A nexus of miR-1271, PAX4 and ALK/RYK influences the cytoskeletal architectures in Alzheimer's Disease and Type 2 Diabetes.

Alzheimer's Disease (AD) and Type 2 Diabetes (T2D) share a common hallmark of insulin resistance. Reportedly, two non-canonical Receptor Tyrosine Kinases (RTKs), ALK and RYK, both targets of the same micro RNA miR-1271, exhibit significant and consistent functional down-regulation in post-mortem AD and T2D tissues. Incidentally, both have Grb2 as a common downstream adapter and NOX4 as a common ROS producing factor. Here we show that Grb2 and NOX4 play critical roles in reducing the severity of both the diseases. The study demonstrates that the abundance of Grb2 in degenerative conditions, in conjunction with NOX4, reverse cytoskeletal degradation by counterbalancing the network of small GTPases. PAX4, a transcription factor for both Grb2 and NOX4, emerges as the key link between the common pathways of AD and T2D. Down-regulation of both ALK and RYK through miR-1271, elevates the PAX4 level by reducing its suppressor ARX via Wnt/ß-Catenin signaling. For the first time, this study brings together RTKs beyond Insulin Receptor (IR) family, transcription factor PAX4 and both AD and T2D pathologies on a common regulatory platform.

Dietary restriction improves proteostasis and increases life span through endoplasmic reticulum hormesis.

Unfolded protein response (UPR) of the endoplasmic reticulum (UPRER) helps maintain proteostasis in the cell. The ability to mount an effective UPRER to external stress (iUPRER) decreases with age and is linked to the pathophysiology of multiple age-related disorders. Here, we show that a transient pharmacological ER stress, imposed early in development on Caenorhabditis elegans, enhances proteostasis, prevents iUPRER decline with age, and increases adult life span. Importantly, dietary restriction (DR), that has a conserved positive effect on life span, employs this mechanism of ER hormesis for longevity assurance. We found that only the IRE-1-XBP-1 branch of UPRER is required for the longevity effects, resulting in increased ER-associated degradation (ERAD) gene expression and degradation of ER resident proteins during DR. Further, both ER hormesis and DR protect against polyglutamine aggregation in an IRE-1-dependent manner. We show that the DR-specific FOXA transcription factor PHA-4 transcriptionally regulates the genes required for ER homeostasis and is required for ER preconditioning-induced life span extension. Finally, we show that ER hormesis improves proteostasis and viability in a mammalian cellular model of neurodegenerative disease. Together, our study identifies a mechanism by which DR offers its benefits and opens the possibility of using ER-targeted pharmacological interventions to mimic the prolongevity effects of DR.

LRSAM1 E3 ubiquitin ligase: molecular neurobiological perspectives linked with brain diseases.

Cellular protein quality control (PQC) plays a significant role in the maintenance of cellular homeostasis. Failure of PQC mechanism may lead to various neurodegenerative diseases due to accumulation of aberrant proteins. To avoid such fatal neuronal conditions PQC employs autophagy and ubiquitin proteasome system (UPS) to degrade misfolded proteins. Few quality control (QC) E3 ubiquitin ligases interplay an important role to specifically recognize misfolded proteins for their intracellular degradation. Leucine-rich repeat and sterile alpha motif-containing 1 (LRSAM1) is a really interesting new gene (RING) class protein that possesses E3 ubiquitin ligase activity with promising applications in PQC. LRSAM1 is also known as RING finger leucine repeat rich (RIFLE) or TSG 101-associated ligase (TAL). LRSAM1 has various cellular functions as it modulates the protein aggregation, endosomal sorting machinery and virus egress from the cells. Thus, this makes LRSAM1 interesting to study not only in protein conformational disorders such as neurodegeneration but also in immunological and other cancerous disorders. Furthermore, LRSAM1 interacts with both cellular protein degradation machineries and hence it can participate in maintenance of overall cellular proteostasis. Still, more research work on the quality control molecular functions of LRSAM1 is needed to comprehend its roles in various protein aggregatory diseases. Earlier findings suggest that in a mouse model of Charcot-Marie-Tooth (CMT) disease, lack of LRSAM1 functions sensitizes peripheral axons to degeneration. It has been observed that in CMT the patients retain dominant and recessive mutations of LRSAM1 gene, which encodes most likely a defective protein. However, still the comprehensive molecular pathomechanism of LRSAM1 in neuronal functions and neurodegenerative diseases is not known. The current article systematically represents the molecular functions, nature and detailed characterization of LRSAM1 E3 ubiquitin ligase. Here, we review emerging molecular mechanisms of LRSAM1 linked with neurobiological functions, with a clear focus on the mechanism of neurodegeneration and also on other diseases. Better understanding of LRSAM1 neurobiological and intracellular functions may contribute to develop promising novel therapeutic approaches, which can also propose new lines of molecular beneficial targets for various neurodegenerative diseases.

Ube3a deficiency inhibits amyloid plaque formation in APPswe/PS1δE9 mouse model of Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by progressive decline in memory and cognitive function. Pathological hallmark of AD includes aberrant aggregation of amyloid beta (Aß) peptide, which is produced upon sequential cleavage of amyloid precursor protein (APP) by ß- and γ -secretases. On the contrary, α-secretase cleaves APP within the Aß sequence and thereby prevents Aß generation. Here, we investigated the role of ubiquitin ligase Ube3a (involved in synaptic function and plasticity) in the pathogenesis of AD using APPswe/PS1δE9 transgenic mouse model and first noticed that soluble pool of Ube3a was age-dependently decreased in AD mouse in comparison with wild type controls. To further explore the role of Ube3a in AD patho-mechanism, we generated brain Ube3a-deficient AD mice that exhibited accelerated cognitive and motor deficits compared with AD mice. Interestingly, these Ube3a-deficient AD mice were excessively obese from their age of 12 months and having shorter lifespan. Biochemical analysis revealed that the Ube3a-deficient AD mice had significantly reduced level of Aß generation and amyloid plaque formation in their brain compared with age-matched AD mice and this effect could be due to the increased activity of α-secretase, ADAM10 (a disintegrin and metalloproteinase-10) that shift the proteolysis of APP towards non-amyloidogenic pathway. These findings suggest that aberrant function of Ube3a could influence the progression of AD and restoring normal level of Ube3a might be beneficial for AD.

Topoisomerase 1 inhibitor topotecan delays the disease progression in a mouse model of Huntington's disease.

Huntington's disease (HD) is a dominantly inherited progressive neurodegenerative disorder caused by the accumulation of polyglutamine expanded mutant huntingtin as inclusion bodies primarily in the brain. After the discovery of the HD gene, considerable progress has been made in understanding the disease pathogenesis and multiple drug targets have been identified, even though currently there is no effective therapy. Here, we demonstrate that the treatment of topotecan, a brain-penetrating topoisomerase 1 inhibitor, to HD transgenic mouse considerably improved its motor behavioural abnormalities along with a significant extension of lifespan. Improvement of behavioural deficits are accompanied with the significant rescue of their progressively decreased body weight, brain weight and striatal volume. Interestingly, topotecan treatment also significantly reduced insoluble mutant huntingtin load in the HD mouse brain. Finally, we show that topotecan treatment to HD mouse not only inhibits the expression of transgenic mutant huntingtin, but also at the same time induces the expression of Ube3a, an ubiquitin ligase linked to the clearance of mutant huntingtin. These findings suggest that topotecan could be a potential therapeutic molecule to delay the progression of HD.

Rescue of altered HDAC activity recovers behavioural abnormalities in a mouse model of Angelman syndrome.

Angelman syndrome (AS) is a neurodevelopmental disorder characterized by severe intellectual and developmental disabilities. The disease is caused by the loss of function of maternally inherited UBE3A, a gene that exhibits paternal-specific imprinting in neuronal tissues. Ube3a-maternal deficient mice (AS mice) display many classical features of AS, although, the underlying mechanism of these behavioural deficits is poorly understood. Here we report that the absence of Ube3a in AS mice brain caused aberrant increase in HDAC1/2 along with decreased acetylation of histone H3/H4. Partial knockdown of Ube3a in cultured neuronal cells also lead to significant up-regulation of HDAC1/2 and consequent down-regulation of histones H3/H4 acetylation. Treatment of HDAC inhibitor, sodium valproate, to AS mice showed significant improvement in social, cognitive and motor impairment along with restoration of various proteins linked with synaptic function and plasticity. Interestingly, HDAC inhibitor also significantly increased the expression of Ube3a in cultured neuronal cells and in the brain of wild type mice but not in AS mice. These results indicate that anomalous HDAC1/2 activity might be linked with synaptic dysfunction and behavioural deficits in AS mice and suggests that HDAC inhibitors could be potential therapeutic molecule for the treatment of the disease.

Dexamethasone induces heat shock response and slows down disease progression in mouse and fly models of Huntington's disease.

Huntington's disease (HD) is an inherited neurodegenerative disorder caused by abnormal expansion of glutamine repeats in the protein huntingtin. In HD brain, mutant huntingtin undergoes proteolytic processing, and its N-terminal fragment containing poly-glutamine repeats accumulate as insoluble aggregates leading to the defect in cellular protein quality control system and heat shock response (HSR). Here we demonstrate that the defective HSR in the brain is due to the down-regulation of heat shock factor 1 (HSF1) in both mice and fly models of HD. Interestingly, treatment of dexamethasone (a synthetic glucocorticoid) to HD mice or flies significantly increased the expression and transactivation of HSF1 and induction of HSR and these effects are mediated through the down-regulation of HSP90. Dexamethasone treatment also significantly decreased the aggregate load and transient recovery of HD-related behavioural phenotypes in both disease models. These results suggest that dexamethasone could be a potential therapeutic molecule for the treatment of HD and related poly-glutamine disorders.

Deficiency of Ube3a in Huntington's disease mice brain increases aggregate load and accelerates disease pathology.

Huntington's disease (HD) is an inherited neurodegenerative disorder caused by abnormal expansion of CAG repeats in the gene encoding huntingtin. Mutant huntingtin undergoes proteolytic processing and its N-terminal fragment containing polyglutamine repeat accumulates as inclusion not only in nucleus but also in cytoplasm and neuronal processes. Here, we demonstrate that removal of ubiquitin ligase Ube3a selectively from HD mice brain resulted in accelerated disease phenotype and shorter lifespan in comparison with HD mice. The deficiency of Ube3a in HD mice brain also caused significant increase in global aggregates load, and these aggregates were less ubiquitinated when compared with age-matched HD mice. These Ube3a-maternal deficient HD mice also showed drastic reduction of DARPP-32, a dopamine-regulated phoshphoprotein in their striatum. These results emphasize the crucial role of Ube3a in the progression of HD and its immense potential as therapeutic target.

Impaired adult hippocampal neurogenesis and its partial reversal by chronic treatment of fluoxetine in a mouse model of Angelman syndrome.

Angelman syndrome (AS) is a neurodevelopmental disorder characterized by severe cognitive and motor deficits, caused by the loss of function of maternally inherited Ube3a. Ube3a-maternal deficient mice (AS model mice) recapitulate many essential features of AS, but how the deficiency of Ube3a lead to such behavioural abnormalities is poorly understood. Here we have demonstrated significant impairment of adult hippocampal neurogenesis in AS mice brain. Although, the number of BrdU and Ki67-positive cell in the hippocampal DG region was nearly equal at early postnatal days among wild type and AS mice, they were significantly reduced in adult AS mice compared to wild type controls. Reduced number of doublecortin-positive immature neurons in this region of AS mice further indicated impaired neurogenesis. Unaltered BrdU and Ki67-positive cells number in the sub ventricular zone of adult AS mice brain along with the absence of imprinted expression of Ube3a in the neural progenitor cell suggesting that Ube3a may not be directly linked with altered neurogenesis. Finally, we show that the impaired hippocampal neurogenesis in these mice can be partially rescued by the chronic treatment of antidepressant fluoxetine. These results suggest that the chronic stress may lead to reduced hippocampal neurogenesis in AS mice and that impaired neurogenesis could contribute to cognitive disturbances observed in these mice.

Neuronatin-mediated aberrant calcium signaling and endoplasmic reticulum stress underlie neuropathology in Lafora disease.

Lafora disease (LD) is a teenage-onset inherited progressive myoclonus epilepsy characterized by the accumulations of intracellular inclusions called Lafora bodies and caused by mutations in protein phosphatase laforin or ubiquitin ligase malin. But how the loss of function of either laforin or malin causes disease pathogenesis is poorly understood. Recently, neuronatin was identified as a novel substrate of malin that regulates glycogen synthesis. Here we demonstrate that the level of neuronatin is significantly up-regulated in the skin biopsy sample of LD patients having mutations in both malin and laforin. Neuronatin is highly expressed in human fetal brain with gradual decrease in expression in developing and adult brain. However, in adult brain, neuronatin is predominantly expressed in parvalbumin-positive GABAergic interneurons and localized in their processes. The level of neuronatin is increased and accumulated as insoluble aggregates in the cortical area of LD brain biopsy samples, and there is also a dramatic loss of parvalbumin-positive GABAergic interneurons. Ectopic expression of neuronatin in cultured neuronal cells results in increased intracellular Ca(2+), endoplasmic reticulum stress, proteasomal dysfunction, and cell death that can be partially rescued by malin. These findings suggest that the neuronatin-induced aberrant Ca(2+) signaling and endoplasmic reticulum stress might underlie LD pathogenesis.

Reversal of reduced parvalbumin neurons in hippocampus and amygdala of Angelman syndrome model mice by chronic treatment of fluoxetine.

Angelman syndrome (AS) is a neuropsychiatric disorder characterized by autism, intellectual disability and motor disturbances. The disease is primarily caused by the loss of function of maternally inherited UBE3A. Ube3a maternal-deficient mice recapitulates many essential feature of AS. These AS mice have been shown to be under chronic stress and exhibits anxiety-like behaviour because of defective glucocorticoid receptor signalling. Here, we demonstrate that chronic stress in these mice could lead to down-regulation of parvalbumin-positive interneurons in the hippocampus and basolateral amygdala from early post-natal days. Down-regulation of parvalbumin-positive interneurons number could be because of decrease in the expression of parvalbumin in these neurons. We also find that treatment with fluoxetine, a selective serotonin reuptake inhibitor, results in restoration of impaired glucocorticoid signalling, elevated serum corticosterone level, parvalbumin-positive interneurons and anxiety-like behaviours. Our findings suggest that impaired glucocorticod signalling in hippocampus and amygdala of AS mice is critical for the decrease in parvalbumin interneurons number, emergence of anxiety and other behavioural deficits and highlights the importance of fluoxetine in the recovery of these abnormalities.

Defective glucocorticoid hormone receptor signaling leads to increased stress and anxiety in a mouse model of Angelman syndrome.

Angelman syndrome (AS) is a neurodevelopmental disorder caused due to deletions or loss-of-function mutations in maternally inherited UBE3A. Ube3a functions as an ubiquitin ligase as well as a transcriptional coactivator of steroid hormone receptors. However, the mechanisms by which maternal Ube3a deficiency gives rise to phenotypic features of AS are not clear. We report here that Ube3a regulates glucocorticoid receptor (GR) transactivation and GR signaling pathway is disrupted in Ube3a-maternal-deficient mice brain. The expression of several GR-dependent genes is down-regulated in multiple brain regions of Ube3a-maternal-deficient mice. AS mice show significantly higher level of blood corticosterone, selective loss of GR and reduced number of parvalbumin-positive inhibitory interneurons in their hippocampus that could ultimately lead to increased stress. These mice also exhibit increased anxiety-like behavior, which could be due to chronic stress. Altogether, our findings suggest that chronic stress due to altered GR signaling might lead to anxiety-like behavior in a mouse of model of AS.

Malin regulates Wnt signaling pathway through degradation of dishevelled2.

Using yeast-two hybrid screening followed by co-immunoprecipitation assay, we have found that the Lafora disease ubiquitin ligase malin interacts with dishevelled2, a key mediator of Wnt signaling pathway. Overexpression of malin enhances the degradation of dishevelled2 and inhibits Wnt signaling, which is evident from the down-regulation of ß-catenin target genes and the decrease in ß-catenin-mediated transcriptional activity. Partial knockdown of malin significantly increases the level of dishevelled2 and up-regulates Wnt signaling. Several malin mutants are found to be ineffective in degrading dishevelled2 and regulating the Wnt pathway. We have also found that malin enhances K48- and K63-linked ubiquitination of dishevelled2 that could lead to its degradation through both proteasome and autophagy. Altogether, our results indicate that malin regulates Wnt signaling pathway through the degradation of dishevelled2 and suggest possible deregulation of Wnt signaling in Lafora disease.

Dysfunction of the ubiquitin ligase Ube3a may be associated with synaptic pathophysiology in a mouse model of Huntington disease.

Huntington disease (HD) is a hereditary neurodegenerative disorder characterized by progressive cognitive, psychiatric, and motor symptoms. The disease is caused by abnormal expansion of CAG repeats in the gene encoding huntingtin, but how mutant huntingtin leads to early cognitive deficits in HD is poorly understood. Here, we demonstrate that the ubiquitin ligase Ube3a, which is implicated in synaptic plasticity and involved in the clearance of misfolded polyglutamine protein, is strongly recruited to the mutant huntingtin nuclear aggregates, resulting in significant loss of its functional pool in different regions of HD mouse brain. Interestingly, Arc, one of the substrates of Ube3a linked with synaptic plasticity, is also associated with nuclear aggregates, although its synaptic level is increased in the hippocampus and cortex of HD mouse brain. Different regions of HD mouse brain also exhibit decreased levels of AMPA receptors and various pre- and postsynaptic proteins, which could be due to the partial loss of function of Ube3a. Transient expression of mutant huntingtin in mouse primary cortical neurons further demonstrates recruitment of Ube3a into mutant huntingtin aggregates, increased accumulation of Arc, and decreased numbers of GluR1 puncta in the neuronal processes. Altogether, our results suggest that the loss of function of Ube3a might be associated with the synaptic abnormalities observed in HD.

MicroRNA-124 targets CCNA2 and regulates cell cycle in STHdh(Q111)/Hdh(Q111) cells.

Mutation in huntingtin (HTT) gene causes Huntington's disease (HD). Expression of many micro RNAs is known to alter in cell, animal models and brains of HD patients, but their cellular effects are not known. Here, we show that expression of microRNA-124 (miR-124) is down regulated in HD striatal mutant STHdh(Q111)/Hdh(Q111) cells, a cell model of HD compared to STHdh(Q7)/Hdh(Q7) cells. STHdh(Q7)/Hdh(Q7) and STHdh(Q111)/Hdh(Q111) cells express endogenously full length wild type and mutant HTT respectively. We confirmed this result in R6/2 mouse, an animal model of HD, expressing mutant HTT. Gene Ontology terms related to cell cycle were enriched significantly with experimentally validated targets of miR-124. We observed that expression of Cyclin A2 (CCNA2), a putative target of miR-124 was increased in mutant STHdh(Q111)/Hdh(Q111) cells and brains of R6/2 mice. Fraction of cells in S phase was higher in asynchronously growing mutant STHdh(Q111)/Hdh(Q111) cells compared to wild type STHdh(Q7)/Hdh(Q7) cells and could be altered by exogenous expression or inhibition of miR-124. Exogenous expression or knock down of CCNA2, a target of miR-124, also alters proportion of cells in S phase of HD cell model. In summary, decreased miR-124 expression could increase CCNA2 in cell and animal model of HD and is involved in deregulation of cell cycle in STHdh(Q111)/Hdh(Q111) cells.

Itraconazole Confers Cytoprotection Against Neurodegenerative Disease-Associated Abnormal Protein Aggregation.

Cells perform regular maintenance to avoid the accumulation of misfolded proteins. Prolonged accumulation of these proteotoxic inclusions generates potential risk of ageing-related diseases such as neurodegenerative diseases. Therefore, removal of such abnormal aggregates can ensure the re-establishment of proteostasis. Ubiquitin proteasome system (UPS) actively participates in the selective removal of aberrantly folded clients with the help of complex proteasome machinery. However, specific induction of proteasome functions to remove abnormal proteins remains an open challenge. Here, we show that Itraconazole treatment induces proteasome activities and degrades the accumulation of bonafide-misfolded proteins, including heat-denatured luciferase. Exposure of Itraconazole elevates the degradation of neurodegenerative disease-associated proteins, e.g. expanded polyglutamine, mutant SOD1, and mutant α-synuclein. Our results suggest that Itraconazole treatment prevents the accumulation of neurodegenerative disease-linked misfolded proteins and generates cytoprotection. These findings reveal that Itraconazole removes abnormal proteins through sequential proteasomal activation and represents a potential protective therapeutic role against protein-misfolding neurodegenerative diseases.

Sequestration of chaperones and proteasome into Lafora bodies and proteasomal dysfunction induced by Lafora disease-associated mutations of malin.

Lafora disease (LD) is an autosomal recessive progressive myoclonic epilepsy characterized by the presence of intracellular polyglucosan inclusions commonly known as Lafora bodies in many tissues, including the brain, liver and skin. The disease is caused by mutations in either EPM2A gene, encoding the protein phosphatase, laforin, or EPM2B gene, encoding the ubiquitin ligase, malin. But how mutations in these two genes cause disease pathogenesis is poorly understood. In this study, we show that the Lafora bodies in the axillary skin and brain stain positively for the ubiquitin, the 20S proteasome and the molecular chaperones Hsp70/Hsc70. Interestingly, mutant malins that are misfolded also frequently colocalizes with Lafora bodies in the skin biopsy sample of the respective LD patient. The expression of disease-causing mutations of malin in Cos-7 cells results in the formation of the profuse cytoplasmic aggregates that colocalize with the Hsp70/Hsc70 chaperones and the 20S proteasome. The mutant malin expressing cells also exhibit proteasomal dysfunction and cell death. Overexpression of Hsp70 decreases the frequency of the mutant malin aggregation and protects from mutant malin-induced cell death. These findings suggest that Lafora bodies consist of abnormal proteins, including mutant malin, targeted by the chaperones or the proteasome for their refolding or clearance, and failure of these quality control systems could lead to LD pathogenesis. Our data also indicate that the Hsp70 chaperone could be a potential therapeutic target of LD.

Understanding the pathogenesis of Angelman syndrome through animal models.

Angelman syndrome (AS) is a neurodevelopmental disorder characterized by severe mental retardation, lack of speech, ataxia, susceptibility to seizures, and unique behavioral features such as easily provoked smiling and laughter and autistic features. The disease is primarily caused by deletion or loss-of-function mutations of the maternally inherited UBE3A gene located within chromosome 15q11-q13. The UBE3A gene encodes a 100 kDa protein that functions as ubiquitin ligase and transcriptional coactivator. Emerging evidence now indicates that UBE3A plays a very important role in synaptic function and in regulation of activity-dependent synaptic plasticity. A number of animal models for AS have been generated to understand the disease pathogenesis. The most widely used model is the UBE3A-maternal-deficient mouse that recapitulates most of the essential features of AS including cognitive and motor abnormalities. This paper mainly discusses various animal models of AS and how these models provide fundamental insight into understanding the disease biology for potential therapeutic intervention.

Long non-coding RNA MALAT1 protects against Aß1-42 induced toxicity by regulating the expression of receptor tyrosine kinase EPHA2 via quenching miR-200a/26a/26b in Alzheimer's disease.

Altered expressions of Receptor Tyrosine Kinases (RTK) and non-coding (nc) RNAs are known to regulate the pathophysiology of Alzheimer's disease (AD). However, specific understanding of the roles played, especially the mechanistic and functional roles, by long ncRNAs in AD is still elusive. Using mouse tissue qPCR assays we observe changes in the expression levels of 41 lncRNAs in AD mice of which only 7 genes happen to have both human orthologs and AD associations. Post validation of these 7 human lncRNA genes, MEG3 and MALAT1 shows consistent and significant decrease in AD cell, animal models and human AD brain tissues, but MALAT1 showed a more pronounced decrease. Using bioinformatics, qRT-PCR, RNA FISH and RIP techniques, we could establish MALAT1 as an interactor and regulator of miRs-200a, -26a and -26b, all of which are naturally elevated in AD. We could further show that these miRNAs target the RTK EPHA2 and several of its downstream effectors. Expectedly EPHA2 over expression protects against Aß1-42 induced cytotoxicity. Transiently knocking down MALAT1 validates these unique regulatory facets of AD at the miRNA and protein levels. Although the idea of sponging of miRNAs by lncRNAs in other pathologies is gradually gaining credibility, this novel MALAT1- miR-200a/26a/26b - EPHA2 regulation mechanism in the context of AD pathophysiology promises to become a significant strategy in controlling the disease.

Design and Development of Benzothiazole-Based Fluorescent Probes for Selective Detection of Aß Aggregates in Alzheimer's Disease.

The formation and accumulation of amyloid beta (Aß) peptide are considered the crucial events that are responsible for the progression of Alzheimer's disease (AD). Herein, we have designed and synthesized a series of fluorescent probes by using electron acceptor-donor end groups interacting with a π-conjugating system for the detection of Aß aggregates. The chemical structure of these probes denoted as RMs, having a conjugated π-system (C═C), showed a maximum emission in PBS (>600 nm), which is the best range for a fluorescent imaging probe. Among all these probes, RM-28 showed an excellent fluorescence property with an emission maximum of >598 nm upon binding to Aß aggregates. RM-28 also showed high sensitivity (7.5-fold) and high affinities toward Aß aggregates (Kd = 175.69 ± 4.8 nM; Ka = 0.5 × 107 M-1). It can cross the blood-brain barrier of mice efficiently. The affinity of RM-28 toward Aß aggregates was observed in 3xTg-AD brain sections of the hippocampus and cortex region using a fluorescent imaging technique, as well as an in vitro fluorescence-based binding assay with Aß aggregates. Moreover, RM-28 is highly specific to Aß aggregates and does not bind with intracellular proteins like bovine serum albumin (BSA) and α-synuclein (α-Syn) aggregates. The results indicate that the probe RM-28 emerges as an efficient and veritable highly specific fluorescent probe for the detection of Aß aggregates in both in vitro and in vivo model systems.