{"gene":"CHRM5","run_date":"2026-06-09T22:57:18","timeline":{"discoveries":[{"year":2021,"finding":"M5 receptor (CHRM5) in the ventral subiculum (vSub) modulates initial motivation for alcohol self-administration; intra-vSub negative allosteric modulation of M5 (ML375) reduced alcohol self-administration but not context-induced alcohol-seeking. Chrm5 mRNA was up-regulated in vSub following long-term alcohol consumption. Chrm5 was localized on vSub cells projecting to the nucleus accumbens shell (AcbSh), as shown by retrograde tracing combined with RNAscope.","method":"RT-qPCR, retrograde tracing, RNAscope in situ hybridization, intra-vSub pharmacology with selective negative allosteric modulator (ML375) in alcohol-preferring rats","journal":"British journal of pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization by retrograde tracing + RNAscope and functional pharmacology in vivo, single lab but two orthogonal methods","pmids":["33942300"],"is_preprint":false},{"year":2025,"finding":"Chrm5-eNOS-nitric oxide signaling in bone marrow arterial cells mediates arterial dilation; age-related decline in non-neurogenic acetylcholine disrupts this Chrm5-eNOS-NO axis, reducing arterial dilation, bone marrow blood flow, sinusoidal wall shear stress, Piezo1 activation, and consequently hematopoietic stem/progenitor cell (HSPC) transendothelial migration and homing efficiency.","method":"In vivo imaging, pharmacological manipulation, integrative metabolomic analyses, genetic/signaling pathway dissection in mouse bone marrow","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo functional imaging plus metabolomics and pharmacological rescue in a single study; replication not yet established","pmids":["40593589"],"is_preprint":false},{"year":1999,"finding":"Mouse Chrm5 was mapped to chromosome 2 by restriction fragment length variant (RFLV) analysis in interspecific backcross mice. Sequencing of Chrm5 in El2 (epilepsy 2) mutant mice did not support Chrm5 as the El2 mutation.","method":"RFLV mapping in interspecific backcross mice; Chrm5 sequencing in El2 mutant mice","journal":"Genes & genetic systems","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct genetic mapping and sequencing; negative finding for El2 is explicit","pmids":["10549128"],"is_preprint":false},{"year":2002,"finding":"Rat Chrm5 was mapped to chromosome 3 by radiation hybrid (RH) mapping.","method":"Radiation hybrid mapping with rat RH map server","journal":"Hearing research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct RH mapping experiment, single lab","pmids":["12433399"],"is_preprint":false},{"year":2012,"finding":"In the murine oviduct, RT-PCR of laser-microdissected epithelium revealed expression of Chrm1 and Chrm3 but not Chrm5 in ampullar epithelium. Mice with targeted deletion of Chrm5 (along with Chrm1, 3, 4) showed normal high basal ciliary beat frequency and particle transport speed in the oviductal ampulla, indicating oviductal ciliary activity is independent of Chrm5.","method":"RT-PCR of laser-microdissected oviductal epithelium; ciliary beat frequency and particle transport speed measurements in Chrm knockout mice","journal":"Biology of reproduction","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct knockout functional measurement; negative result for Chrm5 role in oviductal cilia, single lab","pmids":["22302687"],"is_preprint":false},{"year":2023,"finding":"A homozygous missense variant (p.Gln184Arg) in CHRM5 was identified by exome sequencing in a patient with neurogenic bladder; CHRM5 is expressed in murine and human bladder walls and Chrm5 knockout mice display bladder overactivity. However, in vitro functional studies of the variant did not provide evidence to confirm pathogenicity.","method":"Exome sequencing; expression analysis in bladder tissue; functional in vitro studies of variant (result: negative/inconclusive)","journal":"American journal of medical genetics. Part A","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single case, in vitro functional studies were negative/uninformative; expression data only correlative","pmids":["37213061"],"is_preprint":false},{"year":2021,"finding":"Forced expression of MYOCD in human coronary artery smooth muscle cells increased CHRM5 mRNA approximately 2-fold (along with CHRM2 and CHRM3), as shown by RNA-sequencing and RT-qPCR, indicating CHRM5 transcription is upregulated by myocardin-related transcription factors.","method":"Forced MYOCD expression in human coronary artery SMCs followed by RNA-sequencing and RT-qPCR","journal":"Frontiers in physiology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, single expression-level readout without direct mechanistic follow-up for CHRM5 specifically","pmids":["34539433"],"is_preprint":false}],"current_model":"CHRM5 (M5 muscarinic acetylcholine receptor) is a Gq/11-coupled GPCR expressed in dopaminergic brain regions, bladder, bone marrow vasculature, and other tissues; in the ventral subiculum it controls initial motivation for alcohol self-administration via projections to the nucleus accumbens shell, and in bone marrow arterial cells it operates through a Chrm5–eNOS–nitric oxide axis to regulate arterial dilation, blood flow, wall shear stress, and consequently hematopoietic stem cell homing efficiency."},"narrative":{"mechanistic_narrative":"CHRM5 (M5 muscarinic acetylcholine receptor) is a muscarinic receptor expressed across neural and vascular tissues where it transduces cholinergic signals into region-specific physiological outputs [PMID:33942300, PMID:40593589]. In the ventral subiculum, CHRM5 is expressed on neurons projecting to the nucleus accumbens shell and controls initial motivation for alcohol self-administration: its mRNA is up-regulated after long-term alcohol consumption, and selective negative allosteric modulation reduces alcohol self-administration [PMID:33942300]. In bone marrow arterial cells, CHRM5 operates through an eNOS-nitric oxide axis to drive arterial dilation, and this Chrm5-eNOS-NO signaling controls bone marrow blood flow and sinusoidal wall shear stress, thereby gating Piezo1 activation and hematopoietic stem/progenitor cell transendothelial migration and homing [PMID:40593589]. CHRM5 is also expressed in bladder wall tissue, where Chrm5-knockout mice display bladder overactivity [PMID:37213061]. Beyond these tissue-level roles, the receptor's downstream effector coupling has not been characterized in detail in the available corpus apart from the eNOS-NO axis [PMID:40593589].","teleology":[{"year":1999,"claim":"Establishing the chromosomal location of Chrm5 and testing it as a candidate disease gene clarified its genetic identity and excluded it as the El2 epilepsy locus.","evidence":"RFLV mapping in interspecific backcross mice and Chrm5 sequencing in El2 mutant mice","pmids":["10549128"],"confidence":"Medium","gaps":["No functional or signaling role addressed","Mapping alone does not establish tissue expression or activity"]},{"year":2002,"claim":"Radiation hybrid mapping localized rat Chrm5, extending its genetic characterization across species.","evidence":"Radiation hybrid mapping with rat RH map server","pmids":["12433399"],"confidence":"Medium","gaps":["No functional consequence established","No link to a specific tissue or pathway"]},{"year":2012,"claim":"Testing whether Chrm5 contributes to oviductal ciliary activity defined a tissue where the receptor is dispensable, sharpening the boundaries of its physiological role.","evidence":"RT-PCR of laser-microdissected oviductal epithelium and ciliary measurements in Chrm knockout mice","pmids":["22302687"],"confidence":"Medium","gaps":["Negative result; does not identify where Chrm5 IS required","Compound knockout complicates attribution to Chrm5 specifically"]},{"year":2021,"claim":"Linking CHRM5 in the ventral subiculum to alcohol motivation answered where and how the receptor modulates addiction-relevant behavior, identifying a defined neural circuit.","evidence":"RT-qPCR, retrograde tracing, RNAscope, and intra-vSub pharmacology with the negative allosteric modulator ML375 in alcohol-preferring rats","pmids":["33942300"],"confidence":"Medium","gaps":["Downstream signaling within vSub neurons not defined","Mechanism linking M5 modulation to projection activity unresolved","Single lab"]},{"year":2021,"claim":"Identifying CHRM5 as a transcriptional target of myocardin-related factors hinted at how its expression is regulated in vascular smooth muscle.","evidence":"Forced MYOCD expression in human coronary artery SMCs with RNA-seq and RT-qPCR","pmids":["34539433"],"confidence":"Low","gaps":["Expression-level readout only without mechanistic follow-up for CHRM5","Direct promoter regulation not demonstrated","Functional consequence of increased CHRM5 not tested"]},{"year":2023,"claim":"Searching for a CHRM5 disease link in neurogenic bladder tested causality of a candidate variant, but the variant's pathogenicity could not be confirmed.","evidence":"Exome sequencing, bladder expression analysis, and in vitro functional studies of the p.Gln184Arg variant (inconclusive)","pmids":["37213061"],"confidence":"Low","gaps":["In vitro functional studies were negative/uninformative","Single case; expression data only correlative","Causality of CHRM5 in neurogenic bladder unresolved"]},{"year":2025,"claim":"Defining a Chrm5-eNOS-NO axis in bone marrow arteries answered how cholinergic signaling mechanically regulates the hematopoietic niche through vascular dilation and shear stress.","evidence":"In vivo imaging, pharmacological manipulation, metabolomics, and signaling pathway dissection in mouse bone marrow","pmids":["40593589"],"confidence":"Medium","gaps":["Direct receptor-eNOS coupling mechanism not structurally resolved","Replication not yet established","Source/regulation of non-neurogenic acetylcholine partially defined"]},{"year":null,"claim":"The proximal signal transduction events downstream of CHRM5 receptor activation across its varied tissues remain to be unified mechanistically.","evidence":"","pmids":[],"confidence":"Low","gaps":["No structural model of CHRM5 in the corpus","G-protein coupling and second-messenger cascades not directly characterized here","Tissue-specific effector selection unexplained"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060089","term_label":"molecular transducer activity","supporting_discovery_ids":[0,1]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0,1,5]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[1]}],"complexes":[],"partners":[],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P08912","full_name":"Muscarinic acetylcholine receptor M5","aliases":[],"length_aa":532,"mass_kda":60.1,"function":"The muscarinic acetylcholine receptor mediates various cellular responses, including inhibition of adenylate cyclase, breakdown of phosphoinositides and modulation of potassium channels through the action of G proteins. Primary transducing effect is Pi turnover","subcellular_location":"Cell membrane; Postsynaptic cell membrane","url":"https://www.uniprot.org/uniprotkb/P08912/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/CHRM5","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/CHRM5","total_profiled":1310},"omim":[{"mim_id":"118496","title":"CHOLINERGIC RECEPTOR, MUSCARINIC, 5; CHRM5","url":"https://www.omim.org/entry/118496"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enriched","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"brain","ntpm":7.1}],"url":"https://www.proteinatlas.org/search/CHRM5"},"hgnc":{"alias_symbol":[],"prev_symbol":[]},"alphafold":{"accession":"P08912","domains":[{"cath_id":"1.20.1070.10","chopping":"26-234_447-522","consensus_level":"medium","plddt":92.8209,"start":26,"end":522}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P08912","model_url":"https://alphafold.ebi.ac.uk/files/AF-P08912-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P08912-F1-predicted_aligned_error_v6.png","plddt_mean":68.62},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CHRM5","jax_strain_url":"https://www.jax.org/strain/search?query=CHRM5"},"sequence":{"accession":"P08912","fasta_url":"https://rest.uniprot.org/uniprotkb/P08912.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P08912/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P08912"}},"corpus_meta":[{"pmid":"15292665","id":"PMC_15292665","title":"Linkage of M5 muscarinic and alpha7-nicotinic receptor genes on 15q13 to schizophrenia.","date":"2004","source":"Neuropsychobiology","url":"https://pubmed.ncbi.nlm.nih.gov/15292665","citation_count":57,"is_preprint":false},{"pmid":"17163532","id":"PMC_17163532","title":"Characterization of a 5.3 Mb deletion in 15q14 by comparative genomic hybridization using a whole genome \"tiling path\" BAC array in a girl with heart defect, cleft palate, and developmental delay.","date":"2007","source":"American journal of medical genetics. Part A","url":"https://pubmed.ncbi.nlm.nih.gov/17163532","citation_count":56,"is_preprint":false},{"pmid":"19262686","id":"PMC_19262686","title":"Expression of muscarinic receptor subtypes in tree shrew ocular tissues and their regulation during the development of myopia.","date":"2009","source":"Molecular vision","url":"https://pubmed.ncbi.nlm.nih.gov/19262686","citation_count":45,"is_preprint":false},{"pmid":"27375434","id":"PMC_27375434","title":"Long-Term Estrogen Receptor Beta Agonist Treatment Modifies the Hippocampal Transcriptome in Middle-Aged Ovariectomized Rats.","date":"2016","source":"Frontiers in cellular neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/27375434","citation_count":43,"is_preprint":false},{"pmid":"32650830","id":"PMC_32650830","title":"Clinical features, treatment, and survival outcome of primary pulmonary NUT midline carcinoma.","date":"2020","source":"Orphanet journal of rare diseases","url":"https://pubmed.ncbi.nlm.nih.gov/32650830","citation_count":43,"is_preprint":false},{"pmid":"22302687","id":"PMC_22302687","title":"Ciliary activity in the oviduct of cycling, pregnant, and muscarinic receptor knockout mice.","date":"2012","source":"Biology of reproduction","url":"https://pubmed.ncbi.nlm.nih.gov/22302687","citation_count":27,"is_preprint":false},{"pmid":"17608938","id":"PMC_17608938","title":"Variation in the gene coding for the M5 muscarinic receptor (CHRM5) influences cigarette dose but is not associated with dependence to drugs of addiction: evidence from a prospective population based cohort study of young adults.","date":"2007","source":"BMC genetics","url":"https://pubmed.ncbi.nlm.nih.gov/17608938","citation_count":23,"is_preprint":false},{"pmid":"30564629","id":"PMC_30564629","title":"Evolution of the Muscarinic Acetylcholine Receptors in Vertebrates.","date":"2018","source":"eNeuro","url":"https://pubmed.ncbi.nlm.nih.gov/30564629","citation_count":22,"is_preprint":false},{"pmid":"33942300","id":"PMC_33942300","title":"Muscarinic M4 and M5 receptors in the ventral subiculum differentially modulate alcohol seeking versus consumption in male alcohol-preferring rats.","date":"2021","source":"British journal of pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/33942300","citation_count":21,"is_preprint":false},{"pmid":"22899762","id":"PMC_22899762","title":"Muscarinic acetylcholine receptor 3 is dominant in myopia progression.","date":"2012","source":"Investigative ophthalmology & visual science","url":"https://pubmed.ncbi.nlm.nih.gov/22899762","citation_count":21,"is_preprint":false},{"pmid":"20082457","id":"PMC_20082457","title":"An interstitial 15q11-q14 deletion: expanded Prader-Willi syndrome phenotype.","date":"2010","source":"American journal of medical genetics. Part A","url":"https://pubmed.ncbi.nlm.nih.gov/20082457","citation_count":16,"is_preprint":false},{"pmid":"34539433","id":"PMC_34539433","title":"Regulation of the Muscarinic M3 Receptor by Myocardin-Related Transcription Factors.","date":"2021","source":"Frontiers in physiology","url":"https://pubmed.ncbi.nlm.nih.gov/34539433","citation_count":13,"is_preprint":false},{"pmid":"30198559","id":"PMC_30198559","title":"Inter- and intra-specific differences in muscarinic acetylcholine receptor expression in the neural pathways for vocal learning in songbirds.","date":"2018","source":"The Journal of comparative neurology","url":"https://pubmed.ncbi.nlm.nih.gov/30198559","citation_count":11,"is_preprint":false},{"pmid":"35017609","id":"PMC_35017609","title":"Genetic diversity and selection in Puerto Rican horses.","date":"2022","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/35017609","citation_count":11,"is_preprint":false},{"pmid":"38757095","id":"PMC_38757095","title":"The Role of Aryl Hydrocarbon Receptor in Bone Biology.","date":"2024","source":"International journal of tryptophan research : IJTR","url":"https://pubmed.ncbi.nlm.nih.gov/38757095","citation_count":10,"is_preprint":false},{"pmid":"35370785","id":"PMC_35370785","title":"Mechanistic Clues Provided by Concurrent Changes in the Expression of Genes Encoding the M1 Muscarinic Receptor, β-Catenin Signaling Proteins, and Downstream Targets in Adenocarcinomas of the Colon.","date":"2022","source":"Frontiers in physiology","url":"https://pubmed.ncbi.nlm.nih.gov/35370785","citation_count":10,"is_preprint":false},{"pmid":"10549128","id":"PMC_10549128","title":"Mapping of five subtype genes for muscarinic acetylcholine receptor to mouse chromosomes.","date":"1999","source":"Genes & genetic systems","url":"https://pubmed.ncbi.nlm.nih.gov/10549128","citation_count":8,"is_preprint":false},{"pmid":"38550331","id":"PMC_38550331","title":"Exploring the therapeutic mechanism of potential phytocompounds from Kalanchoe pinnata in the treatment of diabetes mellitus by integrating network pharmacology, molecular docking and simulation approach.","date":"2024","source":"Saudi pharmaceutical journal : SPJ : the official publication of the Saudi Pharmaceutical Society","url":"https://pubmed.ncbi.nlm.nih.gov/38550331","citation_count":6,"is_preprint":false},{"pmid":"25121092","id":"PMC_25121092","title":"Expression of acetylcholine receptors by experimental rat renal allografts.","date":"2014","source":"BioMed research international","url":"https://pubmed.ncbi.nlm.nih.gov/25121092","citation_count":5,"is_preprint":false},{"pmid":"32900274","id":"PMC_32900274","title":"Screening natural inhibitors against upregulated G-protein coupled receptors as potential therapeutics of Alzheimer's disease.","date":"2020","source":"Journal of biomolecular structure & dynamics","url":"https://pubmed.ncbi.nlm.nih.gov/32900274","citation_count":5,"is_preprint":false},{"pmid":"37042962","id":"PMC_37042962","title":"Integrated LC-MS/MS and network pharmacology approach for predictingactive ingredients and pharmacological mechanisms of Tribulus terrestris L. against cardiac diseases.","date":"2023","source":"Journal of biomolecular structure & dynamics","url":"https://pubmed.ncbi.nlm.nih.gov/37042962","citation_count":5,"is_preprint":false},{"pmid":"12433399","id":"PMC_12433399","title":"Radiation hybrid mapping of five muscarinic acetylcholine receptor subtype genes in Rattus norvegicus.","date":"2002","source":"Hearing research","url":"https://pubmed.ncbi.nlm.nih.gov/12433399","citation_count":5,"is_preprint":false},{"pmid":"37213061","id":"PMC_37213061","title":"Recessive CHRM5 variant as a potential cause of neurogenic bladder.","date":"2023","source":"American journal of medical genetics. Part A","url":"https://pubmed.ncbi.nlm.nih.gov/37213061","citation_count":3,"is_preprint":false},{"pmid":"36468138","id":"PMC_36468138","title":"Role of non-neuronal cholinergic system in the early stage response of epithelial-mesenchymal transformation related markers in A549 cells induced by coal particles.","date":"2022","source":"Heliyon","url":"https://pubmed.ncbi.nlm.nih.gov/36468138","citation_count":2,"is_preprint":false},{"pmid":"40405417","id":"PMC_40405417","title":"Modulatory effects of M3 muscarinic acetylcholine receptor on inflammatory profiles of human memory T helper cells.","date":"2025","source":"Journal of immunology (Baltimore, Md. : 1950)","url":"https://pubmed.ncbi.nlm.nih.gov/40405417","citation_count":2,"is_preprint":false},{"pmid":"39379070","id":"PMC_39379070","title":"Acute Hypobaric Hypoxia Causes Alterations in Acetylcholine-Mediated Signaling Through Varying Expression of Muscarinic Receptors in the Prefrontal Cortex and Cerebellum of Rats' Brain.","date":"2024","source":"High altitude medicine & biology","url":"https://pubmed.ncbi.nlm.nih.gov/39379070","citation_count":2,"is_preprint":false},{"pmid":"40129096","id":"PMC_40129096","title":"Monoamine neurotransmitter-related gene-based genome-wide association study of low-dose ketamine in patients with treatment-resistant depression.","date":"2025","source":"Journal of psychopharmacology (Oxford, England)","url":"https://pubmed.ncbi.nlm.nih.gov/40129096","citation_count":2,"is_preprint":false},{"pmid":"39740827","id":"PMC_39740827","title":"Epigenetic Modification of Muscarinic Acetylcholine Receptors in Squamous Cell Carcinoma of the Head and Neck.","date":"2025","source":"Anticancer research","url":"https://pubmed.ncbi.nlm.nih.gov/39740827","citation_count":1,"is_preprint":false},{"pmid":"39938415","id":"PMC_39938415","title":"Repurposing of dipeptidyl peptidase FDA-approved drugs in Alzheimer's disease using network pharmacology and in-silico approaches.","date":"2025","source":"Computational biology and chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/39938415","citation_count":1,"is_preprint":false},{"pmid":"40593589","id":"PMC_40593589","title":"Decreased non-neurogenic acetylcholine in bone marrow triggers age-related defective stem/progenitor cell homing.","date":"2025","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/40593589","citation_count":0,"is_preprint":false},{"pmid":"41153348","id":"PMC_41153348","title":"Exploring Genetic Heterogeneity in Type 2 Diabetes Subtypes.","date":"2025","source":"Genes","url":"https://pubmed.ncbi.nlm.nih.gov/41153348","citation_count":0,"is_preprint":false},{"pmid":"41637787","id":"PMC_41637787","title":"Integrative metabolome and transcriptome analyses provide insights into skeletal muscle development of two duck breeds during embryonic stage.","date":"2026","source":"Poultry science","url":"https://pubmed.ncbi.nlm.nih.gov/41637787","citation_count":0,"is_preprint":false},{"pmid":"32405354","id":"PMC_32405354","title":"Biological behaviors of muscarinic receptors in mesenchymal stem cells derived from human placenta and bone marrow.","date":"2020","source":"Iranian journal of basic medical sciences","url":"https://pubmed.ncbi.nlm.nih.gov/32405354","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":20689,"output_tokens":1745,"usd":0.044121,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":8594,"output_tokens":2268,"usd":0.049835,"stage2_stop_reason":"end_turn"},"total_usd":0.093956,"stage1_batch_id":"msgbatch_01ACAxJQabDsht39JXeWG22d","stage2_batch_id":"msgbatch_01QEU22A9U5sbPTgsMzTmvSf","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2021,\n      \"finding\": \"M5 receptor (CHRM5) in the ventral subiculum (vSub) modulates initial motivation for alcohol self-administration; intra-vSub negative allosteric modulation of M5 (ML375) reduced alcohol self-administration but not context-induced alcohol-seeking. Chrm5 mRNA was up-regulated in vSub following long-term alcohol consumption. Chrm5 was localized on vSub cells projecting to the nucleus accumbens shell (AcbSh), as shown by retrograde tracing combined with RNAscope.\",\n      \"method\": \"RT-qPCR, retrograde tracing, RNAscope in situ hybridization, intra-vSub pharmacology with selective negative allosteric modulator (ML375) in alcohol-preferring rats\",\n      \"journal\": \"British journal of pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization by retrograde tracing + RNAscope and functional pharmacology in vivo, single lab but two orthogonal methods\",\n      \"pmids\": [\"33942300\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Chrm5-eNOS-nitric oxide signaling in bone marrow arterial cells mediates arterial dilation; age-related decline in non-neurogenic acetylcholine disrupts this Chrm5-eNOS-NO axis, reducing arterial dilation, bone marrow blood flow, sinusoidal wall shear stress, Piezo1 activation, and consequently hematopoietic stem/progenitor cell (HSPC) transendothelial migration and homing efficiency.\",\n      \"method\": \"In vivo imaging, pharmacological manipulation, integrative metabolomic analyses, genetic/signaling pathway dissection in mouse bone marrow\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo functional imaging plus metabolomics and pharmacological rescue in a single study; replication not yet established\",\n      \"pmids\": [\"40593589\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Mouse Chrm5 was mapped to chromosome 2 by restriction fragment length variant (RFLV) analysis in interspecific backcross mice. Sequencing of Chrm5 in El2 (epilepsy 2) mutant mice did not support Chrm5 as the El2 mutation.\",\n      \"method\": \"RFLV mapping in interspecific backcross mice; Chrm5 sequencing in El2 mutant mice\",\n      \"journal\": \"Genes & genetic systems\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct genetic mapping and sequencing; negative finding for El2 is explicit\",\n      \"pmids\": [\"10549128\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Rat Chrm5 was mapped to chromosome 3 by radiation hybrid (RH) mapping.\",\n      \"method\": \"Radiation hybrid mapping with rat RH map server\",\n      \"journal\": \"Hearing research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct RH mapping experiment, single lab\",\n      \"pmids\": [\"12433399\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"In the murine oviduct, RT-PCR of laser-microdissected epithelium revealed expression of Chrm1 and Chrm3 but not Chrm5 in ampullar epithelium. Mice with targeted deletion of Chrm5 (along with Chrm1, 3, 4) showed normal high basal ciliary beat frequency and particle transport speed in the oviductal ampulla, indicating oviductal ciliary activity is independent of Chrm5.\",\n      \"method\": \"RT-PCR of laser-microdissected oviductal epithelium; ciliary beat frequency and particle transport speed measurements in Chrm knockout mice\",\n      \"journal\": \"Biology of reproduction\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct knockout functional measurement; negative result for Chrm5 role in oviductal cilia, single lab\",\n      \"pmids\": [\"22302687\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"A homozygous missense variant (p.Gln184Arg) in CHRM5 was identified by exome sequencing in a patient with neurogenic bladder; CHRM5 is expressed in murine and human bladder walls and Chrm5 knockout mice display bladder overactivity. However, in vitro functional studies of the variant did not provide evidence to confirm pathogenicity.\",\n      \"method\": \"Exome sequencing; expression analysis in bladder tissue; functional in vitro studies of variant (result: negative/inconclusive)\",\n      \"journal\": \"American journal of medical genetics. Part A\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single case, in vitro functional studies were negative/uninformative; expression data only correlative\",\n      \"pmids\": [\"37213061\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Forced expression of MYOCD in human coronary artery smooth muscle cells increased CHRM5 mRNA approximately 2-fold (along with CHRM2 and CHRM3), as shown by RNA-sequencing and RT-qPCR, indicating CHRM5 transcription is upregulated by myocardin-related transcription factors.\",\n      \"method\": \"Forced MYOCD expression in human coronary artery SMCs followed by RNA-sequencing and RT-qPCR\",\n      \"journal\": \"Frontiers in physiology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, single expression-level readout without direct mechanistic follow-up for CHRM5 specifically\",\n      \"pmids\": [\"34539433\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"CHRM5 (M5 muscarinic acetylcholine receptor) is a Gq/11-coupled GPCR expressed in dopaminergic brain regions, bladder, bone marrow vasculature, and other tissues; in the ventral subiculum it controls initial motivation for alcohol self-administration via projections to the nucleus accumbens shell, and in bone marrow arterial cells it operates through a Chrm5–eNOS–nitric oxide axis to regulate arterial dilation, blood flow, wall shear stress, and consequently hematopoietic stem cell homing efficiency.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"CHRM5 (M5 muscarinic acetylcholine receptor) is a muscarinic receptor expressed across neural and vascular tissues where it transduces cholinergic signals into region-specific physiological outputs [#0, #1]. In the ventral subiculum, CHRM5 is expressed on neurons projecting to the nucleus accumbens shell and controls initial motivation for alcohol self-administration: its mRNA is up-regulated after long-term alcohol consumption, and selective negative allosteric modulation reduces alcohol self-administration [#0]. In bone marrow arterial cells, CHRM5 operates through an eNOS-nitric oxide axis to drive arterial dilation, and this Chrm5-eNOS-NO signaling controls bone marrow blood flow and sinusoidal wall shear stress, thereby gating Piezo1 activation and hematopoietic stem/progenitor cell transendothelial migration and homing [#1]. CHRM5 is also expressed in bladder wall tissue, where Chrm5-knockout mice display bladder overactivity [#5]. Beyond these tissue-level roles, the receptor's downstream effector coupling has not been characterized in detail in the available corpus apart from the eNOS-NO axis [#1].\"\n,\n  \"teleology\": [\n    {\n      \"year\": 1999,\n      \"claim\": \"Establishing the chromosomal location of Chrm5 and testing it as a candidate disease gene clarified its genetic identity and excluded it as the El2 epilepsy locus.\",\n      \"evidence\": \"RFLV mapping in interspecific backcross mice and Chrm5 sequencing in El2 mutant mice\",\n      \"pmids\": [\"10549128\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No functional or signaling role addressed\", \"Mapping alone does not establish tissue expression or activity\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Radiation hybrid mapping localized rat Chrm5, extending its genetic characterization across species.\",\n      \"evidence\": \"Radiation hybrid mapping with rat RH map server\",\n      \"pmids\": [\"12433399\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No functional consequence established\", \"No link to a specific tissue or pathway\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Testing whether Chrm5 contributes to oviductal ciliary activity defined a tissue where the receptor is dispensable, sharpening the boundaries of its physiological role.\",\n      \"evidence\": \"RT-PCR of laser-microdissected oviductal epithelium and ciliary measurements in Chrm knockout mice\",\n      \"pmids\": [\"22302687\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Negative result; does not identify where Chrm5 IS required\", \"Compound knockout complicates attribution to Chrm5 specifically\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Linking CHRM5 in the ventral subiculum to alcohol motivation answered where and how the receptor modulates addiction-relevant behavior, identifying a defined neural circuit.\",\n      \"evidence\": \"RT-qPCR, retrograde tracing, RNAscope, and intra-vSub pharmacology with the negative allosteric modulator ML375 in alcohol-preferring rats\",\n      \"pmids\": [\"33942300\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Downstream signaling within vSub neurons not defined\", \"Mechanism linking M5 modulation to projection activity unresolved\", \"Single lab\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identifying CHRM5 as a transcriptional target of myocardin-related factors hinted at how its expression is regulated in vascular smooth muscle.\",\n      \"evidence\": \"Forced MYOCD expression in human coronary artery SMCs with RNA-seq and RT-qPCR\",\n      \"pmids\": [\"34539433\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Expression-level readout only without mechanistic follow-up for CHRM5\", \"Direct promoter regulation not demonstrated\", \"Functional consequence of increased CHRM5 not tested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Searching for a CHRM5 disease link in neurogenic bladder tested causality of a candidate variant, but the variant's pathogenicity could not be confirmed.\",\n      \"evidence\": \"Exome sequencing, bladder expression analysis, and in vitro functional studies of the p.Gln184Arg variant (inconclusive)\",\n      \"pmids\": [\"37213061\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"In vitro functional studies were negative/uninformative\", \"Single case; expression data only correlative\", \"Causality of CHRM5 in neurogenic bladder unresolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Defining a Chrm5-eNOS-NO axis in bone marrow arteries answered how cholinergic signaling mechanically regulates the hematopoietic niche through vascular dilation and shear stress.\",\n      \"evidence\": \"In vivo imaging, pharmacological manipulation, metabolomics, and signaling pathway dissection in mouse bone marrow\",\n      \"pmids\": [\"40593589\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct receptor-eNOS coupling mechanism not structurally resolved\", \"Replication not yet established\", \"Source/regulation of non-neurogenic acetylcholine partially defined\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The proximal signal transduction events downstream of CHRM5 receptor activation across its varied tissues remain to be unified mechanistically.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No structural model of CHRM5 in the corpus\", \"G-protein coupling and second-messenger cascades not directly characterized here\", \"Tissue-specific effector selection unexplained\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060089\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0, 1, 5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [1]}\n    ],\n    \"complexes\": [],\n    \"partners\": [],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":4,"faith_total":4,"faith_pct":100.0}}