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Impact of human D398N single nucleotide polymorphism on intracellular calcium response mediated by ?3?4?5 nicotinic acetylcholine receptors.

ABSTRACT: The human CHRNA5 D398N polymorphism (rs16969968) causes an aspartic acid to asparagine change in the nicotinic acetylcholine receptor (nAChR) ?5 subunit gene. The N398 variant of CHRNA5 is linked to increased risk for nicotine dependence. In this study, we explored the effect of the CHRNA5 D398N polymorphism on the properties of human ?3?4* nicotinic acetylcholine receptors in human embryonic kidney (HEK) cells. Addition of either D398 or N398 variant of ?5 subunit in the ?3?4* receptor did not affect total [(125)I]-epibatidine binding or surface expression of the receptor. However, addition of ?5(D398) into ?3?4* receptor decreased the maximal response to agonist without significantly affecting EC(50) in aequorin intracellular calcium assay. ?3?4?5(N398) nAChRs showed further decreased maximal response. The differences in agonist efficacy between the receptor subtypes were found to be dependent upon the concentration of external calcium but independent of external sodium. Moreover, activation of ?3?4?5 nAChRs led to significantly greater intracellular calcium release from IP(3) stores relative to ?3?4 nAChRs although no effect of the ?5 polymorphism was observed. Finally, inclusion of the ?5 variant caused a small shift to the left in IC(50) for some of the antagonists tested, depending upon ?5 variant but did not affect sensitivity of ?3?4* receptors to desensitization in response to incubation with nicotine. In conclusion, addition of either variant of ?5 into an ?3?4?5 receptor similarly effects receptor pharmacology and function. However, the N398 variant exhibits a reduced response to agonists when extracellular calcium is high and it may lead to distinct downstream cellular signaling.

SUBMITTER: Tammimaki A

PROVIDER: S-EPMC3661904 | biostudies-literature | 2012 Nov

REPOSITORIES: biostudies-literature

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Impact of human D398N single nucleotide polymorphism on intracellular calcium response mediated by α3β4α5 nicotinic acetylcholine receptors.

Tammimäki Anne A Herder Penelope P Li Ping P Esch Caroline C Laughlin James R JR Akk Gustav G Stitzel Jerry A JA

Neuropharmacology 20120720 6

The human CHRNA5 D398N polymorphism (rs16969968) causes an aspartic acid to asparagine change in the nicotinic acetylcholine receptor (nAChR) α5 subunit gene. The N398 variant of CHRNA5 is linked to increased risk for nicotine dependence. In this study, we explored the effect of the CHRNA5 D398N polymorphism on the properties of human α3β4* nicotinic acetylcholine receptors in human embryonic kidney (HEK) cells. Addition of either D398 or N398 variant of α5 subunit in the α3β4* receptor did not ...[more]

PMID: 22820273

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Project description:1. SH-SY5Y cells express alpha7 and alpha3* subtypes of nicotinic acetylcholine receptors (AChR). Numbers of these receptors are upregulated by chronic treatment with nicotinic agonists or KCl. In this study we have examined the functional consequences of these drug treatments on nicotine- or KCl-evoked increases in [Ca(2+)](i), in SH-SY5Y cells. 2. In untreated cells, nicotine increased [Ca(2+)](i) (EC(50) 7.5 microM). Responses to 10 microM nicotine were abolished by the non-selective nicotinic antagonist mecamylamine and were partially blocked by alpha7-selective antagonists, the alpha3beta2*-selective antagonist alpha-conotoxin-MII, and by cadmium and verapamil. 3. After treatment for 4 days with nicotinic agonists, nicotine-evoked increases in [Ca(2+)](i) were significantly decreased by about 25%. Nicotine-evoked responses were paradoxically increased in the presence of acute methyllycaconitine (MLA; an alpha7-selective antagonist) although other alpha7-selective antagonists were without effect, while alpha-conotoxin-MII gave a partial inhibition. The increase observed with MLA was abolished by mecamylamine but not by alpha-conotoxin-MII and was still observed 24 h after chronic nicotine treatment. 4. After treatment for 4 days with KCl, nicotine-evoked increases in [Ca(2+)](i) were also decreased by 25%, but acute MLA was without effect. Responses to 20 mM KCl were unchanged by prior treatment with nicotine or KCl. Treatment for 4 days with 5 microM verapamil reduced responses to both nicotine and KCl by about 50%. 5. Multiple nicotinic AChR subtypes contribute to nicotine-evoked increases in [Ca(2+)](i) in SH-SY5Y cells. Responses to acute nicotine are reduced after chronic nicotine or KCl treatment, with loss of the component attributed to the alpha7 subtype. However, in nicotine-treated cells this effect is reversed when nicotine stimulation is applied in the presence of acute MLA. The antagonist may assist in converting a non-functional alpha7 nicotinic AChR to a conducting state.

Dataset Information

Impact of human D398N single nucleotide polymorphism on intracellular calcium response mediated by ?3?4?5 nicotinic acetylcholine receptors.

Publications

Impact of human D398N single nucleotide polymorphism on intracellular calcium response mediated by α3β4α5 nicotinic acetylcholine receptors.

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