Although protein phosphorylation has been characterized more extensively, modulation of the acetylation state of signaling molecules is now being recognized as a key means of signal transduction. The enzymes responsible for mediating these changes include histone acetyl transferases and histone deacetylases (HDACs). Members of the HDAC family of enzymes have been identified as potential therapeutic targets for diseases ranging from cancer to ischemia and neurodegeneration. We initiated a project to conduct comprehensive gene expression mapping of the 11 HDAC isoforms (HDAC1-11) (classes I, II, and IV) throughout the rat brain using high-resolution in situ hybridization (ISH) and imaging technology. Internal and external data bases were employed to identify the appropriate rat sequence information for probe selection. In addition, immunohistochemistry was performed on these samples to separately examine HDAC expression in neurons, astrocytes, oligodendrocytes, and endothelial cells in the CNS. This double-labeling approach enabled the identification of specific cell types in which the individual HDACs were expressed. The signals obtained by ISH were compared to radiolabeled standards and thereby enabled semiquantitative analysis of individual HDAC isoforms and defined relative levels of gene expression in >50 brain regions. This project produced an extensive atlas of 11 HDAC isoforms throughout the rat brain, including cell type localization, providing a valuable resource for examining the roles of specific HDACs in the brain and the development of future modulators of HDAC activity.

The current model to explain the organization of the mammalian nervous system is based on studies of anatomy, embryology, and evolution. To further investigate the molecular organization of the adult mammalian brain, we have built a gene expression-based brain map. We measured gene expression patterns for 24 neural tissues covering the mouse central nervous system and found, surprisingly, that the adult brain bears a transcriptional ''imprint'' consistent with both embryological origins and classic evolutionary relationships. Embryonic cellular position along the anteriorposterior axis of the neural tube was shown to be closely associated with, and possibly a determinant of, the gene expression patterns in adult structures. We also observed a significant number of embryonic patterning and homeobox genes with region-specific expression in the adult nervous system. The relationships between global expression patterns for different anatomical regions and the nature of the observed region-specific genes suggest that the adult brain retains a degree of overall gene expression established during embryogenesis that is important for regional specificity and the functional relationships between regions in the adult. The complete collection of extensively annotated gene expression data along with data mining and visualization tools have been made available on a publicly accessible web site (www.barlow-lockhartbrainmapnimhgrant.org).database ͉ development ͉ evolution ͉ gene expression profiling ͉ inbred strains of mice T he adult nervous system achieves its mature form as the result of neuroectodermal cells committing to a specific fate and then segregating into distinct regional collectives of neurons that become fully functional through establishment of connections to other neurons. Our current understanding of brain architecture and organization is based on studies of embryology, anatomy, and evolution in which direct observation of anatomic structures was the foundation for postulated models of brain structure (1). Recent models of brain development and maturation consider relationships between different regions based on the expression of specific genes in assigning developmental origins of adult structures (2, 3). Here, we have constructed a regional gene expression atlas of the adult mouse brain and analyzed the quantitative results by using molecular classification algorithms.Genome-wide gene expression profiling is a powerful technique for deriving information about specific brain regions (4, 5). This approach has been used to measure gene expression patterns in particular regions, subregions, or cell populations in the brain (6-11). Two previous studies have analyzed gene expression differences across multiple regions of the mammalian brain by using multiple strains or species (12,13). However, the current study is the most extensive to date in terms of the number of genes and the coverage of different neural tissues. Our goal was to create a publicly accessible gene-based brain map with data sets, metadata, datab...

Various transgenic mouse models of Alzheimer's disease (AD) have been developed that overexpress mutant forms of amyloid precursor protein in an effort to elucidate more fully the potential role of ␤-amyloid (A␤) in the etiopathogenesis of the disease. The present study represents the first complete 3D reconstruction of A␤ in the hippocampus and entorhinal cortex of PDAPP transgenic mice. A␤ deposits were detected by immunostaining and thioflavin fluorescence, and quantified by using high-throughput digital image acquisition and analysis. Quantitative analysis of amyloid load in hippocampal subfields showed a dramatic increase between 12 and 15 months of age, with little or no earlier detectable deposition. Three-dimensional reconstruction in the oldest brains visualized previously unrecognized sheets of A␤ coursing through the hippocampus and cerebral cortex. In contrast with previous hypotheses, compact plaques form before significant deposition of diffuse A␤, suggesting that different mechanisms are involved in the deposition of diffuse amyloid and the aggregation into plaques. The dentate gyrus was the hippocampal subfield with the greatest amyloid burden. Sublaminar distribution of A␤ in the dentate gyrus correlated most closely with the termination of afferent projections from the lateral entorhinal cortex, mirroring the selective vulnerability of this circuit in human AD. This detailed temporal and spatial analysis of A␤ and compact amyloid deposition suggests that specific corticocortical circuits express selective, but late, vulnerability to the pathognomonic markers of amyloid deposition, and can provide a basis for detecting prior vulnerability factors. A lzheimer's disease (AD), the most common form of dementia in the aging population, is characterized by the extracellular accumulation of ␤-amyloid (A␤), the intracellular appearance of neurofibrillary tangles, and synaptic and neuronal loss (1). Mounting evidence supports a causal role for A␤ in the pathophysiology of AD (2, 3). Various transgenic models have been developed which overexpress mutant forms of amyloid precursor protein (APP); these models mimic some aspects of AD pathology, including A␤ deposition and synaptic damage (4-9).In AD, amyloid deposition and neurofibrillary tangle formation occur in a spatially and temporally defined pattern in specific neocortical and hippocampal regions that reflects selective vulnerability of certain circuits, particularly corticocortical circuits in neocortex (10, 11) and the perforant path that projects from the entorhinal cortex (EC) to the dentate gyrus (DG) (12, 13). Transgenic mouse models that overexpress mutant APP show an age-dependent accumulation of A␤ (14); however, there has been no comprehensive quantitative analysis of the spatial and temporal progression of amyloid and A␤ accumulation, especially in the most vulnerable regions.Deposits of A␤ that form in AD have been morphologically classified into several types, such as diffuse, fibrillar, dense-cored or classic, compact, or ''burnt-out'' ...

High-resolution magnetic resonance microscopy (MRM) was used to determine regional brain volumetric changes in a mouse model of Alzheimer's disease. These transgenic (Tg) mice overexpress human mutant amyloid precursor protein (APP) V717F under control of platelet-derived growth factor promoter (PDAPP mice), and cortical and hippocampal ␤-amyloid (A␤) deposits accumulate in heterozygotes after 8 -10 mos. We used MRM to obtain 3D volumetric data on mouse brains imaged in their skulls to define genotype-and age-related changes. Hippocampal, cerebellar, and brain volumes and corpus callosum length were quantified in 40-, 100-, 365-, and 630-day-old mice. Measurements taken at age 100 days, before A␤ deposition, revealed a 12.3% reduction of hippocampus volume in Tg mice compared with WT controls. This reduction persisted without progression to age 21 mos. A significant 18% increase in hippocampal volume occurred between 40 and 630 days in WT mice, and no corresponding significant increase occurred in Tg mice. Cavalieri volume estimates of hippocampal subfields from 100-day-old Tg mice further localized a 28% volume deficit in the dentate gyrus. In addition, corpus callosum length was reduced by Ϸ25% in Tg mice at all ages analyzed. In summary, reduced hippocampal volume and corpus callosum length can be detected by MRM before A␤ deposition. We conclude that overexpression of APP and amyloid may initiate pathologic changes before the appearance of plaques, suggesting novel targets for the treatment of Alzheimer's disease and further reinforcing the need for early diagnosis and treatment.T he essential neuropathologic features of Alzheimer's disease (AD) include the progressive deposition of amyloid plaques and neurofibrillary tangles in neocortical and hippocampal structures and a parallel global decrease in cortical volume (1, 2). Extensive data from analysis of postmortem human brain and mouse models of AD associate the neuropathology of AD with alterations in the expression, distribution, and deposition of ␤ amyloid protein (A␤). In AD brains, A␤ levels are increased, and the protein can be found in fibrillar chains within compact plaques, aggregated in diffuse plaques, or as oligomers and monomers in regions outside of plaques (3-6). Some of the known human mutations associated with AD affect the processing or cleavage of amyloid precursor protein (APP) and can cause increased A␤ levels or increase the relative amount of the primary plaque component A␤ 1-42 compared with A␤ 1-40 (3, 5-10). Other human mutations have been identified within the coding region of A␤ 1-42 , which can increase A␤ neurotoxicity (11). Transgenic (Tg) mouse models have shown that high levels of A␤ can cause AD-like amyloid plaque pathology. Tg mice that overexpress APP have some but not all deficits observed in AD (12, 13). Other Tg mouse models that have mutations associated with AD resulting in up-regulation of A␤ production have many, but not all, of the observed deficits seen in AD, including reduced hippocampal volume, reduced synap...

In recent years, several mouse models of amyotrophic lateral sclerosis (ALS) have been developed. One, caused by a G86R mutation in the superoxide dismutase‐1 (SOD‐1) gene associated with familial ALS, has been subjected to extensive quantitative analyses in the spinal cord. However, the human form of ALS includes pathology elsewhere in the nervous system. In the present study, analyses were extended to three motor nuclei in the brainstem. Mutant mice and control littermates were evaluated daily, and mutants, along with their littermate controls, were killed when they were severely affected. Brains were removed after perfusion and processed for Nissl staining, the samples were randomized, and the investigators were blinded to their genetic status. Stereologic methods were used to estimate the number of neurons, mean neuronal volumes, and nuclear volume in three brainstem motor nuclei known to be differetially involved in the human form of the disease, the oculomotor, facial, and hypoglossal nuclei. In the facial nucleus, neuron number consistently declined (48%), an effect that was correlated with disease severity. The nuclear volume of the facial nucleus was smaller in the SOD‐1 mutant mice (45.7% difference from control mice) and correlated significantly with neuron number. The oculomotor and hypoglossal nuclei showed less extreme involvement (<10% neuronal loss overall), with a trend toward fewer neurons in the hypoglossal nucleus of animals with severe facial nucleus involvement. In the oculomotor nucleus, neuronal loss was seen only once in five mice, associated with very severe disease. There was no significant change in the volume of individual neurons in any of these three nuclei in any transgenic mouse. These results suggest that different brainstem motor nuclei are differentially affected in this SOD‐1 mutant model of ALS. The relatively moderate and late involvement of the hypoglossal nucleus indicates that, although the general patterns of neuronal pathology match closely those seen in ALS patients, some differences exist in this transgenic model compared with the progression of the disease in humans. However, these patterns of cellular vulnerability may provide clues for understanding the differential susceptibility of neural structures in ALS and other neurodegenerative diseases. J. Comp. Neurol. 416:112–125, 2000. © 2000 Wiley‐Liss, Inc.

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Increasing evidence from mouse models of Alzheimer's disease shows that overexpression of a mutant form of the amyloid precursor protein (APP) and its product, ␤-amyloid peptide, initiate pathological changes before amyloid deposition. To evaluate the cytological basis for one of these early changes, namely reduced volume of the dentate gyrus (DG), we have used high-throughput diOlistic cell loading and 3D neuronal reconstruction to investigate potential dendritic pathology of granule cells (GCs) in 90-day-old PDAPP mice. Labeled GCs from fixed hippocampal slices were selected randomly and imaged digitally by using confocal laserscanning microscopy. The dendritic complexity of GCs was quantified according to subordinate morphological parameters, including soma position within the granule cell layer (superficial versus deep) and topographic location within the DG (dorsal versus ventral blade) along the anterior-posterior hippocampal axis. Initial analysis, which included all sampled GC types, revealed a 12% reduction of total dendritic length in PDAPP mice compared with littermate controls. Further analysis, performed with refined subgroups, found that superficially located GCs in the dorsal blade were profoundly altered, exhibiting a 23% loss in total dendritic length, whereas neurons in the ventral blade were unaffected. Superficial GCs were particularly vulnerable (a 32% reduction) in the posterior region of the DG. Furthermore, the dendritic reductions of this select group were uniformly localized within middleto-outer portions of the dentate molecular layer. We conclude that substantial dendritic pathology is evident in 90-day-old PDAPP mice for a spatially defined subset of GCs well before amyloid accumulation occurs.T he classical neuropathological hallmarks of Alzheimer's disease (AD) include the presence of neurofibrillary tangles within neurons and extracellular cerebrovascular, diffuse, and neuritic plaques (1). The key molecular constituent of the plaque is ␤-amyloid (A␤), a 39-to 43-amino acid amyloidogenic peptide, which is a product of the proteolytic processing of the amyloid precursor protein (APP) (2). It has been shown that familial APP mutations may facilitate AD-like neuropathological changes by accelerating aberrant APP proteolytic processing. This hypothesis has been reinforced by evidence from transgenic (Tg) mouse models of AD in which overexpression of mutant APP results in amyloid deposition (3-10). Neuritic dystrophy associated with the deposits of A␤ in AD brains is evidenced by progressive dendritic dystrophy within the hippocampal complex (11-17). Although the assumption has been that A␤ deposition leads to neuritic dystrophy that would impair hippocampal function (18,19), the causal links between the deposition and pathology have been largely inferential, with minimal data available on the time course of these events.Tg mouse models have provided a unique opportunity to characterize A␤-induced neuropathology, including the nature and time of onset of the physiological and morpholog...

The primate cingulate gyrus contains multiple cortical areas that can be distinguished by several neurochemical features, including the distribution of neurofilament protein-enriched pyramidal neurons. In addition, connectivity and functional properties indicate that there are multiple motor areas in the cortex lining the cingulate sulcus. These motor areas were targeted for analysis of potential interactions among regional specialization, connectivity, and cellular characteristics such as neurochemical profile and morphology. Specifically, intracortical injections of retrogradely transported dyes and intracellular injection were combined with immunocytochemistry to investigate neurons projecting from the cingulate motor areas to the putative forelimb region of the primary motor cortex, area M1. Two separate groups of neurons projecting to area M1 emanated from the cingulate sulcus, one anterior and one posterior, both of which furnished commissural and ipsilateral connections with area M1. The primary difference between the two populations was laminar origin, with the anterior projection originating largely in deep layers, and the posterior projection taking origin equally in superficial and deep layers. With regard to cellular morphology, the anterior projection exhibited more morphologic diversity than the posterior projection. Commissural projections from both anterior and posterior fields originated largely in layer VI. Neurofilament protein distribution was a reliable tool for localizing the two projections and for discriminating between them. Comparable proportions of the two sets of projection neurons contained neurofilament protein, although the density and distribution of the total population of neurofilament protein-enriched neurons was very different in the two subareas of origin. Within a projection, the participating neurons exhibited a high degree of morphologic heterogeneity, and no correlation was observed between somatodendritic morphology and neurofilament protein content. Thus, although the neurons that provide the anterior and posterior cingulate motor projections to area M1 differ morphologically and in laminar origin, their neurochemical profiles are similar with respect to neurofilament protein. This suggests that neurochemical phenotype may be a more important unifying feature for corticocortical projections than morphology.