Cholesterol movement between bovine rod outer segment disk membranes and phospholipid vesicles

Abstract

The ability of cholesterol to move between bovine rod outer segment disk membranes and phospholipid membranes was examined. Disk membranes were incubated with small unilamellar phospholipid vesicles containing varying amounts of cholesterol. Aliquots were removed at specific times, and then the disks and the vesicles were separated by centrifugation and assayed for phospholipid and cholesterol content. When incubated with vesicles containing no cholesterol, the cholesterol to phospholipid ratio in the disk membrane was reduced due to migration of cholesterol from the disks into the vesicles. The cholesterol content of these cholesterol depleted disks could be readily returned to the normal disk cholesterol content by incubation of the cholesterol-depleted disks with small unilamellar vesicles containing high cholesterol. An apparent partition coefficient K was calculated as the quotient of the cholesterol/phospholipid mole ratio in the donor membranes and the cholesterol/phospholipid mole ratio in the acceptor membranes. The value of K was approximately 1 at cholesterol levels below normal disk cholesterol content, for disk membranes and phosphatidylcholine small unilamellar vesicles. Inclusion of phosphatidylethanolamine in the small unilamellar vesicle acceptor raised K, indicating that phosphatidylethanolamine creates an unfavourable environment for cholesterol. The cholesterol to phospholipid ratio of native disks could be increased by incubation with phosphatidylcholine small unilamellar vesicles (donor) which contained higher amounts of cholesterol than the disk membrane acceptor. In these experiments the distribution of cholesterol between disks and small unilamellar vesicles always favored the vesicles. The apparent partition coefficient was 1.7 at several cholesterol levels above the native disk cholesterol content. Liposomes made from lipid extracted from the disk membrane behaved in the same manner as intact disks with respect to cholesterol distribution at equilibrium. The phospholipid content of the disk membrane may be an important factor in determining the cholesterol content of the disk membrane.