The molecular organization of nerve membranes : VI. The separation of axolemma from schwann cell membranes of giant and retinal squid axons by density gradient centrifugation.

Plasma membranes were isolated from two types of squid nerves which have morphologically, different ratios of axolemma/Schwannlemma (A/S). These membranes were studied by means of differential and density gradient centrifugation.Thoroughly dissected giant axons were used as membrane source having low A/S ratio. Retinal fibers were used as membrane source with high A/S ratio. A similar procedure for the isolation of the plasma membranes was used for both types of squid axons.Differential centrifugation showed that at 1,500×g, the yield of membrane enzymes (Na, K-ATPase and NADH-ferricyanide oxidoreductase) from giant axon homogenates was 2 to 5 times greater than from retinal nerve homogenates, but at 105,000×g the opposite was the case, the yield from retinal axons being about two times greater. Thus, the major part of the membrane material from the retinal nerve seems to be less dense than the membrane material from giant axons.The behavior of the 105,000×g fraction from both types of fibers was studied by determining protein Na, K-ATPase, and NADH-oxidoreductase along a lineal sucrose gradient (10 to 40%; centrifuged at 40,600×g for 90 min). By any of the three measurements, retinal axons yielded a greater amount (2:1) of plasma membranes sedimenting at low sucrose concentration (16 to 25%) as compared to that observed at high sucrose concentration (35 to 38%). Giant axons, on the contrary, yielded a higher proportion of membranes (2.5:1) sedimenting at high sucrose concentrations (over 40%).The experimental data indicate that a different cellular origin can account for the behavior of nerve membranes along lineal gradient centrifugation. The membranes floating at low sucrose concentration ("light membranes") can be tentatively ascribed to the axolemma; the membranes found at high sucrose concentration ("heavy membranes") to the Schwannlemma and basement membranes.In accord with their high A/S morphological ratio, squid retinal axons yielded 5 times more light membranes (axolemma) than dissected giant axons.