Peter A. Paskevich

Decreased Neuronal and Increased Oligodendroglial Densities in Huntington's Disease Caudate Nucleus

Decreased density of neurons was found throughout the head of the caudate nucleus in Huntingtons disease (HD), with the most severe neuronal loss early in the disease in the medial region. The density of reactive astrocytes is inversely proportional to the neuronal loss. In cases of mild Huntingtons disease which had no identifiable abnormality on conventional neuropathologic evaluation (grade 0), there is a reduction in neuron density without an accompanying reactive astrocytosis. The pattern for decrease in neurons and accompanying astrocytosis suggests that the earliest changes occur in the most medial portion of the head of the caudate nucleus and subsequently sweep laterally across the caudate nucleus to the internal capsule. An increased density of oligodendrocytes is observed in the head of the caudate nucleus for the lower grades (0, 1 and 2). The decreased neuronal and increased Oligodendroglial densities may be of significance in understanding the pathogenesis of HD. These altered densities, observed in the absence of reactive astrocytosis, suggest that these changes may not represent recent effects of disease, but rather that HD gene expression may influence brain cell densities from early in the life of the gene carrier.

The amyloid precursor protein is concentrated in neuronal lysosomes in normal and Alzheimer disease subjects.

The 4.2-kilodalton (kDa) polypeptide associated with the cerebral amyloid deposits of Alzheimers disease (AD) derives from a much larger protein that is encoded by a gene on chromosome 21. In the present study, we have used antibodies raised against portions of the amyloid protein precursor (APP) to map its normal distribution and to gain further insights into the events that lead to amyloid deposition. Antibodies raised against several different portions of APP reacted with proteins having apparent molecular sizes of 65, 67, and 132 kDa on Western blots. In sections through the normal human brain, immunocytochemistry revealed punctate concentrations of the protein in pyramidal cells of the neocortex, particularly in associative regions, and intense staining in the CA1 pyramidal cells of the hippocampus. By electron microscopy, this punctate distribution coincided with dense concentrations of the protein in secondary lysosomes. In the hippocampus of several AD cases examined, abnormally dense immunostaining in enlarged intracellular domains accompanied a severe atrophy of the CA1 neurons. These data suggest that accumulations of APP in lysosomes of particular neurons may, in AD, lead to proteolytic events that form the insoluble 4.2-kDa amyloid peptide.

The Effects of Haloperidol on Synaptic Patterns in the Rat Striatum

A morphometric analysis of the corpus striatum of rats chronically treated with haloperidol was performed at the light and electron microscopic levels. Although the density of striatal neurons was unchanged in the haloperidol-treated group, there was a small increase in neuronal size (13%). This change in cell size was paralleled by a trend towards larger dendrite calibres occurring in the drug-treated animals. The distribution curve for axon terminal size indicated that 12% of the overall population was shifted from a range with a median size of 0.8 micron 2 to one with 1.6 micron 2 in the drug-treated group. This increase in size of some striatal terminals was accompanied by a concomitant increase in numbers of their associated synaptic vesicles, resulting in a similar density of vesicles for both control and drug-treated animals.

The Lysosomal System in Neurons

Disturbed lysosomal function may be implicated at several stages of Alzheimers pathogenesis. Lysosomes and acid hydrolases accumulate in the majority of neocortical pyramidal neurons before typical degenerative changes can be detected, indicating that altered lysosome function is among the earliest markers of metabolic dysfunction in Alzheimers disease. These early alterations could reflect accelerated membrane and protein turnover, defective lysosome or hydrolase function, abnormal lysosomal trafficking or any combination of these possibilities. Because APP is partly metabolized in lysosomes, early disturbances in lysosomal function could promote the production of abnormal and/or neurotoxic APP fragments within intact cells. Lysosomal abnormalities progressively worsen as neurons begin to degenerate. Based on existing literature on cell death, increased perturbation and instability of the lysosomal system may be expected to contribute to the atrophy and eventual lysis of the neuron. Finally, the release of hydrolase-filled lysosomes and lipofuscin aggregates from dying neurons accounts for the abundant deposition of enzymatically active acid hydrolases of all classes in the extracellular space--a phenomenon that may be unique to Alzheimers disease. Acting on APP present in surrounding dystrophic neurites, cellular debris and astrocyte processes, dysregulated hydrolases may cleave APP in atypical sequential patterns, thereby generating self-aggregating protease-resistant APP fragments that can be only processed to beta-amyloid. Genetic mutations or posttranslational factors of APP should further enhance the generation of amyloidogenic fragments by a dysregulated lysosomal system. Given that very little, if any, beta-amyloid is detected intracellularly, yet extracellular beta-amyloid is very abundant, our data suggest that the final steps of APP processing and the generation of most beta-amyloid in the brain parenchyma occur extracellularly and may involve one or more lysosomal proteases.

Synaptic rearrangements in medial prefrontal cortex of haloperidol-treated rats.

The effects of daily administration of haloperidol for 16 weeks on the structure of layer VI in medial prefrontal cortex of rat was performed at the light and electron microscopic levels. At the light microscopic level, no difference in either the size or the density of neurons was observed. At the electron microscopic level, the mean dendritic calibre of haloperidol-treated rats was twice that observed in control animals, but this was due to a selective loss of small-calibre dendritic profiles. Rats treated with neuroleptic also showed a reduction in axon terminals with asymmetric postsynaptic membrane specializations, which, in control animals, were preferentially associated with small-calibre dendritic profiles. These small-calibre dendritic profiles were found to be spines rather than small terminal dendritic shafts. An increase in axon terminals showing no membrane specialization on larger dendritic profiles also occurred in rats treated daily with the neuroleptic. The data suggest the possibility that haloperidol may have induced a relocation of asymmetric terminals from resorbed spinous processes to larger dendritic branches with the concomitant loss of their postsynaptic membrane specialization.

Aluminum salts induce the accumulation of neurofilaments in perikarya of NB2a/dl neuroblastoma

NB2a/dl neuroblastoma cells were exposed to aluminum chloride or aluminum lactate (0.1-1 mM) for 3 and 6 days. Additional cultures were exposed to aluminum salts as the cells were stimulated to elaborate axonal neurites by dibutyryl cyclic AMP. By phase-contrast microscopy, aluminum salts had no effect on the morphology of undifferentiated (NB2a(-] or differentiated (NB2a(+] cells, or on neuritic elaboration and maintenance. Silver straining by the Bielschowsky method, however, demonstrated argyrophilic accumulations in perikarya of many NB2a(-) and NB2a(+) cells treated with aluminum salts. At the ultrastructural level, whorls of intermediate filaments were the most prominent abnormalities in neuronal perikarya. Although phosphorylated high-molecular weight neurofilament subunits (NF-H) are normally detected by immunocytochemical analyses only within axonal neurites of NB2a/dl cells, aluminum salt treatment caused the detection of phosphorylated epitopes of NF-H within perikaryal of NB2a(-) and NB2a(+) cytoskeletons, suggesting that the argyrophilic filamentous accumulations are composed at least partly of phosphorylated NF-H.

Abnormalities of the Endosomal-Lysomal System in Alzheimer’s disease

The lysosome is a major component of a dynamic and polymorphic system of acidic vacuolar compartments in the cell that is capable of degrading most large cellular molecules, i.e., nucleic acids, polysaccharides, proteins, and lipids, to low molecular weight products. As the major site of intracellular digestion, lysosomes contain dozens of hydrolytic enzymes that are housed within membrane-bound, low pH compartments (1, 2). The use of electron microscopy, enzyme and immunocytochemistry techniques have provided morphological criteria to identify lysosomes and to distinguish among different lysosomal stages and types. Lysosomal biogenesis begins within the endomembrane system of organelles. Synthesis and glycosylation take place in the endoplasmic reticulum followed by posttranslational modification and acquisition of phosphmannosyl residues within the Golgi apparatus. Mannose phosphate receptors interact with trans Golgi-derived coated vesicles containing nascent hydrolases and deliver these to acidic prelysosomal compartments, the late endosomes (3–5). The membrane-bound compartments that house both enzyme and the material to be digested are quite heterogeneous and include different morphological types of structures such as dense bodies, multivesicular bodies, and autophagic vacuoles. As digestion proceeds, a third type of lysosome-derived structure forms containing accumulating material that is resistant to degradation along with varying amounts of acid hydrolase activities. These residual granules, which are characterized by the color and autofluorescence of the accumulated material include lipofuscin, ceroid, and hemofuscin (1–5).

Huntington’s Disease: Neuropathological Grading

The striatum was found to exhibit marked variation in the severity of neuropathological involvement in post mortem brain specimens from 163 clinically diagnosed cases of Huntington’s disease (HD). A grading system was established by macroscopic and microscopic criteria, resulting in five grades (0–4) designated in ascending order of severity. The earliest changes were seen in the medial caudate nucleus (CN), in the tail of the CN, and in the dorsal part of the putamen. The grade correlated with the extent of clinical disability as assessed by a rating scale. Quantitative measurements revealed a 50% neuronal loss in the CN in grade 1 and a 95% loss in grade 4. Astrocytes are greatly increased in grades 2–4. Analyses of the CN in grade 4 reflect mainly astrocytic composition with a component of remote neurons projecting to the striatum. Because of the relative preservation of the lateral half of the head of the CN in grades 1–2, these regions would reflect early cellular and biochemical changes in HD.