Hypothalamic-pituitary-adrenal (HPA) axis activity and anxiety-like behavior during aging: A test of the glucocorticoid cascade hypothesis in amyloidogenic APPswe/PS1dE9 mice.

Alzheimer's disease (AD) is a progressive, dementing, whole-body disorder that presents with decline in cognitive, behavioral, and emotional functions, as well as endocrine dysregulation. The etiology of AD is not fully understood but stress- and anxiety-related hormones may play a role in its development and trajectory. The glucocorticoid cascade hypothesis posits that levels of glucocorticoids increase with age, leading to dysregulated negative feedback, further elevated glucocorticoids, and resulting neuropathology. We examined the impact of age (from 2 to 10 months) and stressor exposure (predator odor) on hormone levels (corticosterone and ghrelin), anxiety-like behavior (open field and light dark tests), and memory-related behavior (novel object recognition; NOR), and whether these various measures correlated with neuropathology (hippocampus and cortex amyloid beta, Aß) in male and female APPswe/PS1dE9 transgenic and non-transgenic mice. Additionally, we performed exploratory analyses to probe if the open field and light dark test as commonly used tasks to assess anxiety levels were correlated. Consistent with the glucocorticoid cascade hypothesis, baseline corticosterone increased with age. Predator odor exposure elevated corticosterone at each age, but in contrast to the glucocorticoid cascade hypothesis, the magnitude of stressor-induced elevations in corticosterone levels did not increase with age. Overall, transgenic mice had higher post-stressor, but not baseline, corticosterone than non-transgenic mice, and across both genotypes, females consistently had higher (baseline and post-stressor) corticosterone than males. Behavior in the open field test primarily showed decreased locomotion with age, and this was pronounced in transgenic females. Anxiety-like behaviors in the light dark test were exacerbated following predator odor, and female transgenic mice were the most impacted. Compared to transgenic males, transgenic females had higher Aß concentrations and showed more anxiety-like behavior. Performance on the NOR did not differ significantly between genotypes. Lastly, we did not find robust, statistically significant correlations among corticosterone, ghrelin, recognition memory, anxiety-like behaviors, or Aß, suggesting outcomes are not strongly related on the individual level. Our data suggest that despite Aß accumulation in the hippocampus and cortex, male and female APPswePS1dE9 transgenic mice do not differ robustly from their non-transgenic littermates in physiological, endocrine, and behavioral measures at the range of ages studied here.

PET imaging of microglia by targeting macrophage colony-stimulating factor 1 receptor (CSF1R).

While neuroinflammation is an evolving concept and the cells involved and their functions are being defined, microglia are understood to be a key cellular mediator of brain injury and repair. The ability to measure microglial activity specifically and noninvasively would be a boon to the study of neuroinflammation, which is involved in a wide variety of neuropsychiatric disorders including traumatic brain injury, demyelinating disease, Alzheimer's disease (AD), and Parkinson's disease, among others. We have developed [11C]CPPC [5-cyano-N-(4-(4-[11C]methylpiperazin-1-yl)-2-(piperidin-1-yl)phenyl)furan-2-carboxamide], a positron-emitting, high-affinity ligand that is specific for the macrophage colony-stimulating factor 1 receptor (CSF1R), the expression of which is essentially restricted to microglia within brain. [11C]CPPC demonstrates high and specific brain uptake in a murine and nonhuman primate lipopolysaccharide model of neuroinflammation. It also shows specific and elevated uptake in a murine model of AD, experimental allergic encephalomyelitis murine model of demyelination and in postmortem brain tissue of patients with AD. Radiation dosimetry in mice indicated [11C]CPPC to be safe for future human studies. [11C]CPPC can be synthesized in sufficient radiochemical yield, purity, and specific radioactivity and possesses binding specificity in relevant models that indicate potential for human PET imaging of CSF1R and the microglial component of neuroinflammation.

Tau is required for progressive synaptic and memory deficits in a transgenic mouse model of α-synucleinopathy.

Parkinson's disease dementia (PDD) and dementia with Lewy bodies (DLB) are clinically and neuropathologically highly related α-synucleinopathies that collectively constitute the second leading cause of neurodegenerative dementias. Genetic and neuropathological studies directly implicate α-synuclein (αS) abnormalities in PDD and DLB pathogenesis. However, it is currently unknown how αS abnormalities contribute to memory loss, particularly since forebrain neuronal loss in PDD and DLB is less severe than in Alzheimer's disease. Previously, we found that familial Parkinson's disease-linked human mutant A53T αS causes aberrant localization of the microtubule-associated protein tau to postsynaptic spines in neurons, leading to postsynaptic deficits. Thus, we directly tested if the synaptic and memory deficits in a mouse model of α-synucleinopathy (TgA53T) are mediated by tau. TgA53T mice exhibit progressive memory deficits associated with postsynaptic deficits in the absence of obvious neuropathological and neurodegenerative changes in the hippocampus. Significantly, removal of endogenous mouse tau expression in TgA53T mice (TgA53T/mTau-/-), achieved by mating TgA53T mice to mouse tau-knockout mice, completely ameliorates cognitive dysfunction and concurrent synaptic deficits without affecting αS expression or accumulation of selected toxic αS oligomers. Among the known tau-dependent effects, memory deficits in TgA53T mice were associated with hippocampal circuit remodeling linked to chronic network hyperexcitability. This remodeling was absent in TgA53T/mTau-/- mice, indicating that postsynaptic deficits, aberrant network hyperactivity, and memory deficits are mechanistically linked. Our results directly implicate tau as a mediator of specific human mutant A53T αS-mediated abnormalities related to deficits in hippocampal neurotransmission and suggest a mechanism for memory impairment that occurs as a consequence of synaptic dysfunction rather than synaptic or neuronal loss. We hypothesize that these initial synaptic deficits contribute to network hyperexcitability which, in turn, exacerbate cognitive dysfunction. Our results indicate that these synaptic changes present potential therapeutic targets for amelioration of memory deficits in α-synucleinopathies.

Association between serotonin denervation and resting-state functional connectivity in mild cognitive impairment.

Resting-state functional connectivity alterations have been demonstrated in Alzheimer's disease (AD) and mild cognitive impairment (MCI) before the observation of AD neuropathology, but mechanisms driving these changes are not well understood. Serotonin neurodegeneration has been observed in MCI and AD and is associated with cognitive deficits and neuropsychiatric symptoms, but the role of the serotonin system in relation to brain network dysfunction has not been a major focus of investigation. The current study investigated the relationship between serotonin transporter availability (SERT; measured using positron emission tomography) and brain network functional connectivity (measured using resting-state functional MRI) in 20 participants with MCI and 21 healthy controls. Two SERT regions of interest were selected for the analysis: the Dorsal Raphe Nuclei (DRN) and the precuneus which represent the cell bodies of origin and a cortical target of projections of the serotonin system, respectively. Both regions show decreased SERT in MCI compared to controls and are the site of early AD pathology. Average resting-state functional connectivity did not differ between MCI and controls. Decreased SERT in DRN was associated with lower hippocampal resting-state connectivity in MCI participants compared to controls. Decreased SERT in the right precuneus was also associated with lower resting-state connectivity of the retrosplenial cortex to the dorsal lateral prefrontal cortex and higher resting-state connectivity of the retrosplenial cortex to the posterior cingulate and in patients with MCI but not in controls. These results suggest that a serotonergic mechanism may underlie changes in brain functional connectivity in MCI. Hum Brain Mapp 38:3391-3401, 2017. © 2017 Wiley Periodicals, Inc.

Sex-related dimorphism in dentate gyrus atrophy and behavioral phenotypes in an inducible tTa:APPsi transgenic model of Alzheimer's disease.

Sex differences are a well-known phenomenon in Alzheimer's disease (AD), with women having a higher risk for AD than men. Many AD mouse models display a similar sex-dependent pattern, with females showing earlier cognitive deficits and more severe neuropathology than males. However, whether those differences are relevant to human disease is unclear. Here we show that in AD mouse models that overexpress amyloid precursor protein (APP) under control of the prion protein promoter (PrP), female transgenic mice have higher APP expression than males, complicating interpretations of the role of sex-related factors in such models. By contrast, in a tTa:APPsi model, in which APP expression is driven by the tetracycline transactivator (tTa) from the CaMKIIα promoter, there are no sex-related differences in expression or processing of APP. In addition, the levels of Aß dimers and tetramers, as well as Aß peptide accumulation, are similar between sexes. Behavioral testing demonstrated that both male and female tTa:APPsi mice develop age-dependent deficits in spatial recognition memory and conditional freezing to context. These cognitive deficits were accompanied by habituation-associated hyperlocomotion and startle hyper-reactivity. Significant sex-related dimorphisms were observed, due to females showing earlier onsets of the deficits in conditioned freezing and hyperlocomotion. In addition, tTa:APPsi males but not females demonstrated a lack of novelty-induced activation. Both males and females showed atrophy of the dentate gyrus (DG) of the dorsal hippocampus, associated with widening of the pyramidal layer of the CA1 area in both sexes. Ventral DG was preserved. Sex-related differences were limited to the DG, with females showing more advanced degeneration than males. Collectively, our data show that the tTa:APPsi model is characterized by a lack of sex-related differences in APP expression, making this model useful in deciphering the mechanisms of sex differences in AD pathogenesis. Sex-related dimorphisms observed in this model under conditions of equal APP expression between sexes suggest a higher sensitivity of females to the effects of APP and/or Aß production.

Acute stressors do not impair short-term memory or attention in an aged mouse model of amyloidosis.

Memory impairment in Alzheimer's disease patients is thought to be associated with the accumulation of amyloid-beta peptides and tau proteins. However, inconsistent reports of cognitive deficits in pre-clinical studies have raised questions about the link between amyloid-beta and cognitive decline. One possible explanation may be that studies reporting memory deficits often involve behavioral assessments that entail a high stress component. In contrast, in tasks without a high stress component transgenic mice do not consistently show declines in memory. The glucocorticoid cascade hypothesis of aging and the vicious cycle of stress framework suggest that stress exacerbates dementia progression by initiating a cycle of hypothalamic-pituitary-adrenal axis activation and subsequent brain deterioration. Using the APPswe/PS1dE9 mouse model of amyloidosis, we assessed whether stressor exposure prior to testing differentially impaired cognitive performance of aged male and female mice. As part of a larger study, mice performed a delayed match-to-position (DMTP) or a 3-choice serial-reaction time (3CSRT) task. Unexpectedly, these mice did not exhibit cognitive declines during aging. Therefore, at 73 and 74 weeks of age, we exposed mice to a predator odor or forced swim stressor prior to testing to determine if stress revealed cognitive deficits. We predicted stressor exposure would decrease performance accuracy more robustly in transgenic vs. non-transgenic mice. Acute stressor exposure increased accuracy in the DMTP task, but not in the 3CSRT task. Our data suggest that acute stressor exposure prior to testing does not impair cognitive performance in APPswe/PS1dE9 mice.

NMDA Receptor-Arc Signaling Is Required for Memory Updating and Is Disrupted in Alzheimer's Disease.

BACKGROUND: Memory deficits are central to many neuropsychiatric diseases. During acquisition of new information, memories can become vulnerable to interference, yet mechanisms that underlie interference are unknown. METHODS: We describe a novel transduction pathway that links the NMDA receptor (NMDAR) to AKT signaling via the immediate early gene Arc and evaluate its role in memory. The signaling pathway is validated using biochemical tools and transgenic mice, and function is evaluated in assays of synaptic plasticity and behavior. The translational relevance is evaluated in human postmortem brain. RESULTS: Arc is dynamically phosphorylated by CaMKII (calcium/calmodulin-dependent protein kinase II) and binds the NMDAR subunits NR2A/NR2B and a previously unstudied PI3K (phosphoinositide 3-kinase) adapter p55PIK (PIK3R3) in vivo in response to novelty or tetanic stimulation in acute slices. NMDAR-Arc-p55PIK recruits p110α PI3K and mTORC2 (mechanistic target of rapamycin complex 2) to activate AKT. NMDAR-Arc-p55PIK-PI3K-mTORC2-AKT assembly occurs within minutes of exploratory behavior and localizes to sparse synapses throughout hippocampal and cortical regions. Studies using conditional (Nestin-Cre) p55PIK deletion mice indicate that NMDAR-Arc-p55PIK-PI3K-mTORC2-AKT functions to inhibit GSK3 and mediates input-specific metaplasticity that protects potentiated synapses from subsequent depotentiation. p55PIK conditional knockout mice perform normally in multiple behaviors including working memory and long-term memory tasks but exhibit deficits indicative of increased vulnerability to interference in both short-term and long-term paradigms. The NMDAR-AKT transduction complex is reduced in postmortem brain of individuals with early Alzheimer's disease. CONCLUSIONS: A novel function of Arc mediates synapse-specific NMDAR-AKT signaling and metaplasticity that contributes to memory updating and is disrupted in human cognitive disease.

Longitudinal assessment of cognitive function in the APPswe/PS1dE9 mouse model of Alzheimer's-related beta-amyloidosis.

Preclinical models of Alzheimer's disease (AD)-related cognitive decline can be useful for developing therapeutics. The current study longitudinally assessed short-term memory, using a delayed matching-to-position (DMTP) task, and attention, using a 3-choice serial reaction time (3CSRT) task, from approximately 18 weeks of age through death or 72 weeks of age in APPswe/PS1dE9 mice, a widely used mouse model of AD-related amyloidosis. Both transgenic (Tg) and non-Tg mice exhibited improvements in DMTP accuracy over time. Breaks in testing reduced DMTP accuracy but accuracy values quickly recovered in both Tg and non-Tg mice. Both Tg and non-Tg mice exhibited high accuracy in the 3CSRT task with breaks in testing briefly reducing accuracy values equivalently in the 2 genotypes. The current results raise the possibility that deficits in Tg APPswe/PS1dE9 mice involve impairments in learning rather than declines in established performances. A better understanding of the factors that determine whether deficits develop will be useful for designing evaluations of potential pharmacotherapeutics and may reveal interventions for clinical application.

Synthesis and biodistribution of [11C]A-836339, a new potential radioligand for PET imaging of cannabinoid type 2 receptors (CB2).

Recently, A-836339 [2,2,3,3-tetramethylcyclopropanecarboxylic acid [3-(2-methoxyethyl)-4,5-dimethyl-3H-thiazol-(2Z)-ylidene]amide] (1) was reported to be a selective CB2 agonist with high binding affinity. Here we describe the radiosynthesis of [11C]A-836339 ([11C]1) via its desmethyl precursor as a candidate radioligand for imaging CB2 receptors with positron-emission tomography (PET). Whole body and the regional brain distribution of [11C]1 in control CD1 mice demonstrated that this radioligand exhibits specific uptake in the CB2-rich spleen and little specific in vivo binding in the control mouse brain. However, [11C]1 shows specific cerebral uptake in the lipopolysaccharide (LPS)-induced mouse model of neuroinflammation and in the brain areas with Abeta amyloid plaque deposition in a mouse model of Alzheimer's disease (APPswe/PS1dE9 mice). These data establish a proof of principle that CB2 receptors binding in the neuroinflammation and related disorders can be measured in vivo.

Increased Sparsity of Hippocampal CA1 Neuronal Ensembles in a Mouse Model of Down Syndrome Assayed by Arc Expression.

Down syndrome (DS) is the leading chromosomal cause of intellectual disability, yet the neural substrates of learning and memory deficits remain poorly understood. Here, we interrogate neural networks linked to learning and memory in a well-characterized model of DS, the Ts65Dn mouse. We report that Ts65Dn mice exhibit exploratory behavior that is not different from littermate wild-type (WT) controls yet behavioral activation of Arc mRNA transcription in pyramidal neurons of the CA1 region of the hippocampus is altered in Ts65Dn mice. In WT mice, a 5 min period of exploration of a novel environment resulted in Arc mRNA transcription in 39% of CA1 neurons. By contrast, the same period of exploration resulted in only ~20% of CA1 neurons transcribing Arc mRNA in Ts65Dn mice indicating increased sparsity of the behaviorally induced ensemble. Like WT mice the CA1 pyramidal neurons of Ts65Dn mice reactivated Arc transcription during a second exposure to the same environment 20 min after the first experience, but the size of the reactivated ensemble was only ~60% of that in WT mice. After repeated daily exposures there was a further decline in the size of the reactivated ensemble in Ts65Dn and a disruption of reactivation. Together these data demonstrate reduction in the size of the behaviorally induced network that expresses Arc in Ts65Dn mice and disruption of the long-term stability of the ensemble. We propose that these deficits in network formation and stability contribute to cognitive symptoms in DS.

Intranasal post-cardiac arrest treatment with orexin-A facilitates arousal from coma and ameliorates neuroinflammation.

Cardiac arrest (CA) entails significant risks of coma resulting in poor neurological and behavioral outcomes after resuscitation. Significant subsequent morbidity and mortality in post-CA patients are largely due to the cerebral and cardiac dysfunction that accompanies prolonged whole-body ischemia post-CA syndrome (PCAS). PCAS results in strong inflammatory responses including neuroinflammation response leading to poor outcome. Currently, there are no proven neuroprotective therapies to improve post-CA outcomes apart from therapeutic hypothermia. Furthermore, there are no acceptable approaches to promote cortical or cognitive arousal following successful return of spontaneous circulation (ROSC). Hypothalamic orexinergic pathway is responsible for arousal and it is negatively affected by neuroinflammation. However, whether activation of the orexinergic pathway can curtail neuroinflammation is unknown. We hypothesize that targeting the orexinergic pathway via intranasal orexin-A (ORXA) treatment will enhance arousal from coma and decrease the production of proinflammatory cytokines resulting in improved functional outcome after resuscitation. We used a highly validated CA rat model to determine the effects of intranasal ORXA treatment 30-minute post resuscitation. At 4hrs post-CA, the mRNA levels of proinflammatory markers (IL1ß, iNOS, TNF-α, GFAP, CD11b) and orexin receptors (ORX1R and ORX2R) were examined in different brain regions. CA dramatically increased proinflammatory markers in all brain regions particularly in the prefrontal cortex, hippocampus and hypothalamus. Post-CA intranasal ORXA treatment significantly ameliorated the CA-induced neuroinflammatory markers in the hypothalamus. ORXA administration increased production of orexin receptors (ORX1R and ORX2R) particularly in hypothalamus. In addition, ORXA also resulted in early arousal as measured by quantitative electroencephalogram (EEG) markers, and recovery of the associated behavioral neurologic deficit scale score (NDS). Our results indicate that intranasal delivery of ORXA post-CA has an anti-inflammatory effect and accelerates cortical EEG and behavioral recovery. Beneficial outcomes from intranasal ORXA treatment lay the groundwork for therapeutic clinical approach to treating post-CA coma.

Environmental enrichment mitigates cognitive deficits in a mouse model of Alzheimer's disease.

Epidemiological studies suggest that individuals with greater education or more cognitively demanding occupations have diminished risk of developing dementia. We wanted to test whether this effect could be recapitulated in rodents using environmental enrichment, a paradigm well documented to attenuate behavioral deficits induced by various pathological insults. Here, we demonstrate that learning and memory deficits observed in a transgenic mouse model of Alzheimer's disease can be ameliorated by enrichment. Female transgenic mice overexpressing amyloid precursor protein and/or presenilin-1 and nontransgenic controls were placed into enriched or standard cages at 2 months of age and tested for cognitive behavior after 6 months of differential housing. Enrichment significantly improved performance of all genotypes in the radial water maze and in the classic and repeated-reversal versions of the Morris water maze. However, enrichment did not benefit all genotypes equally. Mice overproducing amyloid-beta (Abeta), particularly those with amyloid deposits, showed weaker memory for the platform location in the classic Morris water maze and learned new platform positions in the repeated-reversals task less quickly than their nontransgenic cagemates. Nonetheless, enrichment normalized the performance of Abeta-overproducing mice to the level of standard-housed nontransgenic mice. Moreover, this functional preservation occurred despite increased neuritic plaque burden in the hippocampus of double-transgenic animals and elevated steady-state Abeta levels, because both endogenous and transgene-derived Abeta are increased in enriched animals. These results demonstrate that the generation of Abeta in vivo and its impact on the function of the nervous system can be strongly modulated by environmental factors.

BACE1, a major determinant of selective vulnerability of the brain to amyloid-beta amyloidogenesis, is essential for cognitive, emotional, and synaptic functions.

A transmembrane aspartyl protease termed beta-site APP cleavage enzyme 1 (BACE1) that cleaves the amyloid-beta precursor protein (APP), which is abundant in neurons, is required for the generation of amyloid-beta (Abeta) peptides implicated in the pathogenesis of Alzheimer's disease (AD). We now demonstrate that BACE1, enriched in neurons of the CNS, is a major determinant that predisposes the brain to Abeta amyloidogenesis. The physiologically high levels of BACE1 activity coupled with low levels of BACE2 and alpha-secretase anti-amyloidogenic activities in neurons is a major contributor to the accumulation of Abeta in the CNS, whereas other organs are spared. Significantly, deletion of BACE1 in APPswe;PS1DeltaE9 mice prevents both Abeta deposition and age-associated cognitive abnormalities that occur in this model of Abeta amyloidosis. Moreover, Abeta deposits are sensitive to BACE1 dosage and can be efficiently cleared from the CNS when BACE1 is silenced. However, BACE1 null mice manifest alterations in hippocampal synaptic plasticity as well as in performance on tests of cognition and emotion. Importantly, memory deficits but not emotional alterations in BACE1(-/-) mice are prevented by coexpressing APPswe;PS1DeltaE9 transgenes, indicating that other potential substrates of BACE1 may affect neural circuits related to emotion. Our results establish BACE1 and APP processing pathways as critical for cognitive, emotional, and synaptic functions, and future studies should be alert to potential mechanism-based side effects that may occur with BACE1 inhibitors designed to ameliorate Abeta amyloidosis in AD.

Effectiveness of estrogen replacement in restoration of cognitive function after long-term estrogen withdrawal in aging rats.

Recent studies suggest that some aspects of learning and memory may be altered by a midlife loss of estrogen, indicating a potential causal relationship between the deficiency of ovarian hormones and cognitive aging. In this study, the effects of estrogen withdrawal and replacement were tested in middle-aged Fischer-344 rats using different memory tasks. Estrogen withdrawal accelerated the rate of cognitive aging. A deficit first occurred 4 months after ovariectomy in working memory, which was tested in a delayed-nonmatching-to-position task, and progressed from long-delay to short-delay trials. Reference memory, which was tested in a place discrimination task and a split-stem T-maze, was not affected by aging or ovariectomy. The efficacy of estrogen in ameliorating the cognitive deficit in old rats depended on the type of treatment (acute vs chronic) and whether the aging-related decline in a particular cognitive process was aggravated by estrogen withdrawal. Chronic estrogen treatment (implants) was effective in improving working memory only when primed with repeated injections of estrogen, indicating that simulating the estrogen fluctuations of the estrous cycle may be more effective than the widely used mode of chronic pharmacological treatment. A challenge with scopolamine revealed that ovariectomy-induced cognitive deterioration coincided with a compromised cholinergic system. Importantly, the estrogen treatment that had restored effectively the cognitive abilities of old ovariectomized rats did not reduce their sensitivity to scopolamine. Taking into consideration that estrogen was highly effective against the amnestic action of scopolamine when tested in young-adult rats, these data emphasize that mechanisms of the protective effect of estrogen differ in young and old rats.

Persistent amyloidosis following suppression of Abeta production in a transgenic model of Alzheimer disease.

BACKGROUND: The proteases (secretases) that cleave amyloid-beta (Abeta) peptide from the amyloid precursor protein (APP) have been the focus of considerable investigation in the development of treatments for Alzheimer disease. The prediction has been that reducing Abeta production in the brain, even after the onset of clinical symptoms and the development of associated pathology, will facilitate the repair of damaged tissue and removal of amyloid lesions. However, no long-term studies using animal models of amyloid pathology have yet been performed to test this hypothesis. METHODS AND FINDINGS: We have generated a transgenic mouse model that genetically mimics the arrest of Abeta production expected from treatment with secretase inhibitors. These mice overexpress mutant APP from a vector that can be regulated by doxycycline. Under normal conditions, high-level expression of APP quickly induces fulminant amyloid pathology. We show that doxycycline administration inhibits transgenic APP expression by greater than 95% and reduces Abeta production to levels found in nontransgenic mice. Suppression of transgenic Abeta synthesis in this model abruptly halts the progression of amyloid pathology. However, formation and disaggregation of amyloid deposits appear to be in disequilibrium as the plaques require far longer to disperse than to assemble. Mice in which APP synthesis was suppressed for as long as 6 mo after the formation of Abeta deposits retain a considerable amyloid load, with little sign of active clearance. CONCLUSION: This study demonstrates that amyloid lesions in transgenic mice are highly stable structures in vivo that are slow to disaggregate. Our findings suggest that arresting Abeta production in patients with Alzheimer disease should halt the progression of pathology, but that early treatment may be imperative, as it appears that amyloid deposits, once formed, will require additional intervention to clear.

Cannabinoid CB2 Receptors in a Mouse Model of Aß Amyloidosis: Immunohistochemical Analysis and Suitability as a PET Biomarker of Neuroinflammation.

In Alzheimer's disease (AD), one of the early responses to Aß amyloidosis is recruitment of microglia to areas of new plaque. Microglial receptors such as cannabinoid receptor 2 (CB2) might be a suitable target for development of PET radiotracers that could serve as imaging biomarkers of Aß-induced neuroinflammation. Mouse models of amyloidosis (J20APPswe/ind and APPswe/PS1ΔE9) were used to investigate the cellular distribution of CB2 receptors. Specificity of CB2 antibody (H60) was confirmed using J20APPswe/ind mice lacking CB2 receptors. APPswe/PS1ΔE9 mice were used in small animal PET with a CB2-targeting radiotracer, [11C]A836339. These studies revealed increased binding of [11C]A836339 in amyloid-bearing mice. Specificity of the PET signal was confirmed in a blockade study with a specific CB2 antagonist, AM630. Confocal microscopy revealed that CB2-receptor immunoreactivity was associated with astroglial (GFAP) and, predominantly, microglial (CD68) markers. CB2 receptors were observed, in particular, in microglial processes forming engulfment synapses with Aß plaques. In contrast to glial cells, neuron (NeuN)-derived CB2 signal was equal between amyloid-bearing and control mice. The pattern of neuronal CB2 staining in amyloid-bearing mice was similar to that in human cases of AD. The data collected in this study indicate that Aß amyloidosis without concomitant tau pathology is sufficient to activate CB2 receptors that are suitable as an imaging biomarker of neuroinflammation. The main source of enhanced CB2 PET binding in amyloid-bearing mice is increased CB2 immunoreactivity in activated microglia. The presence of CB2 immunoreactivity in neurons does not likely contribute to the enhanced CB2 PET signal in amyloid-bearing mice due to a lack of significant neuronal loss in this model. However, significant loss of neurons as seen at late stages of AD might decrease the CB2 PET signal due to loss of neuronally-derived CB2. Thus this study in mouse models of AD indicates that a CB2-specific radiotracer can be used as a biomarker of neuroinflammation in the early preclinical stages of AD, when no significant neuronal loss has yet developed.

Treatment with bexarotene, a compound that increases apolipoprotein-E, provides no cognitive benefit in mutant APP/PS1 mice.

BACKGROUND: Though the precise cause(s) of Alzheimer's disease (AD) remain unknown, there is strong evidence that decreased clearance of ß-amyloid (Aß) from the brain can contribute to the disease. Therapeutic strategies to promote natural Aß clearance mechanisms, such as the protein apolipoprotein-E (APOE), hold promise for the treatment of AD. The amount of APOE in the brain is regulated by nuclear receptors including retinoid X receptors (RXRs). Drugs that activate RXRs, including bexarotene, can increase APOE and ABCA1 production, and have been shown to decrease the Aß burden and improve cognition in mouse models of Aß amyloidosis. Although recent bexarotene studies failed to replicate the rapid clearance of Aß from brains, behavioral and cognitive effects of this compound remain controversial. FINDINGS: In efforts to clarify these behavioral findings, mutant APP/PS1 mice were acutely dosed with bexarotene. While ABCA1 was upregulated in mutant APP/PS1 mice treated with bexarotene, this drug failed to attenuate Aß plaques or cognitive deficits in these mice. CONCLUSIONS: We recommend rigorous preclinical study to evaluate the mechanism and utility of such a compound for AD therapy.

Alzheimer's therapeutics: translation of preclinical science to clinical drug development.

Over the past three decades, significant progress has been made in understanding the neurobiology of Alzheimer's disease. In recent years, the first attempts to implement novel mechanism-based treatments brought rather disappointing results, with low, if any, drug efficacy and significant side effects. A discrepancy between our expectations based on preclinical models and the results of clinical trials calls for a revision of our theoretical views and questions every stage of translation-from how we model the disease to how we run clinical trials. In the following sections, we will use some specific examples of the therapeutics from acetylcholinesterase inhibitors to recent anti-Aß immunization and γ-secretase inhibition to discuss whether preclinical studies could predict the limitations in efficacy and side effects that we were so disappointed to observe in recent clinical trials. We discuss ways to improve both the predictive validity of mouse models and the translation of knowledge between preclinical and clinical stages of drug development.

Partial depletion of CREB-binding protein reduces life expectancy in a mouse model of Huntington disease.

Previous studies have reported that mutant huntingtin (htt) interferes with cyclic AMP response element binding protein binding protein (CBP)-mediated transcription, possibly by inhibiting the acetylation of histones. In Drosophila models that express fragments of mutant htt, histone deacetylase inhibitors reverse deficits in histone acetylation, rescue photoreceptor degeneration, and prolong their survival. These compounds also improve motor deficits in a transgenic mouse model of Huntington disease (HD). To determine whether endogenous CBP depletion contributes to HD pathogenesis, we crossed HD-N171-82Q transgenic mice with mice harboring a disrupted CBP gene and produced mice with partial (50%) depletion of CBP. This reduction of CBP levels decreased the life expectancy of the HD-N171-82Q Line 6 mouse model. The loss of CBP had no obvious impact on the severity of motor impairment, degeneration of the striatum, mutant htt inclusion formation, or global levels of acetylated histones H3 or H4 in brain. In cell models, we confirmed that mutant htt inclusions recruit human CBP but found no evidence for interactions between soluble forms of mutant htt and CBP. Although we identified no neurological explanation for the decreased life expectancy of HD-N171-82Q mice with partial depletion of CBP, the data are consistent with the notion that CBP function mitigates mutant htt toxicity by a currently unidentified mechanism.

Modeling an anti-amyloid combination therapy for Alzheimer's disease.

As only symptomatic treatments are now available for Alzheimer's disease (AD), safe and effective mechanism-based therapies remain a great unmet need for patients with this neurodegenerative disease. Although gamma-secretase and BACE1 [beta-site beta-amyloid (Abeta) precursor protein (APP) cleaving enzyme 1] are well-recognized therapeutic targets for AD, untoward side effects associated with strong inhibition or reductions in amounts of these aspartyl proteases have raised concerns regarding their therapeutic potential. Although moderate decreases of either gamma-secretase or BACE1 are not associated with mechanism-based toxicities, they provide only modest benefits in reducing Abeta in the brains of APPswe/PS1DeltaE9 mice. Because the processing of APP to generate Abeta requires both gamma-secretase and BACE1, it is possible that moderate reductions of both enzymes would provide additive and significant protection against Abeta amyloidosis. Here, we test this hypothesis and assess the value of this novel anti-amyloid combination therapy in mutant mice. We demonstrate that genetic reductions of both BACE1 and gamma-secretase additively attenuate the amyloid burden and ameliorate cognitive deficits occurring in aged APPswe/PS1DeltaE9 animals. No evidence of mechanism-based toxicities was associated with such decreases in amounts of both enzymes. Thus, we propose that targeting both gamma-secretase and BACE1 may be an effective and safe treatment strategy for AD.