Muscarinic Cholinergic Receptor Subtypes in Hippocampus in Human Cognitive Disorders

Total muscarinic receptor levels, the levels of the subtypes exhibiting high and low affinity for pirenzepine, and the high- and low-affinity agonist states of the receptor were investigated in hippocampal tissue obtained at autopsy from mentally normal individuals and the following pathological groups: Alzheimer's disease, Parkinson's disease, Down's syndrome, alcoholic dementia, Huntington's chorea, and motor-neurone disease. A moderate decrease in the density of both high-affinity pirenzepine and high-affinity agonist subtypes was found in Alzheimer's disease, whereas a trend towards an increase in the overall muscarinic receptor density was apparent in the parkinsonian patients without dementia, mainly due to an increase in the low-affinity agonist state; the differences between the Alzheimer's disease and nondemented parkinsonian cases were highly significant. As previously reported, the levels of both choline acetyltransferase and acetylcholinesterase were markedly reduced in both Alzheimer's disease and Parkinson's disease--with a greater loss of both enzymes in the demented subgroup of parkinsonian patients. Activities of the cholinergic enzymes were also extensively reduced in Down's syndrome, accompanied by a loss of high-affinity pirenzepine binding. There were no significant receptor or enzyme alterations in the other groups studied. These observations suggest that in the human brain, extensive degeneration of cholinergic axons to the hippocampus, as indicated by a loss of cholinergic enzymes, is not necessarily accompanied by extensive muscarinic receptor abnormalities (as might be expected if a major subpopulation were presynaptic). Moreover, the opposite changes in muscarinic binding in Parkinson's and Alzheimer's diseases may be related to the greater severity of dementia in the latter disease.     

Neuronal [3H]Benzodiazepine Binding and Levels of GABA, Glutamate, and Taurine Are Normal in Huntington''s Disease Cerebellum

Alterations in one subunit of the proposed GABA receptor complex, namely, the GABA receptor, have been observed in Huntington's disease cerebellum. We measured binding to a second subunit, the benzodiazepine binding site, in the autopsied cerebellum of 12 patients dying with adult-onset Huntington's disease. Neuronal benzodiazepine ([3H]flunitrazepam) binding density (Bmax) and affinity in cerebellar cortex of the Huntington's disease patients were not significantly different from control values. Similarly, maximal GABA stimulation of benzodiazepine binding was normal in the Huntington's disease cerebellum. In addition, no significant changes were observed in the concentrations of GABA, glutamate, and taurine in cerebellar cortex, nor of GABA in the dentate nucleus.     

Somatostatin Binding Sites in Human and Monkey Brain: Localization and Characterization

A radioiodinated analogue of somatostatin 28, 125I [Leu8,D-Trp22,Tyr25] SS-28, was used to localize and characterize somatostatin binding sites in both human and monkey brain. High-affinity binding sites (approximately 1 nM) were found in cerebral cortex. The highest binding was in cerebral cortex with intermediate binding found in hippocampus, striatum, and amygdala and low binding in hypothalamus and brainstem. There was a rough correlation between somatostatin receptor binding and concentrations of somatostatin-like immunoreactivity (SLI) in human brain. Somatostatin receptors were stable for up to 24 h in an animal model simulating human autopsy conditions and there was no correlation between postmortem interval and receptor binding in human brain. Pharmacologic characterization in human cortex showed that there was a correlation between the inhibition of receptor binding by somatostatin analogues and their known abilities to inhibit growth hormone secretion. These findings demonstrate that a highly specific membrane-associated receptor for somatostatin is present in both monkey and human brain. Examination of somatostatin receptor binding in Alzheimer's disease and Huntington's disease may improve understanding of the role of somatostatin in both these illnesses.     

Decreased Density of Presynaptic α2-Adrenoceptors in Postmortem Brains of Patients with Alzheimer''s Disease

The full agonist [3H]bromoxidine (UK 14304) was used to quantitate alpha 2-adrenoceptors in postmortem brains of patients with Alzheimer's disease. The effects of aging and human serum Cohn fraction IV on [3H]bromoxidine binding were also assessed. In patients with Alzheimer's disease, the binding capacity (Bmax) of [3H]bromoxidine was lower in the frontal cortex (37%), hypothalamus (33%), and cerebellum (52%) than in matched controls. In the hippocampus, amygdala, and head of caudate, the binding capacities (Bmax) were unchanged. Quantitative autoradiographic analyses with [3H]bromoxidine confirmed the existence of a marked reduction (55-60%) in alpha 2A-adrenoceptor density in the frontal cortex (layers I and III). In patients with dementia who did not meet neuropathological criteria for Alzheimer's disease, the density of alpha 2-adrenoceptors was unchanged. In control subjects, the density of alpha 2A-adrenoceptors in the frontal cortex showed a significant negative correlation with age at death. The inhibitory effect of human serum Cohn fraction IV on [3H]bromoxidine was very similar in control subjects and patients with Alzheimer's disease. The observed decrease in the density of brain alpha 2-adrenoceptors in Alzheimer's disease may represent direct biochemical evidence of a presynaptic location of this receptor on noradrenergic nerve terminals in the human CNS.     

No evidence for increased oxidative damage to lipids, proteins, or DNA in Huntington's disease

It has been proposed that mitochondrial dysfunction and excitotoxic mechanisms lead to oxidative damage in the brain of Huntington;s disease patients. We sought evidence that increased oxidative damage occurs by examining postmortem brain material from patients who had died with clinically and pathologically diagnosed Huntington's disease. Oxidative damage was measured using methods that have already demonstrated the presence of increased oxidative damage in Parkinson's disease, Alzheimer's disease, and senile dementia of the Lewy body type. No alterations in the levels of lipid peroxidation (as measured by lipid peroxides and thiobarbituric acid-malondialdehyde adducts) were found in the caudate nucleus, putamen, or frontal cortex of patients with Huntington's disease compared with normal controls. Similarly, there were no elevations in the levels of 8-hydroxyguanine or of a wide range of other markers of oxidative DNA damage. Levels of protein carbonyls in these tissues were also unaltered. Our data suggest that oxidative stress is not a major component of the degenerative processes occurring in Huntington's disease, or at least not to the extent that occurs in other neurodegenerative disorders.     

Differential Regulation of Molecular Subtypes of Muscarinic Receptors in Alzheimer's Disease

Abstract: Molecular subtypes of muscarinic receptors (m1–m5) are novel targets for cholinergic replacement therapies in Alzheimer's disease. However, the status of these receptors in human brain and Alzheimer's disease is incompletely understood. The m1–m5 receptors in brains from control subjects and Alzheimer's disease patients were examined using a panel of specific antisera and radioligand binding. Quantitative immunoprecipitation demonstrated a predominance of the m1, m2, and m4 receptor subtypes in cortical and subcortical regions in control subjects. In Alzheimer's disease, normal levels of m1 receptors measured by radioligand binding contrasted with decreased m1 receptor immunoreactivity, suggesting that the m1 receptor is altered in Alzheimer's disease. The m2 immunoreactivity was decreased, consistent with the loss of m2 binding sites and the location of this receptor subtype on presynaptic cholinergic terminals. The m4 receptor was up-regulated significantly and may offer a target for new memory-enhancing drugs. Differential alterations of molecular subtypes of muscarinic receptors may contribute to the cholinergic component of Alzheimer's disease dementia.     

Increase in brain 125I-cholecystokinin (CCK) receptor binding following chronic haloperidol treatment, intracisternal 6-hydroxydopamine or ventral tegmental lesions

Specific 125I-CCK receptor binding was significantly increased in brain tissue taken from guinea pig or mouse following chronic (2-3 week) daily administration of haloperidol (2-3 mg/kg/day). Scatchard analysis indicated the increase in CCK binding was due to an increased receptor number (B max) with no change in affinity (Kd). In guinea pigs, the increased CCK binding was observed in the mesolimbic regions and frontal cortex, but not in striatum, hippocampus nor posterior cortex. In mice, however, the increases occurred in both pooled cerebral cortical-hippocampal tissue, and in the remainder of the brain. Enhanced CCK receptor binding was also observed in membranes prepared from whole brain of mice one month following intracisternal injection of 6-hydroxydopamine. Additionally, an increase in CCK binding was observed in mesolimbic regions and frontal cortex, but not striatum or hippocampus, of guinea pigs 3 weeks after an unilateral radiofrequency lesions of the ipsilateral ventral tegmentum. The present studies demonstrate that three different procedures which reduce dopaminergic function in the brain enhance CCK receptor binding. The data provide further support for a functional interrelationship between dopaminergic systems and CCK in some brain regions and raise the possibility that CCK may play a role in the antipsychotic action of neuroleptics.     

Reduced Glycine Stimulation of [3H]MK-801 Binding in Alzheimer''s Disease

The novel N-methyl-D-aspartate receptor channel ligand (+)-[3H]5-methyl-10,11-dihydro-5H-dibenzo[a,d]-cyclohepten-5, 10-imine maleate ([3H]MK-801) has been utilized to label this receptor in human brain tissue. Characteristics of [3H]MK-801 binding to well-washed membranes from 17 control subjects and 16 patients with Alzheimer's disease were determined in frontal, parietal, and temporal cerebral cortex and cerebellar cortex. In control tissue the pharmacological specificity of the binding of this substance is entirely consistent with the profile previously reported for rat brain. Binding could be stimulated by the addition of glutamic acid to the incubation medium; addition of glycine produced further enhancement which was not prevented by strychnine. The specificity of the effects of these and other amino acids on the binding was the same as in the rat. In Alzheimer's disease significantly less binding was observed in the frontal cortex under glutamate- and glycine-stimulated conditions. This appears to be associated with a reduced affinity of the site whereas the pharmacological specificity of the site remained unchanged. The effect did not appear to be due to differences in mode of death between Alzheimer's disease and control subjects and is unlikely to be related to factors for which the groups were matched. In contrast, binding was not altered in the absence of added amino acids and presence of glutamate alone. These results imply that in the cerebral cortex the agonist site and a site in the cation channel of the receptor are not selectively altered, but that their coupling to a strychnine-insensitive glycine recognition site is impaired.     

Insulin-Like Growth Factor I Receptor Binding in Brains of Alzheimer''s and Alcoholic Patients

Patients with chronic alcoholism and/or Alzheimer's disease show degenerative changes in the cerebral cortex and hippocampus. To investigate possible changes in insulin-like growth factor I receptor binding sites in brain tissue of patients with these pathological conditions, the number of 125I-insulin-like growth factor I binding sites was determined in tissues obtained from control patients and those with Alzheimer's and/or with a history of alcoholism. The four experimental groups examined consisted of patients from similar age groups. Postmortem histology and a clinical history were used for the diagnosis of Alzheimer's disease and alcoholism, respectively. Careful clinical records were kept concerning other variables such as immediate cause of death and medications administered before death. Specific binding of 125I-insulin-like growth factor I to homogenates prepared from cerebral cortex of Alzheimer's, alcoholic, alcoholic Alzheimer's, and age-matched control patients was similar, although Alzheimer's patients tended to have slightly higher binding values. No significant differences in insulin-like growth factor I binding in cerebral cortex were found with regard to age of patients, the interval between death and autopsy, and CNS-active medications. No statistical differences in 125I-insulin-like growth factor I binding were noted in hippocampal tissue from the four patient groups. Thus, human insulin-like growth factor I binding sites in cerebral cortex and hippocampus appear unaffected by several variables.     

Acetylcholinesterase in Huntington''s and Alzheimer''s Diseases: Simultaneous Enzyme Assay and Immunoassay of Multiple Brain Regions

A newly developed enzyme-linked immunosorbent assay for acetylcholinesterase (AChE) protein was combined with conventional measures of enzyme activity in a study of 15 brain regions from six control cases (non-neurological deaths), six cases of Alzheimer's disease, and six cases of Huntington's disease. In the control brains, the mean AChE activity varied 100-fold from region to region (cortex lowest, striatum highest). The variation in enzyme activity was exactly paralleled by a variation in protein immunoreactivity. Overall, the homospecific activity of AChE averaged 0.26 +/- 0.007 mU/pg, close to the value for electrophoretically homogeneous enzyme isolated from red blood cells. Similar homospecific activities were observed in samples from Huntington's and Alzheimer's brains. Evidently, AChE that is immunoreactive but enzymatically inactive does not accumulate in any of the three conditions examined. Huntington's brain samples showed normal total contents of AChE, but Alzheimer's brains showed significant decreases of both enzyme activity and immunoreactivity in all seven cortical regions and in two out of the eight subcortical structures examined, hippocampus and nucleus accumbens.     

Increase in Kynurenic Acid in Huntington''s Disease Motor Cortex

Huntington's disease is a neurological disorder characterised by a progressive chorea and dementia. Recent evidence has suggested that dysfunction involving endogenous excitatory amino acids may be important in the pathogenesis of this disease. Following the recent demonstration that kynurenic acid is present in the brain, we examined the levels in various areas of brain from patients who died with Huntington's disease and from age/sex-matched controls. Blocks (100-500 mg) of cortex (Brodmann's areas 4 and 10) and caudate nucleus and globus pallidus (lateral and medial parts) were obtained from the Cambridge Brain Bank. The tissue was then processed for the extraction and analysis of kynurenic acid. Whereas no differences in the content of kynurenic acid were observed in the caudate nucleus, lateral or medial globus pallidus, or prefrontal cortex (area 10) between controls' brains and those from patients who died with Huntington's disease, there was a 94% (p less than 0.01; n = 5) increase in the kynurenic acid content in the motor cortex (area 4) from Huntington's disease brains, relative to those of controls. Some time ago we suggested that a subtle change in the relative concentrations of quinolinic and kynurenic acids might be important in the pathogenesis of neurodegeneration. It is possible that the observation of raised kynurenic acid levels supports this supposition. Further work is now in progress to determine whether the change in kynurenic acid is a primary effect or a compensatory response to an increase in excitatory activity.     

Glycine Receptors in the Human Substantia Nigra as Defined by [3H]Strychnine Binding

Abstract: Specific [³H]strychnine binding was used to identify the glycine receptor macromolecular complex in human spinal cord, substantia nigra, inferior olivary nucleus, and cerebral cortex. In material from control patients a high-affinity K d (3–8 n m ) was observed in the spinal cord and the substantia nigra, both the pars compacta and the pars reticulata. This is very similar to the values observed in the rat and bovine spinal cord (8 and 3 n m , respectively) and rat substantia nigra (12 n m ). In the human brain the distribution of [³H]strychnine binding (at 10 n m ) was: spinal cord – substantia nigra, pars compacta > substantia nigra, pars reticulata = inferior olivary nucleus > cerebral cortex. The binding capacity ( B _max) of the rat brain (substantia nigra or spinal cord) was approximately 10-fold that of the human brain. [ ³ H]Strychnine binding was significantly decreased in the substantia nigra from Parkinson's disease patients, both in the pars compacta (67% of control) and the pars reticulata (50% of control), but not in the inferior olivary nucleus. The results were reproduced in a preliminary experiment in rats with unilateral 6-hydroxydopamine lesions of the medial forebrain bundle. In the substantia nigra from patients who died with Huntington's disease, [³H]strychnine binding tended to be high (150% of control, NS) in both the pars compacta and the reticulata. [³H]Strychnine binding was unaltered in the substantia nigra of patients with senile dementia. Together with previous neurophysiological and neuropharmacological findings, those results support the hypothesis of glycine receptors occurring on dopamine cell bodies and/or dendrites in the substantia nigra.     

Characterization of central cholecystokinin receptors using a radioiodinated octapeptide probe

We have developed a binding assay for 125I-Bolton-Hunter-labeled cholecystokinin octapeptide (125I-(BH)CCK8) using mouse cerebral cortex membrane preparations. This ligand interacts with cortical membrane preparations in a saturable, high-affinity manner, satisfying the requirements for specific cholecystokinin receptor labeling. Salt is required for maximal binding and BSA is specifically inhibitory with cerebral cortical but not with pancreatic sites. Cholecystokinin peptides as small as CCK30-33 displace binding at low nanomolar concentrations. Dissociation of 125I-(BH)CCK8 is biphasic in both mouse and guinea pig cortex. Pretreatment of membranes at 37 degrees C results in a marked loss of recognition sites, suggesting that the sites may be rapidly metabolized in vivo. After 37 degrees C pretreatment, the loss of CCK recognition sites corresponds to a selective loss of the slow component of dissociation curves. This selective elimination of one dissociation population, as well as the biphasic dissociation kinetics, suggests that at least two distinct CCK receptor subtypes exist in the brain.     

Is the neuronal basis of Alzheimer's disease cholinergic or glutamatergic?

The hypothesis that the symptomatology of Alzheimer's disease is attributable to cholinergic dysfunction is supported by postmortem studies that have demonstrated reduced choline acetyltransferase (ChAT) activity across all areas of cerebral cortex and diminished numbers of perikarya in the basal forebrain nucleus basalis of Meynert. Biopsy studies of ChAT activity, choline uptake, and acetylcholine synthesis also suggest that cholinergic denervation occurs relatively early in the course of the disease, and in confirmation of postmortem data, correlates with the severity of cognitive impairment. An alternative hypothesis to explain the dementia of Alzheimer's disease is the glutamatergic hypothesis. This is based largely on postmortem evidence indicating reduced binding and uptake of D[3H]aspartate, as well as loss of a number of other putative markers, such as phosphate-activated glutaminase activity, glutamate concentration, and the number of pyramidal cell perikarya, with this latter change correlating with the severity of dementia. Short-comings of each hypothesis are discussed and the merits of single neuron hypotheses to explain the dementia of Alzheimer's disease are considered.     

Lipid Composition in Different Regions of the Brain in Alzheimer''s Disease/Senile Dementia of Alzheimer''s Type

The lipid compositions of 10 different brain regions from patients affected by Alzheimer's disease/senile dementia of Alzheimer's type were analyzed. The total phospholipid amount decreased somewhat in nucleus caudatus and in white matter. The cortical areas that are morphologically affected by Alzheimer's disease, i.e., frontal and temporal cortex and the hippocampus, showed elevated contents of lipid solvent-extractable phosphatidylinositol. Sphingomyelin content was decreased in regions rich in myelin. There was a 20-50% decrease in dolichol amount in all investigated parts of the brain, but no change was seen in the polyisoprenoid pattern. Levels of alpha-unsaturated polyprenes were decreased in Alzheimer brains. Dolichyl-phosphate content increased in most regions, up to 100%. In both control and Alzheimer tissue almost all of the dolichyl-phosphate was covalently bound, apparently through glycosylation. Cholesterol amounts were highly variable but mostly unchanged, whereas ubiquinone concentrations increased by 30-100% in most regions in brains affected by Alzheimer's disease. These results demonstrate that both phospholipids and neutral lipids are modified in brains affected by Alzheimer's disease/senile dementia of Alzheimer's type.     

Dementia of the Alzheimer's Type: Changes in Hippocampal L-[3H]Glutamate Binding

Abstract: Glutamate or a related excitatory amino acid is thought to be the major excitatory neurotransmitter of hippocampal afferents, intrinsic neurons, and efferents. We have used an autoradiographic technique to investigate the status of excitatory amino acid receptors in the hippocampal formation of patients dying with dementia of the Alzheimer type (DAT). We examined l-[³H]glutamate binding to sections from the hippocampal formation of six patients dying of DAT and six patients without DAT and found marked reductions in total [³H]glutamate binding in all regions of hippocampus and adjacent parahippocampal cortex in DAT brains as compared to controls. When subtypes of excitatory amino acid receptors were assayed, it was found that binding to the N -methyl-d-aspartate (NMDA)-sensitive receptor was reduced by 75–87%, with the greatest loss found in stratum moleculare and stratum pyramidale of CA1. Binding to quisqualate (QA)-sensitive receptors was reduced by 45–69%. There were smaller reductions (21–46%) in GABA_A receptors in DAT cases. Muscarinic cholinergic receptors assayed in adjacent sections of hippocampal formation were unchanged in DAT. Benzodiazepine receptors were reduced significantly only in parahippocampal cortex by 44%. These results suggest that glutamatergic neurotransmission within the hippocampal formation is likely to be severely impaired in Alzheimer's disease. Such impairment may account for some of the cognitive decline and memory deficits that characterize DAT.     

Characterization of l-[3H]Nicotine Binding in Human Cerebral Cortex: Comparison Between Alzheimer''s Disease and the Normal

Putative nicotine receptors in the human cerebral cortex were characterized with L-[3H]nicotine, L-[3H]Nicotine binding was enhanced by the addition of Ca2+ and abolished in the presence of Na3EDTA. Association and dissociation of the ligand were rapid at 25 degrees C with t1/2 values of 2 and 3 min, respectively. Saturation binding analysis revealed an apparent single class of sites with a dissociation constant of 5.6 nM and a Hill coefficient of 1.05. There was no effect of postmortem interval on the density of binding sites assayed up to 24 h in rat frontoparietal cortex. Nicotine binding in human cortical samples was also unaltered by increasing sampling delay. In human cortical membranes, binding site density decreased with normal aging. Receptor affinity and concentration in samples of frontal cortex (Brodmann area 10) from patients with Alzheimer's disease were comparable to age-matched control values. Samples of infratemporal cortex (Brodmann area 38) from patients with Alzheimer's disease had a 50% reduction in the number of L-[3H]nicotine sites. Choline acetyltransferase activity was significantly decreased in both cortical areas. Enzyme activities in the temporal pole were reduced to 20% of control values. These data indicate that postsynaptic nicotine receptors are spared in the frontal cortex in Alzheimer's disease. In the infratemporal cortex, significant numbers of receptors remain despite the severe reduction in choline acetyltransferase activity. Replacement therapy directed at these sites may be warranted in Alzheimer's disease.     

The somatostatin receptor on isolated pancreatic acinar cell plasma membranes. Identification of subunit structure and direct regulation by cholecystokinin

Somatostatin binding to its receptors on rat pancreatic acinar membranes was characterized with [125I-Tyr1]somatostatin. Binding at 24 degrees C was rapid reaching a maximum after 60 min and was reversible upon the addition of 1 microM unlabeled ligand. Scatchard analysis revealed a single class of binding sites, with a Kd of 0.32 +/- 0.03 nM and a binding capacity of 600 +/- 54 fmol/mg of protein. Specificity for the somatostatin was demonstrated with the inhibition of labeled hormone binding by somatostatin analogs in proportion to their biological activities. When [125I-Tyr1]somatostatin was cross-linked to its receptors with the photoreactive cross-linker n-hydroxysuccinimidyl-4-azidobenzoate, the hormone was associated with Mr = 90,000 protein. Similar mobilities of the radioactive band were observed in the presence and absence of dithiothreitol. In contrast to other unrelated peptides, cholecystokinin (CCK) and its analogs directly reduced [125I-Tyr1] somatostatin binding to isolated membranes. The effect of CCK was one-half-maximal at 3 nM and maximal at 100 nM. In the presence of 3 nM CCK8, the binding capacity for somatostatin was decreased to 237 +/- 39 fmol/mg of protein without a significant change in affinity. Dibutyryl cyclic GMP, a CCK receptor antagonist, blocked this action of CCK8 indicating that the CCK receptor mediated the decrease in [125-Tyr1]somatostatin binding. In contrast cerebral cortex membranes, which also possess a somatostatin receptor, were not regulated by CCK. These results indicate, therefore, that 1) purified pancreatic acinar plasma membranes contain specific receptors for somatostatin, 2) the receptor has an apparent Mr of about 90,000, and 3) the binding of somatostatin to its receptor on pancreatic plasma membranes is regulated by CCK analogs acting via the CCK receptor.     

Glycosaminoglycans in Cortical Autopsy Samples from Alzheimer Brain

Each of the known classes of mammalian glycosaminoglycans, with the exception of keratan sulphate, was found in cerebral cortex samples from patients with Alzheimer-type dementia and age-matched controls. These molecules were quantitated, after electrophoresis and staining with Alcian Blue dye, by scanning densitometry. No significant differences were found between the mean levels of each of the above glycosaminoglycans in frontal cortex from patients with dementia compared with controls. An increase (26%; p less than 0.05) in the mean level of hyaluronate, but not of other glycosaminoglycans, was found in temporal cortex samples. On the other hand, the uronic acid content of hyaluronate degradation products following Streptomyces hyaluronidase treatment of brain glycosaminoglycans did not reveal any statistically significant changes in Alzheimer's disease. HPLC of disaccharide products from Arthrobacter chondroitinase AC digests did not reveal any significant changes in sulphate substitution of chondroitin sulphate in Alzheimer brain.     

Enzyme Activities in Relation to pH and Lactate in Postmortem Brain in Alzheimer-Type and Other Dementias

Phosphate-activated glutaminase, glutamic acid decarboxylase, pyruvate dehydrogenase, succinic dehydrogenase, pH, and lactate were measured in frontal cortex and caudate nucleus of postmortem brains from cases of Alzheimer-type dementia (ATD), Down's syndrome, Huntington's disease, and one case of Pick's disease, as well as from sudden death and agonal controls. Lactate levels were higher and pH, phosphate-activated glutaminase, and glutamic acid decarboxylase levels were lower in the agonal controls than in the sudden death controls. Phosphate-activated glutaminase and glutamic acid decarboxylase were correlated with tissue pH and lactate, and also were reduced by in vitro acidification, suggesting that the low activities of these enzymes in agonal controls were related to decreased pH consequent upon lactate accumulation. Compared with control tissues at the same pH, phosphate-activated glutaminase and glutamic acid decarboxylase were unaltered in ATD and Down's frontal cortex and reduced in Huntington's caudate nucleus, and glutamic acid decarboxylase was reduced in Huntington's frontal cortex. These data suggest that GABAergic neurons are not affected in ATD and confirm the GABAergic defect in Huntington's disease. Pyruvate dehydrogenase and succinic dehydrogenase activities were the same in agonal controls and sudden death controls and were unaffected by acid pH and lactate in vitro, and pyruvate dehydrogenase was not correlated with pH or lactate. Reduced pyruvate dehydrogenase in frontal cortex of individual ATD, Down's, and Pick's cases, and in the caudate nucleus of Huntington's and Down's cases, was accompanied by gliosis/neuron loss. We conclude that decreased pyruvate dehydrogenase reflects neuronal loss.