Oxygen at high pressures: a theoretical approach to monoatomic phases.

We have studied the ζ-phase of solid oxygen using the generalized gradient approximation in the density functional approach. Calculations of total energies and pressures have been carried out for the prototype of diatomic ζ-phase and other hypothetical monoatomic crystal structures. The diatomic phase was found to be stable over a wide range of pressure (100-2000 GPa). The stacking of molecular layers is discussed in comparison with the available experimental data.

The effect of copper supplementation on the brindled mouse: a clinico-pathological study.

Brindled mottled is a neurological mutant mouse. Hemizygous males have many clinical and biochemical features in common with kinky hair syndrome (KHS) in humans, and usually die around postnatal day 15, after severe emaciation. Neuronal mitochondrial abnormalities and neuronal degeneration in the cerebrum and cerebellum were constant neuropathological findings in this mutant. A single intraperitoneal injection of cupric chloride, 10 micrograms/g body weight, resulted in an improvement of clinical symptoms and prevention of neuronal degeneration. The degree of improvement was dependent on the date of injection, and day 7 to 10 postnatal appeared to the most effective date. The male hemizygotes which received cupric chloride injections at day 7 or 10 overcame the lethality, and no neuronal degeneration was detected in these mice, although neuronal mitochondrial changes were still persistent. However, following two injections at days 7 and 10, no abnormalities were detected in the cerebral cortical neurons. Even at the ultrastructural level, abnormal mitochondria were very scarce. In the cerebellum, however, mitochondrial changes in the Purkinje cells, particularly in the rostral portion, and generation of white matter were noted in these mice, which were clinically perfectly healthy, judging from the growth rate and behavior. However, cerebellar changes were far less in those which received additional injections later on. These observations indicate that, at least in brindled mutant mice, supplementation of copper is quite beneficial for clinical improvement and the prevention of neuropathological lesions, but the date of administration appears to have crucial importance.

Radial component of central myelin in shiverer mouse.

'Radial component' of the central myelin was investigated in a neurological mutant mouse, shiverer, which is characterized by the lack of myelin basic protein and paucity of the major dense line in the CNS myelin. As has been noted previously in the normal as well as other neurological mutant mice, radial component consisted of rows of interlamellar tight junctions and was accompanied with electron lucent linear structures (ELLS) over the major dense lines. In the areas where major dense line had formed in shiverer CNS myelin, numerous ELLS run across the major dense lines and were not always associated with a fusion of double intraperiod lines. The possible role of ELLS in myelin formation is briefly discussed.

The twitcher mouse: accumulation of galactosylsphingosine and pathology of the central nervous system.

In the twitcher mouse, a murine model of globoid cell leukodystrophy (GLD), pathological changes of various parts of the central nervous system correlated well with the concentration of galactosylsphingosine (psychosine). The development of GLD lesions was more obvious in tracts with a more rapid progression of myelination. It was suggested that accumulation of galactosylsphingosine subsequent to myelin maturation caused suicidal death of myelin forming cells.

The twitcher mouse: accumulation of galactosylsphingosine and pathology of the sciatic nerve.

Morphological and biochemical changes were investigated in the early developmental stages of sciatic nerve of the twitcher mouse, a murine model of human globoid cell leukodystrophy. The concentration of galactosylsphingosine (psychosine) and the chronological changes of the twitcher mouse peripheral nerve pathology correlated well. Galactosylsphingosine had already accumulated at birth and dramatically increased with age. Characteristic inclusions were observed in Schwann cells and macrophages of the twitcher mouse on the 5th postnatal day. Endoneurial edema developed after 10 postnatal days and the hypomyelination was pronounced at 15-20 postnatal days. These findings suggest that galactosylsphingosine is cytotoxic for myelin-forming cells and is closely related to pathogenetic events in the twitcher mouse.

The zitter rat: membranous abnormality in the Schwann cells of myelinated nerve fibers.

Ultrastructural alterations of the Schwann cells were chronologically examined in zitter and in control rats. The membranous abnormalities closely involving the nuclear membranes of the Schwann cells and mesaxons were constantly observed in the zitter rats. These structures consisted of scroll- or whirl-like unit membrane accumulations directly extending to the nuclear envelope. Occasionally, similar membranous structure were observed in association with the endoplasmic reticulum, lysosome or mitochondria in the cytoplasm of the Schwann cells. In the control rats, whirl-like membranous structures in the nuclear membranes of the Schwann cells were observed only intermittently at 1, 5 and at 7 days of age in the developing process. In conclusion, we postulate that the zitter rat, which is an established animal model for hypomyelination of the central nervous system (CNS), has some inherited abnormality related to the membrane biosynthesis and its regulation in Schwann cells as well as in oligodendrocytes.

The twitcher mouse: normal pattern of early myelination in the spinal cord.

The pattern of early myelination was investigated in the dorsal columns of the cervical spinal cord in the twitcher, an authentic murine model of human globoid cell leukodystrophy, and their littermates. There were no differences in the number of myelinated fibers until the day 20 postnatal. However, myelin sheath in the homozygous affected twitchers at the day 20 were thinner than those of heterozygous and normal littermates, while at the day 10 no significant differences were detected. These observations indicated that in the twitcher mouse, despite the genetic deficiency of galactosylceramidase, myelination progresses normally in early stages and then hypomyelination becomes apparent before myelin breakdown.

Thiamine-deficient encephalopathy in rats: effects of deficiencies of thiamine and magnesium.

Thiamine- and vitamin B-deficient encephalopathy was produced in adult Wistar rats and the effect of magnesium deprivation and/or the administration of guanidine, a magnesium antagonist, was studied. These encephalopathic rats had symmetrical lesions, edema, exudation of fibrin, spongy changes, petechial hemorrhages, neuronal degeneration and gliosis in the pontine tegmentum, mainly the lateral and medial vestibular nuclear areas. Mild pathological changes were seen in the purely thiamine- or vitamin B-deficient rats, whereas severe pathological changes occurred in thiamine- or vitamin B-deficient rats combined with magnesium deprivation and/or guanidine administration. However, magnesium-deficient diets or guanidine administration did not produce pathologic changes in rats. These findings suggest that thiamine deficiency is an essential factor in the development of these pathological changes in the brainstem, and that magnesium and guanidine, an antagonist of magnesium, play important roles in the development of these disorders.

The twitcher mouse: attenuated processes of Schwann cells in unmyelinated fibers.

Morphological alterations occurring in Schwann cells of unmyelinated fibers (unmyelinated Schwann cells) were investigated in the sciatic nerve of the twicher mouse, a murine model of human globoid cell leukodystrophy. After postnatal day 10, the number of Schwann cell-axon units gradually increased and the number of unmyelinated axons per unit progressively decreased in the twitcher mouse. However, the total number of unmyelinated axons showed no significant differences between twitcher and normal mice. Thus, these alterations of unmyelinated Schwann cells in the twitcher mouse suggest that attenuated branching of cellular processes develops at an early stage and progresses together with progression of demyelination in this mutant.

Effects of intracarotid hyperosmolar mannitol in triethyl tin (TET)-induced rat brain edema--preservation of blood-brain barrier (BBB) in TET edema.

The effect of intracarotid hyperosmolar mannitol on the blood-brain barrier (BBB) in triethyl tin (TET)-induced rat brain edema was examined by using intravenous Evans blue (EB, MW 68,000) and Adriamycin (ADM, MW 580) as tracers. Three ml of 1.4 M mannitol solution were administered through the right carotid artery during 45 s for BBB opening. The barrier was opened for 60-120 minutes and then re-established. The reversibility was preserved in TET-treated rats and controls. In fact, the intravenous injection of EB stained both TET-treated and non-treated cerebral hemispheres with mannitol-induced transient BBB disruption, but not without BBB disruption. BBB was resistant to both high and low molecular weight substances in TET-induced edema. The importance of this hyperosmotic studies provides the evidence for normal BBB function in TET-induced brain edema.

Neurotoxicity of adriamycin passed through the transiently disrupted blood-brain barrier by mannitol in the rat brain.

Neurotoxicity of adriamycin (ADM) was investigated following the transient disruption of the blood-brain barrier (BBB) in rats. The BBB disruption by the hyperosmotic agent (1.4 M mannitol) was confirmed by the leakage of ADM and Evans blue administered intravenously. Neuropathological changes due to the toxicity of ADM were found as early as day 4. The neurons in the cerebral cortex and nucleus caudatus-putamen showed focal clearing of the nuclear chromatin, increased dense bodies in the cytoplasm and dilatation of the cisternae of the rough endoplasmic reticulum (r-ER) and Golgi apparatus. By day 7, nucleolar segregation and irregular membranous structures appeared in the nuclei with the progression of cytoplasmic changes. By day 10, the cytoplasm of many neurons was vacuolated. Electron-microscopically, the cisternae of the r-ER and Golgi apparatus were prominently dilated in these neurons. Neuronal microtubules were increased in number, in particular in the perinuclear region. Numerous whorl-like membranous structures and separation of nuclear membrane were also observed. Some astrocytic processes surrounding the blood vessels revealed loss of organelles and a few pericytes showed an increased number of lysosomes on days 7 and 10. This experiment clearly demonstrates that ADM has strong neurotoxic effects in the central nervous system when the BBB is disrupted, and provides the warning for the possibilities of neurotoxic side effects when ADM is administered, in combination with a hyperosmotic agent, for the treatment of human malignant tumors, including brain tumors.

Resistance of quaking mouse CNS to triethyl tin edema.

Intraperitoneal injection of triethyl tin (TET) sulfate, 5 or 10 mg/kg body weight did not induce intramyelinic edema without altering water content in quaking mice while in C57BL/6J and littermate control mice, water content was increased and typical intramyelinic edema was induced following TET injection. Even among control mice, however, there were some strain differences in the histological severity of the edema, which were in precise agreement with the quantitative alterations in water content. These observations suggest that CNS myelin in quaking may differ qualitatively from that in controls and the mode of response to TET is under genetic control.

Triethyl tin does not induce intramyelinic vacuoles in the cns of the quaking mouse.

Triethyl tin (TET), when injected intraperitoneally, failed to produce the typical intramyelinic edema in the spinal cord of quaking mice with two different genetic backgrounds (B6C3H-qk and BTBRTF/Nev-qk), while control littermates and normal C57BL/6J mice were susceptible, as expected. The only prominent change in the quaking mice was the presence of spherical vacuoles containing floccular electron-dense materials, some of which were clearly within the oligodendroglial perikarya and the inner and outer tongues. They are likely to represent degenerative responses. Consistent with the lack of edema, no increase in the water content was found in the quaking spinal cord following TET injection. Although the presence of numerous interlamellar tight junctions in quaking CNS myelin may mechanically restrict formation of the intralamellar vacuoles, the unique changes in the oligodendroglia and the lack of edema fluid accumulation suggest more fundamental metabolic abnormality that renders the quaking CNS resistant to the triethyl tin-induced edema.

The twitcher mouse: degeneration of oligodendrocytes in vitro.

Oligodendrocytes were isolated from the brain of the twitcher (twi/twi), an authentic murine model of globoid cell leukodystrophy (GLD), carrier (+/twi) and their littermate controls (+/+) and were maintained in vitro for 24 days. By 4 days in vitro (4 DIV) oligodendrocytes developed thorny processes and their morphology were closely similar to each other regardless of their genetic status. After 17 DIV, however, oligodendrocytes in twi/twi progressively degenerated and only 12% of oligodendrocytes counted at 10 DIV survived at 24 DIV in twi/twi, while in +/twi and +/+, mean survival rates were 85 and 83% respectively. Characteristic inclusions of GLD were detected in the perikarya of degenerating twi/twi oligodendrocytes indicating that metabolic defect was expressed even in isolated oligodendrocytes. These results further support the hypothesis that the primary pathogenetic event in murine GLD, twitcher, is degeneration of oligodendrocytes due to progressive accumulation of the toxic metabolite, galactosylsphingosine (psychosine).

Early myelination in zitter rat: morphological, immunocytochemical and morphometric studies.

Early myelination in zitter rat was investigated by light and electron microscopic observations, by immunostaining for myelin basic protein (MBP), proteolipid protein (PLP) and myelin-associated glycoprotein (MAG), and by morphometric analysis from the 1st to the 28th day of age. Although the commencement of myelination in zitter rats was not delayed in comparison with control rats, the density or the number of myelinated fibers in zitter rats was significantly below that in controls, both in the ventral column of the cervical spinal cord and in the optic nerve. In contrast, the density or the number of aberrant myelin sheaths was increased in zitter rats, and this difference became more marked with increasing age. The persistent presence of abnormal membranous structures associated with the oligodendroglial nuclear membrane and the increased number of aberrant myelin sheaths were characteristic to the zitter mutation, although these alterations were also observed transiently in control rats. Quantitative analysis supported the proposition that hypomyelination in zitter rats is primarily pathological and becomes more prominent with advancing age. However, the fundamental structure of the myelin lamellae appeared to be normal and the immunoreactivities for MBP, PLP and MAG were slightly delayed and weakend in comparison with age-matched controls. Thus, in zitter rat there is the functional abnormality of the oligodendrocytes to integrate the processes of membrane biosynthesis and to expel excessive production of membranous structures associated with the membranous organellae such as nuclear membrane, and it is postulated that this functional abnormality is characteristic to only the zitter mutation.

Effects of methylcobalamin on vitamin B1- and B-deficient encephalopathy in rats.

Vitamin B1- and B-deficient encephalopathy was produced in adult Wistar rats and the effect of methylcobalamin was studied. These experiments were performed in vitamin B1- or B-deficient rats with or without the administration of guanidine, a magnesium antagonist. These encephalopathic rats showed symmetrical lesions in the pontine tegmentum. No pathological changes in the brainstem were seen in guanidine-administered rats. This suggests that guanidine itself doesn't produce these brainstem lesions. Moderate to severe pathological changes occurred in vitamin B-deficient rats with guanidine administration, whereas these pathological changes were milder in vitamin B1-deficient rats with guanidine administration and in vitamin B1- and B-deficient rats with the combination of the guanidine and methylcobalamin administrations. These facts suggest that vitamin B12, methylcobalamin, plays an important preventive role in the development of the pontine lesions in this experiment.

Emery-Dreifuss muscular dystrophy. An autopsy case.

We reported the autopsy findings in a 50-year-old man with typical clinical features of Emery-Dreifuss muscular dystrophy. Special attention was directed to the spinal cord and ventral spinal roots to determine whether cause of the muscle wasting was denervation or myopathy. Neuropathological studies disclosed no abnormality of the spinal cord, and the ventral spinal roots were intact. The skeletal muscles showed dystrophic changes of varying degrees, and marked cardiomyopathy was evident. We consider that muscle wasting in this man was due to muscular dystrophy.

A comparison of erythrocytes, lymphocytes and blood plasma as samples in fatty acid analysis for the diagnosis of adrenoleukodystrophy.

We studied the very-long-chain fatty acids of blood plasma, erythrocyte membranes and lymphocytes in 4 adrenoleukodystrophy patients, 5 adrenoleukodystrophy obligate carriers, 12 normal controls and 81 patients with various neurological disorders by high-performance liquid chromatography and compared the reliabilities in the diagnosis of adrenoleukodystrophy of these 3 components of peripheral blood. Of 81 patients with various neurological disorders, 2 myotonic dystrophy and 2 spinocerebellar degeneration patients showed increased ratios of C26:0 to C22:0 in erythrocyte membranes, but not in blood plasma and lymphocytes. None of the 12 normal controls showed increased ratios of C26:0 to C22:0 in erythrocyte membranes, blood plasma and lymphocytes. These results suggest that fatty acid analysis for the diagnosis of adrenoleukodystrophy is more reliable when blood plasma and lymphocytes are used than when erythrocyte membranes are used.

Formalin fixed brains are useful for magnetic resonance imaging (MRI) study.

We carried out magnetic resonance imaging (MRI) studies on human brains which had been fixed in formalin solution for over 2 years and had been proven neuropathologically to be cases of multiple sclerosis (MS), progressive multifocal leukoencephalopathy (PML), and Balo's concentric sclerosis (Balo). Using spin echo (SE) and inversion recovery (IR) pulse sequences to detect demyelinated lesions in a living person with MS, the demyelinated lesions of the fixed brains in cases of MS, PML and Balo definitely re-appeared, although T1 and T2 in the gray and white matter were reduced following fixation. High signal areas on the SE images corresponded not only to the characteristic distribution of demyelinated lesions in the white matter but also to sparse myelin, gliosis and mild perivascular cuffing in the white matter around the demyelinated foci in cases of the fixed MS, PML and Balo brains. On the IR images, only MS plaques were evident. This MRI study of fixed brains proved useful to elucidate clinicopathological correlations.

Hyperosmotic blood-brain barrier disruption in brains of rats with an intracerebrally transplanted RG-C6 tumor.

To test the results of blood-brain barrier (BBB) disruption in the treatment of brain tumor, RG-C6 glioma was transplanted into the brains of rats. Intracarotid infusions of normal saline and hyperosmotic mannitol were then made, followed by intravenous injection of Evans blue dye plus albumin (EB, MW 68,000), horseradish peroxidase (HRP, MW 40,000), and 5-fluorouracil (5-FU, MW 130). Uptake of the drug and the consistency of drug levels in the normal brain and tumor varied widely among these three agents. Both EB and HRP penetrated the brain tumors but did not stain the normal brain tissues. After BBB opening, penetration of EB and HRP into the normal brain was drastically increased; however, the uptake of EB and HRP in the tumor was not increased. The concentration of 5-FU in the tumor was higher than that in the serum and, although it increased 1.5-fold after BBB opening, the increase was not statistically significant. Conversely, there was a progressive increase in concentrations of 5-FU in the tumor-free brain regions (p less than 0.05). These observations suggest that an intracarotid infusion of hyperosmotic mannitol may increase neurotoxicity because it allows greater delivery of anticancer drugs into the normal brain tissue than into the tumor tissues.