Increased monoamine oxidase B activity in plaque-associated astrocytes of Alzheimer brains revealed by quantitative enzyme radioautography.

The aetiology and pathogenesis of Alzheimer's disease are currently poorly understood, but symptomatic disease is associated with amyloid plaques, neurofibrillary tangles, neuronal loss and numerous alterations of neurotransmitter systems in the CNS. Monoamine oxidase type B is known to be increased in Alzheimer diseased brains. The distribution and abundance of catalytic sites for monoamine oxidases A and B in post mortem human brains of 11 Alzheimer disease cases and five age-matched controls were investigated by quantitative enzyme radioautography. Using tritiated monoamine oxidase inhibitors (Ro41-1049 and lazabemide)--as high affinity substrates selective for monoamine oxidases A and B, respectively--it was found that monoamine oxidase B activity increased up to three-fold exclusively in temporal, parietal and frontal cortices of Alzheimer disease cases compared with controls. This increase was restricted to discrete patches (approximately 185 microns in diameter) which occupied approximately 12% of the cortical areas examined. In other brain regions (hippocampal formation >> caudate-putamen > cerebellum), patches of [3H]lazabemide-enriched binding were less abundant. [3H]Ro41-1049 binding (i.e. monoamine oxidase A) was unchanged in all tissues of diseased versus control brains. The monoamine oxidase B-enriched patches in all cortical regions correlated, in their distribution and frequency, with glial fibrillary acidic protein-immunoreactive clusters of astrocytes. Diffuse and mature beta-amyloid-immunoreactive senile plaques as well as patches of high density binding of [3H]PK-11195--a high-affinity ligand for peripheral-type (mitochondrial) benzodiazepine binding sites in microglia/macrophages--were found throughout Alzheimer diseased cortices. The up-regulation of monoamine oxidase B in plaque-associated astrocytes in Alzheimer's disease--in analogy to its proposed role in neurodegenerative disorders such as Parkinson's disease--might, indirectly, be a potential source of cytotoxic free radicals. Lazabemide, a selective reversible monoamine oxidase B inhibitor, is currently under clinical evaluation for the treatment of Parkinson's and Alzheimer's diseases. We conclude that enzyme radioautography with [3H]lazabemide is a reliable high resolution assay for plaque-associated astroglioses in Alzheimer's disease. Its clinical diagnostic utility for positron emission tomography or single photon emission computer tomography studies is being investigated.

Calbindin D-28k and monoamine oxidase A immunoreactive neurons in the nucleus basalis of Meynert in senile dementia of the Alzheimer type and Parkinson's disease.

In this study, calbindin D-28k (CaBP), monoamine oxidase A (MAO A) and nerve growth factor receptor (NGFr) immunoreactivities were investigated in the nucleus basalis of Meynert (NbM) in patients with senile dementia of the Alzheimer type (SDAT), with Parkinson's disease (PD) with or without dementia, and in controls. Immunocytochemistry using specific antibodies in differing serial sections was employed, and cell counts and NbM nuclear volume measurements were made. Most of the large multipolar NbM neurons showed CaBP immunoreactivity in the cytoplasm of their somata, dendrites and axons. In adjacent, NGFr-reacted sections, the large NbM neurons were also found to be intensely immunoreactive for NGFr on their cellular surfaces. In addition, a subpopulation of large NbM neurons and glial cells were found to be immunoreactive for MAO A. The number of CaBP-immunoreactive (CaBP-i) neurons was decreased by an average of 55% in the 6 SDAT patients, 70% in the 2 nondemented PD patients and 40% in the 1 demented PD patient. The volume calculated for the compact part of the NbM formed by the CaBP-i neuronal somata decreased by an average of 47% in SDAT. On the other hand, measurements in the volume of NGFr-i neurons (including the dendritic arborization) showed an average decrease of 25% in SDAT patients compared to controls. Although all SDAT and PD patients showed a decrease of CaBP-i neurons in the NbM, a loss of MAO-A-i NbM neurons was found only in those patients with dementia. Therefore, the relative proportions of MAO-A-i to CaBP-i neurons were increased in the nondemented PD patients (14.2 and 19.6%) when compared with those in the demented PD patient (2.2%) and with the SDAT patients (0.3-5.6%). These data indicate that a balanced presence of MAO-A-i cholinergic, large NbM neurons may be necessary for the proper maintenance of cognitive function. Functionally this may be translated to mean that dementing changes may cause a decrease from the normal amount of MAO A enzyme activity. This suggests that therapeutic strategies based upon correction of MAO-A activities by MAO-A inhibitors may be important to ameliorating some of the loss in cholinergic function in dementias of SDAT and PD.

Depression and senile dementia of the Alzheimer type: a role for moclobemide.

Depression is common in patients with senile dementia of the Alzheimer type (SDAT) and may precede the onset of the dementia; the underlying biological and neurotransmitter mechanisms may be common to both diseases, so far as norepinephrine lesions are concerned. The major routes of metabolism of amines in the brain utilize the monoamine oxidase (MAO) enzymes. Due to the consistent severity of norepinephrine lesions in the locus coeruleus of patients with pre-senile dementia or SDAT and the fact that MAO-A enzyme is the major metabolizing enzyme present in the locus coeruleus in man, the new specific, reversible MAO-A inhibitors may have a place in the treatment of depression associated with SDAT.

Dopamine D2 receptors in the rat, monkey and the post-mortem human hippocampus. An autoradiographic study using the novel D2-selective ligand 125I-NCQ 298.

The distribution of dopamine D2 receptors in the hippocampal region of the rat, monkey and the postmortem human brain was studied with in vitro receptor autoradiography using the selective salicylamide ligand 125I-NCQ 298. Specific binding was defined in the presence of the D2-selective compound raclopride. In all 3 species, higher densities of specifically bound 125I-NCQ 298 was found in the retrohippocampal structures than in the hippocampus proper. In the rat, layers 1 and 3 of the entorhinal cortex and layer 2 of the presubiculum were found to be rich in specific binding sites. In the monkey, the highest densities were detected in the deep layers (4 through 6) of the entorhinal cortex (EC) and in layer 2 of the presubiculum. Relatively high density of binding was found in the granule cell layer of area dentata. In the human brain, less specific binding was seen as compared to the other two species; the highest densities occurred in the outer layers of the presubiculum and in the hilus of area dentata. These findings show that D2 receptors are present in the hippocampal region and that the retrohippocampal region, including the entorhinal cortex, is enriched in dopamine D2 receptors.

Alterations in the locus coeruleus in dementias of Alzheimer's and Parkinson's disease.

For diagnostic purposes, a differentiation can be made between the locus coeruleus (LC) in normal brain and the LC, in senile dementia of the Alzheimer's type (SDAT) and Parkinson's disease (PD). The differentiation is based on findings concerning the morphological alterations of the TH-immunoreactive; neurons, on the topographical distribution of neuron loss within the length of the LC, and on the total reduction in cell number.

Galanin receptors in the post-mortem human brain. Regional distribution of 125I-galanin binding sites using the method of in vitro receptor autoradiography.

The distribution of putative receptors for the peptide galanin was studied in the normal post-mortem human brain by using 125I-galanin (0.5 nM) in combination with in vitro receptor autoradiography. Specific binding of 125I-galanin was found in a large number of brain areas throughout the neuraxis. Highest binding densities occurred in the basal forebrain and hypothalamus, while the basal ganglia, major parts of the thalamus and the tectum were found to be poor in binding sites. All cortical areas harboured 125I-galanin binding, and in the visual cortex a laminated pattern was present. In the hippocampus, 125I-galanin binding occurred in layer 2 of the entorhinal cortex, in the uncus and in the hippocampal-amygdala area. In the brain-stem, 125I-galanin binding was found in serotoninergic noradrenergic cell groups as well as in the reticular formation and in the parabrachial nuclei. Galanin receptors may, thus, mediate the response of galanin in numerous structures in the human brain.

Quantitation of catecholamine neurons in the locus coeruleus in human brains of normal young and older adults and in depression.

A quantitative study of the morphology and distribution of norepinephrinergic neurons in the human locus coeruleus (LC) is given for normal young and older adult brain. Norepinephrine (NE)-producing neurons are identified by immunocytochemistry of two NE biosynthetic enzymes, tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DBH), visualized by the peroxidase-antiperoxidase and immunogold-silver-staining methods. TH and DBH immunoreactions yield equivalent results. Both immunocytochemical visualization methods allow detailed analysis of neuronal morphology. The neurons of the human LC fall into four classes: large multipolar neurons with round or multiangular somata, large elliptical "bipolar" neurons, small multipolar neurons, and small ovoid "bipolar" neurons. Though most of the neurons contain neuromelanin pigment, some larger neurons lack pigmentation. Dendritic arborization of all neurons is extensive. Computer-assisted quantitative measurements of the parameters somatic size, dendritic arbor length, surface area, and volume are given. Somatic areas of LC neurons of all four classes are decreased in older adult brain, but dendritic arborization is equally extensive as in the younger. The rostrocaudal length of the LC is approximately 15 mm, and no age-dependent decrease is observed. Computer-assisted mapping of immunoreactive neurons and three-dimensional reconstruction allow division of the LC into rostral, middle, and caudal parts with characteristic distribution of neurons. Small neurons predominate in all parts, but the relative contribution of larger cells decreases in a rostrocaudal direction. A cell loss of 27-37% occurs in older adult brains and to 55% in the brain of a chronically depressed patient without dementia. Cell loss is highest in the rostral part, lower in the middle, and absent in the caudal part, and more small cells are lost than larger ones.

Alterations in catecholamine neurons of the locus coeruleus in senile dementia of the Alzheimer type and in Parkinson's disease with and without dementia and depression.

The present study provides qualitative and quantitative investigations of the norepinephrine (NE) neurons in the locus coeruleus (LC) in two neurodegenerative disorders, the senile dementia of the Alzheimer type (SDAT) and Parkinson's disease (PD). The group of PD subjects was subdivided into cases without dementia (P - D), cases with dementia, L-dopa responsive (P + D), and cases with fulminant dementia whose motor disorder symptoms were L-dopa nonresponsive (P + D/L-dopa non-responsive). NE neurons were demonstrated by immunocytochemistry against tyrosine hydroxylase (TH). Quantitations of neuronal parameters and cell numbers and three-dimensional reconstructions of the LC were carried out with a computer-assisted system. In SDAT cases, the rostrocaudal LC length (13 +/- 2.2 mm) is shorter than in controls (14.9 +/- 1.4 mm). The four basic LC neuron classes found in the normal human brain (large multipolar, large "bipolar," small multipolar, and small "bipolar" neurons; Chan-Palay and Asan: J. Comp. Neurol. this issue) are recognizable, but many cell somata are swollen and misshapen with fore-shortened, thick, and less branched dendrites. LC neuron numbers are reduced (between -3.5% and -87.5%). Neuron loss is greatest in the rostral part, less in the middle, and least in the caudal part. In PD cases, the rostrocaudal length (12.4 +/- 1.5 mm) is shorter than in SDAT and controls. The neuronal morphology is more severely altered than in SDAT. The basic neuron classes are hardly distinguishable. Most cell bodies are swollen; they frequently contain Lewy bodies; and the dendrites are short and thin with absent or reduced arborizations. Neuron numbers are more reduced than in SDAT (between -26.4% and -94.4%). Alterations are as severe caudally as rostrally in P - D, and P + D/L-dopa nonresponsive cases. P + D cases are more severely affected rostrally. The presence of depression in SDAT and Parkinson's patients is accompanied by the greatest loss of LC neurons. On the basis of morphological alterations of the TH-immunoreactive neurons, and the degree and topographical distribution of neuron loss, a differentiation is possible between the LC in normal brain and that in SDAT and PD for diagnostic purposes.

Neurons with galanin innervate cholinergic cells in the human basal forebrain and galanin and acetylcholine coexist.

Immunocytochemistry with antibodies against cholinacetyltransferase (ChAT) and the novel peptide galanin (GA) were conducted as a single label or as double label experiments on the basal forebrain nuclei of brains from eleven human subjects without prior history of neurological disease. ChAT positive or cholinergic neurons form the major population of cells in the basal nucleus of Meynert. A minor portion of these ChAT positive neurons demonstrate a coexistence with GA positive immunoreactivity suggesting that they are cholinergic/GA neurons. Small fusiform neurons with long dendrites and complex local axonal networks are GA immunoreactive and are local circuit interneurons. The cholinergic cells in the basal nucleus are innervated by a fine network of GA immunoreactive axons and terminals which enwrap their perikarya and dendrites. It is suggested that GA in local circuit interneurons may provide a significant control or modulation of the cholinergic neurons and of cholinergic functions within the basal nucleus. In human diseases which feature a destruction of the basal forebrain cholinergic neurons, surviving GA neurons may inhibit most remaining cholinergic neurons and their function, with severe consequences.

Cholecystokinin-octapeptide (CCK-8) receptors in the hippocampal region: a comparative in vitro autoradiographic study in the rat, monkey and the postmortem human brain.

Quantitative in vitro receptor autoradiography of 125I-CCK-8 was used to study the regional distribution of CCK-8 receptors in the primate hippocampal region. In the monkey, specific 125I-CCK-8 binding sites were enriched in layer 2 of the presubiculum, layers 1, 2, and 4 of the entorhinal area and in the inner two-thirds of the molecular layer of the area dentata. Moderate to low densities were detected in layer 3 of the entorhinal area, the deep layers of the presubiculum, all layers of subiculum and subfields CA1 and CA3 of Ammon's horn. In the human brain, the highest densities of 125I-CCK-8 binding sites were detected in layer 2 of the presubiculum and layer 2 of the lateral entorhinal area. Moderate to low levels were detected in the Ammon's horn and area dentata. This pattern of receptor distribution overlaps only partly with that found in the rat and indicates phylogenetic differences in the localization of CCK-8 receptors within the hippocampal region.

Galanin hyperinnervates surviving neurons of the human basal nucleus of Meynert in dementias of Alzheimer's and Parkinson's disease: a hypothesis for the role of galanin in accentuating cholinergic dysfunction in dementia.

This study summarizes the findings from postmortem examination of the brains of 22 control cases without neurological deficit, 12 cases of senile dementia of the Alzheimer type (SDAT), and nine cases of Parkinson's disease (three without signs of intellectual deterioration, four with dementia, and two atypical with dementia nonresponsive to L-dopa treatment). The aim of this study was to find the similarities and differences in galanin innervation of the cholinergic basal nucleus neurons in these dementing disorders as compared with controls. Immunocytochemistry with antibodies against galanin peptide and against choline acetyltransferase was applied on perfused brain preparations. Galanin peptide is present in the basal nucleus of Meynert neuron networks in the normal human brain: in local circuit neurons, in a number of galanin/cholinergic neurons, and in a feedback circuit via collaterals) that terminate upon the cholinergic neuronal somata and dendrites. Thus, peptide galanin circuits could function as powerful modulators of the activities of basal nucleus cholinergic neurons, both within the basal forebrain and in their wider projections to the neocortex and amygdala. As galanin has been shown to inhibit cholinergic activity, this galanin network could suppress the activity of cholinergic neurons. In SDAT, there is a primary loss of cholinergic neurons compounded by a secondary reaction of the remaining cholinergic neurons to the terminal degeneration in the cortex. Galanin networks demonstrate an inverse relationship to the cholinergic cell loss. Galanin axons hypertrophy and hyperinnervate the remaining cholinergic neurons. In Parkinson's disease the loss of cholinergic neurons is accentuated by the presence of dementia: the hypertrophy of the galanin axonal networks on cholinergic neurons is dramatic in Parkinson's disease with dementia. These observations throw new light on the neurotransmitter bases for these dementias. Galanin controls cholinergic mechanisms in the basal nucleus of Meynert, and dementia is accompanied by augmentation of galanin innervation onto an already depressed population of cholinergic neurons, thus demonstrating an appreciable amount of plasticity even in aged brain. These findings suggest that the present therapy of cholinergic enhancement as a means to retard intellectual deterioration can by itself have little effect at best, in these dementias. The suppressive effect of galanin peptide has to be reduced or curtailed, perhaps concurrently with the treatment of the cholinergic deficit.(ABSTRACT TRUNCATED AT 400 WORDS)

Simultaneous demonstrations of neuropeptide Y gene expression and peptide storage in single neurons of the human brain.

A combination of in situ hybridization for neuropeptide Y mRNA that used a 32P-labeled complementary RNA probe and immunocytochemistry with polyclonal antibodies against neuropeptide Y were applied to human cortical brain samples to simultaneously localize neuropeptide Y and its mRNA. These two techniques allowed simultaneous identification of neuropeptide Y gene expression and peptide storage in single neurons of the human brain.

Techniques for the simultaneous demonstrations of neuropeptide y gene expression and peptide storage in single neurons of the human brain.

The distribution of mRNA encoding neuropeptide Y (NPY) in neurons of the normal human cerebral cortex in surgical biopsy specimens and postmortem brain was studied by in situ hybridization techniques. A 32P-labeled complementary RNA (cRNA) probe was used on cryostat sections of formaldehyde-fixed cortical biopsy specimens. Hybridization to NPY mRNA was found in all samples: after autoradiography, discrete deposits of silver granules were observed on neuronal cell bodies abundantly distributed in the deep layers of the cortex, particularly laminae IV and VI, and on smaller cell bodies in the white matter. The localization of the neurons hybridized for NPY mRNA was comparable to that of NPY-immunoreactive cells as shown in sections from the same tissue blocks immunostained by using NPY antibodies. The specificity of the in situ hybridization technique was confirmed by hybridization analysis of electrophoretically fractionated RNA. This study clearly demonstrated the consistent localization of NPY gene transcription and expression in normal mature human cortical neurons. A combination of the methods of in situ hybridization for mRNA encoding Neuropeptide Y (NPY) using a 32P-labeled complementary RNA probe and immunocytochemistry using polyclonal antibodies against NPY were applied on human cortical brain samples. The manipulation of these two techniques allowed successful simultaneous demonstration of NPY gene expression and peptide storage in single neurons of the human brain.

Co-localization of neuropeptide tyrosine and somatostatin immunoreactivity in neurons of individual subfields of the rat hippocampal region.

The co-localization of neuropeptide tyrosine (NPY) and somatostatin (SOM) in rat hippocampal cells was studied in double labelling experiments using a combination of antibodies against the two peptides on the same tissue section. The individual hippocampal subfields show large variations in the relative number of NPY- and SOM-immunoreactive (-i) neurons. While the entorhinal area is far richer in SOM as compared to NPY-i cells, NPY-i cells predominate in all subfields (e.g. regio superior, regio inferior) of Ammon's horn. Co-localization of both peptides in single neurons was highest in regio inferior and in the area dentata and lowest in the retrohippocampal structures. In the dorsal hippocampus, the number of SOM-i cells containing NPY-i was higher than the number of NPY-i cells containing SOM-i. This pattern was reversed in the retrohippocampal region. At ventral levels the incidence of colocalization of NPY- and SOM-i in single cells increased in all hippocampal subfields.