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BACKGROUND: Background: Neurodegenerative diseases manifest behavioral dysfunction with disease progression. Intervention with neuropsychiatric drugs is part of most multi-drug treatment paradigms. However, only a fraction of patients responds to the treatments and those responding must deal with drug-drug interactions and tolerance issues generally attributed to off-target activities. Recent efforts have focused on the identification of underexplored targets and exploration of improved outcomes by treatment with selective molecular probes. Objective: As part of ongoing efforts to identify and validate additional targets amenable to therapeutic intervention, we examined levels of the serotonin 5-HT2b receptor (5-HT2bR) in Alzheimer's disease (AD) brains and the potential of a selective 5-HT2bR antagonist to counteract synaptic plasticity and memory damage induced by AD-related proteins, amyloid-β, and tau. Methods: This work used a combination of biochemical, chemical biology, electrophysiological, and behavioral techniques. Biochemical methods included analysis of protein levels. Chemical biology methods included the use of an in vivo molecular probe MW071, a selective antagonist for the 5HT2bR. Electrophysiological methods included assessment of long-term potentiation (LTP), a type of synaptic plasticity thought to underlie memory formation. Behavioral studies investigated spatial memory and associative memory. Results: 5HT2bR levels are increased in brain specimens of AD patients compared to controls. 5HT2bR antagonist treatment rescued amyloid-β and tau oligomer-induced impairment of synaptic plasticity and memory. Conclusions: The increased levels of 5HT-2bR in AD patient brains and the attenuation of disease-related synaptic and behavioral dysfunctions by MW071 treatment suggest that the 5HT-2bR is a molecular target worth pursuing as a potential therapeutic target. [ABSTRACT FROM AUTHOR]

Background: Synaptic dysfunction prevalent in Alzheimer's disease (AD) brain is closely associated with increased accumulation of amyloid-β (Aβ) peptides in the brain parenchyma. It is widely believed that Aβ peptides trigger synaptic dysfunction by interfering with the synaptic vesicular fusion and the release of neurotransmitters, primarily facilitated by the SNARE protein complexes formed by VAMP-2, SNAP-25, and syntaxin-1. However, Aβ interactions with SNARE proteins to ultimately disrupt synaptic vesicular fusion are not well understood.Objective: Our objective is to elucidate mechanisms by which Aβ peptides perturb SNARE complexes.Methods: Intensity (qualitative) and lifetime (quantitative) based measurements involving Forster (fluorescence) resonance energy transfer (FRET) followed by fluorescence lifetime imaging microscopy (FLIM) were employed to investigate the effect of Aβ peptides on dynamic interactions between VAMP-2, labeled with cerulean (Cer) at the N-terminus (FRET donor), and SNAP-25 labeled with citrine (Cit) on the N-terminus (FRET acceptor). The FRET and FLIM interactions at the exocytosis locations on the pre-synaptic membrane were recorded under spontaneous and high potassium evoked conditions. Moreover, cellular accumulation of fluorescein labeled Aβ (F-Aβ) peptides and their co-localization with Cer-VAMP2 was investigated by confocal microscopy.Results: The F-Aβ40 and F-Aβ42 are internalized by differentiated N2A cells, where they colocalize with Cer-VAMP2. Both Aβ40 and Aβ42 decrease interactions between the N-termini of Cer-VAMP2 and Cit-SNAP25 in N2A cells, as determined by FRET/FLIM.Conclusion: By perturbing the N-terminal interactions between VAMP-2 and SNAP-25, Aβ40 and Aβ42, can directly interfere with the SNARE complex formation, which is critical for the docking and fusion of synaptic vesicles. [ABSTRACT FROM AUTHOR]

Serotonin 4 (5-HT4) receptor signaling does not only have the physiological function of improving cognition, but might also be helpful in the therapy of Alzheimer's disease (AD) through regulation of the production of soluble amyloid-β protein precursor alpha (sAβPPα). To analyze the relationship between 5-HT4 receptor signaling and sAβPPα production, we stably transfected H4 cells with AβPP and 5-HT4 receptor (H4/AβPP/5-HT4 cells). We found that 24-h incubation with the 5-HT4 receptor agonist RS-67333 upregulates matrix metalloproteinase-9 (MMP-9). Furthermore, MMP-9 overexpression enhanced sAβPPα levels, whereas knockdown with MMP-9 siRNA decreased sAβPPα levels. When RS-67333 was injected for 10 days in Tg2576 mice, a model of amyloid-β peptide (Aβ) deposition, there was an increase in hippocampal levels of sAβPPα, C-terminal fragment α, and MMP-9, as well as a decrease in hippocampal senile plaque number and levels of the 40 amino acid peptide, Aβ40. Taken all together, these experiments demonstrate that 5-HT4 receptor stimulation induces expression of MMP-9 which cleaves AβPP through α-secretase-like activity, leading to an increase of sAβPPα levels and a reduction of Aβ load. [ABSTRACT FROM AUTHOR]

Oligomeric amyloid-beta (Abeta) interferes with long-term potentiation (LTP) and cognitive processes, suggesting that Abeta peptides may play a role in the neuronal dysfunction which characterizes the early stages of Alzheimer's disease (AD). Multiple lines of evidence have highlighted RAGE (receptor for advanced glycation end-products) as a receptor involved in Abeta-induced neuronal and synaptic dysfunction. In the present study, we investigated the effect of oligomeric soluble Abeta1-42 on LTP elicited by the stimulation of different intracortical pathways in the mouse visual cortex. A variety of nanomolar concentrations (20-200 nM) of Abeta1-42 were able to inhibit LTP in cortical layer II-III induced by either white matter (WM-Layer II/III) or the layer II/III (horizontal pathway) stimulation, whereas the inhibition of LTP was more susceptible to Abeta1-42, which occurred at 20 nM of Abeta, when stimulating layer II-III horizontal pathway. Remarkably, cortical slices were resistant to nanomolar Abeta1-42 in the absence of RAGE (genetic deletion of RAGE) or blocking RAGE by RAGE antibody. These results indicate that nanomolar Abeta inhibits LTP expression in different neocortical circuits. Crucially, it is demonstrated that Abeta-induced reduction of LTP in different cortical pathways is mediated by RAGE. [ABSTRACT FROM AUTHOR]

Alzheimer's disease, one of the most important brain pathologies associated with neurodegenerative processes, is related to overactivation of calpain-mediated proteolysis. Previous data showed a compelling efficacy of calpain inhibition against abnormal synaptic plasticity and memory produced by the excess of amyloid-β, a distinctive marker of the disease. Moreover, a beneficial effect of calpain inhibitors in Alzheimer's disease is predictable by the occurrence of calpain hyperactivation leading to impairment of memory-related pathways following abnormal calcium influxes that might ensue independently of amyloid-β elevation. However, molecules currently available as effective calpain inhibitors lack adequate selectivity. This work is aimed at characterizing the efficacy of a novel class of epoxide-based inhibitors, synthesized to display improved selectivity and potency towards calpain 1 compared to the prototype epoxide-based generic calpain inhibitor E64. Both functional and preliminary toxicological investigations proved the efficacy, potency, and safety of the novel and selective calpain inhibitors NYC438 and NYC488 as possible therapeutics against the disease. [ABSTRACT FROM AUTHOR]

Amyloid-β (Aβ) peptides are constitutively produced in the brain throughout life via mechanisms that can be regulated by synaptic activity. Although Aβ has been extensively studied as the pathological plaque-forming protein species in Alzheimer's disease (AD), little is known about the normal physiological function(s) and signaling pathway(s). We previously discovered that physiologically-relevant, low picomolar amounts of Aβ can enhance synaptic plasticity and hippocampal-dependent cognition in mice. In this study, we demonstrated that astrocytes are cellular candidates for participating in this type of Aβ signaling. Using calcium imaging of primary astrocyte cultures, we observed that picomolar amounts of Aβ peptides can enhance spontaneous intracellular calcium transient signaling. After application of 200 pM Aβ42 peptides, the frequency and amplitude averages of spontaneous cytosolic calcium transients were significantly increased. These effects were dependent on α7 nicotinic acetylcholine receptors (α7-nAChRs), as the enhancement effects were blocked by a pharmacological α7-nAChR inhibitor and in astrocytes from an α7 deficient mouse strain. We additionally examined evoked intercellular calcium wave signaling but did not detect significant picomolar Aβ-induced alterations in propagation parameters. Overall, these results indicate that at a physiologically-relevant low picomolar concentration, Aβ peptides can enhance spontaneous astrocyte calcium transient signaling via α7-nAChRs. Since astrocyte-mediated gliotransmission has been previously found to have neuromodulatory roles, Aβ peptides may have a normal physiological function in regulating neuron-glia signaling. Dysfunction of this signaling process may underlie glia-based aspects of AD pathogenesis. [ABSTRACT FROM AUTHOR]

Amyloid-β peptide (Aβ) is considered a key protein in the pathogenesis of Alzheimer's disease (AD) because of its neurotoxicity and capacity to form characteristic insoluble deposits known as senile plaques. Aβ derives from amyloid-β protein precursor (AβPP), whose proteolytic processing generates several fragments including Aβ peptides of various lengths. The normal function of AβPP and its fragments remains poorly understood. While some fragments have been suggested to have a function in normal physiological cellular processes, Aβ has been widely considered as a 'garbage' fragment that becomes toxic when it accumulates in the brain, resulting in impaired synaptic function and memory. Aβ is produced and released physiologically in the healthy brain during neuronal activity. In the last 10 years, we have been investigating whether Aβ plays a physiological role in the brain. We first demonstrated that picomolar concentrations of a human Aβ42 preparation enhanced synaptic plasticity and memory in mice. Next, we investigated the role of endogenous Aβ in healthy murine brains and found that treatment with a specific antirodent Aβ antibody and an siRNA against murine AβPP impaired synaptic plasticity and memory. The concurrent addition of human Aβ42 rescued these deficits, suggesting that in the healthy brain, physiological Aβ concentrations are necessary for normal synaptic plasticity and memory to occur. Furthermore, the effect of both exogenous and endogenous Aβ was seen to be mediated by modulation of neurotransmitter release and α7-nicotinic receptors. These findings need to be taken into consideration when designing novel therapeutic strategies for AD. [ABSTRACT FROM AUTHOR]