The Purkinje cell and its afferents in human hereditary ataxia.

The cerebellar cortex in patients with autosomal dominant and recessive ataxia was studied by Golgi impregnation and immunocytochemistry in order to gain further insight into the pathogenesis of neuronal atrophy which accompanies these disorders. Monoclonal antisera were used to visualize phosphorylated and non-phosphorylated neurofilament proteins, and a synapse-specific protein (P38; synaptophysin). Golgi stain and immunocytochemistry for non-phosphorylated neurofilament protein revealed partial or complete loss of distal Purkinje cell dendrities in the dominant cases and in one recessive case. Many preserved parallel fibres were shown by the monoclonal antibody to phosphorylated neurofilament protein. This antibody also gave strong reaction product in torpedoes. Axosomatic and axodendritic terminals on Purkinje cells were reduced in number, and loss of mossy fiber terminals was revealed by monoclonal anti-P38. The described methods provided additional morphological evidence of the heterogeneity of the hereditary ataxias. Purkinje cell atrophy progressed from loss and simplification of the dendritic tree to disappearance of the cell body. While these cells appeared to be especially vulnerable, other neurons of the molecular and granular layers were not exempt. There was evidence that at least some extracerebellar afferents, such as mossy fibers, were also affected by the disease process.

The hereditary ataxias.

Efforts to classify the hereditary ataxias by their clinical and neuropathological phenotypes are troubled by excessive heterogeneity. Linkage analysis opened the door to a new approach with the methods of molecular biology. The classic form of autosomal recessive ataxia, Friedreich's ataxia (FA), is now known to be due to an intronic expansion of a guanine-adenine-adenine (GAA)-trinucleotide repeat. The autosomal dominant ataxias such as olivopontocerebellar atrophy (OPCA), familial cortical cerebellar atrophy (FCCA), and Machado-Joseph disease (MJD) have been renamed the spinocerebellar ataxias (SCA). Specific gene loci are indicated as SCA-1, SCA-2, SCA-3, SCA-4, SCA-5, SCA-6, and SCA-7. In 5 of them (SCA-1, SCA-2, SCA-3, SCA-6, and SCA-7), expanded cytosine-adenine-guanine (CAG)-trinucleotide repeats and their abnormal gene products cause the ataxic condition. The most common underlying loci for olivopontocerebellar atrophy (OPCA) are SCA-1 and SCA-2, although other genotypes may be added in the future. A major recent advance was the identification of the gene for SCA-3 and MJD, and the high prevalence of this form of autosomal dominant ataxia. In FA and the SCA with expanded CAG-trinucleotide repeats, clinical and neuropathological severity are inversely correlated with the lengths of the repeats. Anticipation in the dominant ataxias can now be explained by lengthening of the repeats in successive generations. Progress is being made in the understanding of the pathogenesis of FA and SCA as the absent or mutated gene products are studied by immunocytochemistry in human and transgenic murine brain tissue. In FA, frataxin is diminished or absent, and an excess of mitochondrial iron may cause the illness of the nervous system and the heart. In SCA-3, abnormal ataxin-3 is aggregated in neuronal nuclei, and in SCA-6, a mutated alpha1A-calcium channel protein is the likely cause of abnormal calcium channel function in Purkinje cells and in the death of these neurons.

Experimental superficial siderosis of the central nervous system. I. Morphological observations.

Autologous washed red blood cells were injected weekly over a period of three to six months into the cisterna magna of adult New Zealand white rabbits. After three months, the surface of the brain stem, cerebellum, and piriform cortex showed a distinct brown color, and staining of the gross specimens for iron produced an intense blue color which extended for a distance of 1-2 mm into the brain parenchyma. Enhanced iron stains of vibratome sections revealed the accumulation of reaction product in microglia and Bergmann glia of the cerebellar cortex, and in microglia and astrocytes of the piriform cortex. Ferritin immunocytochemistry revealed reaction product in cerebellar microglia and Bergmann glia which strongly resembled that obtained by the enhanced iron stain. In the piriform cortex, only microglia were reactive with anti-ferritin. Electron microscopy confirmed the accumulation of electron-dense ferritin granules only in the cytoplasm of microglia. Bergmann glia in the cerebellum and astrocytic processes in the piriform cortex were replete with intermediate filaments and contained an excess of glycogen. After six months, small granules of hemosiderin began to appear in cerebellar and piriform cortices. The observations support that the sequence of conversion of hemoglobin to ferritin and hemosiderin occurs in brain as in other organs.

Brain hemosiderin and superficial siderosis of the central nervous system.

Brain tissue from five patients with superficial siderosis of the central nervous system was examined by immunocytochemistry for ferritin, glial fibrillary acidic protein (GFAP), alpha 1-antitrypsin, and alpha 1-antichymotrypsin, and by lectin affinity cytochemistry with biotinylated Ricinus communis agglutinin-1 (RCA-1). The sections were pretreated with 2,2'-dipyridyl and sodium hydrosulfite to remove iron and to reveal the antigenic sites. In siderotic cerebellar cortex, ferritin reaction product occurred in the hemosiderin matrix, the cell bodies and processes of Bergmann glia, and in microglia. Astrocytes other than Bergmann glia did not contain ferritin reaction product. RCA-1 stained microglia and hemosiderin whereas antisera to alpha 1-antitrypsin and alpha 1-antichymotrypsin only reacted with iron-depleted granules. The selective vulnerability of the eighth cranial nerve was explained by the presence of ferritin-reactive and lectin-positive microglia. Hemosiderin isolated from frozen cerebellum contained ferritin, GFAP, and vimentin. The presence of the intermediate filament proteins was likely due to co-localization with hemosiderin granules in Bergmann glia. The ability of the brain to biosynthesize ferritin in response to prolonged contact with hemoglobin iron is thought to be the most important factor in the pathogenesis of superficial siderosis. The great severity of the lesion in the exposed cerebellar cortex is readily explained by accelerated ferritin biosynthesis in Bergmann glia.

The heterogeneous distribution of brain transferrin.

Immunocytochemistry with antisera to transferrin has often been used to identify oligodendroglia in tissue sections and cultures, but reaction product also occurs in blood vessel walls and nerve cells. There is considerable species variation. Serum transferrin is largely biosynthesized in the liver, and its established physiological role is the transport of iron to tissue sites and delivery of the metal to the interior of cells that have transferrin receptors on their surfaces. In sections of the central nervous system, the visualization of iron and transferrin generally does not coincide, and transferrin may have importance to normal brain function beyond iron transport. For a comparative analysis of transferrin in rabbit and rat brain, polyclonal antisera were raised against purified serum transferrins of these species. The antisera were used for transferrin immunocytochemistry on vibratome sections and for immunochemical detection on electroblots. Transferrin immunocytochemistry and iron histochemistry were compared. The electrophoretic separation of brain extracts and transfer to nitrocellulose membranes permitted the quantitation of the protein and the study of the carbohydrate chains of tissue-bound transferrins by biotinylated lectins. An unexpected result in the rabbit was the dense immunocytochemical reaction product in Bergmann glia and Golgi epithelial cells. Reaction in the cytoplasm of oligodendrocytes was relatively faint in this species except for some selected white matter tracts, e.g. the inferior cerebellar peduncles. In sections of rat brain, oligodendrocytes and vessel walls reacted vigorously in all locations. Transferrin levels in rat brain were substantially higher than in rabbit brain. In the rabbit, maximum transferrin levels occurred in the cerebellum.(ABSTRACT TRUNCATED AT 250 WORDS)

Fine structure of neurons of the hypertrophied human inferior olive.

Ultrastructural study of hypertrophied inferior olives from three cases of palatal myoclonus revealed tha nerve cells not infrequently contained numerous round, homogeneously electron-dense granules (mean diameter, 360 nm) located within expanded cisternal profiles of rough endoplasmic reticulum (RER). Control tissue did not contain similar structures. These granules may consist of proteinaceous secretion of the RER which, for reason unknown, accumulates during transsynaptic degeneration of the inferior olive. Additional noteworthy electron microscopic features of neurons of the hypertrophied olive were as follows: 1. neurofilamentous hyperplasia, greater in dendrites than in perikarya; 2. vacuoles in intermediate and large (up to 15 mum) size, derived from RER; and 3. prominent intracytoplasmic protrusions by boutons containing dense core vesicles (mean diameter, 98 nm). Glomeruloids were identified ultrastructurally and consisted of numerous boutons interspersed among neurofilament-rich dendrites and occasional, filament-packed astrocytic profiles. Frequently, these boutons also contained dense-core vesicles. It seems possible that the boutons mentioned may arise from collateral axonal sprouts.

Synapses in the hereditary ataxias.

The goal of this investigation was the systematic assessment of synapses in the hereditary ataxias by the immunocytochemical and immunofluorescent visualization of SNAP-25, a protein of the presynaptic membrane. Sections were prepared from the cerebellar cortex, dentate nucleus, basis pontis, inferior olivary nuclei, and the spinal cord in 57 cases of autosomal dominant and recessive ataxia. The neuropathological phenotype included 18 cases of olivopontocerebellar atrophy (OPCA), 14 cases of familial cortical cerebellar atrophy (FCCA), 4 cases of Machado-Joseph disease (MJD), and 21 cases of Friedreich's ataxia (FA). Among the autosomal dominant ataxias, spinocerebellar ataxia type 1 (SCA-1), SCA-2, MJD/SCA-3, and SCA-6 were represented. Expanded guanine-adenine-adenine trinucleotide repeats were confirmed in 7 patients with FA. The abundance of SNAP-25 was estimated by comparing the fluorescence of the regions of interest to that of the frontal cortex, which was considered unaffected by the disease process. Despite severe Purkinje cell loss, abundant SNAP-25 reaction product remained in the molecular layer of FCCA and OPCA. Among the cases of OPCA, those identified as SCA-2 showed the most severe overall synaptic destruction in cerebellum and brain stem. In SCA-1, which caused either OPCA or FCCA, significant synaptic loss was restricted to the inferior olivary nuclei. Sparing of cerebellar cortex and inferior olivary nuclei was the rule for MJD/SCA-3 and FA, though the dentate nucleus showed reduced SNAP-25 immunoreactivity in both ataxias. In FA, preservation of SNAP-25 in the dentate nucleus was characteristic of long survival. Severe cases with short survival revealed synaptic depletion of the dentate nucleus. At the level of the spinal cord, synaptic loss in the dorsal nuclei of Clarke characterized FA and MJD/SCA-3. The inexorable clinical progression of the hereditary ataxias could not be attributed to synaptic loss in a single anatomic structure of cerebellum, brain stem, or spinal cord. Nevertheless, synaptic loss in dentate and inferior olivary nuclei correlated more precisely with the severity of the ataxia than the changes in the cerebellar cortex.

Oligodendroglioma of the medulla oblongata in a neonate.

A 4-month-old boy had recurrent attacks of apnea before progressive quadriparesis, ocular motility disturbance, and depressed corneal reflexes led to the discovery of a posterior fossa mass. Death occurred postoperatively during prolonged apnea. Necropsy disclosed an oligodendroglioma of the medulla oblongata. A review of this rare neoplasm in the neonatal period is given.

Hereditary ataxia and the sixth chromosome.

Possible linkage of the gene or genes for dominant hereditary ataxia and three genetic markers on the short arm of the sixth chromosome (HLA, properdin factor B [Bf], and glyoxalase I) was investigated in five families. Logarithmic odds (lod scores) were calculated for the linkages and found to be either inconclusive or in favor of nonlinkage. Caution is advised in the summing of lod scores for separate families because of the wide spectrum of clinical and anatomical manifestations of dominant hereditary ataxia. Three families with recessive hereditary ataxia were also studied. Identical haplotypes occurred in affected and unaffected siblings. It did not appear likely that the recessive genes of the parents were transmitted in linkage with the markers on the short arm of the sixth chromosome.

Familial oculoleptomeningeal amyloidosis. Report of a new family with unusual features.

A family had a dominantly inherited amyloid angiopathy that involved the meninges of the brain and spinal cord, retina, vitreous humor, peripheral nerves, and systemic organs. Clinical features included hemiplegic migraine, periodic obtundation, psychosis, seizures, intracerebral hemorrhage, myelopathy, visual impairment, deafness, and peripheral neuropathy. Pathological findings consisted of amyloid deposition in the leptomeningeal and retinal vessels, in the vitreous humor, and in perivascular tissue throughout the body. Evaluation of the amyloid showed it to be a transthyretin (prealbumin). A brief course of plasmapheresis produced a short-lived decrease concentration in circulating transthyretin.

Oculoleptomeningeal amyloidosis associated with a new transthyretin variant Ser64.

BACKGROUND: A Canadian family with oculoleptomeningeal amyloidosis with both central and peripheral nervous system disorders was described in 1988. Death of affected family members resulted from recurrent cerebral hemorrhage. OBJECTIVE: To determine if oculoleptomeningeal amyloidosis is caused by a mutation in transthyretin (prealbumin). METHODS: DNA isolated from peripheral blood and archival tissues of affected members of the kindred was studied by direct DNA sequencing and restriction fragment length polymorphism analysis. RESULTS: Direct DNA sequencing identified a thymine-to-cytosine transition at the second base of codon 64, which resulted in a replacement of serine for phenylalanine. This mutation, which creates an additional HinfI site was detected by restriction fragment length polymorphism analysis in each affected individual. CONCLUSION: In this kindred, oculoleptomeningeal amyloidosis is related to a mutation in transthyretin (Phe64Ser).

Adult-onset hereditary ataxia in Scotland.

A systematic search for cases of adult-onset hereditary ataxia was conducted on location in Scotland. The investigation resulted in the discovery of eight pedigrees with 42 patients of whom 16 were alive in 1975. Nine patients were examined by the authors and recent hospital records were available on the remaining seven. The clinical features were quite variable. In declining order of frequency, findings were gait and limb ataxia, dysarthria, hyperreflexia, extrapyramidal motor disturbances, impaired vibratory sense, spasticity, defects of extraocular movements and nystagmus, reflex depression, Babinski signs, impaired joint position sense, muscle weakness, optic atrophy, and mental abnormalities. Foot deformity occurred only once. Inheritance was compatible with autosomal dominant transmission, but complicated by consanguinity in two families. The minimum prevalence was calculated as 0.31/100,000. Autopsy in two members in one family revealed olivopontocerebellar degeneration.

Marchiafava-Bignami disease.

Marchiafava-Bignami disease was diagnosed postmortem in a 39-year-old man who drank excessive amounts of white port wine. This is the fifth report of the disease in a native North-American with no Italian ancestry. The lesion involved the corpus callosum and hippocampal commissure but spared the anterior commissure, middle cerebellar peduncles, optic chiasm, and centrum semiovale. Wernicke-Korsakoff encephalopathy and pellagroid neuronal changes were also present.

Supranuclear ophthalmoplegia in olivopontocerebellar degeneration.

Autosomal dominant olivopontocerebellar degeneration was diagnosed in a family of Scottish ancestry by clinical examination and autopsy. In addition to having progressive cerebellar ataxia, head titubation, and severe dysarthria, the patients are unable to initiate saccadic eye movements. Slow pursuit movements are normal. Reflex movements of the eyes caused by passive rotation or caloric labyrinthine stimulation are not impaired but are not associated with nystagmus. The phenomenon can be classified as supranuclear pseudo-ophthalmoplegia. It differs from congenital ocular motor apraxia in age at onset and the absence of random eye movements. The anatomic lesion responsible for the defect of saccadic eye movements remains to be established.

Olivary hypertrophy: histochemical demonstration of hydrolytic enzymes.

Of four patients with palatal myoclonus, three had infarcts resulting from atherosclerosis, and one had cerebral emboli from a left atrial myxoma. Three specimens showed lesions in the brainstem and bilateral hypertrophy of the inferior olivary nuclei; the fourth revealed unilateral olivary changes caused by an infarct in the contralateral dentate nucleus. After incubation for acetylcholinesterase, neuropilar and capillary wall staining were absent or much reduced, but there was increased denisty of reaction product in the neuronal cell bodies and in numerous tortuous dendrites. Methods for acid phosphatase showed strong activity in the dendrites and glomeruloid structures of the diseased olives. Reactions for nonspecific esterase indicated dendritic expansion and reduced staining density in nerve cell bodies, but augmented glial reactivity.

Olivopontocerebellar atrophy: immunocytochemical and Golgi observations.

Brain tissue was obtained promptly after death from a patient with autosomal dominant olivopontocerebellar atrophy and studied by immunocytochemistry and a Golgi technique. Antiglutamic acid decarboxylase showed severe loss of Purkinje cells and their terminals in the dentate nucleus. Stains for neuron-specific enolase (NSE) and microtubule-associated proteins (MAP) confirmed the integrity of the dentate nucleus. Basket and stellate cells revealed secondary changes, but Golgi neurons were intact. Methods for NSE and MAP disclosed dendritic alterations and loss of neurons in the basis pontis and inferior olivary nuclei. Golgi impregnation of Purkinje cells showed loss of major dendrites, paucity of spiny branchlets, and axonal expansions.

Amyloid fibril protein in familial amyloidosis with cranial neuropathy and corneal lattice dystrophy (FAP type IV) is related to transthyretin.

Immunocytochemical methods were used to study the nature of the amyloid deposits in the Finnish type-familial amyloid polyneuropathy (FAP) type IV, which is characterized by cranial neuropathy and corneal lattice dystrophy. Commercial antisera to human plasma transthyretin (prealbumin) did not stain the amyloid deposits, but in every case a positive staining was obtained with antibodies raised against transthyretin-related amyloid fibril whole protein isolated from the myocardium of a patient with familial amyloid polyneuropathy from the state of New York. The FAP type IV amyloid deposits stained also with antiserum to serum amyloid P component, but did not stain with antisera to retinol-binding protein, amyloid A protein, gamma-trace protein, beta 2-microglobulin, or immunoglobulin light chains. The serum level of serum transthyretin was significantly decreased in FAP type IV patients (256 +/- 75 (SD) mg/L, n = 15) as compared with Finnish control subjects (360 +/- 56 mg/L, n = 30, P less than 0.001), whereas the level of retinol-binding protein was within the normal range. The results of this study strongly suggest that the amyloid fibril protein in FAP type IV amyloidosis is related to transthyretin.

Enzyme activity of human central nervous system myelin.

Following ultracentrifugation in sucrose and hypotonic exposure ('osmotic shock'), myelin fractions were prepared from homogenates of human centrum ovale obtained post-mortem. Non-specific esterase (NsE), succinic dehydrogenase and acid phosphatase activities were determined. One isolation procedure utilized aqueous sucrose solutions only. Other preparative technics involved addition of phosphate buffer (pH 6.6 or 8.0) or varying concentrations of calcium or magnesium ions to preparative media. The NsE activity of myelin increased greatly when electrolytes were included in these preparative solutions. Morphologic homogeneity and protein and lipid contents of myelin were similar whatever the isolation procedure.

The nucleus pontis centralis caudalis in Huntington's disease.

Slow saccadic eye movements occur in some patients with Huntington's disease (HD), and minor defects of supranuclear eye movement control can be demonstrated in the majority by neuroophthalmological laboratory methods. In the pathogenesis of slowed saccades, a lesion of the paramedian pontine reticular formation and specifically the nucleus pontis centralis caudalis was considered likely due to similar eye movement disturbances in well documented degenerative and vascular lesions of the lower pontine tegmentum. A systematic morphometric study was performed on the nucleus pontis centralis caudalis in 9 patients with HD. Two of them had grossly defective saccades during life, and 7 had normal eye movements on routine examination. In 8 patients, the nucleus was reduced in size, revealed a higher than normal neuronal density, and a striking loss of large neurons. One patient with HD and normal morphometric results had died 2 years after the onset of chorea from an unrelated illness. It is proposed that the nucleus pontis centralis caudalis is regularly affected in HD and that progressive loss of large neurons is the cause of saccadic slowing.

The history of iron in the brain.

Brain iron research began in the late nineteenth century when Zaleski (1886) made a quantitative analysis of one human brain and correlated iron levels with observations on stained slices and some microscopic sections. Gradually, the realization grew that the central nervous system (CNS) contained iron which was different from hemoglobin-iron. This non-heme iron was found in highest concentrations in globus pallidus, substantia nigra, red nucleus, and dentate nucleus. The enhancement of the traditional histochemical stain, potassium ferrocyanide in hydrochloric acid, by incubating the reacted sections in a solution of diaminobenzidine and hydrogen peroxide, revealed iron in many cell types of the CNS, including neurons, microglia, oligodendroglia, and some astrocytes. A large proportion of the soluble brain iron was shown to be present in ferritin. Brain ferritin was found to be very similar to the protein from other organs in that it contained heavy and light subunits. Several investigators reported the presence of other iron-related proteins in the central nervous system, including transferrin, transferrin receptor, and the ferritin repressor protein. Brain was shown to respond to the extravasation of blood by converting the iron in heme to hemosiderin by a sequence of steps which was quite similar to the process in extracerebral organs. The methods of molecular biology have contributed greatly to our understanding of brain iron but many questions remain about its unique anatomical distribution and its role in degenerative diseases such as Parkinson's disease and Alzheimer's dementia.