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Lipids of plasma membrane and outer acrosomal membrane from bovine spermatozoa
J E Parks1, J W Arion, R H Foote
1Department of Animal Science, Cornell University, Ithaca, New York 14853.
Abstract:
Plasma membrane (PM), primarily from the anterior sperm head, and outer acrosomal membrane (OAM), were isolated from ejaculated bovine spermatozoa, and the major lipid classes were characterized. Whole sperm (WS) lipids were analyzed for comparison. PM was removed by nitrogen cavitation and purified by sucrose density-gradient centrifugation. The OAM was removed by centrifugation through hyperosmotic sucrose and recovered by sucrose density-gradient centrifugation. The PM contained primarily spherical vesicles from the region overlying the OAM and was enriched 9- and 13-fold in 5'-nucleotidase and alkaline phosphatase activity, respectively, compared to the original cavitate. The OAM was recovered as caplike structures with associated ground substance. Protein, phospholipid, and cholesterol (PR, PL, and CH as micrograms/5 x 10(9) sperm) were 300, 467, and 93 for PM and 276, 111, and 25 for OAM, respectively. Corresponding values for WS (mg/5 x 10(9) sperm) were 31.4, 6.63, and 0.72. The PR/PL (w/w) and CH/PL (mol/mol) ratios were 0.66 and 0.38 for PM; 2.48 and 0.26 for OAM; and 4.39 and 0.22 for WS. Cholesterol was the only free sterol detected by gas/liquid chromatography in WS, PM, and OAM, with traces of CH sulfate present in all three preparations. Glycolipid tentatively identified as sulfogalactolipid was detected by thin-layer chromatography (TLC) in PM but not OAM. Phospholipid composition of WS and membranes was determined by TLC. Cardiolipin (3% of total PL) was present in WS only. Choline, ethanolamine, and inositol phosphoglycerides (CP, EP, PI, PIP, PIPP); sphingomyelin (SP); phosphatidylserine (PS); and lysophosphatidylcholine (LPC) were present in WS, PM, and OAM. Approximately 50% of total PL was CP in all preparations; SP was 13% of PL in PM and 17% in OAM (p less than 0.05); EP was 7% of PL in PM and 10% in OAM (p less than 0.05). The differences in composition between PM and OAM is discussed with respect to capacitation and ability of sperm to undergo the acrosome reaction.
Insights
This study characterizes lipids in bovine sperm membranes, revealing distinct compositions in the plasma membrane (PM) and outer acrosomal membrane (OAM). These lipid differences are crucial for understanding sperm capacitation and the acrosome reaction.
Area of Science:
- Reproductive Biology
- Cell Biology
- Biochemistry
Background:
- The plasma membrane (PM) and outer acrosomal membrane (OAM) of spermatozoa are critical for fertilization.
- Understanding their lipid composition is essential for elucidating sperm function, including capacitation and the acrosome reaction.
Purpose of the Study:
- To isolate and characterize the major lipid classes of the PM and OAM from ejaculated bovine spermatozoa.
- To compare the lipid profiles of the PM and OAM with whole sperm (WS) lipids.
Main Methods:
- Isolation of PM and OAM using nitrogen cavitation and hyperosmotic sucrose centrifugation.
- Purification of membranes via sucrose density-gradient centrifugation.
- Analysis of lipid classes, including phospholipids and sterols, using gas/liquid chromatography and thin-layer chromatography (TLC).
Main Results:
- The PM and OAM exhibited distinct lipid compositions compared to WS.
- Cholesterol was the predominant free sterol, with traces of cholesterol sulfate present in all preparations.
- Specific phospholipid differences were noted, with sphingomyelin and ethanolamine phosphoglycerides showing higher percentages in OAM compared to PM.
Conclusions:
- The distinct lipid profiles of the bovine sperm PM and OAM suggest functional specializations.
- These compositional differences likely play a significant role in regulating sperm capacitation and the acrosome reaction process.