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The M2 muscarinic G-protein-coupled receptor is voltage-sensitive
Yair Ben-Chaim1, Oded Tour, Nathan Dascal
1The Otto Loewi Minerva Center for Cellular and Molecular Neurobiology, the Hebrew University, Jerusalem 91904, Israel.
Abstract:
G-protein coupled receptors are not considered to exhibit voltage sensitivity. Here, using Xenopus oocytes, we show that the M2 muscarinic receptor (m2R) is voltage-sensitive. The m2R-mediated potassium channel (GIRK) currents were used to assay the activity of m2R. We found that the apparent affinity of m2R toward acetylcholine (ACh) was reduced upon depolarization. Binding experiments of [3H]ACh to individual oocytes expressing m2R confirmed the electrophysiological findings. When the GIRK channels were activated either by overexpression of Gbetagamma subunits or by injection of GTPgammaS, the ratio between the currents measured at -60 mV and +40 mV was the same as for the basal activity of the GIRK channel. Thus, the steps downstream to agonist activation of m2R are not voltage-sensitive. We further found that, in contrast to m2R, the apparent affinity of m1R was increased upon depolarization. We also found that the voltage sensitivity of binding of [3H]ACh to oocytes expressing m2R was greatly diminished following pretreatment with pertussis toxin. The cumulative results suggest that m2R is, by itself, voltage-sensitive. Furthermore, the voltage sensitivity does not reside in the ACh binding site, rather, it most likely resides in the receptor region that couples to the G-protein.
Insights
The M2 muscarinic receptor (m2R), a G-protein coupled receptor, demonstrates voltage sensitivity. This voltage sensitivity is independent of acetylcholine binding and G-protein activation.
Area of Science:
- Neuroscience
- Molecular Biology
- Biophysics
Background:
- G-protein coupled receptors (GPCRs) are traditionally not considered voltage-sensitive.
- The M2 muscarinic receptor (m2R) is a key GPCR involved in various physiological processes.
Purpose of the Study:
- To investigate the potential voltage sensitivity of the M2 muscarinic receptor (m2R).
- To determine the specific regions of m2R responsible for voltage sensitivity.
Main Methods:
- Electrophysiological recordings of GIRK channel currents in Xenopus oocytes expressing m2R.
- Radioligand binding assays using [3H]ACh to measure receptor-ligand interactions.
- Pertussis toxin treatment to assess the role of G-protein coupling.
Main Results:
- M2 muscarinic receptor (m2R) activity showed voltage dependence, with reduced apparent affinity upon depolarization.
- Downstream signaling pathways (Gbetagamma subunits, GTPgammaS) were not voltage-sensitive.
- Voltage sensitivity was diminished after pertussis toxin treatment, indicating involvement of the G-protein coupling region.
Conclusions:
- The M2 muscarinic receptor (m2R) exhibits intrinsic voltage sensitivity, challenging previous assumptions about GPCRs.
- Voltage sensitivity is localized to the receptor's G-protein coupling domain, not the acetylcholine binding site.
- These findings reveal a novel mechanism of GPCR regulation by membrane potential.