Contribution of valine 7' of TMD2 to gating of neuronal alpha3 receptor subtypes.

Imagen de Manuel F Navedo
TítuloContribution of valine 7' of TMD2 to gating of neuronal alpha3 receptor subtypes.
Publication TypeJournal Article
Year of Publication2006
AutoresNieves-Cintrón, M, Caballero-Rivera, D, Navedo, MF, Lasalde-Dominicci, JA
JournalJ Neurosci Res
Date Published2006 Dec
Palabras claveAcetylcholine, Aconitine, Animals, Dihydro-beta-Erythroidine, Dose-Response Relationship, Drug, Dose-Response Relationship, Radiation, Electric Stimulation, Ion Channel Gating, Membrane Potentials, Mutagenesis, Nicotine, Nicotinic Agonists, Nicotinic Antagonists, Oocytes, Patch-Clamp Techniques, Protein Structure, Tertiary, Rats, Receptors, Nicotinic, Time Factors, Valine, Xenopus laevis

The second transmembrane domain (TMD2) of the Cys-loop family of ligand-gated ion channels forms the channel pore. The functional role of the amino acid residues contributing to the channel pore in neuronal nicotinic alpha3 receptors is not well understood. We characterized the contribution of TMD2 position V7' to channel gating in neuronal nicotinic alpha3 receptors. Site-directed mutagenesis was used to substitute position alpha3 (V7') with four different amino acids (A, F, S, or Y) and coexpressed each mutant subunit with wild-type (WT) beta2 or beta4 subunits in Xenopus oocytes. Whole-cell voltage clamp experiments show that substitution for an alanine, serine, or phenylalanine decreased by 2.3-6.2-fold the ACh-EC(50) for alpha3beta2 and alpha3beta4 receptor subtypes. Interestingly, mutation V7'Y did not produce a significant change in ACh-EC(50) when coexpressed with the beta2 subunit but showed a significant approximately two-fold increase with beta4. Similar responses were obtained with nicotine as the agonist. The antagonist sensitivity of the mutant channels was assessed by using dihydro-beta-erythroidine (DHbetaE) and methyllycaconitine (MLA). The apparent potency of DHbetaE as an antagonist increased by approximately 3.7- and 11-fold for the alpha3beta2 V7'S and V7'F mutants, respectively, whereas no evident changes in antagonist potency were observed for the V7'A and V7'Y mutants. The V7'S and V7'F mutations increase MLA antagonist potency for the alpha3beta4 receptor by approximately 6.2- and approximately 9.3-fold, respectively. The V7'A mutation selectively increases the MLA antagonist potency for the alpha3beta4 receptor by approximately 18.7-fold. These results indicate that position V7' contributes to channel gating kinetics and pharmacology of the neuronal nicotinic alpha3 receptors.

Alternate JournalJ. Neurosci. Res.
PubMed ID17044037
Grant ListGM08102-27 / GM / NIGMS NIH HHS / United States
R01 GM56371-5 / GM / NIGMS NIH HHS / United States
R25 GM61151-01A1 / GM / NIGMS NIH HHS / United States