Alterations in dopaminergic receptors in Huntington's disease.

To detect variations in dopaminergic receptors and cholinergic activity in regions of postmortem Huntington's diseased brains, ³H-spiroperidol binding assays and choline acetyltransferase (ChAc) activities were carried out. A significant reduction in ³H-spiroperidol binding in the caudate nucleus, putamen and frontal cortex of choreic brains was detected which appeared to be due to a decrease in the total number of binding sites rather than to a decrease in affinity of ³H-spiroperidol for the dopaminergic receptor. In choreic brains, there were also significant reductions in ChAc activity in the caudate nucleus and putamen. The decreases of both ³H-spiroperidol binding and ChAc activity in the neostriatum suggest that the dopaminergic receptors are localized postsynaptically on cholinergic interneurons. Dopaminergic receptor alterations in the basal ganglia may be one of the causes of the abnormal motor movements found in HD while alterations of these receptors in the frontal cortex may be associated with the neuronal degeneration found in that area of choreic brains.     

Neurotransmitter receptor alterations in Parkinson's disease.

Neurotransmitter receptor binding for GABA, serotonin, cholinergic muscarinic and dopamine receptors and choline acetyltransferase (ChAc) activity were measured in the frontal cortex, caudate nucleus, putamen and globus pallidus from postmortem brains of 10 Parkinsonian patients and 10 controls. No changes in any of these systems were observed in the frontal cortex. In the caudaye nucleus, only the apparent dopamine receptor binding was altered with a significant 30% decrease in the Parkinsonian brain. Both cholinergic muscarinic and serotonin receptor binding were significantly altered in the putamen, the former increasing and the latter decreasing with respect to controls. In addition, ChAc activity was decreased in the putamen. In the globus pallidus, only ChAc activity was significantly changed, decreasing about 60%, with no change in neurotransmitter receptor binding. The results suggest that a progressive loss of dopaminergic receptors in the caudate nucleus may contribute to the decreased response of Parkinsonian patients to L-dopa and dopamine agonist therapy.     

Huntington's disease and its animal model: alterations in kainic acid binding.

The density of ³H-kainic acid (KA) binding was determined in several regions of Huntington's Diseased (HD) and control human brains. ³H-Kainic acid binding was significantly reduced by 55% in the caudate nucleus and by 53% in the putamen of HD brains. In addition, ³H-KA binding was determined in rat striatum at various intervals following lesion with KA, a procedure which produces an animal model of HD. After KA lesion, ³H-KA binding in the rat striatum underwent a slow reduction, reaching 25% of control after 6 weeks. Several properties of ³H-KA binding to rat brain membranes were also investigated, including inhibition by ions, regional distribution and displacement by various compounds. The findings confirm the validity of the KA-lesioned model for HD and suggest a post-synaptic location for kainic acid receptors in the striatum.     

CHOLINE ACETYLTRANSFERASE ACTIVITY OF HUMAN BRAIN TISSUE DURING DEVELOPMENT AND AT MATURITY

Abstract— (1) On analysis of human brain tissue to determine its choline acetyltransferase (ChAc) content the recovery of enzyme from many regions is very poor when the tissue is acetone-dried and then extracted in the standard manner; for this reason the method is unsuitable when quantitative recoveries are required; it is preferable to prepare sucrose homogenates and activate these with ether before incubation.
(2) From measurements made on homogenates of one adult brain the highest concentration of ChAc was found in the putamen and the lowest in the corpus callosum. The caudate nucleus also had a high activity. As in other mammals, the concentration of enzyme in the cerebellum was found to be low. Analogous results were obtained on a nine-year-old brain but the level of ChAc activity was generally higher than in the older brain.
(3) During foetal development up to thirty-two weeks, ChAc is higher in the cerebellum than in the caudate, the thalamus, corpora quadrigemina, medulla and spinal cord. In all regions the concentration and total amount of enzyme rise fairly steadily up to this time; between 24 and 32 weeks, however, its concentration in the cerebellum and corpora quadrigemina falls slightly although the total increases considerably.
(4) Comparison of the results with the data of other authors indicates general agreement between the distribution of the enzyme in the human brain and its distribution in other mammals, especially the rhesus monkey. The corpus callosum may be an exception since in man it contains little ChAc while in lower mammals it seems to have relatively high concentrations of both ACh and ChAc.
(5) In comparing the values for ChAc reported here with the values for AChE reported by others, three tissues, the globus pallidus, substantia nigra and cerebellum are found to be exceptional in that relative to their concentration in the caudate the activity of ChAc is only about one-tenth that of AChE.     

Changes of Biogenic Amines and Their Metabolites in Postmortem Brains from Patients with Alzheimer-Type Dementia

Noradrenaline (NA), 3,4-dihydroxyphenylethylamine (dopamine, DA), 5-hydroxytryptamine (serotonin, 5-HT), homovanillic acid (HVA), and 5-hydroxyindoleacetic acid (5-HIAA) were measured in 22 regions of postmortem brains from four histologically verified cases with Alzheimer-type dementia (ATD) and nine histologically normal controls. Compared with the controls, concentrations of 5-HT and 5-HIAA in the ATD brains were significantly reduced in nine regions (superior frontal gyrus, insula, cingulate gyrus, amygdala, putamen, medial and lateral segments of globus pallidus, substantia nigra, lateral nucleus of thalamus) and in eight regions (amygdala, substantia innominata, caudate, putamen, medial and lateral segments of globus pallidus, medial and lateral nuclei of thalamus), respectively. NA concentrations of the ATD brains were significantly reduced in six regions (cingulate gyrus, substantia innominata, putamen, hypothalamus, medial nucleus of thalamus, raphe area). In contrast, significant reductions of DA and HVA concentrations in the ATD brains were found only in putamen and amygdala, respectively. The 5-HIAA/5-HT ratio in the ATD brains decreased significantly in locus coeruleus, while the HVA/DA ratio increased significantly in putamen and medial segment of globus pallidus. These findings suggest that the serotonergic and noradrenergic systems are affected, while the dopaminergic system is relatively unaffected in ATD brains.     

Decreased Proline Endopeptidase Activity in the Basal Ganglia in Huntington''s Disease

Soluble proline endopeptidase (EC 3.4.21.26) activity was measured by a fluorometric assay in eight human brain areas (caudate nucleus, lateral globus pallidus, medial globus pallidus, substantia nigra-zona compacta, substantia nigra-zona reticulata, frontal cortex-Brodmann area 10, temporal cortex-Brodmann area 38, and hippocampus), in 10 control and 10 Huntington's disease brains. An abnormally low activity (22% of control activity) was found in the caudate nucleus of Huntington's disease brains; significantly decreased activity was also detected in the lateral globus pallidus and medial globus pallidus (37% and 40% of control, respectively).     

GABA content and glutamic acid decarboxylase activity in brain of Huntington's chorea patients and control subjects.

—GABA contents are significantly decreased in the caudate nucleus, putamen-globus pallidus, substantia nigra, and occipital cortex in autopsied brain from Huntington's chorea patients, as compared to values in the same regions from control subjects who have died without neurological disease. Homocarnosine levels are lower in choreic than in control brain, but only in the putamen-globus pallidus and the cerebellar cortex are the differences significant. Activity of the enzyme which synthesizes GABA, glutamic acid decarboxylase, is reduced in the brains of some choreic patients, but may be equally low in brain of control subjects, even though the latter exhibit normal brain GABA content. Low glutamic acid decarboxylase activity in autopsied human brain is not uniquely characteristic of Huntington's chorea. No evidence was found in this study for an inhibitor of glutamic acid decarboxylase in choreic brain, nor for the presence of an isoenzyme with decreased affinity for glutamate. GABA aminotransferase, the enzyme which degrades GABA, was equally active in control and choreic brain; therefore, increased activity of this enzyme cannot account for the low brain GABA levels in Huntington's chorea.     

Differential Changes in the Content of Amino Acid Neurotransmitters in Discrete Regions of the Rat Brain Prior to the Onset and During the Course of Homocysteine-Induced Seizures

Changes in amino acid concentrations were investigated in selected regions of rat brain prior to the onset and during the course of epileptiform seizures induced by L-homocysteine. The concentration of gamma-aminobutyric acid (GABA) decreased preictally in substantia nigra (-18%), caudate putamen (-26%), and inferior colliculus (-46%). After seizure onset, the GABA content was further reduced in substantia nigra (-31%) and additionally in hippocampus (-18%). Preictal taurine levels were elevated in globus pallidus (+26%) and caudate putamen (+13%) but returned to normal after seizure onset. However, in hippocampus, taurine decreased both preictally (-22%) and after seizure onset (-56%). Glycine was reduced preictally only in globus pallidus (-13%). After seizure onset the direction of its concentration change varied in the brain regions studied. Glutamate levels decreased preictally in hippocampus (-10%) and hypothalamus (-46%) but increased in globus pallidus (+14%). Normal levels were detectable after seizure onset in hypothalamus and globus pallidus but a further reduction in hippocampus (-59%) and significant reductions in substantia nigra (-15%) and caudate putamen (-17%) were detected. Aspartate was elevated in hippocampus, both preictally (+49%) and after seizure onset (+21%) while at the same phases in globus pallidus a consistent reduction (-30%) was observed. The glutamine content increased preictally in globus pallidus (+41%) and hypothalamus (+36%), and in all brain areas during the ictal phase of seizure, the hippocampus exhibiting a dramatic increase (approximately 300%). The contents of serine and alanine were altered in most regions studied only after seizure onset, with the exception of the hippocampus, where a decrease (-41%) of serine was observed preictally.     

Alterations in 3H-GABA binding in Huntington's chorea.

³H-GABA binding was measured in the caudate nucleus, putamen, parietal cortex and cerebellar cortex of control patients and patients with Huntington's chorea. The density of ³H-GABA binding in the parietal cortex was similar in both patient groups. In the striatal regions there was a significant large (70–80 percent) decrease in the density of GABA binding which is consistent with the severe atrophy and cell loss observed in these regions. In contrast the membranes prepared from the cerebellar cortex exhibited both an increased binding capacity (at 25 nM ³H-GABA) and an apparently increased affinity (decreased KD) for ³H-GABA. The decreased capacity of the striatum to bind ³H-GABA may partially explain the reported lack of clinical efficacy of GABAmimetic compounds in Huntington's chorea.     

GABA receptor binding in the parkinsonian brain

The binding of GABA to postsynaptic receptors was studied in the cerebral and cerebellar cortex, caudate nucleus, putamen, pallidum and substantia nigra from autopsy brains of 12 parkinsonian patients and 11 controls. GABA receptor binding in the substantia nigra was significantly decreased in the parkinsonian brain. In the other brain regions, however, GABA binding was unchanged. There was no correlation between GABA binding and sex, age, duration or severity of the disease. The results suggest the involvement of nigral GABA receptor in Parkinson's disease.     

The regional distribution of dopamine and serotonin uptake and transmitter concentrations in the human brain

The regional distribution of the dopamine and serotonin uptake sites in human brain have been assessed and compared with the distribution of the transmitters and their metabolites measured in the same brains and also with a limited regional distribution of the uptake sites in rat and sheep brain. The affinity of the uptake sites for both transmitters was determined and found to be c. 0.2 μ M in all 3 species. Most dopamine uptake in all species was in caudate and putamen samples. Many regions of the human brain showed no dopamine uptake and little dopamine uptake was seen in sheep cortex or nigral preparations. Dopamine and metabolite concentrations were highest in the caudate, putamen and substantia nigra. Most serotonin uptake was seen in the hypothalamus in all 3 species; less was observed in the striatal regions; the cortical and nigral preparations of sheep brain showed little serotonin uptake though cortical preparations of rat brain had high levels of uptake. In the human brain, other regions did not show serotonin uptake. Highest concentrations of serotonin were found in the substantia nigra and medulla, intermediate concentrations in the putamen, globus pallidus, hypothalamus, olfactory tubercle and thalamus; very low concentrations of serotonin were found in other regions. The use of the human uptake site for pharmacological studies and as a marker for monoaminergic afferents in human health and disease is discussed.     

Manganese accumulates in iron-deficient rat brain regions in a heterogeneous fashion and is associated with neurochemical alterations

Previous studies have shown that iron deficiency (ID) increases brain manganese (Mn), but specific regional changes have not been addressed. Weanling rats were fed one of three semipurified diets: control (CN), iron deficient (ID), or iron deficient/manganese fortified (IDMn+). Seven brain regions were analyzed for Mn concentration and amino acid (glutamate, glutamine, taurine, γ-aminobutyric acid) concentrations. Both ID and IDMn+ diets caused significant (p<0.05) increases in Mn concentration across brain regions compared to CN. The hippocampus was the only brain region in which the IDMn+ group accumulated significantly more Mn than both the CN and ID groups. ID significantly decreased GABA concentration in hippocampus, caudate putamen, and globus pallidus compared to CN rats. Taurine was significantly increased in the substantia nigra of the IDMn+ group compared to both ID and CN. ID also altered glutamate and glutamine concentrations in cortex, caudate putamen, and thalamus compared to CN. In the substantia nigra, Mn concentration positively correlated with increased taurine concentration, whereas in caudate putamen, Mn concentration negatively correlated with decreased GABA. These data show that ID is a significant risk factor for central nervous system Mn accumulation and that some of the neurochemical alterations associated with ID are specifically attributable to Mn accumulation.     

Globus pallidus: a target brain region for divalent metal accumulation associated with dietary iron deficiency

Recently, iron deficiency has been connected with a heterogeneous accumulation of manganese in the rat brain. The striatum is particularly vulnerable, for there is a significant negative correlation between accumulated manganese and gamma-aminobutyric acid levels. The effect of dietary iron deficiency on the distribution of zinc and copper, two other divalent metals with essential neurobiological roles, is relatively unexplored. Thus, the primary goal of this study was to examine the effect of manipulating dietary iron and manganese levels on the concentrations of copper, iron, manganese and zinc in five rat brain regions as determined with inductively coupled plasma mass spectrometry analysis. Because divalent metal transporter has been implicated as a transporter of brain iron, manganese, and to a lesser extent zinc and copper, another goal of the study was to measure brain regional changes in transporter levels using Western blot analysis. As expected, there was a significant effect of iron deficiency (P < 0.05) on decreasing iron concentrations in the cerebellum and caudate putamen; and increasing manganese concentrations in caudate putamen, globus pallidus and substantia nigra. Furthermore, there was a significant effect of iron deficiency (P < 0.05) on increasing zinc concentration and a statistical trend (P = 0.08) toward iron deficiency-induced copper accumulation in the globus pallidus. Transporter protein in all five regions increased due to iron deficiency compared to control levels (P < 0.05); however, the globus pallidus and substantia nigra revealed the greatest increase. Therefore, the globus pallidus appears to be a target for divalent metal accumulation that is associated with dietary iron deficiency, potentially caused by increased transporter protein levels.     

Monoamine Oxidase Activity and Monoamine Metabolism in Brains of Parkinsonian Patients Treated with l-Deprenyl

Monoamine oxidase (MAO) type A and type B were measured using kynuramine, 3,4-dihydroxyphenylethylamine (dopamine, DA), and 5-hydroxytryptamine (5-HT, serotonin) in 20 brain areas. The highest activities were found in the striatum (caudate nucleus, putamen, globus pallidus, and substantia nigra), hypothalamus, and c-mammilare. The ratio of DA to 5-HT deamination varied in the different regions, being in favor of DA in the striatum. With kynuramine as the substrate IC50 values of a number of inhibitors indicated that l-deprenyl was far more potent an inhibitor of human brain MAO than clorgyline or harmaline. N-Desmethylpropargylindane hydrochloride (AGN 1135) was also shown to have MAO-B inhibitory selectivity similar to that of l-deprenyl. Brains obtained at autopsy from l-deprenyl-treated Parkinsonian patients showed that, whereas MAO-B was fully inhibited by the therapeutic doses of l-deprenyl, substantial MAO-A activity was still evident. These results are matched by the significant increases of DA noted in caudate nucleus, globus pallidus, putamen, and substantia nigra and the unaltered 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) in the same regions. These data indicate that the therapeutic actions of l-deprenyl may lie in its selective inhibition of MAO-B resulting in increased brain levels of DA formed from L-dihydroxyphenylacetic acid (L-DOPA).     

Altered Muscarinic and Nicotinic Receptor Densities in Cortical and Subcortical Brain Regions in Parkinson's Disease

Abstract: Muscarinic and nicotinic cholinergic receptors and choline acetyltransferase activity were studied in postmortem brain tissue from patients with histopathologically confirmed Parkinson's disease and matched control subjects. Using washed membrane homogenates from the frontal cortex, hippocampus, caudate nucleus, and putamen, saturation analysis of specific receptor binding was performed for the total number of muscarinic receptors with [³H]quinuclidinyl benzilate, for muscarinic M₁ receptors with [³H]pirenzepine, for muscarinic M₂ receptors with [³H]oxotremorine-M, and for nicotinic receptors with (–)-[³H]nicotine. In comparison with control tissues, choline acetyltransferase activity was reduced in the frontal cortex and hippocampus and unchanged in the caudate nucleus and putamen of parkinsonian patients. In Parkinson's disease the maximal binding site density for [³H]quinuclidinyl benzilate was increased in the frontal cortex and unaltered in the hippocampus, caudate nucleus, and putamen. Specific [³H]pirenzepine binding was increased in the frontal cortex, unaltered in the hippocampus, and decreased in the caudate nucleus and putamen. In parkinsonian patients B_max values for specific [³H]oxotremorine-M binding were reduced in the cortex and unchanged in the hippocampus and striatum compared with controls. Maximal (–)-[³H]nicotine binding was reduced in both the cortex and hippocampus and unaltered in both the caudate nucleus and putamen. Alterations of the equilibrium dissociation constant were not observed for any ligand in any of the brain areas examined. The present results suggest that both the innominatocortical and the septohippocampal cholinergic systems degenerate in Parkinson's disease. The reduction of cortical [³H]oxotremorine-M and (–)-[³H]nicotine binding is compatible with the concept that significant numbers of the binding sites labelled by these ligands are located on presynaptic cholinergic nerve terminals, whereas the increased [³H]pirenzepine binding in the cortex may reflect postsynaptic denervation supersensitivity.     

Regional distribution of enkephalinase in the rat brain by autoradiography

The first visualization of enkephalinase (neutral metalloendopeptidase, E.C.3.4.24.11) in rat brain was obtained by autoradiography, using a new tritiated inhibitor: [3H]N-[( R,S )3-(N-hydroxy) carboxamido-2-benzyl propanoyl]glycine (3H-HCBP-Gly). The preliminary analysis of sections clearly showed a discrete localization of enkephalinase in enkephalin enriched regions, such as caudate nucleus, putamen, globus pallidus, and substantia nigra. Moreover 3H-HCBP-Gly binding also occurred in choroid plexus and spinal cord.     

Characterization of specific in vivo binding of neuroleptic drugs in rat brain.

Lesioning of the rat striatum with kainic acid may provide a useful animal model with which to study Huntington's Disease since, in both situations, changes in several neurochemical parameters appear similar. In this study, we examined the time course of dopaminergic (DA) and muscarinic cholinergic (MCHOL) receptor alterations after kainic acid injection into the rat striatum. As early as two days after unilateral, intrastriatal injection of kainic acid, most striatal perikaya in the injected area had been destroyed as seen by histological examination. A progressive decrease in the DA and MCHOL receptors continued which was not due to changes in their affinity for their respective receptors. By 48 days after injection, there was about 75% decrease in DA receptors and about a 65% decrease in MCHOL receptors. The DA receptor loss is similar in extent to the reported loss in activity of striatal, dopamine-stimulated adenylate cyclase after kainic acid lesion. The DA and MCHOLreceptor loss is similar to the reported loss of neostriatal DA and MCHOL receptors in Huntington's Disease.     

Dopamine receptor binding: differentiation of agonist and antagonist states with 3H-dopamine and 3H-haloperidol.

³H-Dopamine and ³H-haloperidol bind with high affinity and selectivity to synaptic dopamine receptors in membrane preparations of the calf caudate. Binding of both ligands shows marked regional variations with greatest density in caudate, putamen, globus pallidus, nucleus accumbens and olfactory tubercle, areas rich in dopamine nerve terminals. The rank-order of phenothiazines and related agents as well as catecholamines in displacing both dopamine and haloperidol binding closely parallels their pharmacological potencies and affinities for the dopamine-sensitive adenylate cyclase. Dopamine's affinity for specific ³H-dopamine binding sites is 100 times its apparent affinity for the dopamine sensitive adenylate cyclase. Agonists have about 50 times more affinity for dopamine than haloperidol sites, whereas antagonists display about 100 times greater affinity for haloperidol than dopamine sites.     

L-Glutamate toxicity in Huntington's disease fibroblasts

Brain degeneration in Huntington's Disease is thought to occur primarily in the regions of high L-glutamate concentrations. Huntington's Disease fibroblast cultures have a sensitivity to these high concentrations of L-glutamate. These cells show degeneration and loss of viability, within 12 hrs, following treatment with 30 mM L-glutamate. This effect appears to be specific for L-glutamate, can be prevented by glutamine, and is not observed in matched control cultures. The observed glutamate sensitivity may serve as a biochemical genetic marker and the excessive effect of glutamate on neuronal membranes could initiate the characteristic neuronal degeneration observed in Huntington's Disease brains.     

Effect of fenamin and haloperidol on conditioned activity of neurons of the motor cortex and striopallidal system

The activity of neurones in the motor cortex, caudate nucleus, putamen and globus pallidus was studied during elaboration of motor conditioned reflexes to time in rabbits, treated with 1-amphetamine and haloperidol. Mechanisms of reproduction of cells trace activity in the reflex to time at the omission of trials, reacted to 1-amphetamine by increasing the intensity of reactions in the motor cortex and inactivation in putamen cells. The curve of dynamics of intensity changes of trace discharges in the course of a series of trials omissions remained unaltered only in motor cortex; in the other structures it significantly differed from the norm of intact animals. Haloperidol depressed the mechanisms of reproduction of trace reactions of the globus pallidus cells, and made them almost fully inactive in the motor cortex; the putamen neurones reacted to haloperidol by an increase of trace reactions intensity. Against the background of the animal chronic 1-amphetamine intoxication, haloperidol normalized the dynamics and intensity of trace activity. "Therapeutic" effect of haloperidol was most distinctly expressed in the motor cortex and putamen cells, less--in the caudate nucleus and was completely absent in the globus pallidus.