Cholinergic dilation of cerebral blood vessels is abolished in M ₅ muscarinic acetylcholine receptor knockout mice

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 M 5 receptor-selective ligands. To circumvent these difficulties, we used gene targeting technology to generate M 5 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 M 5 receptors play a role in facilitating muscarinic agonist-induced dopamine release in the striatum. Because M 5 receptor mRNA has been detected in several blood vessels, we also investigated whether the lack of M 5 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 extracerebral arteries remained fully intact in M5R ؊/؊ mice. Our findings provide direct evidence that M 5 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 M 5 receptors may represent an attractive therapeutic target. M olecular cloning studies have revealed the existence of five molecularly distinct muscarinic acetylcholine receptor subtypes (M 1 -M 5 ) (1, 2). During the past decade, considerable progress has been made in delineating the physiological roles of the M 1 -M 4 muscarinic receptors (3, 4). In contrast, the physiological relevance of the M 5 receptor subtype, which is the most recent member of the muscarinic receptor family to be cloned (5, 6), remains unknown at present (7,8). However, expression of the cloned M 5 muscarinic receptor gene in cultured mammalian cells has shown that the encoded receptor protein is functional and efficiently couples to G proteins of the G q family, similar to the M 1 and M 3 receptor subtypes (5-8).Immunoprecipitation and in situ mRNA hybridization studies have demonstrated the presence of M 5 receptor protein͞mRNA in different areas of the brain including hippocampus, hypothalamus, and distinct midbrain regions (substantia nigra pars compacta and ventral tegmental area) (9-11). However, M 5 receptors are expressed at very low levels, representing less than 2% of the total muscarinic receptor population (M 1 -M 5 ) expressed in the brain (9). Interestingly, the use of highly sensitive reverse transcriptase-PCR (RT-PCR) techniques suggests that M 5 receptors are expressed in all major brain regions (12). More recently, M 5 receptors also have been detected in several peripheral tissues or cells including peripheral blood l...

Selective activation of the M 1 muscarinic acetylcholine receptor achieved by allosteric potentiation

“…4), a function of ACh attributed to M 5 based on receptor localization and knockout mouse experiments (31,32). Our data indicate that M 1 also regulates CBF perhaps via neurovascular coupling.…”

Muscarinic regulation of dopamine and glutamate transmission in the nucleus accumbens

“…Earlier studies show that muscarinic agonists potentiate DA efflux in the striatum (5)(6)(7)(8). However, other studies reported that mAChR agonists depress DA transients measured with fast-scan cyclic voltammetry (FSCV) and evoked by electrical stimulation in the striatum (9)(10)(11).…”

Dysregulated hippocampal acetylcholine neurotransmission and impaired cognition in M2, M4 and M2/M4 muscarinic receptor knockout mice

“…Antisense 14,15 and gene targeting technologies have emerged as powerful new tools in identifying the receptor subtypes involved in various muscarinic cholinergic functions. [16][17][18][19][20][21] Although pharmacological, neuroanatomical and clinical studies have suggested an important role of muscarinic receptors in synaptic plasticity and memory, the functions of the individual muscarinic receptor subtypes remain unclear. In this study, we have used M2 and M4 receptor single knockout (KO) as well as M2/M4 receptor double KO mice to investigate the functional importance of M2 and M4 receptors in the regulation of acetylcholine release in the hippocampus and in cognitive processes.…”