In this study we examined the effect of cholesterol (Diet 2), cholesterol and fish oil (FO) polyunsaturated acid (Diet 3), and polyunsaturated acid (Diet 4) enriched diets upon the acrosome reaction (AR) of New Zealand White rabbit spermatozoa. Male rabbits fed with cholesterol alone or with FO increased their cholesterol and LDL-cholesterol serum levels after 15 days of diet. Ten semen samples were obtained after 2 months of diet. Our results suggest that hypercholesterolemia and hypertriglyceridemia in male rabbits could produce a decreased capacity of sperm AR after 4 h (0%, 0%, and 60% lower than the control), 6 h (0%, 68%, and 44%), or 8 h (58%, 52% and 32%) of incubation in capacitating medium. Another set of experiments were made with lysophosphatidylcholine (LPC), 80 micrograms/ml, and the same pattern of AR was seen. Nevertheless, the high cholesterol and total lipids (TL) levels in serum did not affect the cholesterol levels in seminal plasma (SP) but affect the SP total lipids. The diminished capacity of rabbit sperm to undergo the AR was not reverted by in vitro incubation with the Shinitsky medium for cholesterol depletion (MDC). These results indirectly suggest that the cholesterol/phospholipid ratio in hypercholesterolemic sperm is similar to that of controls and are in agreement with preliminary studies made in our laboratory that evidenced the same cholesterol/phospholipid ratio in rabbit sperm from hypercholesterolemic animals than from controls.(ABSTRACT TRUNCATED AT 250 WORDS)

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Summary. The distribution of membrane filipin‐sterol complexes (FSC) was examined ultra‐structurally in cauda epididymal sperm from normal and hypercholesterolaemic rabbits. Membrane FSC were quantitatively analysed on replicas of filipin‐treated cells. We determined a significant difference in FSC concentration in the plasma membrane of the acrosome region (PMAR) of hypercholesterolaemic animals compared to normal rabbits. Hypercholesterolaemic animals had 0.56 ± 0.05 FSC complex per μm2 (enriched Cholesterol diet: Diet 2) in the marginal segment of PMAR; 0.62 ± 0.05 FSC complex per μm2 (enriched Cholesterol and fish oil diet: Diet 3) and only 0.28 ± 0.01 FSC complex per μm2 for normal animals (Control Diet 1). In the principal (anterior) segment we found 0.54 ± 0.10 FSC complex per μm2 (Diet 2), 0.56 ± 0.03 FSC complex per μm2 (Diet 3) and 0.30 ± 0.04 FSC complex per μm2 (Control Diet 1). We also counted 0.47 ± 0.1 FSC complex per μm2 in the equatorial segment of PMAR for Diet 2, 0.27 ± 0.05 and 0.28 ± 0.04 FSC complex per μm2 in Diet 1 and Diet 3 respectively. Diet 4 (fish oil) did not differ from the control. An increase in the Cholesterol (Chol) level in biological membranes or a difference in the Chol membrane domains could cause a variation in the membrane rigidity that could modify the sperm membrane fusion capacity and functionality. The results presented in this paper are in agreement and could explain the decrease in the kinetic of the sperm acrosome reaction that we have observed in experimentally hypercholesterolaemic rabbits (Díaz‐Fontdevila & Bustos‐Obregón, 1992).

The distribution of membrane filipin sterol complexes (FSC) in the plasma membrane of the acrosomal region (PMAR) of rabbit sperm from epididymis and testis, in normal and hypercholesterolaemic rabbits, was examined at ultrastructural level. Membrane FSC were quantitatively analysed on freeze fracture replicas of filipin-treated cells. Cauda epididymal sperm shows a significant increase in filipin sterol complexes concentration in PMAR of hypercholesterolaemic animals compared to normal rabbits. Hypercholesterolaemic animals had 0.53 k0.08 FSC pm-2 in the marginal segment of PMAR and 0.26f0.03 FSC pmP2 for normal animals. In the principal piece we found 0.70 f 0.07 FSC pm-' for hypercholesterolaemic and 0.43 f0.03 FSC pm-' for control animals. We also counted 0.58k0.04 FSC pm-' in the equatorial segment of PMAR for hypercholesterolaemic and 0.38f0.03 FSC pmP2 for normal animals respectively. The FSC concentration of testicular sperm, like sperm from corpus and caput of epididymis in hypercholesterolaemic animals, did not differ from the controls. Cholesterol, phospholipids and cholesterol : phospholipid ratio in caudal epididymal sperm from treated males did not differ from controls. Only the sphingomyelin concentration decreases in cauda epididymal sperm from hypercholesterolaemic males compared to controls.The results presented in this paper suggest that the lipidic domains in PMAR of hypercholesterolaemic rabbits changes when the gametes go through the epididymis. Correspondence: Dra. Diaz-Fontdevila Marina, Dpto. Biologia del Desarrollo INSIBIO (CONICET-UNT), Universidad Nacional de Tucuman. Chacabuco 46 1 (4000) Tucuman, Republica Argentina.