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Role of M2, M3, and M4 muscarinic receptor subtypes in the spinal cholinergic control of nociception revealed using siRNA in rats.

ABSTRACT: Muscarinic acetylcholine receptors (mAChRs) are involved in the control of nociception in the spinal cord. The M(2), M(3), and M(4) mAChR subtypes are present in the spinal dorsal horn. However, the role of the individual subtypes in the anti-nociceptive effect produced by mAChR agonists is uncertain. Here, we determined the contribution of M(2), M(3), and M(4) subtypes to spinal muscarinic analgesia by using small-interference RNA (siRNA) targeting specific mAChR subtypes in rats. The neuronal uptake and distribution of a chitosan-siRNA conjugated fluorescent dye in the spinal cord and dorsal root ganglion were confirmed after intrathecal injection. The control and gene-specific siRNA-chitosan complexes were injected intrathecally for three consecutive days. Quantitative reverse-transcription polymerase chain reaction analysis showed that treatment with siRNA targeting M(2), M(3), or M(4) subtype produced a large reduction in the corresponding mRNA levels in the dorsal root ganglion and dorsal spinal cord. Also, the protein levels of the mAChR subtypes in the spinal cord were significantly down-regulated by siRNA treatment, as determined by the immunoprecipitation and receptor-binding assay. Treatment with the M(2)-siRNA caused a large reduction in the inhibitory effect of muscarine on the nociceptive withdrawal threshold. Furthermore, M(4) knockdown at the spinal level significantly reduced the anti-nociceptive effect of muscarine. However, the anti-nociceptive effect of muscarine was not significantly changed by the M(3)-specific siRNA. Our study suggests that chitosan nanoparticles can be used for efficient delivery of siRNA into the neuronal tissues in vivo. Our findings also provide important functional evidence that M(2) and M(4), but not M(3), contribute to nociceptive regulation by mAChRs at the spinal level.

SUBMITTER: Cai YQ

PROVIDER: S-EPMC4435670 | biostudies-literature | 2009 Nov

REPOSITORIES: biostudies-literature

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Role of M2, M3, and M4 muscarinic receptor subtypes in the spinal cholinergic control of nociception revealed using siRNA in rats.

Cai You-Qing YQ Chen Shao-Rui SR Han Hee-Dong HD Sood Anil K AK Lopez-Berestein Gabriel G Pan Hui-Lin HL

Journal of neurochemistry 20090923 4

Muscarinic acetylcholine receptors (mAChRs) are involved in the control of nociception in the spinal cord. The M(2), M(3), and M(4) mAChR subtypes are present in the spinal dorsal horn. However, the role of the individual subtypes in the anti-nociceptive effect produced by mAChR agonists is uncertain. Here, we determined the contribution of M(2), M(3), and M(4) subtypes to spinal muscarinic analgesia by using small-interference RNA (siRNA) targeting specific mAChR subtypes in rats. The neuronal ...[more]

PMID: 19780895

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Project description:UnlabelledOver the last decade, striatal cholinergic interneurons (ChIs) have reemerged as key actors in the pathophysiology of basal-ganglia-related movement disorders. However, the mechanisms involved are still unclear. In this study, we address the role of ChI activity in the expression of parkinsonian-like motor deficits in a unilateral nigrostriatal 6-hydroxydopamine (6-OHDA) lesion model using optogenetic and pharmacological approaches. Dorsal striatal photoinhibition of ChIs in lesioned ChAT(cre/cre) mice expressing halorhodopsin in ChIs reduces akinesia, bradykinesia, and sensorimotor neglect. Muscarinic acetylcholine receptor (mAChR) blockade by scopolamine produces similar anti-parkinsonian effects. To decipher which of the mAChR subtypes provides these beneficial effects, systemic and intrastriatal administration of the selective M1 and M4 mAChR antagonists telenzepine and tropicamide, respectively, were tested in the same model of Parkinson's disease. The two compounds alleviate 6-OHDA lesion-induced motor deficits. Telenzepine produces its beneficial effects by blocking postsynaptic M1 mAChRs expressed on medium spiny neurons (MSNs) at the origin of the indirect striatopallidal and direct striatonigral pathways. The anti-parkinsonian effects of tropicamide were almost completely abolished in mutant lesioned mice that lack M4 mAChRs specifically in dopamine D1-receptor-expressing neurons, suggesting that postsynaptic M4 mAChRs expressed on direct MSNs mediate the antiakinetic action of tropicamide. The present results show that altered cholinergic transmission via M1 and M4 mAChRs of the dorsal striatum plays a pivotal role in the occurrence of motor symptoms in Parkinson's disease.Significance statementThe striatum, where dopaminergic and cholinergic systems interact, is the pivotal structure of basal ganglia involved in pathophysiological changes underlying Parkinson's disease. Here, using optogenetic and pharmacological approaches, we investigated the involvement of striatal cholinergic interneurons (ChIs) and muscarinic receptor subtypes (mAChRs) in the occurrence of a wide range of motor deficits such as akinesia, bradykinesia, motor coordination, and sensorimotor neglect after unilateral nigrostriatal 6-hydroxydopamine lesion in mice. Our results show that photoinhibition of ChIs in the dorsal striatum and pharmacological blockade of muscarinic receptors, specifically postsynaptic M1 and M4 mAChRs, alleviate lesion-induced motor deficits. The present study points to these receptor subtypes as potential targets for the symptomatic treatment of parkinsonian-like motor symptoms.

Dataset Information

Role of M2, M3, and M4 muscarinic receptor subtypes in the spinal cholinergic control of nociception revealed using siRNA in rats.

Publications

Role of M2, M3, and M4 muscarinic receptor subtypes in the spinal cholinergic control of nociception revealed using siRNA in rats.

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