[Changes in the neurite-glial interrelations under the influence of antiserum against galactocerebrosides (a vital tissue culture study)]. / Izmeneniia neirit-glial'nykh vzaimootnoshenii pri vozdeistvii antisyvorotki k galaktotserebrozidam (prizhiznennoe issledovanie kul'tury tkani).

Two--five-day-old culture of 10-11-day-old chick embryos has been used. Antiserum against galactocerebrosides (anti-GalC) is added to the nutrition medium before cultuvation. In the presence of anti-GalC the growth and gliocyte migration zone decreases, the area index becomes essentially small (3.5 +/- 0.7 in the control, 1.5 +/- 0.5 in the experiment). This is connected with inhibition of migration and proliferation of gliocytes during first 48 h of cultivation, while intensity of neuron regeneration remains unchanged. Alterations of the neurit-glial relations are investigated by means of the vital phase-contrast microscopy. Effect of anti-GalC to peripheral gliocytes is accompanied with a decreasing adhesive ability of their plasmolemma. This makes difficult their flattening on the neurit membrane, formation of contact membranous neurit-glial relations and formation of glial membranes. On the 3d day formation of nervous fasciculi is retarded, as well as their fusion into trunks and plexuses. On the 5th day, unlike the control, a continuous "epineural" covering of neuritic plexuses does not form. Round retractile and defective (with protrusions) forms of gliocytes predominate. These data demonstrate inhibitory effect of anti-GalC on the structural-functional maturation of the glia and on formation of neurit-glial relations in the culture of the sensitive ganglion.

Axonal transport of antibodies to subcellular and protein fractions of rat brain.

Experiments examined the feasibility of using the axonal transport of antibodies as a possible means to characterize nerve membrane composition and the fate of internalized macromolecules. Polyspecific antibodies were generated in rabbits against rat brain synaptosomal and microsomal subcellular fractions and against wheat germ agglutinin-binding proteins isolated by lectin affinity chromatography. Antisera were injected into the vitreal chamber of the eye and into the facial musculature of anesthetized rats to test, respectively, for anterograde transport in retinotectal neurons and for retrograde transport in facial motoneurons. Control injections of preimmune serum were made into the opposite side. After survival for 4-168 h, animals were perfused and the axonally transported rabbit immunoglobulins detected in frozen sections of the brainstem using a modified peroxidase-antiperoxidase immunocytochemical procedure. Antisera against all 3 classes of neuronal antigens contained antibodies that underwent retrograde axonal transport. No evidence of anterograde transport was seen. Neurons containing retrogradely transported immunoglobulins exhibited punctate as well as diffuse staining of the cytoplasm and proximal dendrites, exclusive of the nucleus. Following retrograde transport of antibodies to the synaptosomal fraction, staining of the neuropil around motoneurons was also observed, suggesting transcellular transport of these antibodies. Concentrations of injected antibodies as low as 1% of whole antiserum led to detectable retrograde transport. Increasing concentrations of antibodies above the amount in whole antiserum did not increase the intensity of staining in retrogradely labeled neurons, suggesting saturation. The findings support the view that antibodies to neural membranes are taken up and transported by binding to specific sites on nerve terminals.(ABSTRACT TRUNCATED AT 250 WORDS)

Rapid alterations of the axon membrane in antibody-mediated demyelination.

Alterations of nodal and paranodal axolemma of the rat sciatic nerve were investigated in antigalactocerebroside serum-induced demyelination. A ferric ion-ferrocyanide (FeFCN) stain that appears to stain the regions with a high sodium channel density in nerve fibers was applied. When acute conduction block was initiated 20 to 180 minutes after the antiserum injection, myelin terminal loops began to be detached from the paranodal axolemma and reaction product of FeFCN stain originally localized at the nodes decreased in density and extended to the paranodal axolemma. By the time that complete conduction block was established, 5 hours after the injection, FeFCN stain was barely detectable around the nodal area. The loss of staining was associated with detachment and vesiculovacuolar degeneration of the paranodal myelin. This rapid deterioration and disappearance of normal cytochemical characteristics of the axolemma in the presence of only modest paranodal demyelination could be a morphological correlate of the loss of excitability of the axon membrane.

Enzyme-linked immunosorbent assay (ELISA) for antibodies to human myelin and axolemma-enriched fractions.

The Enzyme-Linked Immunosorbent Assay (ELISA) is a well established procedure for antibody determination which has gained wide acceptance, particularly in diagnostic virology. We have adapted the method for use with the lipid rich antigens of human myelin and axolemma enriched fractions. Adsorption of the antigen onto the assay plates was rapid and relatively independent of pH. Antibodies to myelin and axolemma cross-reacted extensively. Little antibody reaction was noted using human liver microsomes, indicating the antibodies were specific but that myelin and axolemma shared at least one strong common antigen. With further purification of the antigen, this method should be useful in evaluating immunogenicity and antigenic purity of these membrane fractions.