Autopsy Specimens Of Human Brain Research Articles

Are myotonia and epilepsy linked by a chloride channel?

Chen et al.1 report 2 novel findings: a truncation mutation in the CLCN1 gene, encoding the chloride channel protein ClC-1, in a patient with myotonic dystrophy and generalized epilepsy; and an excess of variation in CLCN1 in patients with epilepsy compared to controls. They also demonstrate expression of C1C-1 mRNA and protein in human brain autopsy specimens and mouse brain. Mutations in the CLCN1 gene have been long recognized as the cause of dominant (Thomsen-type) and recessive (Becker-type) myotonia but an association with epilepsy has not been reported. Until now it was believed that ClC-1 protein and channels were not expressed in the brain; therefore, the mutation would have no effect on the CNS. The patient reported in this study had generalized epilepsy and generalized seizures, which involve thalamocortical circuits. Interestingly, the authors found that ClC-1 mRNA was moderately expressed in the mouse thalamus and cortex. The clinical phenotype, combined with the animal data, strongly suggests a role for ClC-1 in the brain.

Expression and cellular distribution of multidrug resistance-related proteins in patients with focal cortical dysplasia

Recent arouse of interest indicated that drug resistant proteins are markedly over-expressed in the epileptogenic tissue and they may be responsible for the one-third of the epileptic patients who were refractory to anti-epileptic drugs (AEDs). Since several AEDs may act as substrates for these drug resistant proteins, the enhanced function of such proteins may increase drug extrusion, resulting in inadequate response to drug therapy in patients with epilepsy. We studied expression of the multidrug resistance protein 1 (MDR1) and multidrug resistance-associated protein 1 (MRP1) in the epileptic tissues resected surgically in 28 patients with focal cortical dysplasia (FCD) by immunohistochemistry. The results were compared with 10 normal necropsy brain tissues. Normal brain showed no MDR1 expression in neurons and astrocytes, while MRP1 expression was very weak, which were encountered in a few samples. MDR1 expression was mainly localized on the vascular endothelial cells. In contrast to normal brain, we found intense MDR1 and MRP1 expression in both neurons and reactive astrocytes in the vast majority of dysplastic tissues. The majority of the dysplastic neurons demonstrated moderate to strong MRP1 immunoreactivity. Endothelial cells showed both MDR1 and MRP1 expression in the majority of the specimens studied. Multidrug transporters are over-expressed in the epileptogenic zone in patients with FCD. These results are concordant with previous studies, in which over-expression of multidrug proteins were shown in epileptogenic brain tissue in patients with FCD, that the over-expression of drug transport proteins in tissue from patients with refractory epilepsy may explain one possible mechanism for drug resistant in these pathologies.

NPC2, the Protein Deficient in Niemann-Pick C2 Disease, Consists of Multiple Glycoforms That Bind a Variety of Sterols

Niemann-Pick C disease is a fatal neurodegenerative disorder characterized by an endolysosomal accumulation of cholesterol and other lipids. One form of the disease is caused by a deficiency in NPC2, a soluble lysosomal glycoprotein that binds cholesterol. To better understand the biological function of NPC2 and how its deficiency results in disease, we have characterized the structural and functional properties of recombinant human protein. Highly purified NPC2 consists of a complex mixture of glycosylated isoforms, similar to that observed in human brain autopsy specimens. Mass spectrometric analysis revealed that of the three potential N-linked glycosylation sites present in the mature protein, Asn-19 is not utilized; Asn-39 is linked to an endoglycosidase H (Endo H)-sensitive oligosaccharide, and Asn-116 is variably utilized, either being unmodified or linked to Endo H-sensitive or Endo H-resistant oligosaccharides. All glycoforms are endocytosed and ameliorate the cholesterol storage phenotype of NPC2-deficient fibroblasts. In addition, the purified preparation contains a mixture of both free and lipid-bound protein. All glycoforms bind cholesterol, and sterol binding to NPC2 significantly alters its behavior upon cation-exchange chromatography. Based on this observation, we developed chromatography-based binding assays and determined that NPC2 forms an equimolar complex with the fluorescent cholesterol analog dehydroergosterol. In addition, we find that NPC2 binds a range of cholesterol-related molecules (cholesterol precursors, plant sterols, some oxysterols, cholesterol sulfate, cholesterol acetate, and 5-alpha-cholestan-3-one) and that 27-hydroxysterol accumulates in NPC2-deficient mouse liver. Binding was not detected for various glycolipids, phospholipids, or fatty acids. These biochemical properties support a direct and specialized function of NPC2 in lysosomal sterol transport.

Immunohistochemical localization of calmodulin-dependent cyclic phosphodiesterase in the human brain.

The amplification of cyclic nucleotide 'second messenger' signals within neurons is controlled by phosphodiesterases which are responsible for their degradation. Calmodulin-dependent phosphodiesterase (CaMPDE) is an abundant enzyme in brain which carries out this function. For the first time, we have localized CaMPDE in the normal human brain at various ages, using a mononoclonal antibody designated A6. This antibody was generated using standard techniques, purified, and applied to tissue sections. Autopsy specimens of human brain with no neuropathological abnormalities were selected representing a range of pre- and postnatal ages. Sections of various brain regions were evaluated for immunoreactivity, graded as nil, equivocal, or definite. We demonstrated definite CaMPDE immunohistochemical staining in neocortex, especially in neurons in layers 2 and 5. There was definite neuronal immunoreactivity in the hippocampus, and in the subiculum. The striatum had definite patchy neuronal staining. Definite terminal staining in the globus pallidus externa and substantia nigra pars reticulata outlined resident neurons, interpreted as axonal terminal staining. Cerebellar Purkinje cells showed definite immunoreactivity. In the developing brain, definite immunohistochemical staining was seen in the cerebellar external granular layer. The expression of CaMPDE in specific subsets of neurons suggests they may correlate with cells having dopaminergic innervation and/or high levels of neuronal integration.

A cell-free approach to the study of membrane trafficking in frozen rat brains potentially applicable to frozen autopsy material

A cell-free transfer system was used to measure capacity of brain membranes to support membrane renewal. To study transfer in brain, radiolabeled donor microsome fractions were prepared using brain slices from rats or frozen human brain autopsy specimens. Acceptor fractions, prepared from fresh or frozen rat brain or frozen human brain autopsy specimens, were immobilized on nitrocellulose. The complete reconstituted transfer system contained ATP plus ATP-regenerating system (or NADH) as a source of energy and brain cytosol. Slices of frozen brain incorporated acetate into membrane lipids with approximately the same efficiency as fresh brains. This efficiency declined with storage at 4 °C but only slowly. Donor fractions labeled with acetate from frozen slices exhibited specific transfer (37 °C minus 4 °C) of labeled membrane lipids with efficiencies comparable to fresh. The acceptor fraction could be prepared either from fresh or frozen material. Transfer was on the average two-fold stimulated by ATP at 37 °C compared to no ATP. Transfer also was stimulated by NADH. Analysis of linear transfer rates between 0 and 30 min revealed no significant effect of delay time or of time of prolonged storage on transfer efficiency beyond an initial decline of ca. 25% observed within the first two weeks after freezing. A decline of transfer was obtained with brains as the animals aged.

Subregional and Intracellular Distribution of NADP-Linked Malic Enzyme in Human Brain

High total activity (expressed as mumol/min/g of wet tissue or per milligram of DNA) and differential subregional distribution of NADP-linked malic enzyme was found in autopsy specimens of human brain. Striatum showed the highest activity of malic enzyme, which was two- to five-fold higher than that in other human organs tested. High activity was also found in frontal cortex, while the lowest activity of the enzyme in the central nervous system was found in cerebellum, substantia alba, and corpus callosum. In striatum, frontal cortex, pons, and cerebellum more than 80% of total malic enzyme activity was localized in the mitochondrial fraction, while in substantia alba and corpus callosum approximately 60% of the enzyme activity was present in the mitochondrial fraction. Relatively high specific activity of malic enzyme was found in a crude mitochondrial fraction isolated from various regions of human brain. The highest specific activity was found in the mitochondria isolated from striatum (more than 100 nmol/min/mg of mitochondrial protein); the lowest, but still high (approximately 32 nmol/min/mg of mitochondrial protein) was present in corpus callosum. These data and the different ratios of citrate synthase to mitochondrial malic enzyme activities found in different regions of brain suggest that human brain mitochondria, like the mitochondria isolated from other mammalian brains, are extremely heterogenous. A possible role of mitochondrial malic enzyme in human brain metabolism is discussed.

A study on peritumoral brain edema around meningiomas by MRI and contrast CT.

In the present study upon 9 meningiomas, the volume of peritumoral brain edema was calculated by integration of the cross-sectional edematous areas on serial MRI slices. It was zero in 3 cases and ranged from 11 to 176.4 ml in the other cases. There was disruption of the cortex in all cases, ranging from slight to severe. T1 and T2 measurements were made at the level of maximum edema extension (at 1.5 T), using a mixed sequence. From the T2 values tissue water content in % was calculated using equations, which had been obtained by correlating water content with relaxation rates. These had been measured on human brain autopsy specimens subjected to hydration or dehydration. Assuming that spread of contrast agent marks the advancement of the front of edema produced by the tumor, calculation of the excess of edema production per unit tumor volume Veff/VN was performed on the basis of CT-studies made before, and at 1.5, 3 and 6 hr after contrast infusion. Of 6 tumors with edema (mean water content of peritumoral white matter of 91% and mean edema volume of 69.2 ml) the Veff/VN was 0.18-1.08 ml/h/cm3 tumor volume, whereas 3 tumors without associated edema had a Veff/VN of 0.03-0.12 ml/h/cm3. In meningiomas cortex penetration seemed to be a separate factor determining the spread of edema.

A study on peritumoural brain oedema around meningiomas by CT and MRI scanning.

There is a great variability in the amount of peritumoural brain oedema accompanying meningiomas. In a previous study it was found that the degree of brain oedema in the white matter around meningiomas correlated with disruption of the layers (especially the cerebral cortex), which separate the tumour from the white matter, as well as with the size and histological subtype of the tumour. In the present study comprising 9 meningiomas, the volume of oedema was calculated by integration of the cross-sectional oedematous areas on serial MRI slices. The volume of oedema was zero in 3 cases and ranged from 11 to 176.4 ml in the other 6 cases. The MRI-scans also showed disruption of the cortex in all cases, ranging from slight to severe. T1 and T2 measurements were made at the level of maximum extension, using a mixed sequence at a field strength of 1.5 T. From the T2 values tissue water content in % was calculated using the equations: WC = 39.36/(R2 + 37.2) for cortex, and WC = 29.63/(R2 + 27.8) for white matter. These had been obtained by correlating water content with relaxation rates, measured in vitro on human brain autopsy specimens which were subjected to hydration with distilled water or dehydration by hyperosmolar solutions. Mean water content amounted to 82.53% for normal cortex, 74.72% for normal white matter, and 84.59% for oedematous white matter around the tumour.(ABSTRACT TRUNCATED AT 250 WORDS)

Phenylethanolamine N-methyltransferase activity is decreased in Alzheimer's disease brains.

Phenylethanolamine N-methyltransferase (PNMT) is the rate-limiting enzyme involved in the synthesis of epinephrine and a specific marker for epinephrine neurons. We have previously described the stability of this enzyme in a variety of pre- and postmortem conditions in human brain autopsy specimens and demonstrated its presence in areas of the human brain that are associated with memory and attention. We now report on 5 patients with Alzheimer's disease (AD) who had a decrease in PNMT activity of 37 to 48% in areas of the brain affected by the disease but not in the cerebellum, an area of the brain unaffected by the disease. The degree of decrease in PNMT activity in the hippocampus correlated significantly with the degree of dementia. We have provided direct immunochemical evidence that the decrease in PNMT activity in AD is due to the loss of this specific enzyme protein.

Reduced and oxidized glutathione in human and monkey brain

Prior animal studies have indicated that levels of oxidized glutathione (GSSG) in brain and other organs are low, comprising several percent, or less, of the total glutathione. An exception is seen in reports for autopsy and biopsy specimens of human brain, where very high levels of GSSG have been indicated. The latter observations imply an unusual redox state for human brain. In the current study, GSSG and reduced glutathione (GSH) were measured in monkey brain and autopsy specimens of human brain. Levels of GSSG were 1.2% or less, of the total glutathione. These results are similar to those for rodent brain, but disagree with earlier reports concerning human brain.

Distribution and concentration of mercury in autopsy specimens of human brain.

THE dangers of mercury, a pollutant of natural bodies of water and a metal with a strong tendency to accumulate in the nervous tissues, have recently caused considerable concern. We have studied the distribution and concentration of mercury in the brain in order to determine the levels normally present.

STUDIES ON BRAIN GANGLIOSIDES. IV. THE EFFECT OF HYPERCAPNIA ON GANGLIOSIDES IN VIVO.

Gangliosides are relatively stable acidic glycolipid components of neuronal membranes. When isolated and purified from normal human, rat, or cat brains, they contain 1 μmole of N-acetylneuraminic acid (NANA) per mg. Gangliosides, isolated and purified from fresh autopsy specimens of human brain of patients who were cyanotic before death, contained considerably less NANA and N-acetyl-galactosamine than normal brain gangliosides. Also, gangliosides isolated from cerebral tissue adjacent to a variety of intracerebral lesions contained less NANA than gangliosides from uninvolved tissue from the same patient. Gangliosides from the brains of cats which had been asphyxiated for periods up to 8 minutes also contained less NANA and hexosamine. Experiments in which cats were treated with various gas mixtures showed that hypercapnia and the consequent respiratory acidosis were the major causes of the change in ganglioside composition in vivo. Treatment with 100% N2produced a respiratory alkalosis but had no effect on brain ganglioside composition. The mechanism by which hypercapnia causes the change in the composition of the membrane gangliosides is unclear. An intact cerebral circulation appeared necessary to elicit the changes. The possibility that hypercapnia releases enzyme activators (e.g. adrenalin) or activates brain glycosidases is discussed. The loss of anionic groups located in the neuronal membrane would alter the behavior of these lipids towards cations and may, in part, explain the altered excitability of neurones following hypercapnia.