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The fifth subunit of the (?4?2)2 ?2 nicotinic ACh receptor modulates maximal ACh responses.

ABSTRACT: BACKGROUND AND PURPOSE:The fifth subunit in the (?4?2)2 ?4 nicotinic ACh receptor (nAChR) plays a determining role in the pharmacology of this nAChR type. Here, we have examined the role of the fifth subunit in the ACh responses of the (?4?2)2 ?2 nAChR type. EXPERIMENTAL APPROACH:The role of the fifth subunit in receptor function was explored using two-electrode voltage clamp electrophysiology, along with subunit-targeted mutagenesis and the substituted cysteine scanning method applied to fully linked (?4?2)2 ?2 receptors. KEY RESULTS:Covalent modification of the cysteine-substituted fifth subunit with a thiol-reactive agent (MTS) caused irreversible inhibition of receptor function. ACh reduced the rate of the reaction to MTS, but the competitive inhibitor dihydro-?-erythroidine had no effect. Alanine substitution of conserved residues that line the core of the agonist sites on ?4(+)/?2(-) interfaces did not impair receptor function. However, impairment of agonist binding to ?4(+)/?2(-) agonist sites by mutagenesis modified the effect of ACh on the rate of the reaction to MTS. The extent of this effect was dependent on the position of the agonist site relative to the fifth subunit. CONCLUSIONS AND IMPLICATIONS:The fifth subunit in the (?4?2)2 ?2 receptor isoform modulates maximal ACh responses. This effect appears to be driven by a modulatory, and asymmetric, association with the ?4(+)/?2(-) agonist sites. LINKED ARTICLES:This article is part of a themed section on Nicotinic Acetylcholine Receptors. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.11/issuetoc.

SUBMITTER: New K

PROVIDER: S-EPMC5978951 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature

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The fifth subunit of the (α4β2)<sub>2</sub> β2 nicotinic ACh receptor modulates maximal ACh responses.

New Karina K Del Villar Silvia Garcia SG Mazzaferro Simone S Alcaino Constanza C Bermudez Isabel I

British journal of pharmacology 20170714 11

<h4>Background and purpose</h4>The fifth subunit in the (α4β2)<sub>2</sub> α4 nicotinic ACh receptor (nAChR) plays a determining role in the pharmacology of this nAChR type. Here, we have examined the role of the fifth subunit in the ACh responses of the (α4β2)<sub>2</sub> β2 nAChR type.<h4>Experimental approach</h4>The role of the fifth subunit in receptor function was explored using two-electrode voltage clamp electrophysiology, along with subunit-targeted mutagenesis and the substituted cyste ...[more]

PMID: 28600847

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Project description:Background and purposeThe mechanisms causing spontaneous epileptic seizures, including carbamazepine-resistant/zonisamide -sensitive seizures and comorbidity in autosomal dominant sleep-related hypermotor epilepsy (ADSHE) are unclear. This study investigated functional abnormalities in thalamocortical transmission in transgenic rats bearing rat S286L-mutant Chrna4 (S286L-TG) of α4 subunit of the nicotinic ACh receptor (nAChR) that corresponds to the human S284L-mutant CHRNA4.Experimental approachEffects of carbamazepine and zonisamide on epileptic discharges of S286L-TG rat were measured using telemetry electrocorticogram. Transmission abnormalities of L-glutamate and GABA in thalamocortical pathway of S286L-TG rats were investigated using multiprobe microdialysis and ultra-high-performance liquid-chromatography.Key resultsEpileptic discharges in S286L-TG rats were reduced by zonisamide but not by carbamazepine, similar to that of S284L-ADSHE patients. Carbamazepine unaffected functional abnormality in transmission of S286L-TG rats. However, zonisamide was able to compensate for the attenuated S286L-mutant nAChR induced GABA release in frontal-cortex, without affecting attenuated thalamocortical glutamatergic transmission. Excitatory effects of S286L-mutant nAChR on thalamocortical transmission were attenuated compared with those of wild-type nAChR. Loss-of-function of S286L-nAChR enhanced transmission in thalamocortical motor pathway by predominantly attenuating GABAergic transmission. However, it attenuated transmission in thalamocortical cognitive pathway by reducing inhibitory GABAergic and excitatory glutamatergic transmission.Conclusion and implicationsOur results suggest that functional abnormalities of S286L-nAChR are associated with intra-frontal and thalamocortical transmission, possibly contributing to the pathogenesis of ADSHE-seizure and comorbidity of S284L-ADSHE. Selective compensation of impaired GABAergic transmission by zonisamide (but not by carbamazepine) in frontal cortex may be involved, at least partially, in carbamazepine-resistant ADSHE-seizure of S284L-ADSHE patients.

Project description:Nicotine, the principal reinforcing compound in tobacco, acts in the brain by activating neuronal nicotinic acetylcholine receptors (nAChRs). This review summarizes our current knowledge regarding how the α5 accessory nAChR subunit, encoded by the CHRNA5 gene, differentially modulates α4β2* and α3β4* receptors at the cellular level. Genome-wide association studies have linked a gene cluster in chromosomal region 15q25 to increased susceptibility to nicotine addiction, lung cancer, chronic obstructive pulmonary disease, and peripheral arterial disease. Interestingly, this gene cluster contains a non-synonymous single-nucleotide polymorphism (SNP) in the human CHRNA5 gene, causing an aspartic acid (D) to asparagine (N) substitution at amino acid position 398 in the α5 nAChR subunit. Although other SNPs have been associated with tobacco smoking behavior, efforts have focused predominantly on the D398 and N398 variants in the α5 subunit. In recent years, significant progress has been made toward understanding the role that the α5 nAChR subunit-and the role of the D398 and N398 variants-plays on nAChR function at the cellular level. These insights stem primarily from a wide range of experimental models, including receptors expressed heterologously in Xenopus oocytes, various cell lines, and neurons derived from human induced pluripotent stem cells (iPSCs), as well as endogenous receptors in genetically engineered mice and-more recently-rats. Despite providing a wealth of available data, however, these studies have yielded conflicting results, and our understanding of the modulatory role that the α5 subunit plays remains incomplete. Here, we review these reports and the various techniques used for expression and analysis in order to examine how the α5 subunit modulates key functions in α4β2* and α3β4* receptors, including receptor trafficking, sensitivity, efficacy, and desensitization. In addition, we highlight the strikingly different role that the α5 subunit plays in Ca2+ signaling between α4β2* and α3β4* receptors, and we discuss whether the N398 α5 subunit variant can partially replace the D398 variant.

Dataset Information

The fifth subunit of the (?4?2)2 ?2 nicotinic ACh receptor modulates maximal ACh responses.

Publications

The fifth subunit of the (α4β2)<sub>2</sub> β2 nicotinic ACh receptor modulates maximal ACh responses.

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