Robert Vassar

Topographic organization of sensory projections to the olfactory bulb

The detection of odorant receptor mRNAs within the axon terminals of sensory neurons has permitted us to ask whether neurons expressing a given receptor project their axons to common glomeruli within the olfactory bulb. In situ hybridization with five different receptor probes demonstrates that axons from neurons expressing a given receptor converge on one, or at most, a few glomeruli within the olfactory bulb. Moreover, the position of specific glomeruli is bilaterally symmetric and is constant in different individuals within a species. These data support a model in which exposure to a given odorant may result in the stimulation of a spatially restricted set of glomeruli, such that the individual odorants would be associated with specific topographic patterns of activity within the olfactory bulb.

Intraneuronal β-Amyloid Aggregates, Neurodegeneration, and Neuron Loss in Transgenic Mice with Five Familial Alzheimer's Disease Mutations: Potential Factors in Amyloid Plaque Formation

Mutations in the genes for amyloid precursor protein (APP) and presenilins (PS1, PS2) increase production of β-amyloid 42 (Aβ42) and cause familial Alzheimers disease (FAD). Transgenic mice that express FAD mutant APP and PS1 overproduce Aβ42 and exhibit amyloid plaque pathology similar to that found in AD, but most transgenic models develop plaques slowly. To accelerate plaque development and investigate the effects of very high cerebral Aβ42 levels, we generated APP/PS1 double transgenic mice that coexpress five FAD mutations (5XFAD mice) and additively increase Aβ42 production. 5XFAD mice generate Aβ42 almost exclusively and rapidly accumulate massive cerebral Aβ42 levels. Amyloid deposition (and gliosis) begins at 2 months and reaches a very large burden, especially in subiculum and deep cortical layers. Intraneuronal Aβ42 accumulates in 5XFAD brain starting at 1.5 months of age (before plaques form), is aggregated (as determined by thioflavin S staining), and occurs within neuron soma and neurites. Some amyloid deposits originate within morphologically abnormal neuron soma that contain intraneuronal Aβ. Synaptic markers synaptophysin, syntaxin, and postsynaptic density-95 decrease with age in 5XFAD brain, and large pyramidal neurons in cortical layer 5 and subiculum are lost. In addition, levels of the activation subunit of cyclin-dependent kinase 5, p25, are elevated significantly at 9 months in 5XFAD brain, although an upward trend is observed by 3 months of age, before significant neurodegeneration or neuron loss. Finally, 5XFAD mice have impaired memory in the Y-maze. Thus, 5XFAD mice rapidly recapitulate major features of AD amyloid pathology and may be useful models of intraneuronal Aβ42-induced neurodegeneration and amyloid plaque formation.

Mice deficient in BACE1, the Alzheimer's β-secretase, have normal phenotype and abolished β-amyloid generation

Mice deficient in BACE1 (beta-site APP cleaving enzyme 1) are healthy, fertile and appear normal in gross anatomy, tissue histology, hematology and clinical chemistry. BACE1−/− mice also hemizygous for an amyloid precursor protein (APP) transgene lack brain β-amyloid (Aβ) and β-secretase-cleaved APP C-terminal fragments (CTFs). These results provide validation of BACE1 as the major β-secretase in vivo and suggest that therapeutic inhibition of BACE1 for the treatment of Alzheimers disease may be free of mechanism-based toxicity.

Spatial segregation of odorant receptor expression in the mammalian olfactory epithelium

The signal elicited by the interaction of odorous ligands with receptors on olfactory sensory neurons must be decoded by the brain to determine which of the numerous receptors have been activated. We have examined the patterns of odorant receptor expression in the rat olfactory epithelium to determine whether the mammalian olfactory system employs spatial segregation of sensory input to encode the identity of an odorant stimulus. In situ hybridization experiments with probes for 11 different odorant receptors demonstrate that sensory neurons expressing distinct receptors are topologically segregated into a small number of broad, yet circumscribed, zones within the olfactory epithelium. Within a given zone, however, olfactory neurons expressing a specific receptor appear to be randomly distributed, rather than spatially localized. The complex mammalian olfactory system may therefore compartmentalize the epithelium into anatomically and functionally discrete units, such that each zone expresses only a subset of the entire receptor repertoire.

The secretases: enzymes with therapeutic potential in Alzheimer disease

The amyloid hypothesis has yielded a series of well-validated candidate drug targets with potential for the treatment of Alzheimer disease (AD). Three proteases that are involved in the processing of amyloid precursor protein—α-secretase, β-secretase and γ-secretase—are of particular interest as they are central to the generation and modulation of amyloid-β peptide and can be targeted by small compounds in vitro and in vivo. Given that these proteases also fulfill other important biological roles, inhibiting their activity will clearly be inherently associated with mechanism-based toxicity. Carefully determining a suitable therapeutic window and optimizing the selectivity of the drug treatments towards amyloid precursor protein processing might be ways of overcoming this potential complication. Secretase inhibitors are likely to be the first small-molecule therapies aimed at AD modification that will be fully tested in the clinic. Success or failure of these first-generation AD therapies will have enormous consequences for further drug development efforts for AD and possibly other neurodegenerative conditions.

BACE1 Deficiency Rescues Memory Deficits and Cholinergic Dysfunction in a Mouse Model of Alzheimer's Disease

beta-site APP cleaving enzyme 1 (BACE1) is the beta-secretase enzyme required for generating pathogenic beta-amyloid (Abeta) peptides in Alzheimers disease (AD). BACE1 knockout mice lack Abeta and are phenotypically normal, suggesting that therapeutic inhibition of BACE1 may be free of mechanism-based side effects. However, direct evidence that BACE1 inhibition would improve cognition is lacking. Here we show that BACE1 null mice engineered to overexpress human APP (BACE1(-/-).Tg2576(+)) are rescued from Abeta-dependent hippocampal memory deficits. Moreover, impaired hippocampal cholinergic regulation of neuronal excitability found in the Tg2576 AD model is ameliorated in BACE1(-/-).Tg2576(+) bigenic mice. The behavioral and electrophysiological rescue of deficits in BACE1(-/-).Tg2576(+) mice is correlated with a dramatic reduction of cerebral Abeta40 and Abeta42 levels and occurs before amyloid deposition in Tg2576 mice. Our gene-based approach demonstrates that lower Abeta levels are beneficial for AD-associated memory impairments, validating BACE1 as a therapeutic target for AD.

The β-Secretase Enzyme BACE in Health and Alzheimer's Disease: Regulation, Cell Biology, Function, and Therapeutic Potential

The β-amyloid (Aβ) peptide is the major constituent of amyloid plaques in Alzheimers disease (AD) brain and is likely to play a central role in the pathogenesis of this devastating neurodegenerative disorder. The β-secretase, β-site amyloid precursor protein cleaving enzyme (BACE1; also called Asp2, memapsin 2), is the enzyme responsible for initiating Aβ generation. Thus, BACE is a prime drug target for the therapeutic inhibition of Aβ production in AD. Since its discovery 10 years ago, much has been learned about BACE. This review summarizes BACE properties, describes BACE translation dysregulation in AD, and discusses BACE physiological functions in sodium current, synaptic transmission, myelination, and schizophrenia. The therapeutic potential of BACE will also be considered. This is a summary of topics covered at a symposium held at the 39th annual meeting of the Society for Neuroscience and is not meant to be a comprehensive review of BACE.

Mutant keratin expression in transgenic mice causes marked abnormalities resembling a human genetic skin disease

To explore the relationship between keratin gene mutations and genetic disease, we made transgenic mice expressing a mutant keratin in the basal layer of their stratified squamous epithelia. These mice exhibited abnormalities in epidermal architecture and often died prematurely. Blistering occurred easily, and basal cell cytolysis was evidence at the light and electron microscopy levels. Keratin filament formation was markedly altered, with keratin aggregates in basal cells. In contrast, terminally differentiating cells made keratin filaments and formed a stratum corneum. Recovery of outer layer cells was attributed to down-regulation of mutant keratin expression and concomitant induction of differentiation-specific keratins as cells terminally differentiate, and the fact that these cells arose from basal cells developing at a time when keratin expression was relatively low. Collectively, the pathobiology and biochemistry of the transgenic mice and their cultured keratinocytes bore a resemblance to a group of genetic disorders known as epidermolysis bullosa simplex.

Phosphorylation of the Translation Initiation Factor eIF2α Increases BACE1 Levels and Promotes Amyloidogenesis

beta-site APP cleaving enzyme-1 (BACE1), the rate-limiting enzyme for beta-amyloid (Abeta) production, is elevated in Alzheimers disease (AD). Here, we show that energy deprivation induces phosphorylation of the translation initiation factor eIF2alpha (eIF2alpha-P), which increases the translation of BACE1. Salubrinal, an inhibitor of eIF2alpha-P phosphatase PP1c, directly increases BACE1 and elevates Abeta production in primary neurons. Preventing eIF2alpha phosphorylation by transfection with constitutively active PP1c regulatory subunit, dominant-negative eIF2alpha kinase PERK, or PERK inhibitor P58(IPK) blocks the energy-deprivation-induced BACE1 increase. Furthermore, chronic treatment of aged Tg2576 mice with energy inhibitors increases levels of eIF2alpha-P, BACE1, Abeta, and amyloid plaques. Importantly, eIF2alpha-P and BACE1 are elevated in aggressive plaque-forming 5XFAD transgenic mice, and BACE1, eIF2alpha-P, and amyloid load are correlated in humans with AD. These results strongly suggest that eIF2alpha phosphorylation increases BACE1 levels and causes Abeta overproduction, which could be an early, initiating molecular mechanism in sporadic AD.

Aβ-Generating Enzymes: Recent Advances in β- and γ-Secretase Research

is higher in neurons of the brain. (2) BACE is localized within acidic intracellular compartments. (3) Overex-pression of BACE in cells increases ␤-secretase cleav-age products; these products start only at known ␤-secre-tase cleavage sites. (4) Antisense inhibition of BACE in cells decreases ␤-secretase cleavage. (5) Purified forms of BACE cleave APP substrates in vitro with correct ␤-secretase specificity. (6) BACE has an acidic pH opti-Alzheimers Disease (AD) is characterized by two hallmark mum and is not inhibited by the common aspartic prote-lesions in the brain: the extracellular amyloid plaques, pri-ase inhibitor pepstatin. Taken together, all the properties marily composed of the 40-42 amino acid A␤ peptide, and of BACE match one-to-one with those of ␤-secretase. the intracellular neurofibrillary tangles made of abnormally BACE2 phosphorylated tau, a microtubule-associated protein. Al-Soon after the discovery of BACE, a homologous gene, though the cause of AD is controversial, most evidence BACE2 (also called Asp1 or Memapsin1), was identified supports a central role for A␤ in the pathogenesis of the by searching genome databases (Saunders et al., 1999; disease (for review, see Selkoe, 1999). Bennett et al., 2000). BACE and BACE2 share 64% amino A␤ is generated by endoproteolytic processing of the acid similarity and both have a C-terminal transmem-large type I transmembrane protein, amyloid precursor brane domain. However, BACE and BACE2 are only protein (APP; Figure 1). Enzymes called ␤-and ␥-secre-‫%04ف‬ similar to other aspartic proteases in the pepsin tase cleave APP to form the N and C termini, respec-family, suggesting that BACE and BACE2 define a novel tively, of the A␤ peptide. ␤-secretase is the rate-limiting family (Figure 2). enzyme in the production of A␤ and cleaves APP first The chromosomal localization of the BACE gene is to form the membrane-bound C99 fragment, which in 11q23.3, a locus not associated with AD, while that of turn is the substrate of ␥-secretase (Figure 1). Clearly, the BACE2 gene is chromosome 21 within the critical the ␤-and ␥-secretases are excellent therapeutic tar-Downs syndrome region (Saunders et al., 1999). There-gets, and major efforts to identify these enzymes have fore, Downs patients, who all develop AD by middle been pursued for over 12 years. Last fall, the long-sought age, have three copies each of the APP gene (also on ␤-secretase was identified as the novel transmembrane chromosome 21) and the BACE2 gene. This observation aspartic protease BACE (for beta-site APP cleaving en-suggested that BACE2 may be involved …