ABSTRACT
Recent observations in our laboratory have indicated substantial and systematic regional variations in the loss of cortical cholinergic fibers in Alzheimer disease (AD). Previous attempts to study the relationship between cortical cholinergic loss and the density of cortical pathological lesions have resulted in conflicting findings. Furthermore, most reports have correlated density of plaques and tangles with the residual level of cholinergic innervation rather than its loss. The purpose of the present study was to determine the relationship between loss of cholinergic axons and density of tangles and beta-amyloid (Abeta) deposits in various cortical areas of AD brains. Abeta deposits and tangles were observed throughout the cerebral cortex. Quantitative analysis revealed almost no correlation between loss of cholinergic fibers and the density of Abeta deposits. Qualitative observations revealed similar results when cored and neuritic plaques were considered separately. By contrast, cholinergic fiber loss displayed a significant correlation with the density of tangles (r = 0.52-0.79). However, in a few areas, such as the cingulate cortex, tangle density appeared to be unrelated to the loss of cholinergic fibers. These results indicate that cortical cholinergic denervation in AD is related to cytoskeletal pathology. However, the lack of a perfect relationship with cytoskeletal pathology implicates additional factors in the cholinergic pathology of AD.
Subject(s)
Alzheimer Disease/pathology , Amyloid beta-Peptides/analysis , Cerebral Cortex/pathology , Choline O-Acetyltransferase/analysis , Nerve Fibers/pathology , Neurofibrillary Tangles/pathology , Plaque, Amyloid/pathology , Aged , Benzothiazoles , Cerebral Cortex/ultrastructure , Fluorescent Dyes , Humans , Nerve Fibers/ultrastructure , Neurofibrillary Tangles/ultrastructure , Organ Specificity , Plaque, Amyloid/ultrastructure , Reference Values , Regression Analysis , ThiazolesABSTRACT
Neurotoxic effects of excitatory amino acid (EAA) receptor agonist N-methyl-D-aspartic acid (NMDA) and its antagonists on ventral horn cholinergic neurons were studied in organotypic rollertube cultures of spinal cord (OTC-SCs) using biochemical assays of choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) activity, and AChE histochemistry. NMDA exposure decreased ChAT and AChE activity by 83% and 66%, respectively. Cultures treated with NMDA also showed a marked loss of AChE staining in both dorsal and ventral horns and a significant, dose-dependent decrease in the number of ventral horn AChE-positive neurons (VHANs). NMDA treatment primarily resulted in the loss of small VHANs (< 300 microns2). VHANs with a size and distribution typical of alpha-motoneurons were relatively well preserved. The effects of NMDA on OTC-SCs appeared to be independent of the age of the cultures. The NMDA antagonist DL-AP5 completely prevented the NMDA-induced loss of ChAT activity, but only attenuated the effect of NMDA on ChE activity. The antagonists DL-AP5, D-AP5 and MK-801, used alone, caused significant loss and/or shrinkage of VHANs. These effects appeared to be distinct from the NMDA mediated toxicity. The results indicate that NMDA and its antagonists exert powerful toxic effects on ventral horn cholinergic neurons. The large cholinergic alpha-motoneurons, however, appear to be relatively immune to these toxic effects.
Subject(s)
Anterior Horn Cells/drug effects , Cholinergic Fibers/drug effects , N-Methylaspartate/antagonists & inhibitors , N-Methylaspartate/drug effects , Neurons/drug effects , Acetylcholinesterase/chemistry , Acetylcholinesterase/drug effects , Acetylcholinesterase/metabolism , Animals , Anterior Horn Cells/pathology , Anterior Horn Cells/physiology , Cellular Senescence/drug effects , Cellular Senescence/physiology , Choline O-Acetyltransferase/chemistry , Choline O-Acetyltransferase/drug effects , Choline O-Acetyltransferase/metabolism , Cholinergic Fibers/physiology , Dose-Response Relationship, Drug , Excitatory Amino Acid Agonists/pharmacology , Excitatory Amino Acid Agonists/toxicity , N-Methylaspartate/toxicity , Organ Culture Techniques , RatsABSTRACT
Peripheral neuropathy is the major dose-limiting toxicity of the hypoxic cell sensitiser, etanidazole. Previous work from this laboratory using culture neuronal cell lines suggested that nitroimidazole-induced degradation of neurofilament proteins might be the critical biological event mediating this neurotoxicity. The purpose of the present study was to develop the neurofilament degradation assay in an organotypic spinal cord culture system with the goal of developing strategies for optimising sensitiser efficacy as well as ameliorating nitroimidazole-induced neurotoxicity. Spinal cord cultures were treated with etanidazole and neurofilament protein degradation was analysed by immunoblot analysis. Spinal cord cultures exposed to etanidazole exhibited a dose-dependent loss of parent neurofilament proteins, with concomitant appearance of low molecular weight degradation products. The potential neurotoxic effect of L, S-buthionine sulphoximine (BSO), a compound that enhances the radiosensitising effectiveness of 2-nitroimidazoles, was also screened in this assay system. BSO alone, at concentrations up to 100 microM, did not promote neurofilament degradation. BSO (20 microM) enhanced the effect of etanidazole on neurofilament degradation by a dose-modifying factor of 1.6 +/- 0.5. Since 20 microM BSO is expected to enhance etanidazole radiosensitisation of hypoxic cells by a larger factor, this suggests that a therapeutic gain could be achieved using BSO in combination with etanidazole in radiation therapy.
Subject(s)
Antineoplastic Agents/pharmacology , Buthionine Sulfoximine/pharmacology , Etanidazole/pharmacology , Neurofilament Proteins/metabolism , Radiation-Sensitizing Agents/pharmacology , Spinal Cord/drug effects , Animals , Animals, Newborn , Drug Synergism , Organ Culture Techniques , Rats , Spinal Cord/metabolism , Superoxides/metabolismABSTRACT
PURPOSE: To determine the feasibility of modifying the aerobic cytotoxicity of etanidazole without interfering with the tumoricidal action of radiation plus etanidazole. METHODS AND MATERIALS: The aerobic cytotoxicity of etanidazole was studied using two different models: (1) Induction of apoptosis in EL4 cells: apoptotic DNA fragmentation was analyzed by agarose gel electrophoresis following 24 h treatment with etanidazole alone or in combination with various modifiers. (2) Spinal cord neuronal loss in organotypic roller tube cultures: Survival of acetylcholinesterase positive ventral horn neurons was analyzed morphometrically following 72 h treatment with etanidazole alone or in combination with vitamin E succinate. RESULTS: Etanidazole (10 mM) induced apoptosis in EL4 cells. This effect was suppressed by 24 h treatment with TPA, IBMX, the free radical scavenger TEMPOL or vitamin E succinate. Vitamin E succinate also protected spinal cord cultures from etanidazole-induced neuronal loss. CONCLUSION: These results suggest that it might be possible to modify the neurotoxicity of etanidazole with agents that would not be expected to interfere with the tumoricidal action of radiation plus etanidazole.
Subject(s)
Etanidazole/pharmacology , Aerobiosis , Animals , Apoptosis , Calcium/metabolism , Cell Survival/drug effects , Lymphoma, T-Cell/pathology , Mice , Superoxides/metabolism , Tocopherols , Tumor Cells, Cultured , Vitamin E/analogs & derivatives , Vitamin E/pharmacologyABSTRACT
We studied the direct effect of platelet secretory products on rat spinal cord explants. Morphological changes in the ventral horn neurons were assessed after staining for acetylcholinesterase (AChE). Compared to control, exposure to platelet secretory products was associated with a significant decrease in the number of AChE-positive neurons per ventral horn. Aspirin appeared to partially prevent this neurotoxicity. These results suggest that platelet secretory products may contribute to neuronal injury.
