Filipin-sterol complexes in disrupted myelin in the rat.

Using filipin as a cytochemical probe for cholesterol we have compared the distribution of filipin labelling in mildly disrupted myelin and normal myelin. The myelin lamellae in rat sciatic nerve were separated either by hypotonic saline (0.035-0.07 M) or nerve section (24-32 h) before aldehyde fixation and filipin treatment. Myelin separation was assessed in ultrathin sections and filipin distribution in freeze-fracture replicas. In separated myelin lamellae filipin labelling was similar throughout the myelin sheath while in normal control myelin filipin occurred most in the outer (abaxonal), least in the inner (adaxonal) and intermediate in the middle lamellae. It is concluded that this heterogeneous filipin labelling in normal myelin is a result of diffusion gradients to filipin within the myelin sheath and that in vivo cholesterol is uniformly distributed throughout all the lamellae of the myelin sheath. The site of the diffusion barrier to filipin within normal myelin is considered.

Distribution of filipin-sterol complexes in the unmyelinated nerve fibre.

The filipin-sterol technique was employed, together with freeze-fracture, to investigate the fine structure of the unmyelinated nerve fibre in the peripheral nerve. No heterogeneity was observed in the distribution of filipin labelling either in the Schwann cell plasma membrane or along the axolemma. The distribution of labelling is contrasted with that in the myelinated nerve fibre and related to the relative morphology and electrophysiology of the two types of fibre.

Filipin-sterol complexes at Schmidt-Lanterman incisures.

Employing the freeze-fracture technique, the distribution of filipin-sterol complexes was determined for membranes of peripheral nerve myelin. A heterogeneous distribution of complexes was observed with the greatest abundance on membranes associated with the cytoplasmic channels of Schmidt-Lanterman and longitudinal incisures. In addition there was an irregular network of well-labelled membrane bands in compact myelin. The results are related to a possible role for these channels and bands in the biochemical turnover of cholesterol in myelin.

Filipin-sterol complexes at nodes of Ranvier.

Using the filipin-sterol technique, regional heterogeneity in the axonal and Schwann cell plasma membranes was investigated at the node of Ranvier and paranodes. Filipin-sterol complexes were abundant at the nodal axolemma but infrequent throughout the paranodal axolemma. The paranodal Schwann cell plasma membrane was rich in complexes which extended over the nodal Schwann cell microvilli. There were no regional differences in filipin labelling of the nodal-paranodal Schwann cell plasma membrane in relation to features such as paranodal cytoplasmic columns or mesaxonal furrows. However, the paranodes of adjacent Schwann cells were sometimes markedly different from each other in the amount of filipin labelling. The extent to which filipin labelling is indicative of cholesterol membrane content is discussed and the findings are related to current concepts of distribution, mobility and interaction of protein and lipid in biomembranes, with particular reference to the nodal axolemma.

Distribution of filipin-sterol complexes in the myelinated nerve fiber.

Using filipin as a cytochemical probe to reveal the distribution of cholesterol, myelinated peripheral nerve fibers were examined in freeze-fracture replicas. Filipin-sterol complexes were most abundant in the Schwann cell and axonal plasma membranes. In the Schwann cell plasma membrane there was no heterogeneity in complex distribution in relation to the subjacent cytoplasmic network. In myelin lamellae there was a decrease in complexes from outer to inner lamellae and some aggregation of complexes in individual lamellae. The density of complexes in cytoplasmic organelles varied from absent in mitochondria to high in lysosome-like bodies. The results are interpreted in terms of the related biochemical composition and biophysical properties of cell membranes, with particular reference to the myelinated nerve fiber. The influence of diffusion barriers and gradients on the formation of complexes by filipin is considered.