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Jurgen Wess - One of the best experts on this subject based on the ideXlab platform.
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Muscarinic Acetylcholine Receptor regulates self renewal of early erythroid progenitors
Science Translational Medicine, 2019Co-Authors: Gaurang Trivedi, Jurgen Wess, Daichi Inoue, Cynthia Chen, Lillian Bitner, Young Rock Chung, Justin Taylor, Mithat Gonen, Omar Abdelwahab, Lingbo ZhangAbstract:Adult stem and progenitor cells are uniquely capable of self-renewal, and targeting this process represents a potential therapeutic opportunity. The early erythroid progenitor, burst-forming unit erythroid (BFU-E), has substantial self-renewal potential and serves as a key cell type for the treatment of anemias. However, our understanding of mechanisms underlying BFU-E self-renewal is extremely limited. Here, we found that the Muscarinic Acetylcholine Receptor, cholinergic Receptor, Muscarinic 4 (CHRM4), pathway regulates BFU-E self-renewal and that pharmacological inhibition of CHRM4 corrects anemias of myelodysplastic syndrome (MDS), aging, and hemolysis. Genetic down-regulation of CHRM4 or pharmacologic inhibition of CHRM4 using the selective antagonist PD102807 promoted BFU-E self-renewal, whereas deletion of Chrm4 increased erythroid cell production under stress conditions in vivo. Moreover, Muscarinic Acetylcholine Receptor antagonists corrected anemias in mouse models of MDS, aging, and hemolysis in vivo, extending the survival of mice with MDS relative to that of controls. The effects of Muscarinic Receptor antagonism on promoting expansion of BFU-Es were mediated by cyclic AMP induction of the transcription factor CREB, whose targets up-regulated key regulators of BFU-E self-renewal. On the basis of these data, we propose a model of hematopoietic progenitor self-renewal through a cholinergic-mediated "hematopoietic reflex" and identify Muscarinic Acetylcholine Receptor antagonists as potential therapies for anemias associated with MDS, aging, and hemolysis.
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role for the m1 Muscarinic Acetylcholine Receptor in top down cognitive processing using a touchscreen visual discrimination task in mice
ACS Chemical Neuroscience, 2015Co-Authors: Robert W Gould, Jurgen Wess, Craig W Lindsley, Ditte Dencker, Michael Bubser, Michael Grannan, Xiaoyan Zhan, Zixiu Xiang, Charles W Locuson, P J ConnAbstract:The M1 Muscarinic Acetylcholine Receptor (mAChR) subtype has been implicated in the underlying mechanisms of learning and memory and represents an important potential pharmacotherapeutic target for the cognitive impairments observed in neuropsychiatric disorders such as schizophrenia. Patients with schizophrenia show impairments in top-down processing involving conflict between sensory-driven and goal-oriented processes that can be modeled in preclinical studies using touchscreen-based cognition tasks. The present studies used a touchscreen visual pairwise discrimination task in which mice discriminated between a less salient and a more salient stimulus to assess the influence of the M1 mAChR on top-down processing. M1 mAChR knockout (M1 KO) mice showed a slower rate of learning, evidenced by slower increases in accuracy over 12 consecutive days, and required more days to acquire (achieve 80% accuracy) this discrimination task compared to wild-type mice. In addition, the M1 positive allosteric modulator B...
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novel insights into m3 Muscarinic Acetylcholine Receptor physiology and structure
Journal of Molecular Neuroscience, 2014Co-Authors: Andrew C Kruse, Brian K Kobilka, Jurgen WessAbstract:Recent studies with M3 Muscarinic Acetylcholine Receptor (M3R) mutant mice suggest that drugs selectively targeting this Receptor subtype may prove useful for the treatment of various pathophysiological conditions. Moreover, the use of M3R-based designer G protein-coupled Receptors (GPCRs) has provided novel insights into how Gq-coupled GPCRs can modulate whole-body glucose homeostasis by acting on specific peripheral cell types. More recently, we succeeded in using X-ray crystallography to determine the structure of the M3R bound to the bronchodilating drug tiotropium, a Muscarinic antagonist (inverse agonist). This new structural information should facilitate the development of orthosteric or allosteric M3R-selective drugs that are predicted to have considerable therapeutic potential.
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m4 Muscarinic Acetylcholine Receptor modulation of associative learning and behavioral flexibility in a novel touchscreen cognitive assessment 845 8
The FASEB Journal, 2014Co-Authors: Ditte Dencker, Thomas M Bridges, Anders Finkjensen, Robert W Gould, Michael Bubser, Michael R Wood, Mark E Duggan, Jurgen WessAbstract:Recent findings indicate that selective activation of the M4 Muscarinic Acetylcholine Receptor (M4 mAChR) may represent a novel treatment approach for the psychotic symptoms and cognitive disturbances observed in several neuropsychiatric disorders. In the present studies, cognitive performance of M4 mAChR knock-out mice (M4-KO mice) was examined across a series of computer touchscreen-based pairwise discrimination tasks assessing susceptibility to interference from previously learned competing sets of information. While M4-KO mice acquired the initial discrimination task in a similar number of sessions as wildtype mice, the M4-KO mice showed impaired acquisition on all subsequent visual discriminations. In addition, the novel M4 positive allosteric modulator VU0467154 reversed disruptions in pairwise discrimination induced by MK-801, an antagonist of the N-methyl-D-aspartate subtype of glutamate Receptors. These data reveal that selective M4 mAChR activation is critical for modulation of normal associativ...
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Muscarinic Acetylcholine Receptor X-ray structures: potential implications for drug development
Current opinion in pharmacology, 2014Co-Authors: Andrew C Kruse, Brian K Kobilka, Jurgen WessAbstract:Muscarinic Acetylcholine Receptor antagonists are widely used as bronchodilating drugs in pulmonary medicine. The therapeutic efficacy of these agents depends on the blockade of M3 Muscarinic Receptors expressed on airway smooth muscle cells. All Muscarinic antagonists currently used as bronchodilating agents show high affinity for all five Muscarinic Receptor subtypes, thus increasing the likelihood of unwanted side effects. Recent X-ray crystallographic studies have provided detailed structural information about the nature of the orthosteric Muscarinic binding site (the conventional Acetylcholine binding site) and an 'outer' Receptor cavity that can bind allosteric (non-orthosteric) drugs. These new findings should guide the development of selective M3 Receptor blockers that have little or no effect on other Muscarinic Receptor subtypes.
Allan I. Levey - One of the best experts on this subject based on the ideXlab platform.
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disease modifying effects of m1 Muscarinic Acetylcholine Receptor activation in an alzheimer s disease mouse model
ACS Chemical Neuroscience, 2017Co-Authors: Evan P Lebois, Jeffrey P Conn, Craig W Lindsley, Thomas M Bridges, Jason P Schroeder, Thomas J Esparza, David L Brody, Scott J Daniels, Allan I. LeveyAbstract:Alzheimer’s disease (AD) is the leading cause of dementia worldwide, and currently no disease-modifying therapy is available to slow or prevent AD, underscoring the urgent need for neuroprotective therapies. Selective M1 Muscarinic Acetylcholine Receptor (mAChR) activation is an attractive mechanism for AD therapy since M1 mediates key effects on memory, cognition, and behavior and has potential for disease-modifying effects on Aβ formation and tau phosphorylation. To validate M1 as a neuroprotective treatment target for AD, the M1-selective agonist, VU0364572, was chronically dosed to 5XFAD mice from a young age preceding Aβ pathology (2 months) to an age where these mice are known to display memory impairments (6 months). Chronic M1 activation prevented mice from becoming memory-impaired, as measured by Morris water maze (MWM) testing at 6 months of age. Additionally, M1 activation significantly reduced levels of soluble and insoluble Aβ40,42 in the cortex and hippocampus of these animals, as measured b...
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Muscarinic Acetylcholine Receptor subtypes in cerebral cortex and hippocampus
Progress in Brain Research, 2004Co-Authors: Laura A Volpicelli, Allan I. LeveyAbstract:Publisher Summary This chapter focuses on the Muscarinic Acetylcholine Receptor (mAChR) family, whose five members participate in critical cholinergic functions including learning, memory, and attention, also presents their distribution. The five subtypes of mAChRs include M 1 , M 2 , M 3 , M 4 , and M 5 . Quantitative analysis of mAChR protein distribution performed in the rat brain using immunoprecipitations with subtype selective antibodies reveals that the M 1 , M 2 , and M 4 mAChRs are the predominate mAChR subtypes expressed in the brain. M 3 and M 5 are also expressed throughout the brain, but in low abundance. In the human brain, M 1 is the primary Receptor in the frontal, temporal, parietal, and occipital cortical areas, representing 35–60% of total mAChRs. M 2 is more abundant in the occipital cortex of human brain (36%) than in the frontal, temporal or parietal cortex (approximately 20%). M 4 represents only approximately 20% of total the mAChRs in the human brain cortex. Electron microscopic studies in primate brain demonstrate that M 1 localizes to postsynaptic dendrites and spines that associate with both asymmetric synapses and symmetric cholinergic synapses, indicating that M 1 can modulate excitatory as well as cholinergic transmission. mAChRs expressed in the hippocampus, play a role in learning and memory, and degeneration of cholinergic projections to the hippocampus has been implicated in Alzheimer's disease.
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rab11a and myosin vb regulate recycling of the m4 Muscarinic Acetylcholine Receptor
The Journal of Neuroscience, 2002Co-Authors: Laura A Volpicelli, James J Lah, Guofu Fang, James R Goldenring, Allan I. LeveyAbstract:Agonist-induced internalization followed by subsequent return to the cell surface regulates G-protein-coupled Receptor (GPCR) activity. Because the cellular responsiveness to ligand depends on the balance between Receptor degradation and recycling, it is crucial to identify the molecules involved in GPCR recovery to the cell surface. In this study, we identify mechanisms involved in the recycling of the M4 subtype of Muscarinic Acetylcholine Receptor. M4 is highly expressed in the CNS, plays a role in locomotor activity, and is a novel therapeutic target for neurologic and psychiatric disorders. Previous studies show that, after cholinergic stimulation, M4 internalizes from the cell surface to endosomes in cell culture and the rat brain. Here, we show that, after activation, M4 traffics to transferrin Receptor- and Rab11a-positive perinuclear endosomes. Expression of the constitutively GDP-bound, inactive mutant Rab11aS25N inhibits M4 trafficking to recycling endosomes. Expression of the C-terminal tail of myosin Vb, a Rab11a effector, enhances M4 accumulation in perinuclear endosomes. Both Rab11aS25N and the myosin Vb tail impair M4 recycling. The results demonstrate that GPCR recycling is mediated through a discrete pathway using both Rab11a and myosin Vb.
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Evaluation of Muscarinic Agonist-Induced Analgesia in Muscarinic Acetylcholine Receptor Knockout Mice
Molecular pharmacology, 2002Co-Authors: Alokesh Duttaroy, Jesus Gomeza, Christian C Felder, Jai-wei Gan, Nasir A. Siddiqui, Anthony S. Basile, W. Dean Harman, Philip L. Smith, Allan I. Levey, Jurgen WessAbstract:Centrally active Muscarinic agonists display pronounced analgesic effects. Identification of the specific Muscarinic Acetylcholine Receptor (mAChR) subtype(s) mediating this activity is of considerable therapeutic interest. To examine the roles of the M 2 and M 4 Receptor subtypes, the two G i /G o -coupled mAChRs, in mediating agonist-dependent antinociception, we generated a mutant mouse line deficient in both M 2 and M 4 mAChRs [M 2 /M 4 double-knockout (KO) mice]. In wild-type mice, systemic, intrathecal, or intracerebroventricular administration of centrally active Muscarinic agonists resulted in robust analgesic effects, indicating that Muscarinic analgesia can be mediated by both spinal and supraspinal mechanisms. Strikingly, Muscarinic agonist-induced antinociception was totally abolished in M 2 /M 4 double-KO mice, independent of the route of application. The nonselective Muscarinic agonist oxotremorine showed reduced analgesic potency in M 2 Receptor single-KO mice, but retained full analgesic activity in M 4 Receptor single-KO mice. In contrast, two novel Muscarinic agonists chemically derived from epibatidine, CMI-936 and CMI-1145, displayed reduced analgesic activity in both M 2 and M 4 Receptor single-KO mice, independent of the route of application. Radioligand binding studies indicated that the two CMI compounds, in contrast to oxotremorine, showed >6-fold higher affinity for M 4 than for M 2 Receptors, providing a molecular basis for the observed differences in agonist activity profiles. These data provide unambiguous evidence that Muscarinic analgesia is exclusively mediated by a combination of M 2 and M 4 mAChRs at both spinal and supraspinal sites. These findings should be of considerable relevance for the development of Receptor subtype-selective Muscarinic agonists as novel analgesic drugs.
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novel interaction between the m4 Muscarinic Acetylcholine Receptor and elongation factor 1a2
Journal of Biological Chemistry, 2002Co-Authors: Daniel B Mcclatchy, Charlotte R Knudsen, Brian F C Clark, Richard A Kahn, Randy A Hall, Allan I. LeveyAbstract:The activation of the Muscarinic Acetylcholine Receptor (mAChR) family, consisting of five subtypes (M1-M5), produces a variety of physiological effects throughout the central nervous system. However, the role of each individual subtype remains poorly understood. To further elucidate signal transduction pathways for specific subtypes, we used the most divergent portion of the subtypes, the intracellular third (i3) loop, as bait to identify interacting proteins. Using a brain pull-down assay, we identify elongation factor 1A2 (eEF1A2) as a specific binding partner to the i3 loop of M4, and not to M1 or M2. In addition, we demonstrate a direct interaction between these proteins. In the rat striatum, the M4 mAChR colocalizes with eEF1A2 in the soma and neuropil. In PC12 cells, endogenous eEF1A2 co-immunoprecipitates with the endogenous M4 mAChR, but not with the endogenous M1 mAChR. In our in vitro model, M4 dramatically accelerates nucleotide exchange of eEF1A2, a GTP-binding protein. This indicates the M4 mAChR is a guanine exchange factor for eEF1A2. eEF1A2 is an essential GTP-binding protein for protein synthesis. Thus, our data suggest a novel role for M4 in the regulation of protein synthesis through its interaction with eEF1A2.
Christian C Felder - One of the best experts on this subject based on the ideXlab platform.
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Pharmacological Characterization of LY593093, an M1 Muscarinic Acetylcholine Receptor-Selective Partial Orthosteric Agonist □ S
2020Co-Authors: Marla L Watt, Douglas A Schober, Stephen Hitchcock, Bin Liu, Amy K Chesterfield, David Mckinzie, Christian C FelderAbstract:ABSTRACT Alzheimer's disease and schizophrenia are characterized by expression of psychotic, affective, and cognitive symptoms. Currently, there is a lack of adequate treatment for the cognitive symptoms associated with these diseases. Cholinergic signaling and, in particular, M1 Muscarinic Acetylcholine Receptor (m1AChR) signaling have been implicated in the regulation of multiple cognitive domains. Thus, the M1AChR has been identified as a therapeutic drug target for diseases, such as schizophrenia and Alzheimer's disease, that exhibit marked cognitive dysfunction as part of their clinical manifestation. Unfortunately, the development of selective M1 agonist medications has not been successful, mostly because of the highly conserved orthosteric Acetylcholine binding site among the five Muscarinic Receptor subtypes. More recent efforts have focused on the development of allosteric M1AChR modulators that target regions of the Receptor distinct from the orthosteric site that are less conserved between family members. However, orthosteric and allosteric ligands may differentially modulate Receptor function and ultimately downstream signaling pathways. Thus, the need for highly selective M1AChR orthosteric agonists still exists, not only as a potential therapeutic but also as a pharmacological tool to better understand the physiologic consequences of M1AChR orthosteric activation. Here, we describe the novel, potent and selective M1AChR orthosteric partial agonist . This compound demonstrates modest to no activity at the other Muscarinic Receptor subtypes, stimulates G␣ qcoupled signaling events as well as -arrestin recruitment, and displays significant efficacy in in vivo models of cognition
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activation and allosteric modulation of a Muscarinic Acetylcholine Receptor
Nature, 2013Co-Authors: Andrew C Kruse, Patrick M Sexton, Arthur Christopoulos, Celine Valant, Katrin Eitel, Harald Hubner, Aaron M Ring, Aashish Manglik, Els Pardon, Christian C FelderAbstract:Despite recent advances in crystallography and the availability of G-protein-coupled Receptor (GPCR) structures, little is known about the mechanism of their activation process, as only the β2 adrenergic Receptor (β2AR) and rhodopsin have been crystallized in fully active conformations. Here we report the structure of an agonist-bound, active state of the human M2 Muscarinic Acetylcholine Receptor stabilized by a G-protein mimetic camelid antibody fragment isolated by conformational selection using yeast surface display. In addition to the expected changes in the intracellular surface, the structure reveals larger conformational changes in the extracellular region and orthosteric binding site than observed in the active states of the β2AR and rhodopsin. We also report the structure of the M2 Receptor simultaneously bound to the orthosteric agonist iperoxo and the positive allosteric modulator LY2119620. This structure reveals that LY2119620 recognizes a largely pre-formed binding site in the extracellular vestibule of the iperoxo-bound Receptor, inducing a slight contraction of this outer binding pocket. These structures offer important insights into the activation mechanism and allosteric modulation of Muscarinic Receptors.
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pharmacological characterization of ly593093 an m1 Muscarinic Acetylcholine Receptor selective partial orthosteric agonist
Journal of Pharmacology and Experimental Therapeutics, 2011Co-Authors: Marla L Watt, Douglas A Schober, Stephen Hitchcock, Bin Liu, Amy K Chesterfield, David L Mckinzie, Christian C FelderAbstract:Alzheimer's disease and schizophrenia are characterized by expression of psychotic, affective, and cognitive symptoms. Currently, there is a lack of adequate treatment for the cognitive symptoms associated with these diseases. Cholinergic signaling and, in particular, M1 Muscarinic Acetylcholine Receptor (m1AChR) signaling have been implicated in the regulation of multiple cognitive domains. Thus, the M1AChR has been identified as a therapeutic drug target for diseases, such as schizophrenia and Alzheimer's disease, that exhibit marked cognitive dysfunction as part of their clinical manifestation. Unfortunately, the development of selective M1 agonist medications has not been successful, mostly because of the highly conserved orthosteric Acetylcholine binding site among the five Muscarinic Receptor subtypes. More recent efforts have focused on the development of allosteric M1AChR modulators that target regions of the Receptor distinct from the orthosteric site that are less conserved between family members. However, orthosteric and allosteric ligands may differentially modulate Receptor function and ultimately downstream signaling pathways. Thus, the need for highly selective M1AChR orthosteric agonists still exists, not only as a potential therapeutic but also as a pharmacological tool to better understand the physiologic consequences of M1AChR orthosteric activation. Here, we describe the novel, potent and selective M1AChR orthosteric partial agonist LY593093 [N-[(1R,2R)-6-({(1E)-1-[(4-fluorobenzyl)(methyl)amino]ethylidene})amino)-2-hydroxy-2,3-dihydro-1H-inden-1-yl]biphenyl-4-carboxamide]. This compound demonstrates modest to no activity at the other Muscarinic Receptor subtypes, stimulates Gα(q)-coupled signaling events as well as β-arrestin recruitment, and displays significant efficacy in in vivo models of cognition.
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Evaluation of Muscarinic Agonist-Induced Analgesia in Muscarinic Acetylcholine Receptor Knockout Mice
Molecular pharmacology, 2002Co-Authors: Alokesh Duttaroy, Jesus Gomeza, Christian C Felder, Jai-wei Gan, Nasir A. Siddiqui, Anthony S. Basile, W. Dean Harman, Philip L. Smith, Allan I. Levey, Jurgen WessAbstract:Centrally active Muscarinic agonists display pronounced analgesic effects. Identification of the specific Muscarinic Acetylcholine Receptor (mAChR) subtype(s) mediating this activity is of considerable therapeutic interest. To examine the roles of the M 2 and M 4 Receptor subtypes, the two G i /G o -coupled mAChRs, in mediating agonist-dependent antinociception, we generated a mutant mouse line deficient in both M 2 and M 4 mAChRs [M 2 /M 4 double-knockout (KO) mice]. In wild-type mice, systemic, intrathecal, or intracerebroventricular administration of centrally active Muscarinic agonists resulted in robust analgesic effects, indicating that Muscarinic analgesia can be mediated by both spinal and supraspinal mechanisms. Strikingly, Muscarinic agonist-induced antinociception was totally abolished in M 2 /M 4 double-KO mice, independent of the route of application. The nonselective Muscarinic agonist oxotremorine showed reduced analgesic potency in M 2 Receptor single-KO mice, but retained full analgesic activity in M 4 Receptor single-KO mice. In contrast, two novel Muscarinic agonists chemically derived from epibatidine, CMI-936 and CMI-1145, displayed reduced analgesic activity in both M 2 and M 4 Receptor single-KO mice, independent of the route of application. Radioligand binding studies indicated that the two CMI compounds, in contrast to oxotremorine, showed >6-fold higher affinity for M 4 than for M 2 Receptors, providing a molecular basis for the observed differences in agonist activity profiles. These data provide unambiguous evidence that Muscarinic analgesia is exclusively mediated by a combination of M 2 and M 4 mAChRs at both spinal and supraspinal sites. These findings should be of considerable relevance for the development of Receptor subtype-selective Muscarinic agonists as novel analgesic drugs.
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cholinergic dilation of cerebral blood vessels is abolished in m 5 Muscarinic Acetylcholine Receptor knockout mice
Proceedings of the National Academy of Sciences of the United States of America, 2001Co-Authors: Masahisa Yamada, Christian C Felder, Alokesh Duttaroy, Yinghong Cui, David L Mckinzie, Kathryn G Lamping, Weilie Zhang, Frank P Bymaster, Chuxia Deng, Frank M FaraciAbstract:Abstract The M5 Muscarinic Receptor is the most recent member of the Muscarinic Acetylcholine Receptor family (M1-M5) to be cloned. At present, the physiological relevance of this Receptor subtype remains unknown, primarily because of its low expression levels and the lack of M5 Receptor-selective ligands. To circumvent these difficulties, we used gene targeting technology to generate M5 Receptor-deficient mice (M5R−/− mice). M5R−/− mice did not differ from their wild-type littermates in various behavioral and pharmacologic tests. However, in vitro neurotransmitter release experiments showed that M5 Receptors play a role in facilitating Muscarinic agonist-induced dopamine release in the striatum. Because M5 Receptor mRNA has been detected in several blood vessels, we also investigated whether the lack of M5 Receptors led to changes in vascular tone by using several in vivo and in vitro vascular preparations. Strikingly, Acetylcholine, a powerful dilator of most vascular beds, virtually lost the ability to dilate cerebral arteries and arterioles in M5R−/− mice. This effect was specific for cerebral blood vessels, because Acetylcholine-mediated dilation of extra-cerebral arteries remained fully intact in M5R−/− mice. Our findings provide direct evidence that M5 Muscarinic Receptors are physiologically relevant. Because it has been suggested that impaired cholinergic dilation of cerebral blood vessels may play a role in the pathophysiology of Alzheimer's disease and focal cerebral ischemia, cerebrovascular M5 Receptors may represent an attractive therapeutic target.
Norbert Pfeiffer - One of the best experts on this subject based on the ideXlab platform.
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the m1 Muscarinic Acetylcholine Receptor subtype is important for retinal neuron survival in aging mice
Scientific Reports, 2019Co-Authors: Panagiotis Laspas, Norbert Pfeiffer, Mayagozel B Zhutdieva, Christoph Brochhausen, Aytan Musayeva, Jenia Kouchek Zadeh, Huige Li, J Wess, Adrian GerickeAbstract:Muscarinic Acetylcholine Receptors have been implicated as potential neuroprotective targets for glaucoma. We tested the hypothesis that the lack of a single Muscarinic Receptor subtype leads to age-dependent neuron reduction in the retinal ganglion cell layer. Mice with targeted disruption of single Muscarinic Acetylcholine Receptor subtype genes (M1 to M5) and wild-type controls were examined at two age categories, 5 and 15 months, respectively. We found no differences in intraocular pressure between individual mouse groups. Remarkably, in 15-month-old mice devoid of the M1 Receptor, neuron number in the retinal ganglion cell layer and axon number in the optic nerve were markedly reduced. Moreover, mRNA expression for the prooxidative enzyme, NOX2, was increased, while mRNA expression for the antioxidative enzymes, SOD1, GPx1 and HO-1, was reduced in aged M1 Receptor-deficient mice compared to age-matched wild-type mice. In line with these findings, the reactive oxygen species level was also elevated in the retinal ganglion cell layer of aged M1 Receptor-deficient mice. In conclusion, M1 Receptor deficiency results in retinal ganglion cell loss in aged mice via involvement of oxidative stress. Based on these findings, activation of M1 Receptor signaling may become therapeutically useful to promote retinal ganglion cell survival.
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the m 1 Muscarinic Acetylcholine Receptor subtype is important for retinal neuron survival in aging mice
Scientific Reports, 2019Co-Authors: Panagiotis Laspas, Norbert Pfeiffer, Mayagozel B Zhutdieva, Christoph Brochhausen, Aytan Musayeva, Jenia Kouchek Zadeh, J Wess, Ning Xia, Adrian GerickeAbstract:Muscarinic Acetylcholine Receptors have been implicated as potential neuroprotective targets for glaucoma. We tested the hypothesis that the lack of a single Muscarinic Receptor subtype leads to age-dependent neuron reduction in the retinal ganglion cell layer. Mice with targeted disruption of single Muscarinic Acetylcholine Receptor subtype genes (M1 to M5) and wild-type controls were examined at two age categories, 5 and 15 months, respectively. We found no differences in intraocular pressure between individual mouse groups. Remarkably, in 15-month-old mice devoid of the M1 Receptor, neuron number in the retinal ganglion cell layer and axon number in the optic nerve were markedly reduced. Moreover, mRNA expression for the prooxidative enzyme, NOX2, was increased, while mRNA expression for the antioxidative enzymes, SOD1, GPx1 and HO-1, was reduced in aged M1 Receptor-deficient mice compared to age-matched wild-type mice. In line with these findings, the reactive oxygen species level was also elevated in the retinal ganglion cell layer of aged M1 Receptor-deficient mice. In conclusion, M1 Receptor deficiency results in retinal ganglion cell loss in aged mice via involvement of oxidative stress. Based on these findings, activation of M1 Receptor signaling may become therapeutically useful to promote retinal ganglion cell survival.
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role of the m3 Muscarinic Acetylcholine Receptor subtype in murine ophthalmic arteries after endothelial removal
Investigative Ophthalmology & Visual Science, 2014Co-Authors: Adrian Gericke, Jurgen Wess, Andreas Steege, Caroline Manicam, Tobias Bohmer, Norbert PfeifferAbstract:Purpose. We tested the hypothesis that the M3 Muscarinic Acetylcholine Receptor subtype mediates cholinergic responses in murine ophthalmic arteries after endothelial removal.
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cholinergic responses of ophthalmic arteries in m3 and m5 Muscarinic Acetylcholine Receptor knockout mice
Investigative Ophthalmology & Visual Science, 2009Co-Authors: Adrian Gericke, Jurgen Wess, Andreas Steege, Veronique G A Mayer, Andreas Patzak, Ulrike Neumann, Franz H Grus, Stephanie C Joachim, Lars Choritz, Norbert PfeifferAbstract:PURPOSE. To determine the functional role of M 3 and M 5 Muscarinic Acetylcholine Receptor subtypes in ophthalmic arteries using gene-targeted mice. METHODS. Muscarinic Receptor gene expression was quantified in murine ophthalmic arteries using real-time PCR. To test the functional relevance of M 3 and M 5 Receptors, ophthalmic arteries from mice deficient in either subtype (M3R -/- , M5R -/- , respectively) and wild-type controls were isolated, cannulated with micropipettes, and pressurized. Changes in luminal vessel diameter in response to Muscarinic and nonMuscarinic Receptor agonists were measured by video microscopy. RESULTS. With the use of real-time PCR, all five Muscarinic Receptor subtypes were detected in ophthalmic arteries. However, mRNA levels of M 1 , M 3 , and M 5 Receptors were higher than those of M 2 , and M 4 Receptors. In functional studies, after preconstriction with phenylephrine, Acetylcholine and carbachol produced concentration-dependent dilations of ophthalmic arteries that were similar in M5R -/- and wild-type mice. Strikingly, cholinergic dilation of ophthalmic arteries was almost completely abolished in M3R -/- mice. Deletion of either M 3 or M 5 Receptor did not affect responses to nonMuscarinic vasodilators such as bradykinin or nitroprusside. CONCLUSIONS. These findings provide the first evidence that M 3 Receptors are critically involved in cholinergic regulation of diameter in murine ophthalmic arteries.
Arthur Christopoulos - One of the best experts on this subject based on the ideXlab platform.
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restoring agonist function at a chemogenetically modified m1 Muscarinic Acetylcholine Receptor
ACS Chemical Neuroscience, 2020Co-Authors: Elham Khajehali, Craig W Lindsley, Patrick M Sexton, Sophie J Bradley, Emma T Van Der Westhuizen, Colin Molloy, Celine Valant, Lisa Finlayson, Andrew B Tobin, Arthur ChristopoulosAbstract:Designer Receptors exclusively activated by designer drugs (DREADDs) have been successfully employed to activate signaling pathways associated with specific Muscarinic Acetylcholine Receptor (mAChR) subtypes. The M1 DREADD mAChR displays minimal responsiveness to the endogenous agonist Acetylcholine (ACh) but responds to clozapine-N-oxide (CNO), an otherwise pharmacologically inert ligand. We have previously shown that benzyl quinolone carboxylic acid (BQCA), an M1 mAChR positive allosteric modulator (PAM), can rescue ACh responsiveness at these Receptors. However, whether this effect is chemotype specific or applies to next-generation M1 PAMs with distinct scaffolds is unknown. Here, we reveal that new M1 PAMs restore ACh function at the M1 DREADD while modulating ACh binding at the M1 wild-type mAChR. Importantly, we demonstrate that the modulation of ACh function by M1 PAMs is translated in vivo using transgenic M1 DREADD mice. Our data provide important insights into mechanisms that define allosteric ligand modulation of agonist affinity vs efficacy and how these effects play out in the regulation of in vivo responses.
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novel fused arylpyrimidinone based allosteric modulators of the m1 Muscarinic Acetylcholine Receptor
ACS Chemical Neuroscience, 2016Co-Authors: Shailesh N Mistry, Manuela Jorg, Ben Capuano, Arthur Christopoulos, Robert J Lane, Peter J ScammellsAbstract:Benzoquinazolinone 1 is a positive allosteric modulator (PAM) of the M1 Muscarinic Acetylcholine Receptor (mAChR), which is significantly more potent than the prototypical PAM, 1-(4-methoxybenzyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (BQCA). In this study, we explored the structural determinants that underlie the activity of 1 as a PAM of the M1 mAChR. We paid particular attention to the importance of the tricyclic scaffold of compound 1, for the activity of the molecule. Complete deletion of the peripheral fused benzene ring caused a significant decrease in affinity and binding cooperativity with Acetylcholine (ACh). This loss of affinity was rescued with the addition of either one or two methyl groups in the 7- and/or 8-position of the quinazolin-4(3H)-one core. These results demonstrate that the tricyclic benzo[h]quinazolin-4(3H)-one core could be replaced with a quinazolin-4(3H)-one core and maintain functional affinity. As such, the quinazolin-4(3H)-one core represents a novel scaffold to fur...
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4 phenylpyridin 2 one derivatives a novel class of positive allosteric modulator of the m1 Muscarinic Acetylcholine Receptor
Journal of Medicinal Chemistry, 2016Co-Authors: Shailesh N Mistry, Manuela Jorg, Natalie B Vinh, Patrick M Sexton, Ben Capuano, Arthur Christopoulos, Robert J Lane, Peter J ScammellsAbstract:Positive allosteric modulators (PAMs) of the M1 Muscarinic Acetylcholine Receptor (M1 mAChR) are a promising strategy for the treatment of the cognitive deficits associated with diseases including Alzheimer’s and schizophrenia. Herein, we report the design, synthesis, and characterization of a novel family of M1 mAChR PAMs. The most active compounds of the 4-phenylpyridin-2-one series exhibited comparable binding affinity to the reference compound, 1-(4-methoxybenzyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid (BQCA) (1), but markedly improved positive cooperativity with Acetylcholine, and retained exquisite selectivity for the M1 mAChR. Furthermore, our pharmacological characterization revealed ligands with a diverse range of activities, including modulators that displayed both high intrinsic efficacy and PAM activity, those that showed no detectable agonism but robust PAM activity and ligands that displayed robust allosteric agonism but little modulatory activity. Thus, the 4-phenylpyridin-2-one scaff...
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mechanistic insights into allosteric structure function relationships at the m1 Muscarinic Acetylcholine Receptor
Journal of Biological Chemistry, 2014Co-Authors: Alaa Abdulridha, Shailesh N Mistry, Patrick M Sexton, Arthur Christopoulos, Robert J Lane, Peter J Scammells, Laura Lopez, Meritxell CanalsAbstract:Abstract Benzylquinolone carboxylic acid (BQCA) is the first highly selective positive allosteric modulator (PAM) for the M1 Muscarinic Acetylcholine Receptor (mAChR), but it possesses low affinity for the allosteric site on the Receptor. More recent drug discovery efforts identified 3-((1S,2S)-2-hydroxycyclohexyl)-6-((6-(1-methyl-1H-pyrazol-4-yl)pyridin-3-yl)methyl)benzo[h]quinazolin-4(3H)-one (referred to herein as benzoquinazolinone 12) as a more potent M1 mAChR PAM with a structural ancestry originating from BQCA and related compounds. In the current study, we optimized the synthesis of and fully characterized the pharmacology of benzoquinazolinone 12, finding that its improved potency derived from a 50-fold increase in allosteric site affinity as compared with BQCA, while retaining a similar level of positive cooperativity with Acetylcholine. We then utilized site-directed mutagenesis and molecular modeling to validate the allosteric binding pocket we previously described for BQCA as a shared site for benzoquinazolinone 12 and provide a molecular basis for its improved activity at the M1 mAChR. This includes a key role for hydrophobic and polar interactions with residues Tyr-179, in the second extracellular loop (ECL2) and Trp-4007.35 in transmembrane domain (TM) 7. Collectively, this study highlights how the properties of affinity and cooperativity can be differentially modified on a common structural scaffold and identifies molecular features that can be exploited to tailor the development of M1 mAChR-targeting PAMs.
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mechanistic insights into allosteric structure function relationships at the m1 Muscarinic Acetylcholine Receptor
Journal of Biological Chemistry, 2014Co-Authors: Alaa Abdulridha, Shailesh N Mistry, Patrick M Sexton, Arthur Christopoulos, Robert J Lane, Peter J Scammells, Laura Lopez, Meritxell CanalsAbstract:Benzylquinolone carboxylic acid (BQCA) is the first highly selective positive allosteric modulator (PAM) for the M1 Muscarinic Acetylcholine Receptor (mAChR), but it possesses low affinity for the allosteric site on the Receptor. More recent drug discovery efforts identified 3-((1S,2S)-2-hydroxycyclohexyl)-6-((6-(1-methyl-1H-pyrazol-4-yl)pyridin-3-yl)methyl)benzo[h]quinazolin-4(3H)-one (referred to herein as benzoquinazolinone 12) as a more potent M1 mAChR PAM with a structural ancestry originating from BQCA and related compounds. In the current study, we optimized the synthesis of and fully characterized the pharmacology of benzoquinazolinone 12, finding that its improved potency derived from a 50-fold increase in allosteric site affinity as compared with BQCA, while retaining a similar level of positive cooperativity with Acetylcholine. We then utilized site-directed mutagenesis and molecular modeling to validate the allosteric binding pocket we previously described for BQCA as a shared site for benzoquinazolinone 12 and provide a molecular basis for its improved activity at the M1 mAChR. This includes a key role for hydrophobic and polar interactions with residues Tyr-179, in the second extracellular loop (ECL2) and Trp-4007.35 in transmembrane domain (TM) 7. Collectively, this study highlights how the properties of affinity and cooperativity can be differentially modified on a common structural scaffold and identifies molecular features that can be exploited to tailor the development of M1 mAChR-targeting PAMs.
