The interstitial nuclei of the human anterior hypothalamus: an investigation of variation with sex, sexual orientation, and HIV status.

The interstitial nuclei of the human anterior hypothalamus (INAH1-4) have been considered candidates for homology with the sexually dimorphic nucleus of the preoptic area of the rat. Volumetric sexual dimorphism has been described for three of these nuclei (INAH1-3), and INAH3 has been reported to be smaller in homosexual than heterosexual men. The current study measured the INAH in Nissl-stained coronal sections in autopsy material from 34 presumed heterosexual men (24 HIV- and 10 HIV+), 34 presumed heterosexual women (25 HIV- and 9 HIV+), and 14 HIV+ homosexual men. HIV status significantly influenced the volume of INAH1 (8% larger in HIV+ heterosexual men and women relative to HIV- individuals), but no other INAH. INAH3 contained significantly more neurons and occupied a greater volume in presumed heterosexual males than females. No sex difference in volume was detected for any other INAH. No sexual variation in neuronal size or density was observed in any INAH. Although there was a trend for INAH3 to occupy a smaller volume in homosexual men than in heterosexual men, there was no difference in the number of neurons within the nucleus based on sexual orientation.

Glycation and microglial reaction in lesions of Alzheimer's disease.

Single, double, and triple immunostaining of cryostat sections of elderly normal and Alzheimer disease (AD) brain was performed with monoclonal and polyclonal antibodies to advanced glycation end products (AGE). The sections were counterstained with thioflavin-S or with immunocytochemistry for A beta and also stained with markers for microglia. AGE-immunoreactivity was detected in senile plaques and neurofibrillary tangles (NFT). AGE immunoreactivity was most intense in dense or reticular amyloid deposits and extracellular NFT, while intracellular NFT and diffuse amyloid had less AGE immunoreactivity. This pattern of immunoreactivity was similar to that noted in previous studies with antibodies to apolipoprotein-E (apo-E). Therefore, double labeling with antibodies to apo-E and AGE was performed. AGE immunoreactivity colocalized to a very high degree with apo-E immunoreactivity, except that relatively more intense apo-E immunoreactivity was detected in amyloid deposits and more intense AGE immunoreactivity in NFT. The lesions that were immunostained with antibodies to AGE and apo-E were often, but not always, associated with a local microglial reaction. The results raise the possibility that apo-E or a fragment of apo-E may be glycated. Biochemical studies are needed to determine the extent of possible apo-E glycation in AD. The present results raise the possibility that glycation may serve as one of the signals for activation of microglia associated with amyloid deposits and extracellular NFT.

Cytoskeletal pathology in non-Alzheimer degenerative dementia: new lesions in diffuse Lewy body disease, Pick's disease, and corticobasal degeneration.

Increasing use of immunocytochemistry for evaluation of dementia disorders has revealed histopathological alterations that were previously unknown, even with sensitive silver techniques. Disorders [Pick's disease (PD), diffuse Lewy body disease (DLBD) and corticobasal degeneration (CBD)] in which immunocytochemistry has revealed occult pathology are discussed. All three disorders have neurofilament (NF) immunoreactive neuronal alterations in the neocortex. In DLBD round, eosinophilic cytoplasmic inclusions referred to as cortical Lewy bodies are neurofilament-positive, while in both PD and CBD neurofilament epitopes are expressed in irregularly swollen neurons and their proximal cell processes, which are referred to as ballooned neurons. Interestingly, the cortical neuronal population that is vulnerable to Lewy bodies is similar to that which is vulnerable to ballooned neurons. Furthermore, Lewy bodies can occasionally be detected within the cytoplasm of ballooned neurons. Besides neurofilament-immunoreactivity, Lewy bodies are immunoreactive for ubiquitin, while ballooned neurons are inconsistently stained with antibodies to ubiquitin. Both Lewy bodies and ballooned neurons can be appreciated with routine histology, but they are much easier to detect with immunocytochemistry. In contrast, a new type of neuritic alteration in the hippocampal CA2/3 region has been recognized in DLBD. These dystrophic neurites cannot be appreciated with routine histology and are only optimally seen with immunocytochemistry for ubiquitin. Their presence is a certain indication of the presence of cortical Lewy bodies. The microtuble associated protein tau is the major constituent of neurofibrillary tangles in Alzheimer's disease (AD). Biochemical studies have shown that Pick bodies, argyrophilic neuronal inclusions that are highly characteristic of, if not pathognomonic for PD are also composed of abnormal tau protein. Along with Pick bodies, tau has recently been detected in glial cells in PD. Similar so-called "gliofibrillary tangles" are increasingly recognized in progressive supranuclear palsy. Previously, CBD was considered to be free of such lesions, but recent studies have revealed widespread tau-positive neuronal and glial cytoskeletal lesions in CBD. A distinctive type of tau-positive glial lesion in CBD is characterized by annular clusters of grain-like tau immunoreactivity reminiscent of a neuritic plaque in AD, except that the clusters are devoid of amyloid. The tau-positive profiles are consistently located around a central astrocyte cell body. Double labeling studies with glial fibrillary acidic protein, vimentin and CD44, which are markers for reactive astrocytes, demonstrates tau immunoreactivity within astrocytic processes; these "astrocytic plaques" appear to be specific for CBD. Although NF, ubiquitin and tau proteins are present in diverse neuronal and glial inclusions in these disorders, the morphology and distribution of these lesions differentiate non-AD dementias.

Neuropathologic overlap of progressive supranuclear palsy, Pick's disease and corticobasal degeneration.

Several neurodegenerative disorders contain tau-immunoreactive neuronal and glial inclusions throughout the cerebral cortex and brainstem. Although these diseases have been considered distinct clinicopathological entities, recent recognition of many neuropathological and clinical parallels has raised the question of overlap between the disorders. In addition, histopathological similarities sometimes complicate neuropathological diagnosis. To address these issues, we examined the morphology and differential distribution of pathologic lesions in three disorders: progressive supranuclear palsy, Pick's disease, and corticobasal degeneration. We found considerable similarity in the anatomical regions affected by the three entities; however, semiquantitative analysis revealed differential anatomical susceptibility. Similarly, although overlap existed in the morphology of tau-immunoreactive inclusions, characteristic differences remained and may be useful in differential diagnosis. In particular, glial inclusions varied dramatically between the disorders. Despite significant overlap among the three neurodegenerative diseases examined, the morphological and regional differences suggest that each is a distinct pathophysiological entity.

Ultrastructure and biochemical composition of paired helical filaments in corticobasal degeneration.

Corticobasal degeneration (CBD) is a neurodegenerative disorder associated with extensive cytoskeletal abnormalities. These include tau-positive neuropil threads and grains, ballooned or swollen neurons, neurofibrillary tangles, and glial inclusions. Given the presence of tau-positive structures in CBD, we investigated whether abnormalities in tau proteins associated with CBD were similar to those in Alzheimer's disease (AD). Fractions of abnormal tau proteins were isolated as Sarkosyl-insoluble pellets. By electron microscopic examination, the fraction from CBD contained twisted filaments that differed from paired helical filaments of AD. In CBD, filaments were shorter in length, rarely longer than 400 nm, 10 to 20% wider in the maximum and minimum widths (26 to 28 nm and 13 to 14 nm, respectively), and the periodic twist (169 to 202 nm) was twice as long as that in AD. Immunogold labeling with a panel of tau-reactive antibodies (Alz 50, Tau 14, AH-1, E-11, PHF-1, and Tau 46) showed no apparent differences in the pattern of tau immunoreactivity between filaments of CBD and AD. Western blots revealed that polypeptides of abnormal tau were present in both fractions; however, only two polypeptides (68 and 64 kd) were present in CBD as compared with three (68, 64, and 60 kd) in AD. Both of these polypeptides were reactive with additional antibodies (E-9, Tau-1 after dephosphorylation, AT8, and NP8). Only one polypeptide (68 kd) bound an antibody to adult-specific tau sequence encoded by exon 2, but neither was reactive with antibodies to adult-specific sequences encoded by exons 3 and 10. The results suggest that abnormalities in the number and heterogeneity of isoforms of tau may be one of the factors contributing to ultrastructural differences in pathological filaments of CBD and AD.

TG-1: a marker for neuronal nuclei in Alzheimer's disease.

TG-1 is a monoclonal antibody (mAb) raised against paired helical filaments purified from Alzheimer's Disease (AD) brain by immunoaffinity chromatography. By immunocytochemistry, TG-1 reveals abundant staining of neuronal nuclei in AD brain, but little or no staining in normal brain. TG-1 stained nuclei are observed in areas of AD brain with neurofibrillary pathology and in certain neurones that are not normally affected. Biochemical studies with TG-1 show antigens of 32-38 kDa in pellets and 50 kDa in supernatants from brain, with no obvious differences between normal and AD. TG-1 also recognizes an unusual structure, i.e., a 'starburst' in brain tissue from AD and elderly normals. Starbursts are not immunoreactive for the astrocytic marker, glial fibrillary acidic protein, and are not associated with amyloid. Widespread nuclear staining is observed with TG-1 in rat brain, and the immunoreactive antigens in purified nuclei are similar to those in human brain. Thus, TG-1 identifies neuronal nuclear antigens that are altered in AD, and provides a new avenue for studying pathogenic mechanisms in the disease.

Microglia and cytokines in neurological disease, with special reference to AIDS and Alzheimer's disease.

Microglia are associated with central nervous system (CNS) pathology of both Alzheimer's disease (AD) and the acquired immunodeficiency syndrome (AIDS). In AD, microglia, especially those associated with amyloid deposits, have a phenotype that is consistent with a state of activation, including immunoreactivity with antibodies to class II major histocompatibility antigens and to inflammatory cytokines (interleukin-1-beta and tumor necrosis factor-alpha). Evidence from other studies in rodents indicate that microglia can be activated by neuronal degeneration. These results suggest that microglial activation in AD may be secondary to neurodegeneration and that, once activated, microglia may participate in a local inflammatory cascade that promotes tissue damage and contributes to amyloid formation. In AIDS, microglia are the primary target of retroviral infection. Both ramified and ameboid microglia, in addition to multinucleated giant cells, are infected by the human immunodeficiency virus (HIV-1). The mechanism of microglial infection is not known since microglia lack CD4, the HIV-1 receptor. Microglia display high affinity receptors for immunoglobulins, which makes antibody-mediated viral uptake a possible mechanism of infection. In AIDS, the extent of active viral infection and cytokine production may be critically dependent upon other factors, such as the presence of coinfecting agents. In the latter circumstance, very severe CNS pathology may emerge, including necrotizing lesions. In other circumstances, HIV infection of microglia probably leads to CNS pathology by indirect mechanisms, including release of viral proteins (gp120) and toxic cytokines. Such a mechanism is the best hypothesis for the pathogenesis of vacuolar myelopathy in adults and the diffuse gliosis that characterizes pediatric AIDS, in which very little viral antigen can be detected.

Regional localization of the regulatory subunit (RII beta) of the type II cAMP-dependent protein kinase in human brain.

The distribution of the regulatory (RII beta) subunits of type II cAMP-dependent protein kinase in cortical and subcortical areas was examined in human control and Alzheimer's disease (AD) brains. Four monoclonal antibodies generated against bovine brain RII, which cross-reacted with human brain RII beta, detected RII-immunoreactivity in pyramidal neurons of the hippocampus and frontal, occipital, parietal and superior temporal cortices and in non-pyramidal neurons of the amygdala and putamen. RII beta immunoreactivity was localized to neuronal perikarya, proximal dendrites and cell processes. With the exception of rare processes in the ventroposterior lateral nucleus, RII-immunoreactivity was not seen in the thalamus. Other areas lacking RII-immunoreactivity included the midbrain, caudate nucleus and globus pallidus. RII-immunoreactivity was not detected in endothelia or glia. Except for the neocortex, the distribution of RII beta immunoreactivity was the same in AD and non-demented control brains; however, cell bodies and their processes stained more intensely and uniformly in the neocortical regions of non-demented controls compared to AD. In the neocortex of AD, RII beta immunoreactivity was substantially decreased in the superior temporal and occipital cortices, but not in the frontal cortex. Our data suggest that RII subunits are regionally distributed in the human brain. RII-immunoreactivity was decreased in some regions of neocortex in AD, but it did not preferentially colocalize with neurofibrillary tangles (NFT), senile plaques, or neuropil threads.

Identification of normal and pathological aging in prospectively studied nondemented elderly humans.

Results of a standardized histochemical and immunocytochemical analysis of the brains of 14 nondemented elderly humans for whom prospective neurological and neuropsychological data had been collected for 3 to 8 years before death suggested that nondemented elderly humans fall into two pathological subgroups that are not clinically distinguishable. One was associated with moderate to marked cerebral amyloid deposition ("pathological aging"), while the other had either minimal or no amyloid deposition ("normal aging"). Neocortical and hippocampal neurofibrillary degeneration was either completely absent or of very limited degree in both subgroups. Both subgroups had ubiquitin-immunoreactive dystrophic neurites in the cerebral cortex and granular degeneration of myelin in white matter. These ubiquitin-immunoreactive structures seem to be a universal and invariant manifestation of brain aging, but the same cannot be said for amyloid deposition and neurofibrillary degeneration. Pathological aging might be preclinical Alzheimer's disease, but it currently cannot be distinguished from normal aging by even sensitive neuropsychological measures. These findings provide strong support for the hypothesis that cerebral amyloid deposition is not necessarily associated with clinically apparent cognitive dysfunction and that additional factors, such as neuronal or synaptic loss or widespread cytoskeletal aberrations, are necessary for dementia in AD.

Ubiquitin-immunoreactive dystrophic neurites in Down's syndrome brains.

Ubiquitin-immunoreactivity was studied in Down's syndrome brains ranging in age from two days to sixty years. Numerous randomly distributed ubiquitin-immunoreactive dot-like structures in the white matter were shown to correspond to granular degeneration of myelin. Granular degeneration of myelin was first detected at age 21 and increased thereafter with age. Other larger and more coarsely granular ubiquitin-immunoreactive structures, most numerous in the middle and upper cortical layers, were consistent with dystrophic neurites. Immunoelectron microscopy demonstrated that the dystrophic neurites contained non-filamentous, membranous, dense bodies. In Down's syndrome, ubiquitin-immunoreactive dystrophic neurites were first detected at age six in the hippocampus, and were consistently more numerous in comparison to age-matched control subjects. In the presence of amyloid, either as diffuse or as compact deposits, ubiquitin-immunoreactive dystrophic neurites frequently formed aggregates consistent with senile plaques. Although apparently independent events, these data suggest that amyloid deposition is associated with local accentuation of ubiquitin-immunoreactive neuritic dystrophy. In addition, since dystrophic neurites appeared substantially earlier in the grey matter in Down's syndrome than in age-matched normals, this may be further evidence that selective aspects of aging are accelerated in Down's syndrome.

Microglia in human disease, with an emphasis on acquired immune deficiency syndrome.

In conclusion, there is overwhelming evidence that within the CNS the primary sites of active HIV-1 infection are microglia. CNS infection may be related to the normal repopulation of the CNS by monocytes (microglial turnover) that carry latent infection into the CNS. Activation of viral infection may depend upon microglial differentiation, soluble factors (cytokines), and/or coexistent infections. Infection of microglia may disturb the normal hemostatic balance that exists between microglia and other glia, and between microglia and neurons, processes that are only recently being explored at the molecular level. The impact that HIV infection of microglia may have on synaptic integrity is unknown. Cytokines appear to be prime candidates as mediators of some of the adverse effects of microglial infection on other CNS cells, myelin and endothelial cells.

Detection of HLA-DR on microglia in the human brain is a function of both clinical and technical factors.

Detection of HLA-DR, a class II major histocompatibility antigen, on glial cells is dependent not only on duration and type of tissue fixation and processing, but also on clinical factors. Glial cells labeled by anti-HLA-DR were consistent with microglia by light microscopic and ultrastructural criteria, and were colabeled with other microglial markers, including LN-1, Leu-M5, and leukocyte common antigen (LCA). In young and elderly subjects who died suddenly, anti-HLA-DR labeled microglia in the white matter, but far fewer cells in the gray matter. In subjects who died of chronic debilitating illness, such as Alzheimer's disease and carcinomatosis, anti-HLA-DR labeled numerous microglia throughout both the gray and white matter. In Alzheimer's disease, microglia were aggregated in compact senile plaques, but loosely associated with diffuse amyloid deposits. These results suggest that HLA-DR may be constitutively expressed in white matter, but induced in gray matter microglia in chronic disease states or in association with amyloid deposits.

Ubiquitin immunoelectron microscopy of dystrophic neurites in cerebellar senile plaques of Alzheimer's disease.

Senile plaques are present in the cerebellum of most Alzheimer patients. They are composed of beta-amyloid deposits lacking neurites detectable with immunocytochemistry for neurofilament, tau and paired helical filament proteins. Recent studies, however, have shown that cerebellar plaques usually contain round structures that are reactive with ubiquitin antibodies. In this immunoelectron microscopic study, the nature of these structures is explored. Ubiquitin-positive structures in cerebellar senile plaques were composed of degenerating neurites that contained membranous and vesicular dense bodies, but no paired helical filaments. A minority of the neurites contained finely granular material. Thus, cerebellar plaques are associated with neuritic degeneration, and the neurites in cerebellar plaques resemble dystrophic neurites in senile plaques of non-demented elderly subjects and subjects with non-Alzheimer dementias. They differ from some of the neurites in senile plaques in the neocortex in Alzheimer's disease by the absence of paired helical filaments. These results suggest that the same mechanisms involved in the generation of dystrophic neurites in pathological aging are involved in generating dystrophic neurites in the cerebellum in Alzheimer's disease.

Microglia in cerebellar plaques in Alzheimer's disease.

Cerebellar amyloid deposits in Alzheimer's disease were studied by immunocytochemistry and with a series of antibodies that recognize human microglia, including anti-HLA-DR, LN-1, Leu-M5 and leukocyte common antigen. Microglia formed a dense reticular array throughout the cerebellum in areas with and without amyloid deposits. In areas with compact and reticular amyloid deposits, microglia had morphological features consistent with activation, such as cytoplasmic swelling and shortening and thickening of cell processes. In areas with diffuse amyloid deposits, microglia had delicate and highly ramified processes. Nevertheless, microglial cells or their processes were detected in association with amyloid deposits of all morphological types. These results raise the possibility that microglia may play a fundamental role in the pathogenesis of amyloid deposition in the cerebellum in Alzheimer's disease.

Astrocytes and microglia in human brain share an epitope recognized by a B-lymphocyte-specific monoclonal antibody (LN-1).

A B-lymphocyte-specific mouse monoclonal antibody, LN-1, recognizes two morphologic classes of glial cells in human brain. The nature and duration of tissue fixation and processing are critical in the detection of the two cell types. In tissue that is lightly fixed, LN-1 recognizes astrocytes. The astrocytic nature of the LN-1 reactive glial cell was confirmed by cytologic features, tissue distribution, immunoelectron microscopy, double labeling immunofluorescent microscopy, and staining of serial sections with antibodies to glial fibrillary acidic protein. In tissue that is fixed for longer periods or in Bouin's fixative, two glial cell types are recognized: astrocytes and microglia. The identity of the latter cell type as microglia was confirmed by morphologic features, tissue distribution, immunoelectron microscopy, and double staining with monoclonal antibodies or lectins to macrophage markers, including class II major histocompatibility antigens. The two cell types had different disposition in senile plaques of elderly individuals and of those with Alzheimer's disease. Astrocytes were present at the periphery of the plaques, whereas microglial cells were centrally placed, often in juxtaposition to amyloid. The results are discussed with respect to ontogeny of glial cells and the ability of monoclonal antibodies to recognize epitopes on unrelated proteins.

Mesostriatal projections in BALB/c and CBA mice: a quantitative retrograde neuroanatomical tracing study.

A major portion of the midbrain dopamine (DA)-containing neurons project to the striatum and make up the mesostriatal DA system. The purpose of the present experiment was to map the location and quantify the density of mesostriatal neurons within two inbred mouse strains (BALB/c and CBA) known to possess different numbers of midbrain DA neurons. Computer-assisted reconstructions were made of both the wheat germ agglutinin-conjugated horseradish peroxidase (WGA-HRP) striatal injection site and the retrogradely labeled midbrain cells. There was no strain differences in the major source or topographical pattern of innervation of the striatum from the midbrain cellular regions. Even following small striatal injections, the labeled midbrain cells were found throughout most of the rostrocaudal extent of the midbrain DA nuclei; some labeled cells were found within the substantia nigra, the ventral tegmental area and the adjacent retrorubral field. Although the BALB/c strain has 20-25% more midbrain DA neurons than the CBA, given comparable striatal injection volumes, there was no significant difference in the number of HRP-filled mesostriatal neurons between the two mouse strains. These data suggest that the mesostriatal neurons give rise to comparable axonal branching within the striatum in the two mouse strains.

Diffuse Lewy body disease: light and electron microscopic immunocytochemistry of senile plaques.

The nature of senile plaques (SP) in 27 cases of diffuse Lewy body disease (LBD) was investigated using immunocytochemistry and antibodies to beta amyloid protein synthetic peptides (BetaSP), ubiquitin (UBQ), paired helical filaments (PHF; Ab39) and a 68-kDa protein in Alzheimer brains (Alz50). Lewy bodies were present in widespread areas of the neocortex of all cases and were more easily detected with ubiquitin immunocytochemistry than with conventional stains. All cases had neocortical SP, but only six cases had neocortical neurofibrillary tangles (NFT). SP were very numerous in most cases and were usually "pale", "diffuse" or "very primitive" plaques with thioflavin S fluorescent microscopy. SP in diffuse LBD were immunostained with BetaSP. Several cases had extensive amyloid angiopathy that was also immunoreactive with BetaSP. SP in diffuse LBD were characterized by amyloid deposits with few or no neuritic elements that could be detected with thioflavin S, Bielschowsky's stain or double staining with BetaSP and Bodian's silver stain. They differed from plaques in Alzheimer's disease by lack of PHF-type neurites that could be stained with Ab39. In diffuse LBD, SP contained PHF-type neurites only in areas coexistent with NFT. Some SP had round, granular neurites that were immunoreactive with UBQ, but weakly argyrophilic with Bodian's stain and nonfluorescent with thioflavin S. Diffuse LBD lacked significant neuritic change in the neuropil that could be detected with UBQ, Ab39 and Alz50. The latter finding is a characteristic feature that distinguishes Alzheimer's disease from diffuse LBD.

Effect of ventral noradrenergic bundle lesions on concentrations of monoamine neurotransmitters and metabolites in several discrete areas of the rat brain.

1. The effects of radiofrequency lesions of the ventral noradrenergic bundle (VNB) on monoamine and metabolite concentrations in several discrete areas of the rat hypothalamus were examined. Monoamines and metabolites were analyzed utilizing high-performance liquid chromatography coupled with electrochemical detection. 2. VNB lesions decreased the concentrations of norepinephrine (NE) and 3-methoxy-4-hydroxyphenylethylene glycol in all areas examined except in the ventromedial nucleus (VMN). Dopamine and 3,4-dihydroxyphenylacetic acid concentrations were selectively decreased in the dorsomedial nucleus (DMN) and also slightly decreased in the medial forebrain bundle following VNB lesions. Serotonin and 5-hydroxyindole-3-acetic acid concentrations were not altered by VNB lesion in any area examined. 3. The results indicate that the NE innervation to the hypothalamus is extensive and that NE in the VMN may not be derived from the VNB. The source of the DA innervation to the DMN may be located in or pass through the area affected by the VNB lesion.

Comparative neuropharmacology of buspirone and MJ-13805, a potential anti-anxiety drug.

Buspirone is a clinically efficacious anti-anxiety drug without any other benzodiazepine-like activity. Although buspirone can displace ligands for dopamine (DA) receptors, its equipotent analog, MJ-13805, cannot. Buspirone can potently increase dopaminergic impulse flow and metabolism, primarily due to inhibition of DA autoreceptors. However, MJ-13805 does not block striatal nerve ending DA autoreceptors and slightly increases striatal DA metabolism. Both drugs potently reverse catalepsy due to either DA receptor blockade or DA depletion which indicates an effect within the extrapyramidal system efferent from the DA neuron. Amantadine is at least ten fold less potent than these drugs for reversal of catalepsy. These data indicate that altered dopaminergic neurotransmission may not be important for the anti-anxiety effect of buspirone and that buspirone should be tested for efficacy in various models of movement disorders. The site and mechanism of action for buspirone and MJ-13805 remains obscure. A metabolite of buspirone, 1-piperazinylpyrimidine, does not reverse catalepsy although this drug is known to be active in anti-anxiety screening tests. Thus, buspirone may have separate mechanisms of action for reduction of anxiety and reversal of catalepsy.