Normal basal laminas are realized on dystrophic Schwann cells in dystrophic in equilibrium shiverer chimera nerves.

Multiple discontinuities are observed in the basal laminas of Schwann cells in mature dystrophic mice. To explore the pathogenesis of this abnormality we have exploited a dystrophic in equilibrium shiverer mouse chimera preparation in which both the basal lamina phenotype and the genotype of myelin-forming Schwann cells can be determined. If the basal lamina abnormality were to arise from an intrinsic deficiency of the dystrophic Schwann cell itself, only those Schwann cells of dystrophic genotype could express the mutant phenotype, whereas the coexisting population of shiverer Schwann cells should express typically normal basal laminas. No such distinction was observed; rather both dystrophic and shiverer Schwann cells were found to express relatively normal basal laminas and two pathogenetic mechanisms remain theoretical possibilities. The dystrophic Schwann cell population may be intrinsically defective but also may be rescued by obtaining the normal product of the dy locus synthesized by the coexisting shiverer cells. Alternatively, an extra Schwann cell deficiency existing within dystrophic mice may be normalized by shiverer cells and the normal intrinsic potential of both dystrophic and shiverer Schwann cells can then be realized. Regardless of the exact mechanism underlying these findings, some extracellularly mediated influence, emanating in vivo from shiverer cells, is capable of ameliorating the basal lamina deficiency typically expressed by dystrophic Schwann cells.

Quantitative ultrastructural studies of the axon Schwann cell abnormality in spinal nerve roots from dystrophic mice.

Lumbo-sacral spinal roots and the nerve to the medial gastrocnemius muscle (NMG) from normal and dystrophic mice were examined by quantitative ultrastructural techniques. It was demonstrated that, although many axons at this level were totally devoid of Schwann cells, totatl axonal numbers were approximately normal. Some axons in these roots were surrounded by Schwann cells but their myelin sheaths were abnormally thin. In addition, cells resembling oligodendrocytes were observed in the dorsal roots of the dystrophic mice. In contrast, Schwann cells and myelin sheaths were normal in the more peripherally situated NMG and the regenerative response of these nerves to crush injury was not significantly different from that of control nerves. Thus, the main abnormality of neural morphology in dystrophic mice is a localized absence of Schwann cells. Such a deficiency does not appear to have influenced axonal growth or the capacity of these axons to regenerate when crushed distally.

Three dimensional analysis of unmyelinated fibers in normal and pathologic autonomic nerves.

The technique of serial-section electron microscopy and diagrammatic three dimensional reconstructions has been used to assess normal and pathological unmyelinated nerve fibers from a peripheral autonomic nerve: the rat cervical sympathetic trunk. Within this predominantly unmyelinated nerve, there is a complex arrangement of axons into longitudinally oriented bundles brought together by chains of Schwann cells. Each bundle is subdivided into smaller components by the cytoplasmic processes of Schwann cells; such subdivisions, which are basal lamina-enclosed masses of Schwann cell cytoplasm, when viewed on cross-sectional electron micrographs, are termed Schwann cell units. The size and shape of each Schwann cell unit varies along the length of fibres, but the diameter of individual axons shows little variation over the segments studied. Axonal branching was not observed in normal unmyelinated nerves. Crush injury and x-irradiation produces different patterns of alteration in the axon-Schwann cell relationships of unmyelinated nerves. Following crush injury, Schwann cell processes increase in diameter and contain numerous small diameter axonal sprouts. Many of the regenerating axons remain thin while others reacquire a normal diameter. X-irradiation affects Schwann cells leading to retraction of their processes and the appearance of naked axonal segments.

Hypomyelination in the peripheral nervous system of shiverer mice and in shiverer in equilibrium normal chimaera.

In shiverer mice, the P1 component of myelin basic protein (MBP) is deficient in both the central nervous system (CNS) and peripheral nervous system (PNS) but compact myelin is more grossly defective in the CNS. In the PNS, myelin exhibits a normal periodic structure, and although examples of subtle abnormalities of shiverer Schwann cell ultrastructure have been described previously, myelin thickness has been reported as unremarkable when observed by light microscopy. We report a quantitative investigation of the myelin sheath thickness of shiverer Schwann cells in which a mild but apparently consistent hypomyelination of axons ensheathed by shiverer Schwann cells was observed. This abnormality was expressed both in the peripheral nerves of a homozygous shiverer mouse and in the shiverer Schwann cells populating the mosaic nerves of a mature shiverer in equilibrium normal mouse chimaera. In addition, multiple interlamellar gaps was found to be a highly consistent feature of shiverer myelin. These observations extend the description of the peripheral nerve defects expressed in shiverer mice and further define these abnormalities as direct consequences of the shiverer Schwann cells' intrinsic genotype. In light of these results, a significant role for P1 in the formation and/or maintenance of normal myelin in the PNS is suggested.

Retinal ganglion cell terminals in the hamster superior colliculus: an ultrastructural study.

The distribution, cross-sectional area, and presynaptic and postsynaptic characteristics of retinal ganglion cell axon terminals in the superior colliculus of normal adult female Syrian hamsters were investigated by quantitative ultrastructural techniques. After an intravitreal injection of horseradish peroxidase, most labelled axon terminals were found in the stratum griseum superficiale and stratum opticum of the contralateral superior colliculus. However, a small proportion (approximately 2%) of retinal ganglion cell axon terminals were located in deeper layers of the superior colliculus between the stratum opticum and the periaqueductal grey matter. Terminals were smaller in the upper two-thirds of the stratum griseum superficiale than in the lower one-third of this layer, the stratum opticum, and the stratum griseum intermedium. Presynaptic characteristics such as the length and number of contacts and the postsynaptic neuronal domains (somata, dendritic spines, or shafts) contacted by retinal ganglion cell axons in the superior colliculus were similar in all layers.

Schwann cell multiplication after crush injury of unmyelinated fibers.

The intensity and duration of Schwann cell multiplication in unmyelinated fibers after crush injury of cervical sympathetic trunks (CSTs) in adult mice was studied using radioautography and electron microscopy. At the level of crush, labeling indexes rose to 22.5% on the second day after injury, but distal to this level, labeling reached a peak of only 2.5%. By the ninth day labeling declined to 1% or less at both sites. Electron microscopy confirmed that the increase in nuclei at the crush was mainly an increase in Schwann cells. Thus, the intensity of Schwann cell multiplication differed between crush and distal sites along the same unmyelinated nerve. The Schwann cell proliferation in the distal CSTs was less intense than that reported in previous studies on myelinated nerves.

Colocalization of TrkB and brain-derived neurotrophic factor proteins in green-red-sensitive cone outer segments.

PURPOSE: To examine the distribution of neurotrophins (NTs) and their catalytic receptors in adult rat photoreceptors. METHODS: Immunocytochemistry and Western blot analyses were performed using primary antibodies raised against NTs (nerve growth factor [NGF], brain-derived neurotrophic factor [BDNF], NT-3, and NT-4/5) and NT receptors (TrkA, TrkB, TrkC, and p75NTR). Double-labeling of retinal sections with opsin-specific antibodies was performed to identify each photoreceptor type. Competitive experiments using excess recombinant NT or Trk receptors confirmed the binding specificity of each antibody. RESULTS: TrkB and BDNF immunoreactivity was colocalized in cone outer segments. TrkB and BDNF were detected in all green-red-sensitive cones, but not in blue-UV cones or rods, and other NTs and NT receptors were not detected in any of the photoreceptor types. CONCLUSIONS: The findings suggest a specific role for BDNF through its signaling receptor TrkB in the function and maintenance of green-red cones, the predominant cone type in the rat retina.

Neuronal and nonneuronal influences on retinal ganglion cell survival, axonal regrowth, and connectivity after axotomy.

In contrast to the abortive regrowth that occurs when axons are interrupted in the adult mammalian CNS, exposure of injured CNS axons to the nonneuronal milieu of a peripheral nerve can lead to extensive axonal elongation. With the application of this experimental approach to the retinocollicular pathway in adult rodents, it has been possible to investigate the influences of neuron-glia and other interactions on the capacity of axotomized CNS neurons to survive injury, to elongate the distances necessary to reach specific targets, and to form connections in the CNS in adult rodents. The results of these investigations indicate that the changed glial environment provided by peripheral nerve grafts permits the guided regeneration of RGC axons to their CNS targets. Back in the CNS glial environment, regenerated axons penetrate their targets for short distances and re-form normal appearing synapses that can excite or inhibit postsynaptic neurons. Further studies will require a better understanding of intrinsic neuronal properties and of the interactions of these neurons with other neurons and with the cellular and noncellular components of the extraneural milieu.

Conduction of nervous impulses in spinal roots and peripheral nerves of dystrophic mice.

Conduction was studied in the sacral ventral roots and ventral tail nerves of dystrophic mice (dy/dy) and phenotypically normal littermates. In myelinated ventral root fibers of normal mice, conduction velocity was uniform with internodal conduction time 45 +/- 5 musec (26 degrees C). In ventral root fibers of dystrophic mice, conduction velocity was decreased and strikingly non-uniform; both saltatory and continuous conduction were observed in different portions of the same nerve fiber. Continuous conduction with velocity less than 2 m/sec (26 degrees C) was characteristically observed in mid-root where the axons are bare; conduction was saltatory close to the exit from the spinal canal and near the spinal cord where the axons are myelinated. Maximum conduction velocity in ventral tail nerves was 21 +/- 3 m/sec for dystrophic mice and 31 +/- 4 m/sec for littermate controls (37 degrees C). Internodal lengths were somewhat decreased in the dystrophic peripheral nerves but there was no significant difference in maximum fiber diameters, myelin thickness or nodal morphology between dystrophic and normal nerves.

Ongoing block of Schwann cell differentiation and deployment in dystrophic mouse spinal roots.

In the spinal roots of dystrophic mice, there are bundles of unensheathed axons and two populations of axon-associated cells: the typical Schwann cells of myelinated fibers and 'uncommitted' cells at the margin of the bundles. Because these 'uncommitted' cells continue to divide in adult animals but fail to ensheath the axons they appose, they can be labelled with tritiated thymidine. In the present experiments, we show that these cells may differentiate into typical Schwann cells of myelinated or unmyelinated fibers when spinal roots from [3H]thymidine-labelled dystrophic mice are grafted into the sciatic nerves of non-dystrophic animals. Thus, this study demonstrates that the 'uncommitted' cells of dystrophic spinal roots are undifferentiated Schwann cells whose differentiation in the intact spinal roots is continuously prevented by some unknown mechanism.

Multipotentiality of Schwann cells in cross-anastomosed and grafted myelinated and unmyelinated nerves: quantitative microscopy and radioautography.

Cross-anastomoses and autogenous grafts of unmyelinated and myelinated nerves were examined by electron microscopy and radioautography to determine if Schwann cells are multipotential with regard to their capacity to produce myelin or to assume the configuration seen in unmyelinated fibres. Two groups of adult white mice were studied. (A) In one group, the myelinated phrenic nerve and the unmyelinated cervical sympathetic trunk (CST) were cross-anastomosed in the neck. From 2 to 6 months after anastomosis, previously unmyelinated distal stumps contained many myelinated fibres while phrenic nerves joined to proximal CSTs became largely unmyelinated. Radioautography of distal stumps indicated that proliferation of Schwann cells occurred mainly in the first few days after anastomosis but was also present to a similar extent in isolated stumps. (B) In other mice, CSTs were grafted to the myelinated sural nerves in the leg. One month later, the unmyelinated CSTs became myelinated and there was no radioautographic indication of Schwann cell migration from the sural nerve stump to the CST grafts. Thus, Schwann cell proliferation in distal stumps is an early local response independent of axonal influence. At later stages, axons from the proximal stumps cause indigenous Schwann cells in distal stumps from the previously unmyelinated nerves to produce myelin while Schwann cells from the previously unmyelinated nerves to produce myelin while Schwann cells from the previously myelinated nerves become associated with unmyelinated fibres. Consequently, the regenerated distal nerve resembled the proximal stump. It is suggested that this change is possible because Schwann cells which divide after nerve injury reacquire the developmental multipotentiality which permits them to respond to aoxonal influences.

Node-like areas of intramembraneous particles in the unensheathed axons of dystrophic mice.

The unensheathed axons in the spinal roots of adult dystrophic mice were examined by freeze-fracture electron microscopy. In most areas of these abnormal fibers the distribution of intramembraneous particles was similar to that of the internodal segments of normal axons with many more particles on the PF (internal) leaflets of these axonal surface membranes than on their EF (external) leaflets. However, patches of axonal membranes were also observed in which the distribution of intramembraneous particles resembled that seen in nodes of Ranvier.

Ensheathment and myelination of regenerating PNS fibres by transplanted optic nerve glia.

Interactions between PNS axons and CNS glia were studied morphologically by transplanting optic nerves into peripheral nerves in groups of rats and mice. Four to 11 months after grafting, small numbers of axons from the peripheral nerves had penetrated the CNS grafts where they became ensheathed and myelinated by CNS glia. Glial protuberances observed at the CNS-PNS interfaces suggested that there had been an active glial response to innervation by PNS axons. These findings provide experimental evidence that denervated CNS glia can be reinnervated and form myelin.

Ouabain binding sites in skeletal muscle from normal and dystrophic mice.

The specific binding of tritiated ouabain was used to estimate the density of Na+-K+-ATPase sites ("Na+-pump" sites) in segments of skeletal muscle from normal and dystrophic mice. Ouabain binding was approximately 4 times greater in red (soleus) muscle than in white (superficial gastrocnemius) muscle from normal animals. In dystrophic soleus muscles, ouabain binding was decreased by nearly one-half. Because Na+-K+-ATPase activity is associated with plasma membranes, these observations constitute further evidence for a sarcolemmal abnormality in dystrophic mice.

Prolonged administration of NT-4/5 fails to rescue most axotomized retinal ganglion cells in adult rats.

The survival of axotomized RGCs was increased by intravitreal NT-4/5 given by repeated injections or osmotic minipumps, but the effects were less complete than predicted. Compared to a single injection of the neurotrophin on day 0, second injections on days 3 or 7 only sustained an additional 10-20% of the RGCs on day 10. Minipumps augmented RGC survival up to 4-fold (50%) at 2 weeks but most RGCs were lost by 1 month. Thus, specific neurotrophins can rescue many RGCs soon after injury but long-term neuronal survival may require a better understanding of changes in neurotrophin receptors and interactions with other molecules.

Familial autoimmune myastenia gravis: four patients involving three generations.

BACKGROUND: Familial autoimmune myasthenia gravis (MG) is rare, although a genetic role for the development of autoimmune MG is suggested by concordance in monozygotic twins and the increased frequency of other autoimmune diseases in family members of myasthenics. METHODS: A patient with a family history of MG was evaluated in hospital. Relatives were interviewed and medical records examined for details regarding the diagnosis of MG in three other family members. RESULTS: The index case first experienced symptoms of MG at age 75 years. She developed generalized MG and required corticosteroids and immunosuppressive therapy to control her disease. Her father developed predominantly bulbar symptoms of MG at age 75 years. He died of complications experienced following a gastrostomy placed for continued difficulty swallowing. His brother developed similar symptoms of MG in his early 60s and died shortly after thymectomy. A 46-year-old nephew of the index case is also beginning to exhibit signs of generalized MG. Acetylcholine receptor antibodies were strongly positive in the index case and her nephew. (The assay was not available for her father and uncle). CONCLUSIONS: Four individuals in three successive generations had diagnoses of autoimmune MG. Study of familial cases such as these may clarify the contribution of genetic factors to the development of this disease.

Regrowth and connectivity of injured central nervous system axons in adult rodents.

The capacity of injured nerve cells to regrow and form terminal connections in the CNS of adult mammals was investigated in axotomized retinal ganglion cells (RGCs) of rodents whose optic nerves were substituted by an autologous segment of peripheral nerve. While many RGCs died after axotomy approximately 20% of the surviving RGCs regenerated axons several cm in length. Some of the regenerated RGC axons entered the superior colliculus where they arborized and formed well differentiated synapses that transynaptically excited or inhibited tectal neurons.