Chronic exposure to manganese decreases striatal dopamine turnover in human alpha-synuclein transgenic mice.

Interaction of genetic and environmental factors is likely involved in Parkinson's disease (PD). Mutations and multiplications of alpha-synuclein (α-syn) cause familial PD, and chronic manganese (Mn) exposure can produce an encephalopathy with signs of parkinsonism. We exposed male transgenic C57BL/6J mice expressing human α-syn or the A53T/A30P doubly mutated human α-syn under the tyrosine hydroxylase promoter and non-transgenic littermates to MnCl2-enriched (1%) or control food, starting at the age of 4 months. Locomotor activity was increased by Mn without significant effect of the transgenes. Mice were sacrificed at the age of 7 or 20 months. Striatal Mn was significantly increased about three-fold in those exposed to MnCl2. The number of tyrosine hydroxylase positive substantia nigra compacta neurons was significantly reduced in 20 months old mice (-10%), but Mn or transgenes were ineffective (three-way ANOVA with the factors gene, Mn and age). In 7 months old mice, striatal homovanillic acid (HVA)/dopamine (DA) ratios and aspartate levels were significantly increased in control mice with human α-syn as compared to non-transgenic controls (+17 and +11%, respectively); after Mn exposure both parameters were significantly reduced (-16 and -13%, respectively) in human α-syn mice, but unchanged in non-transgenic animals and mice with mutated α-syn (two-way ANOVA with factors gene and Mn). None of the parameters were changed in the 20 months old mice. Single HVA/DA ratios and single aspartate levels significantly correlated across all treatment groups suggesting a causal relationship between the rate of striatal DA metabolism and aspartate release. In conclusion, under our experimental conditions, Mn and human α-syn, wild-type and doubly mutated, did not interact to induce PD-like neurodegenerative changes. However, Mn significantly and selectively interacted with human wild-type α-syn on indices of striatal DA neurotransmission, the neurotransmitter most relevant to PD.

Age-related gene-specific changes of A-to-I mRNA editing in the human brain.

A-to-I editing is an adenosine-to-inosine modification of mRNA particularly widespread in the human brain, where it affects thousands of genes. A growing body of evidence suggests that A-to-I RNA editing is necessary for normal development and maintenance in mammals and that its deficiencies contribute to a number of pathological states. In this study, we examined whether mRNA editing levels of two mRNA species, CYFIP2 and GABRA3, change with aging. CYFIP2 has been implicated in synaptic maintenance, while GABRA3 is a GABA receptor subunit, a part of the major inhibitory neurotransmitter system in the CNS. The levels of mRNA editing were assessed in cortex samples of 20 subjects 22-102 years old. The data show an age-dependent statistically significant decrease in editing in CYFIP2. GABRA3 editing remained much more stable with age, implying that age-related decline of RNA editing is gene-specific. This is the first report of age-dependent decline in A-to-I editing. Further examination of these and other vulnerable genes may reveal specific RNA editing mechanisms that contribute to the aging phenotype.

Heritability, correlations and in silico mapping of locomotor behavior and neurochemistry in inbred strains of mice.

The midbrain dopamine system mediates normal and pathologic behaviors related to motor activity, attention, motivation/reward and cognition. These are complex, quantitative traits whose variation among individuals is modulated by genetic, epigenetic and environmental factors. Conventional genetic methods have identified several genes important to this system, but the majority of factors contributing to the variation remain unknown. To understand these genetic and environmental factors, we initiated a study measuring 21 behavioral and neurochemical traits in 15 common inbred mouse strains. We report trait data, heritabilities and genetic and non-genetic correlations between pheno-types. In general, the behavioral traits were more heritable than neurochemical traits, and both genetic and non-genetic correlations within these trait sets were high. Surprisingly, there were few significant correlations between the behavioral and the individual neurochemical traits. However, striatal serotonin and one measure of dopamine turnover (DOPAC/DA) were highly correlated with most behavioral measures. The variable accounting for the most variation in behavior was mouse strain and not a specific neurochemical measure, suggesting that additional genetic factors remain to be determined to account for these behavioral differences. We also report the prospective use of the in silico method of quantitative trait loci (QTL) analysis and demonstrate difficulties in the use of this method, which failed to detect significant QTLs for the majority of these traits. These data serve as a framework for further studies of correlations between different midbrain dopamine traits and as a guide for experimental cross designs to identify QTLs and genes that contribute to these traits.

Risk factors for dopaminergic neuron loss in human alpha-synuclein transgenic mice.

Genetic background, pesticide exposure, age, gender, diet and lifestyle are implicated risk factors in Parkinson's disease. We demonstrate dopamine neuron loss and other features of Parkinsonism based on the interaction of several of these human risk factors in transgenic mice expressing human alpha-synuclein. Mice expressing different forms of human alpha-synuclein had progressive declines in locomotor activity and abnormal responses to apomorphine that were modified by transgenic status. Stereological counts of tyrosine hydroxylase-positive neurons significantly declined with age only in the transgenic lines, consistent with a constant or decreasing risk, with the line expressing a double-mutant form of human alpha-synuclein more severely affected than the line expressing wild-type human alpha-synuclein. Treatment with Mn2+-ethylenebisdithiocarbamate and paraquat resulted in significantly greater effects in the double-mutant line than the other lines. Inclusions were not identified in the transgenic lines. Overexpression of human alpha-synuclein had adverse effects on substantia nigra pars compacta dopaminergic neurons that were modified by risk factors interacting in humans, including human alpha-synuclein mutations, ageing, and exposure to pesticides.

The dorsal root ganglia in adrenomyeloneuropathy: neuronal atrophy and abnormal mitochondria.

Adrenomyeloneuropathy (AMN), a disease of spinal cord, brain, adrenal, and testis, mostly affects men with spastic paraparesis or ataxia beginning in their second or third decade. The spinal cord displays bilateral, usually symmetrical, long tract degeneration particularly of the gracile tract in a "dying-back" pattern. The available data strongly indicate that the fundamental lesion in AMN is an axonopathy or neuronopathy. We compared lumbar dorsal root ganglia (DRG) from 3 AMN patients to 6 age-matched controls histologically, morphometrically, immunohistochemically, and ultrastructurally. There was no apparent neuronal loss, necrosis or apoptosis, nor obvious atrophy; nodules of Nageotte were sparse in both groups. The morphometric studies, however, did reveal neuronal atrophy with a decrease in the number of large neurons and a corresponding increase in neurons less than 2,000 microm2, especially in the 1,500-1,999 microm2 range. No consistent immunohistochemical differences were observed, and no specific cell type appeared to be lost. Many mitochondria in the AMN neurons demonstrated lipidic inclusions; this raises the possibility that, in addition to the well-known peroxisomal defect, impaired mitochondrial function may lead to a failure of ATP-dependent axoplasmic transport in AMN spinal tracts with consequent "dying-back" axonal degeneration. The observation that the DRG parent neurons of the degenerate gracile tracts in AMN undergo atrophy and do not display appreciable evidence of cell death, even at autopsy, provides a wide window of opportunity for the development of therapeutic strategies to combat or prevent this myeloneuropathy.

Potentiated and preferential effects of combined paraquat and maneb on nigrostriatal dopamine systems: environmental risk factors for Parkinson's disease?

The absence of any compelling basis for a heritable basis of idiopathic Parkinson's disease (PD) has focused attention on environmental exposures as causative agents. While the herbicide paraquat has repeatedly been implicated, its impact on dopamine systems following systemic exposures is equivocal. The restricted focus on paraquat also ignores the extensive geographical overlap of its use with other agrichemicals known to adversely impact dopamine systems, including ethylenebisdithiocarbamate fungicides such as maneb. The present study sought to determine whether combined exposures to paraquat and maneb would produce additive effects and support a multiple-hit environmental contribution to PD. C57BL/6 mice were exposed to either paraquat (5-10 mg/kg) or maneb (15-30 mg/kg) i.p. alone or in combination once a week for 4 weeks. Sustained decreases in motor activity immediately following injections were consistently observed only with combined exposures, with activity levels returning to control values 24 h later. Concurrently, levels of dopamine and metabolites and dopamine turnover were increased immediately post-injection only by combined exposures, and returned to control levels or below within 48 h. Reductions in tyrosine hydroxylase immunoreactivity, measured 3 days after the last injection, resulted only from combined exposure and were detected in dorsal striatum, but not in the nucleus accumbens. The fact that combined exposures resulted in potentiated effects that appear to target nigrostriatal dopamine systems suggests that these combinations may be important environmental risk factors for Parkinsonism. These findings also raise questions about the adequacy of current risk assessment guidelines for these chemicals which are based on effect levels derived from exposures to single agents.

The nigrostriatal dopaminergic system as a preferential target of repeated exposures to combined paraquat and maneb: implications for Parkinson's disease.

Experimental evidence supporting 1,1'-dimethyl-4,4'-bipyridinium [paraquat (PQ)] as a risk factor for Parkinson's disease (PD) is equivocal. Other agricultural chemicals, including dithiocarbamate fungicides such as manganese ethylenebisdithiocarbamate [maneb (MB)], are widely used in the same geographical regions as paraquat and also impact dopamine systems, suggesting that mixtures may be more relevant etiological models. This study therefore proposed that combined PQ and MB exposures would produce greater effects on dopamine (DA) systems than would either compound administered alone. Male C57BL/6 mice were treated twice a week for 6 weeks with intraperitoneal saline, 10 mg/kg paraquat, 30 mg/kg maneb, or their combination (PQ + MB). MB, but not PQ, reduced motor activity immediately after treatment, and this effect was potentiated by combined PQ + MB treatment. As treatments progressed, only the combined PQ + MB group evidenced a failure of motor activity levels to recover within 24 hr. Striatal DA and dihydroxyphenylacetic acid increased 1-3 d and decreased 7 d after injections. Only PQ + MB reduced tyrosine hydroxylase (TH) and DA transporter immunoreactivity and did so in dorsal striatum but not nucleus accumbens. Correspondingly, striatal TH protein levels were decreased only by combined PQ + MB 5 d after injection. Reactive gliosis occurred only in response to combined PQ + MB in dorsal-medial but not ventral striatum. TH immunoreactivity and cell counts were reduced only by PQ + MB and in the substantia nigra but not ventral tegmental area. These synergistic effects of combined PQ + MB, preferentially expressed in the nigrostriatal DA system, suggest that such mixtures could play a role in the etiology of PD.

Antipsychotic drug regulation of AMPA receptor affinity states and GluR1, GluR2 splice variant expression.

We recently reported that chronic administration of antipsychotic drugs dramatically elevated [3H]AMPA binding, with minimal elevation of [3H]CNQX binding in rat brain. The aim of the current study was to examine the mechanism of this effect. Chronic haloperidol minimally increased the total number of binding sites (total Bmax) compared to saline-injected animals. Specifically, haloperidol dramatically increased the proportion of high-affinity-site AMPA receptors (approximately 30% increase) without inducing a significant change in the low-affinity constant. In situ hybridization for flip and flop isoforms of GluR1 and GluR2 (AMPA receptors) was not altered in a pattern or degree that compared to the changes seen in AMPA receptor binding. These findings suggest that the long-term action of antipsychotic drugs may be to regulate AMPA receptor responsiveness to agonist stimulation via posttranscriptional means, and is unlikely to be related to GluR1 or GluR2 splice variant expression. This effect may have relevance to both the therapeutic effects and side effects of antipsychotic drugs in humans.

Regional decreases in alpha-[3H]amino-3-hydroxy-5-methylisoxazole-4-propionic acid ([3H]AMPA) and 6-[3H]cyano-7-nitroquinoxaline-2,3-dione ([3H]CNQX) binding in response to chronic low-level lead exposure: reversal versus potentiation by chronic dopamine agonist treatment.

This study evaluated the hypotheses that in vivo lead (Pb) exposure would alter alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor binding and, based on known glutamate-dopamine interactions and Pb-induced changes in dopamine (DA) systems, that AMPA binding might be differentially influenced by DA agonist treatment under conditions of Pb exposure. Alterations in high-affinity ([3H]AMPA) versus total AMPA [6-[3H]cyano-7-nitroquinoxaline-2,3-dione ([3H]CNQX)] receptor binding were determined in medial frontal cortex, dorsal striatum, and nucleus accumbens of rats exposed to 0, 50, or 150 ppm of Pb acetate for 2 weeks or 8 months. Additional 8-month groups received chronic intermittent treatment with saline, the D1 agonist SKF82958, or the general DA agonist apomorphine. Two-week exposures increased AMPA receptor densities, whereas robust decreases occurred after 8 months of Pb; at the latter time point changes were more pronounced for high-affinity than total AMPA receptor binding, with high-affinity effects expressed preferentially in dorsal striatum and nucleus accumbens. DA agonist treatments almost fully reversed Pb-related declines in [3H]AMPA binding but either had no effect (apomorphine) or even further potentiated (SKF82958) the decreases in [3H]CNQX binding. One possible basis for the long-term (8-month) decrease in AMPA binding is a postsynaptic glutamatergic stimulation of non-NMDA receptors.

Time course and regional basis of Pb-induced changes in MK-801 binding: reversal by chronic treatment with the dopamine agonist apomorphine but not the D1 agonist SKF-82958.

In the present study we attempted to further define the time course and regional specificity of lead (Pb)-induced changes in the NMDA receptor complex and the influence of dopaminergic system modulations on these changes. Autoradiographic measurements of alterations in MK-801 binding, as evaluated under four different activation conditions (none, spermidine, glycine, or maximal activation), were performed in medial frontal cortex, dorsal striatum, and nucleus accumbens of male rats after 2 weeks or 8 months of chronic postweaning (from 21 days of age on) exposure to 0, 50, or 150 ppm Pb acetate in drinking water. The 8-month groups also received chronic intermittent intraperitoneal injections of saline, or of the dopamine (DA) agonist apomorphine or the D1 agonist SKF-82958 2-3 times per week beginning at 60 days of age. Two weeks of 50 ppm Pb exposure resulted in small but significant increases in MK-801 binding under conditions of glycine or spermidine activation, whereas decreases were observed in response to 150 ppm under conditions of no or maximal activation in all regions. After 8 months of Pb, concentration-dependent decreases in MK-801 binding were observed across regions under all activation conditions. These effects were noted at blood Pb concentrations averaging as low as 16 microg/dl. Pb-induced decreases in MK-801 binding were either partially or fully reversed by chronic intermittent treatment with the DA agonist apomorphine but not by the D1 agonist SKF-82958, implicating D2-based mechanisms in this reversal. Combined findings from this and previous studies based on this exposure protocol indicate a Pb-induced pattern of widespread hypoglutamatergic function accompanied by increased DA function in mesolimbic systems, a pattern of changes reminiscent of those proposed to underlie schizophrenia. Such findings suggest that Pb exposure, even at current environmental levels, could be a risk factor for behavioral and/or neurological disturbances arising from imbalances of glutamate/dopamine function in mesocorticolimbic systems.

Densitometrical analysis of opioid receptor ligand binding in the human striatum--I. Distribution of mu opioid receptor defines shell and core of the ventral striatum.

Changes in opioid neurotransmission have been implicated in several basal ganglia-related neurological and psychiatric disorders. To gain a better insight into the opioid receptor distribution in the normal human striatum, we examined in post mortem brain the distribution of the mu opioid receptor using ligand binding of [3H]O-ala2-N-methyl-phe4, gly-ol5-enkephalin. Our results indicate at the regional level the presence of a dorsal-to-ventral high-to-low density gradient in the striatum, with lowest densities in the ventral one-third of the putamen and in the nucleus accumbens. At the subregional level, the nucleus accumbens shows two major types of heterogeneities. First, low vs intermediate binding densities distinguish the core and shell subdivisions, respectively. The low-density core and intermediate-density shell regions extend into the putamen and are therefore characteristic for the entire ventral striatum. The second type of heterogeneity is formed by small areas located along the ventral contours of the nucleus accumbens and putamen that display the highest binding density of the entire striatum. Since these areas can also be recognized in the distribution patterns of other markers and in the cytoarchitecture, they appear to possess a separate identity. To emphasize their special neurochemical characteristics we propose the description "neurochemically unique domains in the accumbens and putamen". The present results, with the difference between core and shell of the ventral striatum as the most prominent outcome, together with the notion that the connectional relationships and neurochemical organization of the striatum are very heterogeneous, suggest a strong regional functional differentiation for mu receptor function in the human striatum.

Chronic treatment with typical and atypical antipsychotics increases the AMPA-preferring form of AMPA receptor in rat brain.

We assessed the effects of chronic (21 day) administration of antipsychotic drugs on the density of the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor in rat brain. We used two typical antipsychotic drugs, haloperidol and pimozide, and two atypical antipsychotic drugs, risperidone and clozapine. Antipsychotic drugs as a group significantly elevated the density of the AMPA receptor measured with an AMPA receptor agonist ([3H]AMPA), but not with an AMPA receptor antagonist, 6-cyano-7-nitro-quinoxaline-2,3-dione ([3H]CNQX). In all regions studied, the magnitude of the increase seen with chronic typical antipsychotic drugs was significantly greater than that seen with chronic atypical antipsychotic drugs. In frontal cortex and striatum, typical antipsychotics but not atypical antipsychotics elevated AMPA receptor binding over control. These findings suggest that antipsychotic drugs alter the agonist affinity of the AMPA receptor without altering the number of AMPA receptors. Typical antipsychotic drugs may be more potent in this effect than atypical antipsychotic drugs, especially in critical corticostriatal circuits.

Opioid receptor ligand binding in the human striatum: II. Heterogeneous distribution of kappa opioid receptor labeled with [3H]bremazocine.

Selective kappa opioid receptor autoradiography with [3H]bremazocine (BRM) was used to examine regional and subregional kappa receptor distribution patterns at five rostrocaudal levels through the human striatum. [3H]BRM binding densities were measured in the individual striatal nuclei and in subregions therein. The distribution of [3H]BRM binding sites was found to have a strongly heterogeneous character. At the regional level a rostral-to-caudal decrease in [3H]BRM binding densities was observed. Also, a dorsal-to-ventral differentiation was seen, with higher values in the ventral striatum, especially in the nucleus accumbens, and lower values in the dorsal parts of the caudate nucleus and putamen. These findings suggest an association of kappa receptor function with limbic-related processes in the ventral striatum. Along the ventral edge of the nucleus accumbens and putamen, specific domains with extremely high [3H]BRM binding values were identified.

Preferential vulnerability of nucleus accumbens dopamine binding sites to low-level lead exposure: time course of effects and interactions with chronic dopamine agonist treatments.

This study examined the hypotheses that low-level lead (Pb) exposure would increase dopamine (DA) binding sites, would do so preferentially in nucleus accumbens, and that such effects would be modified by concurrent DA agonist treatment. D1-like and D2-like binding sites and the dopamine transporter (DT) were measured autoradiographically in caudate-putamen and nucleus accumbens of rats exposed from weaning to 0, 50, or 150 ppm Pb acetate drinking solutions with or without concurrent chronic intermittent intraperitoneal injections of the D1-like agonist SKF 82958 or the DA agonist apomorphine after 2 weeks (no injections), 8 months, or 12 months of Pb exposure. Pb selectively decreased DA binding in nucleus accumbens. Decreases in D2-like and DT sites were sustained across the 12-month exposure, whereas D1-like sites evidenced recovery at 12 months. Chronic intermittent DA agonist treatments reversed these effects of Pb in nucleus accumbens, restoring receptor and DT binding levels to normal, despite decreasing binding sites of non-Pb-treated rats. These studies implicate increased DA availability as a mechanism of Pb-induced DA system changes. They also raise the possibility that Pb exposure could serve as a predisposing factor in neurodegenerative diseases associated with DA system dysfunction or could alter the course of DA-based therapeutic treatments.

Chronic antipsychotic treatment alters glycine-stimulated NMDA receptor binding in rat brain.

In this study the chronic effect of antipsychotic drugs (APDs) on N-methyl-D-aspartate (NMDA) receptor binding was evaluated. Rats were treated for 21 days with i.p. injections of haloperidol (0.5 mg/ kg), pimozide (0.5 mg/kg), clozapine (20 mg/kg), risperidone (1 mg/kg) or water vehicle (1 ml/kg). Brain tissue sections underwent different [3H]MK-801 assay conditions. Following a short preincubation wash, there were no effects of APDs on either unenhanced or agonist enhanced [3H]MK-801 binding. Following a prolonged preincubation wash, APDs resulted in a reduction in both unenhanced binding and glycine enhanced binding. The attenuation of glycine stimulation in the APD treated animals was selective as neither NMDA nor spermidine enhanced binding was significantly affected. The present data suggest specific actions at the glycine regulatory site on the NMDA receptor as part of the chronic effects of APDs.

Washable endogenous substances and regional heterogeneity in agonist enhanced [3H]MK-801 binding in rat brain.

NMDA receptor/ion channel function is modulated through a number of distinct sites that regulate channel opening. Published studies report widely varying results in modulatory site agonist effects due to assay conditions and technique. Also, NMDA receptor regulation at these sites by endogenous substances remains poorly characterized. The objectives of the present study in Sprague-Dawley rat forebrain sections were: (i) determine the contribution of various prewash variables on agonist stimulation of the NMDA receptor, (ii) compare regional differences in functional glycine, spermidine and NMDA binding sites under optimized prewash conditions, and (iii) define the influence of endogenous substances at each modulatory site by analyzing changes in binding at different prewash durations. We demonstrate that prewash conditions have a critical influence on [3H]MK-801 binding in rat tissue sections and that this effect was differentially expressed across brain regions. An extended prewash duration caused a regionally specific decrease in unenhanced [3H]MK-801 binding, while a short prewash caused a regionally specific biphasic effect on enhanced [3H]MK-801 binding. After prolonged prewash, binding was restored to previous (unwashed) binding levels with exogenously added glycine, NMDA, or spermidine alone or combinations of agonists. These data suggest that washable endogenous substances contribute to the full functionality of the NMDA receptor and the regional heterogeneity in [3H]MK-801 binding is dependent on the interaction of receptor protein subtypes and the presence of one or more endogenous substances.

Preferential loss of preproenkephalin versus preprotachykinin neurons from the striatum of Huntington's disease patients.

Preferential loss of basal ganglia neurons and terminals occurs in Huntington's disease (HD). Terminals of preproenkephalin medium-size spiny neurons are more vulnerable than terminals of preprotachykinin neurons, but the peptidergic neurons of origin have not yet been shown to die preferentially. We sought to determine, in the striatum, whether preproenkephalin neurons were lost to a greater extent than preprotachykinin neurons and to determine whether there were decreases in specific messenger RNA (mRNA) levels of preproenkephalin, preprotachykinin, and calbindin D28k. We found a grade-related decrease in the number of preprotachykinin- and calbindin D28k-labeled neurons per measuring field in the caudate nucleus of patients with HD. Three measures of the neuronal level of preprotachykinin mRNA were all significantly reduced (6-65% of control values) in HD caudate nucleus. No decline in calbindin D28k mRNA levels per neuron were found in HD striata compared to control striata. We found a greater loss of preproenkephalin neurons per field than preprotachyknin neurons per field in the caudate nucleus of HD brains compared to control brains. Preprotachykinin neurons are lost in HD in a grade-related manner and surviving preprotachykinin neurons are impaired in function. However, preproenkephalin neurons are lost to a greater extent than preprotachykinin neurons, which may explain preferential changes found in projection regions of the striatum. Declines in neuropeptide mRNA may be specific in HD, since calbindin D28k mRNA levels were unchanged. Alterations in the levels of expression of preproenkephalin and preprotachykinin mRNA may be direct or indirect effects of the HD mutation.

DNA end labeling (TUNEL) in Huntington's disease and other neuropathological conditions.

Deoxyribonucleic acid of cells undergoing apoptosis is cleaved by a calcium-dependent endonuclease into oligonucleosomal-sized fragments. These fragments can be labeled using the enzyme terminal deoxynucleotidyl transferase so that the cells can be visualized immunohistochemically. Few investigators have evaluated this method in disease processes of the human central nervous system. The Tdt-mediated dUTP-biotin nick end labeling (TUNEL) technique has been investigated in preliminary studies of a variety of pathologic conditions of the human brain (e.g., gliomas, traumatic brain injury, Parkinson's disease, Parkinson's-Alzheimer's complex, multisystem atrophy, striatonigral degeneration). We focus, however, on Huntington's disease (HD) because of the availability of well-characterized pathological stages for study, and also because of the neurodegenerative diseases studied to date, only Huntington's disease revealed significant and consistent labeling with this method. This implies a possibly unique nature to the mechanism of cell death in Huntington's disease compared to the other neurodegenerative diseases studied. TUNEL+ neurons were found in Grade 1-4 HD neostriatum, while labeled astrocytes were found predominantly in the Grade 1 and 2 cases studied to date. TUNEL+ cells were also found in glioblastoma multiforme and traumatic brain injury. We conclude that while there appear to be several limitations associated with this technique, it may be useful for identifying both apoptosis and necrosis in certain neuropathological conditions.

Reduced expression of preproenkephalin in striatal neurons from Huntington's disease patients.

Differential loss of neurons and terminals occurs in Huntington's disease. Neurons expressing preproenkephalin (PPE) appear to be more vulnerable than neurons expressing preprotachykinin and terminals in the lateral pallidum (containing enkephalin) are more affected than terminals in the medial pallidum (containing substance P). We used in situ hybridization histochemistry and emulsion autoradiography to quantify the number of PPE expressing neurons and the neuronal levels of PPE mRNA in striatum of individuals who died with Huntington's disease and normal controls. We found a grade-related decline in the number of PPE-labeled neurons per field in the striatum of individuals with Huntington's disease compared with controls. Three measures of the neuronal level of PPE mRNA, the mean number of silver grains per PPE neuron, the median number of grains per PPE neuron, and the percentage of PPE neurons with more than 30 grains, were all significantly reduced (41 to 80% of control) in Huntington's disease striatum. The magnitude of the reduction in levels of PPE mRNA per neuron was related to the grade of lesions. These data support the notion that decreased levels of PPE mRNA may account, in part, for the greater loss of enkephalin staining in lateral pallidal terminals compared with substance P staining in medial pallidal terminals. Decreased levels of PPE mRNA may result in clinical symptoms prior to the loss of neurons. The reduction in expression of PPE mRNA suggests that surviving striatal neurons may be affected by the expression of the Huntington's disease gene prior to their imminent cell death.

Substantia nigra lesions in Alzheimer disease and normal aging.

Clinical and pathological overlap between Alzheimer disease (AD) and Parkinson's disease (PD) has been well described; however, the mechanisms of overlap between these two disorders remain unknown. We retrospectively examined clinical and neuropathological features from 66 individuals participating in the Rochester Alzheimer Disease Center to determine the association of AD with substantia nigra (SN) pathology. SN pathology, identified by a loss of pigmented neurons and the presence of gliosis, pigment-laden macrophages, and Lewy bodies, was blindly scored in 48 AD cases and 18 normal elderly controls. We found moderate or severe pathology in the SN in 2 control brains (11%) and 29 AD brains (60%). The numbers of neocortical and hippocampal neurofibrillary tangles (NFTs) and senile plaques (SPs) were not associated with nigral pathology. There was also no significant association of SN pathology with NFTs or SPs in the striatum, the site to which these neurons project. There was no significant association of increasing SN pathology with aging among AD patients, nor with increasing severity and duration of AD. The signs and symptoms of an extrapyramidal movement disorder were, however, associated with increasing SN pathology. We confirm that pathological lesions in the SN are a common feature of AD and an uncommon feature in normal aging. AD is a significant risk factor for SN lesions and PD, but the pathologic severity of AD, as measured by NFTs and SPs, was not associated with SN lesions.