Related Experiment Video

Updated: Jun 19, 2026

A Kinetic Fluorescence-based Ca2+ Mobilization Assay to Identify G Protein-coupled Receptor Agonists, Antagonists, and Allosteric Modulators
07:41

A Kinetic Fluorescence-based Ca2+ Mobilization Assay to Identify G Protein-coupled Receptor Agonists, Antagonists, and Allosteric Modulators

Published on: February 20, 2018

The Ca2+ channel beta subunit determines whether stimulation of Gq-coupled receptors enhances or inhibits N current

John F Heneghan1, Tora Mitra-Ganguli, Lee F Stanish

  • 1Department of Physiology, University of Massachusetts Medical School, Worcester, MA 01655, USA.

The Journal of General Physiology
|October 28, 2009
PubMed

Insights

The beta subunit (Ca(V)beta) of calcium channels acts as a molecular switch. Its palmitoylation state determines whether M(1) receptor stimulation enhances or inhibits N-type calcium channel activity.

Area of Science:

  • Neuroscience
  • Molecular Biology
  • Cell Physiology

Background:

  • M(1) receptor (M(1)R) stimulation modulates N-type calcium channel activity in superior cervical ganglion neurons.
  • Arachidonic acid (AA) is a proposed downstream messenger of M(1)Rs, replicating observed modulation patterns.
  • Previous studies suggested phosphatidylinositol-4,5-bisphosphate depletion also contributes to M(1)R-mediated modulation.

Purpose of the Study:

  • To investigate the physiological mechanisms underlying M(1)R-mediated N-type calcium channel modulation using an expression system.
  • To determine the role of the calcium channel beta subunit (Ca(V)beta) in regulating the enhancement versus inhibition of channel activity.
  • To elucidate the specific contribution of Ca(V)beta subunit palmitoylation to observed modulatory effects.

Main Methods:

  • Utilized an expression system (human embryonic kidney 293 cells) to co-express Ca(V)2.2 channels with various Ca(V)beta subunits.
  • Stimulated M(1)Rs and neurokinin-1 receptors (NK-1Rs) to observe effects on N-channel activity.
  • Investigated the impact of exogenously applied arachidonic acid (AA) and free palmitic acid.
  • Employed mutated and chimeric Ca(V)beta subunits to identify key functional domains, particularly related to palmitoylation.

Main Results:

  • The Ca(V)beta subunit acts as a molecular switch, dictating whether M(1)R/NK-1R stimulation inhibits (Ca(V)beta1b, beta3, beta4) or enhances (Ca(V)beta2a) N-channel activity.
  • Exogenous AA mimicked the pattern of modulation observed with different Ca(V)beta subunits.
  • Palmitoylation of Ca(V)beta2a was found to be essential for the loss of inhibition and unmasking of enhancement.
  • The findings suggest heterogeneous association of Ca(V)beta subunits with Ca(V)2.2 channels in SCG neurons.

Conclusions:

  • Ca(V)beta2a, when palmitoylated, blocks inhibition, thereby unmasking enhancement of N-current.
  • Palmitoylation of Ca(V)beta2a represents a novel mechanism that toggles M(1)R/NK-1R-mediated N-current modulation between enhancement and inhibition.
  • Synaptic N-channel activity modulation by M(1)Rs or NK-1Rs may fluctuate based on the presence of palmitoylated Ca(V)beta2a.

More Related Videos

Characterization of G Protein-coupled Receptors by a Fluorescence-based Calcium Mobilization Assay11:49

Characterization of G Protein-coupled Receptors by a Fluorescence-based Calcium Mobilization Assay

Published on: July 28, 2014

Measuring Fast Calcium Fluxes in Cardiomyocytes12:10

Measuring Fast Calcium Fluxes in Cardiomyocytes

Published on: November 29, 2011

Related Concept Videos

G-Protein Gated Ion Channels01:21

G-Protein Gated Ion Channels

GPCRs are primarily responsible for our sense of smell, taste, and vision.  The binding of a sensory stimulus activates GPCR to stimulate effector proteins, many of which are ion channels in the sensory organs. GPCRs modulate the opening and closing of the target ion channels either directly by binding them, or by releasing second messengers that activate these channels. As ions move across the membrane, the membrane potential is altered, which induces an appropriate response.
Sensory organs,...
Calmodulin-dependent Signaling01:16

Calmodulin-dependent Signaling

Calmodulin (CaM) is a calcium-binding protein in eukaryotes that controls various calcium-regulated cellular processes. It has four calcium-binding sites that bind calcium to form the calcium-calmodulin ( Ca2+-CaM) complex. GPCR stimulation increases the calcium levels in the cells that bind to CaM and induces a conformational change.
The Ca2+-CaM complex does not have enzymatic activity by itself. Instead, the complex binds downstream target proteins, including membrane proteins or enzymes,...
Activation and Inactivation of G Proteins01:22

Activation and Inactivation of G Proteins

Heterotrimeric G proteins are guanine nucleotide-binding proteins. As the name suggests, heterotrimeric G proteins are composed of three subunits: alpha, beta, and gamma. They remain GDP-bound or GTP-bound inside the cells and switch between inactive/active states. The Gα subunit possesses the nucleotide-binding pocket that binds guanine nucleotides and switches between GDP or GTP-bound states. In contrast, the Gꞵ and Gγ subunits are always bound together with high affinity and are together...
Ligand-Gated Ion Channel Receptor: Gating Mechanism01:30

Ligand-Gated Ion Channel Receptor: Gating Mechanism

Ligand-gated ion channels are transmembrane proteins that play a vital role in intercellular communication and functions of the nervous system. They allow the influx of ions across the membrane once the neurotransmitter binds, allowing the subsequent transmission of electrical excitation across the neurons. Other ligand-gated ion channels, like the γ-aminobutyric acid (GABA) receptor, permit anions like chloride into the cells on the binding of the GABA molecule. Their entry into the cell...
GPCRs Regulate Adenylyl Cylase Activity01:09

GPCRs Regulate Adenylyl Cylase Activity

Some GPCRs transmit signals through adenylyl cyclase (AC), a transmembrane enzyme. AC helps synthesize second messenger cyclic adenosine monophosphate (cAMP). AC catalyzes cyclization reaction and converts ATP to cAMP by releasing a pyrophosphate. The pyrophosphate is further hydrolyzed to phosphate by the enzyme pyrophosphatase, which drives cAMP synthesis to completion. However, cAMP is rapidly degraded to 5′ AMP by the enzymes phosphodiesterase (PDE), preventing overstimulation of cells.
Two...
Feedback Regulation of Calcium Concentration01:27

Feedback Regulation of Calcium Concentration

Calcium is an essential signaling molecule required for various cellular functions. Calcium pumps and ion channels on cell and organellar membranes, such as those on the endoplasmic reticulum (ER), regulate calcium concentrations inside the cell. They remain closed, keeping the cytosolic calcium levels low at a resting state.
Various transmembrane receptors, such as G protein-coupled receptors (GPCRs), elicit a response to extracellular signals by increasing cytosolic calcium. Activated GPCRs...