranes, with the plasma membrane containing the highest concentration [16]. Consistent with this distribution, the C/ PL ratio of isolated sperm plasma membrane fraction is higher than that of the acrosomal membrane fraction [17]. Studies employing filipin, a polyene antibiotic that binds to 3b-hydroxysterols to form complexes visible by electron microscopy, confirm this distribution and also suggest that sterols are not uniformly distributed in the plane of the plasma membrane. Filipin-sterol complexes are concentrated in the plasma membrane overlying the acrosome, with less in the posterior region of the head and the flagellum [18]. These observations should be interpreted with caution, however, because filipin-sterol complexes may redistribute in the plane of the membrane [19]. The concentration of cholesterol in the sperm plasma membrane varies considerably among species. The C/PL molar ratios for isolated plasma membrane fractions are about 0.20 in boar sperm [20], 0.36 in stallion sperm [20], about 0.40 in bovine sperm [17], 0.43 in ram sperm [21], and 0.83 in human sperm [22]. Sperm plasma membrane fractions usually originate principally from the anterior head, so these ratios may be more typical of that region than of the entire plasma membrane. It is important to note that these C/PL ratios are not unusual for plasma membranes. The red blood cell membrane has a C/PL molar ratio of about 0.9, and the ratios in plasma membranes of nucleated cells range from 0.4 to 0.8 [23]. These data argue against the view that a recently ejaculated sperm is incapable of acrosomal exocytosis because its plasma membrane is somehow ‘‘frozen’’ by an extremely high concentration of cholesterol. What is the source of sperm cholesterol? Human sperm Accepted February 22, 1998. Received November 20, 1997. 1 Supported by NIH grant HD30763 and the Oklahoma Center for the Advancement of Science and Technology. 2 Correspondence: FAX: (405) 744-8263; e-mail: 7 a minireview, not all of the available literature is cited, and preference is given to citing recent papers; their bibliographies will lead to earlier work. INTRODUCTION TC # 216 8 CROSS CHOLESTEROL AND SPERM FUNCTION IN VITRO When sperm are incubated in a capacitating medium in vitro, the sperm cholesterol content gradually decreases [25, 35]. The loss of cholesterol from human sperm is initially linear, but acrosomal responsiveness appears only after a delay of several hours, suggesting that cholesterol loss precedes the development of responsiveness [25]. Incubation reduces the density of filipin-sterol complexes in the plasma membrane of the anterior sperm head [36], or in some cases, causes clearing of filipin-sterol complexes from small areas of the membrane [37]. Where does the sperm cholesterol go? It must either be converted to another molecule or exit the cell. Davis et al. [38] reported that sperm can convert a substantial portion of radiolabeled cholesterol to another form, but the material was not identified, and little is known about the ability of sperm to carry out such reactions. Current thinking is that most cholesterol loss is due to slow diffusion from the cell, and a net transfer of cholesterol from rat and bovine sperm to the medium has been demonstrated [38, 39]. Most incubation media contain serum albumin, which binds cholesterol efficiently, reducing (...truncated)