Gene and Protein Information
class A G protein-coupled receptor
Species	TM	AA	Chromosomal Location	Gene Symbol	Gene Name	Reference
Human	7	479	11p11.2	CHRM4	cholinergic receptor muscarinic 4	3,34
Mouse	7	479	2 50.63 cM	Chrm4	cholinergic receptor, muscarinic 4	58
Rat	7	478	3q24	Chrm4	cholinergic receptor, muscarinic 4	45,91,105

Previous and Unofficial Names

HM3 [73-74] | Chrm-4 | cholinergic receptor, muscarinic 4 | cholinergic receptor

Database Links
Specialist databases
GPCRdb	acm4_human (Hs), acm4_mouse (Mm)
Other databases
Alphafold	P08173 (Hs), P32211 (Mm), P08485 (Rn)
ChEMBL Target	CHEMBL1821 (Hs), CHEMBL4572 (Mm), CHEMBL317 (Rn)
DrugBank Target	P08173 (Hs)
Ensembl Gene	ENSG00000180720 (Hs), ENSMUSG00000040495 (Mm), ENSRNOG00000017556 (Rn)
Entrez Gene	1132 (Hs), 12672 (Mm), 25111 (Rn)
Human Protein Atlas	ENSG00000180720 (Hs)
KEGG Gene	hsa:1132 (Hs), mmu:12672 (Mm), rno:25111 (Rn)
OMIM	118495 (Hs)
Pharos	P08173 (Hs)
RefSeq Nucleotide	NM_000741 (Hs), NM_007699 (Mm), NM_031547 (Rn)
RefSeq Protein	NP_000732 (Hs), NP_031725 (Mm), NP_113735 (Rn)
UniProtKB	P08173 (Hs), P32211 (Mm), P08485 (Rn)
Wikipedia	CHRM4 (Hs)

Selected 3D Structures
	Description:  Structure of the M4 muscarinic acetylcholine receptor (M4-mT4L) bound to tiotropium. PDB Id:  5DSG Ligand:  tiotropium Resolution:  2.6Å Species:  Human References:  89

Natural/Endogenous Ligands
acetylcholine

Download all structure-activity data for this target as a CSV file

Agonists
Key to terms and symbols	View all chemical structures	Click column headers to sort
Ligand Sp. Action Value Parameter Reference [3H]acetylcholine Hs Agonist 8.2 pKd 51 ⤷ pKd 8.2 [51] pentylthio-TZTP Hs Full agonist 8.7 pKi 43 ⤷ pKi 8.7 [43] NNC 11-1585 Hs Full agonist 8.6 pKi 16 ⤷ pKi 8.6 [16] NNC 11-1607 Hs Full agonist 8.1 pKi 16 ⤷ pKi 8.1 [16] xanomeline Hs Partial agonist 7.4 – 7.7 pKi 99,106 ⤷ pKi 7.4 – 7.7 [99,106] NNC 11-1314 Hs Full agonist 7.3 pKi 16 ⤷ pKi 7.3 [16] sabcomeline Hs Partial agonist 7.2 pKi 106 ⤷ pKi 7.2 [106] McN-A-343 Hs Partial agonist 5.6 – 6.7 pKi 48 ⤷ pKi 5.6 – 6.7 [48] cevimeline Hs Agonist 6.0 pKi 57 ⤷ pKi 6.0 (Ki 1.012x10-6 M) [57] Description: Displacement of [3H]QNB from cloned receptor. arecaidine propargyl ester Hs Full agonist 5.9 pKi 43 ⤷ pKi 5.9 [43] methacholine Rn Agonist 5.8 pKi 72,76 ⤷ pKi 5.8 (Ki 1.6x10-6 M) [72,76] arecoline Hs Full agonist 5.5 pKi 43,71,76 ⤷ pKi 5.5 [43,71,76] milameline Hs Partial agonist 5.5 pKi 106 ⤷ pKi 5.5 [106] oxotremorine Hs Full agonist 5.2 pKi 43,76 ⤷ pKi 5.2 [43,76] oxotremorine-M Hs Full agonist 5.2 pKi 43 ⤷ pKi 5.2 [43] pilocarpine Hs Partial agonist 5.2 pKi 43,76 ⤷ pKi 5.2 [43,76] acetylcholine Hs Full agonist 4.5 – 5.6 pKi 43,46 ⤷ pKi 4.5 – 5.6 [43,46] methylfurmethide Hs Full agonist 4.7 pKi 43 ⤷ pKi 4.7 [43] carbachol Hs Full agonist 4.3 – 4.9 pKi 43,106 ⤷ pKi 4.3 – 4.9 [43,106] furtrethonium Hs Full agonist 4.3 pKi 43 ⤷ pKi 4.3 [43] bethanechol Hs Full agonist 4.0 pKi 43,76 ⤷ pKi 4.0 [43,76] (+)-aceclidine Hs Full agonist 5.4 pEC50 26 ⤷ pEC50 5.4 [26] (-)-aceclidine Hs Partial agonist 4.8 pEC50 26 ⤷ pEC50 4.8 [26] iperoxo Hs Agonist - - 79 ⤷ [79]
View species-specific agonist tables
Agonist Comments
The binding data for McN-A-343 [48] is found on rat striatum. Please consult references [5,52,76,98] for further details of the activity of some of the ligands in this list. McN-A-343 and pilocarpine have been found to be partial agonists [52] and full agonists [76,98] at the M4 receptor.

Antagonists
Key to terms and symbols	View all chemical structures	Click column headers to sort
Ligand Sp. Action Value Parameter Reference [3H]QNB Hs Antagonist 9.7 – 10.5 pKd 42,73 ⤷ pKd 9.7 – 10.5 (Kd 2x10-10 – 3.16x10-11 M) [42,73] [3H]N-methyl scopolamine Hs Antagonist 9.9 – 10.2 pKd 10,42-43,47,50,98 ⤷ pKd 9.9 – 10.2 (Kd 1.4x10-10 – 6x10-11 M) [10,42-43,47,50,98] [3H]AF DX-384 Hs Antagonist 8.7 pKd 11,62,94 ⤷ pKd 8.7 [11,62,94] biperiden Hs Antagonist 8.6 pKd 2 ⤷ pKd 8.6 (Kd 2.4x10-9 M) [2] otenzepad Hs Antagonist 7.0 pKd 27 ⤷ pKd 7.0 [27] tiotropium Hs Antagonist 10.2 – 10.6 pKi 86,88 ⤷ pKi 10.2 – 10.6 [86,88] 3-quinuclidinyl-benzilate Hs Antagonist 10.4 pKi 9 ⤷ pKi 10.4 (Ki 4x10-11 M) [9] umeclidinium Hs Antagonist 10.3 pKi 78 ⤷ pKi 10.3 [78] propantheline Hs Antagonist 10.2 pKi 41 ⤷ pKi 10.2 [41] aclidinium Hs Antagonist 10.0 pKi 88 ⤷ pKi 10.0 [88] oxyphenonium Hs Antagonist 9.8 pKi 9 ⤷ pKi 9.8 (Ki 1.45x10-10 M) [9] atropine Rn Antagonist 9.7 pKi 45 ⤷ pKi 9.7 [45] glycopyrrolate Hs Antagonist 9.1 – 10.0 pKi 84,86 ⤷ pKi 9.1 – 10.0 [84,86] AE9C90CB Hs Antagonist 9.5 pKi 81 ⤷ pKi 9.5 [81] mepenzolic acid Hs Antagonist 9.4 pKi 9 ⤷ pKi 9.4 (Ki 3.6x10-10 M) [9] scopolamine Hs Antagonist 9.1 – 9.5 pKi 2,41 ⤷ pKi 9.1 – 9.5 [2,41] revefenacin Hs Antagonist 9.3 pKi 38 ⤷ pKi 9.3 (Ki 5.5x10-10 M) [38] Description: Determined from a radioligand binding assay using membranes from CHO‐K1 cells expressing the hM4 receptor, and displacement of [3H]NMS tracer. ipratropium Hs Antagonist 9.2 pKi 39 ⤷ pKi 9.2 [39] 4-DAMP Rn Antagonist 9.1 pKi 45 ⤷ pKi 9.1 [45] atropine Hs Antagonist 8.7 – 9.5 pKi 18,39,41,74 ⤷ pKi 8.7 – 9.5 [18,39,41,74] 4-DAMP Hs Antagonist 8.9 pKi 24 ⤷ pKi 8.9 [24] oxybutynin Hs Antagonist 8.4 – 8.7 pKi 23,81 ⤷ pKi 8.4 – 8.7 [23,81] muscarinic toxin 3 Hs Antagonist 8.5 pKi 9 ⤷ pKi 8.5 (Ki 3x10-9 M) [9] silahexocyclium Hs Antagonist 8.5 pKi 6 ⤷ pKi 8.5 [6] tolterodine Hs Antagonist 8.3 – 8.4 pKi 31,81 ⤷ pKi 8.3 – 8.4 [31,81] UH-AH 37 Hs Antagonist 8.3 – 8.4 pKi 31,104 ⤷ pKi 8.3 – 8.4 [31,104] hexocyclium Hs Antagonist 8.3 pKi 6 ⤷ pKi 8.3 [6] dicyclomine Hs Antagonist 8.3 pKi 9 ⤷ pKi 8.3 (Ki 5x10-9 M) [9] PCS1055 Hs Antagonist 8.2 pKi 18 ⤷ pKi 8.2 [18] himbacine Hs Antagonist 7.9 – 8.4 pKi 9,24,44,61 ⤷ pKi 8.4 (Ki 4x10-9 M) [9] pKi 7.9 – 8.2 [24,44,61] amitriptyline Hs Antagonist 8.1 pKi 82 ⤷ pKi 8.1 (Ki 7.2x10-9 M) [82] AQ-RA 741 Hs Antagonist 7.8 – 8.2 pKi 24,31 ⤷ pKi 7.8 – 8.2 [24,31] tripitramine Hs Antagonist 7.9 pKi 63 ⤷ pKi 7.9 [63] darifenacin Hs Antagonist 7.3 – 8.1 pKi 31,39,81 ⤷ pKi 7.3 – 8.1 [31,39,81] hexahydrosiladifenidol Rn Antagonist 7.7 pKi 45 ⤷ pKi 7.7 [45] AFDX384 Hs Antagonist 7.3 – 8.0 pKi 18,24 ⤷ pKi 7.3 – 8.0 [18,24] pirenzepine Hs Antagonist 7.0 – 8.1 pKi 9,24,37,41,44,104 ⤷ pKi 8.1 (Ki 8x10-9 M) [9] pKi 7.0 – 7.6 [24,37,41,44,104] PD 102807 Hs Antagonist 7.4 – 7.6 pKi 18,68 ⤷ pKi 7.4 – 7.6 [18,68] p-F-HHSiD Hs Antagonist 7.1 – 7.5 pKi 27,41 ⤷ pKi 7.1 – 7.5 [27,41] dosulepin Hs Antagonist 7.2 pKi 82 ⤷ pKi 7.2 (Ki 6.1x10-8 M) [82] hexahydrosiladifenidol Hs Antagonist 6.5 – 7.7 pKi 6,27 ⤷ pKi 6.5 – 7.7 [6,27] muscarinic toxin 1 Hs Antagonist 7.1 pKi 35 ⤷ pKi 7.1 [35] hexahydrodifenidol Hs Antagonist 7.1 pKi 6 ⤷ pKi 7.1 [6] pirenzepine Rn Antagonist 7.1 pKi 45 ⤷ pKi 7.1 [45] methoctramine Hs Antagonist 6.6 – 7.5 pKi 6,24,27,37 ⤷ pKi 6.6 – 7.5 [6,24,27,37] tropicamide Hs Antagonist 6.9 pKi 9 ⤷ pKi 6.9 [9] solifenacin Hs Antagonist 6.8 pKi 81 ⤷ pKi 6.8 [81] (S)-dimetindene Hs Antagonist 6.5 pKi 8 ⤷ pKi 6.5 (Ki 2.951x10-7 M) [8] Description: Binding to hM4 receptors expressed in CHO cells. otenzepad Rn Antagonist 6.5 pKi 45 ⤷ pKi 6.5 [45] guanylpirenzepine Rn Antagonist 6.2 pKi 97 ⤷ pKi 6.2 [97] muscarinic toxin 2 Hs Antagonist 5.9 pKi 35 ⤷ pKi 5.9 [35] VU0255035 Hs Antagonist 5.9 pKi 80 ⤷ pKi 5.9 [80] lithocholylcholine Hs Antagonist 5.3 pKi 13 ⤷ pKi 5.3 [13] muscarinic toxin 7 Hs Antagonist <5.0 pKi 64 ⤷ pKi <5.0 [64] ML381 Hs Antagonist >4.5 pKi 30 ⤷ pKi >4.5 (Ki <3x10-5 M) [30]
View species-specific antagonist tables
Antagonist Comments
Biperiden is an approved drug antagonist of muscarinic acetylcholine receptors. We have tagged the M1 subtype as the drug's primary target as affinity is 10-fold higher at this receptor subtype [2].

Allosteric Modulators
Key to terms and symbols	View all chemical structures	Click column headers to sort
Ligand Sp. Action Value Parameter Reference LY2033298 Hs Positive 4.9 – 5.5 pKB 12,85 ⤷ pKB 4.9 – 5.5 [12,85] KT 5720 Hs Neutral 6.4 pKd 54 ⤷ pKd 6.4 [54] WIN 51,708 Hs Negative 6.2 pKd 55 ⤷ pKd 6.2 [55] WIN 62,577 Hs Negative 5.9 pKd 55 ⤷ pKd 5.9 [55] Gö 7874 Hs Neutral 5.7 pKd 54 ⤷ pKd 5.7 [54] alcuronium Hs Negative 5.6 pKd 43 ⤷ pKd 5.6 [43] LY2119620 Hs Positive 5.5 pKd 19 ⤷ pKd 5.5 [19] brucine Hs Negative 4.7 – 6.0 pKd 43,53 ⤷ pKd 4.7 – 6.0 [43,53] staurosporine Hs Neutral 5.3 pKd 54 ⤷ pKd 5.3 [54] strychnine Hs Positive 4.8 – 5.0 pKd 43,50 ⤷ pKd 4.8 – 5.0 [43,50] vinburnine Hs Positive 4.6 pKd 43 ⤷ pKd 4.6 [43] N-benzyl brucine Hs Negative 4.5 pKd 53 ⤷ pKd 4.5 [53] N-benzyl brucine Hs Neutral 4.5 pKd 53 ⤷ pKd 4.5 [53] N-chloromethyl-brucine Hs Neutral 4.4 pKd 53 ⤷ pKd 4.4 [53] vincamine Hs Positive 4.2 pKd 43 ⤷ pKd 4.2 [43] thiochrome Hs Positive 4.0 pKd 51 ⤷ pKd 4.0 [51] brucine N-oxide Hs Neutral 3.6 pKd 53 ⤷ pKd 3.6 [53] brucine N-oxide Hs Positive 3.6 pKd 53 ⤷ pKd 3.6 [53] muscarinic toxin 3 Hs Negative 8.7 pKi 44,67 ⤷ pKi 8.7 [44,67] VU0152100 Rn Positive 6.4 pEC50 4 ⤷ pEC50 6.4 (EC50 3.8x10-7 M) [4] VU0152099 Rn Positive 6.4 pEC50 4 ⤷ pEC50 6.4 (EC50 4.03x10-7 M) [4] VU0010010 Hs Positive - - 17 ⤷ [17]
View species-specific allosteric modulator tables

Primary Transduction Mechanisms
Transducer	Effector/Response
Gi/Go family	Adenylyl cyclase inhibition
References:  60,73

Tissue Distribution
Esophageal smooth muscle. Species:  Human Technique:  Radioligand binding. References:  75
Bladder. Species:  Human Technique:  RT-PCR. References:  92
CNS: forebrain. Species:  Mouse Technique:  immunocytochemistry. References:  40
CNS: cerebral cortex, corpus striatum, thalamus, hypothalamus, pons-medulla. Species:  Mouse Technique:  Radioligand binding. References:  65
Heart: intrinsic neurons. Species:  Rat Technique:  in situ hybridisation. References:  36
CNS: basal forebrain, pedunculopontine and laterodorsal tegmental nuclei. Species:  Rat Technique:  in situ hybridisation. References:  95
CNS: cerebral cortex, hippocampus, corpus striatum, olfactory tubercle, midbrain, pons-medulla, cerebellum. Species:  Rat Technique:  Immunoprecipitation. References:  107
CNS: caudate putamen, nucleus accumbens, olfactory tubercle. Species:  Rat Technique:  in situ hybridisation. References:  100
CNS: hippocampus. Species:  Rat Technique:  immunocytochemistry. References:  56
Vestibular system. Species:  Rat Technique:  RT-PCR. References:  96

Expression Datasets
Show »« Hide Log average relative transcript abundance in mouse tissues measured by qPCR from Regard, J.B., Sato, I.T., and Coughlin, S.R. (2008). Anatomical profiling of G protein-coupled receptor expression. Cell, 135(3): 561-71. [PMID:18984166] [Raw data: website] There should be a chart of expression data here, you may need to enable JavaScript!

Functional Assays
Measurement of IP1 levels in murine fibroblast cells (B82) transfected with the rat M4 receptor. Species:  Rat Tissue:  B82 cells. Response measured:  Stimulation of IP1 accumulation. References:  45
Measurement of cAMP levels in murine fibroblast cells (B82) transfected with the rat M4 receptor. Species:  Rat Tissue:  B82 cells. Response measured:  Inhibition of cAMP accumulation. References:  45
Measurement of cAMP levels in JEG-3 cells transfected with the human M2 receptor, using a cAMP response element (CRE)-coupled luciferase construct as the reporter. Species:  Human Tissue:  JEG-3 cells. Response measured:  Inhibition of cAMP accumulation. References:  60
Measurement of cAMP levels in CHO cells transfected with the human M4 receptor. Species:  Human Tissue:  CHO cells. Response measured:  Inhibition of cAMP accumulation. References:  16
Measurement of ERK1/2 activity in COS-7 cells transfected with the human M4 receptor. Species:  Human Tissue:  COS-7 cells. Response measured:  Increase in ERK1/2 activity. References:  77
Measurement of Ca2+ channel activity in rat superior cervical ganglion neurons endogenously expressing the M4 receptor. Species:  Rat Tissue:  Superior cervical ganglion neurons. Response measured:  Inhibition of Ca2+ channels. References:  28-29
Measurement of AC activity in rat striatal homogenates endogenously expressing the M4 receptor. Species:  Rat Tissue:  Corpus striatum. Response measured:  Inhibition of AC activity. References:  25,66
Measurement of the effects of a ligand on the level, or rate, of binding of GTPγ35S to membranes. Species:  Human Tissue:  CHO cells. Response measured:  The binding of GTPγ35S to G proteins coupled to the receptor. References:  1,49-52,54-55
Measurement of the effects of a ligand on the rate of hydrolysis of GTP by G proteins in membranes. Species:  Human Tissue:  CHO cell membranes. Response measured:  Generation of 32Pi from [γ-32P]GTP. References:  1,52

Physiological Functions
Autoreceptor: modulation of acetylcholine release. Species:  Rat Tissue:  Urinary bladder. References:  20
Autoreceptor: modulation of acetylcholine release. Species:  Human Tissue:  Urinary bladder detrusor muscle. References:  21
Contraction. Species:  Rat Tissue:  Urinary bladder detrusor muscle. References:  70
Inhibition of dopaminergic (D1 receptor) signalling. Species:  Rat Tissue:  Nucleus accumbens. References:  69
Cell migration. Species:  Human Tissue:  Keratinocytes. References:  15

Physiological Consequences of Altering Gene Expression
M4 receptor knockout mice exhibit an increase in anxiolysis compared to wild-type mice. Species:  Mouse Tissue:  Technique:  Gene targeting in embryonic stem cells. References:  22
Hippocampal, cortical and striatal brain slices from M2/M4 double knockout mice lack muscarinic agonist-induced inhibition of acetylcholine release that is seen with wild-type brain slices. Striatal slices from M4 receptor single knockout mice exhibit the same loss of inhibition of acetylcholine release, but the inhibition remains intact in the hippocampal and cortical slices. Species:  Mouse Tissue:  Technique:  Gene targeting in embryonic stem cells. References:  108
Smooth muscle preparations (gallbladder) from M4 receptor knockout mice exhibit reduced agonist-induced contractions compared to wild-type mice. Species:  Mouse Tissue:  Technique:  Gene targeting in embryonic stem cells. References:  83
Vas deferens tissue from M4 receptor knockout mice exhibits reduced agonist-induced inhibition of noradrenaline release compared to wild-type mice. Species:  Mouse Tissue:  Technique:  Gene targeting in embryonic stem cells. References:  90
Striatal slices from M4 receptor knockout mice exhibit abolished agonist-induced potentiation of dopamine release. Species:  Mouse Tissue:  Technique:  Gene targeting in embryonic stem cells. References:  109
M4 receptor knockout mice exhibit enhanced dopaminergic locomotor activity. Species:  Mouse Tissue:  Technique:  Gene targeting in embryonic stem cells. References:  32-33
Epidermal keratinocytes from M4 receptor knockout mice exhibit reduced agonist-induced migration compared to keratinocytes from wild-type mice. Species:  Mouse Tissue:  Technique:  Gene targeting in embryonic stem cells. References:  15
M4 receptor knockout mice exhibit increased basal hippocampal acetylcholine levels compared to wild-type mice. M2/M4 double knockout mice exhibit a further increase in basal acetylcholine levels. In addition, M4 and M2/M4 knockout mice exhibit an increase in hippocampal acetylcholine release in response to a novel environment. Species:  Mouse Tissue:  Technique:  Gene targeting in embryonic stem cells. References:  93
M4 receptor knockout mice exhibit a significantly reduced wound epithelialisation rate compared to wild-type mice. Species:  Mouse Tissue:  Technique:  Gene targeting in embryonic stem cells. References:  14
Smooth muscle from the small intestine of M4 receptor knockout mice do not exhibit any alteration in EFS-induced acetylcholine release. However, M2/M4 double knockout mice exhibit an increase in acetylcholine release. Overall, it is thought that both M2 and M4 receptors mediate the autoinhibitory control of acetylcholine release in the mouse ileum, and that each can compensate for loss of the other.