GSBS Dissertations and Theses

Title

The Role of Ca2+ Channel Subunit Composition in G Protein-Mediated Inhibition of Ca2+ Channels: a Disstertation

Approval Date

5-1-1997

Document Type

Doctoral Dissertation

Department

Graduate School of Biomedical Sciences, Department of Pharmacology

Subjects

Calcium Channels; GTP-Binding Proteins; Signal Transduction; Academic Dissertations

Abstract

Modulation of Ca2+ channels is an important mechanism for regulation of synaptic strength. However, it is clear that some Ca2+ current types are insensitive to inhibitory modulation mediated by heterotrimeric G proteins (G protein inhibition), and among currents which are sensitive to G protein inhibition, there is great variation in the magnitude of Ca2+ current inhibition between cells of different origin. For the experiments in this dissertation, I utilized recently cloned Ca2+ channels to determine the minimal combination of Ca2+ channel subunits which would confer G protein sensitivity to the recombinant channels. In addition, I examined the role Ca2+ channel auxiliary subunits play in regulation of Ca2+ channel sensitivity to inhibitory G proteins, and whether channels which were sensitive to G protein inhibition were regulated equivalently by the auxiliary subunits. Finally, I investigated possible mechanisms by which these auxiliary subunits modulate G protein-mediated inhibition of Ca2+ channels.

I found that α1A and α1B Ca2+ currents, when expressed in Xenopus oocytes, were sensitive to modulation by G proteins in the absence of any Ca2+ channel auxiliary subunits, while α1C currents were not modulated under the same conditions. I conclude from this data that Ca2+ channel α1 subunits are differentially sensitive to G protein modulation, and the α1 subunit of the class A and B Ca2+ channels is sufficient for G protein modulation.

I also tested the ability of Ca2+ channel auxiliary subunits to modulate the magnitude of G protein-mediated inhibition Ca2+ currents. I found that the Ca2+ channel α2 subunit had no effect on the magnitude of G protein inhibition of α1A and α1B currents. However, the Ca2+ channel β3 subunit eliminated tonic G protein inhibition and sharply reduced the magnitude of muscarinic M2 receptor induced G protein inhibition of both α1A and α1B currents. I found, however, that while the magnitude of α1A and α1B current inhibition was equivalent in the absence of auxiliary subunits, the magnitude of inhibition was greater for the α1B channel after co-expression of the Ca2+ channel β3 subunit. These results indicate that the Ca2+ channel β3 subunit reduces the sensitivity of α1A and α1B Ca2+ channels to voltage-dependent G protein modulation, and does so to a greater extent for α1A channels when compared to α1B Ca2+ channels.

I found that M2 receptor induced inhibition of α1B currents was more voltage-dependent after expression of the Ca2+ channel β3 subunit. Additionally, the rate relief of G protein inhibition dramatically increased after co-expression of the Ca2+ channel β3 subunit. I also co-expressed G protein subunits, and determined that inhibition of both α1B and α1Bβ3 currents was mediated by the G protein βγ subunit. Furthermore, the rate of voltage-dependent relief of G protein βγ subunit induced inhibition increased after co-expression of the Ca2+ channel β3 subunit, similar to the increased rate of relief of the M2 receptor induced G protein inhibition. These data, along with data which demonstrates that G protein inhibition results from the binding of the G protein βγ subunit to the Ca2+ channel (De Waard et al., 1997), indicate that the Ca2+ channel β3 subunit subunit reduces the magnitude of G protein inhibition of α1B Ca2+ currents by increasing the rate of dissociation of the G protein βγ subunit, such that moderate depolarizations used to activate the channel also relieve a large portion of the G protein inhibition.

Comments

In the process of seeking author's permission to provide full text.

Rights and Permissions

Copyright is held by the author, with all rights reserved.

This document is currently not available here.

Share

COinS