Targeted inactivation of α i2 or α i3 disrupts activation of the cardiac muscarinic K + channel, I K + Ach , in intact cells

• Published in: Proceedings of the National Academy of Sciences - PNAS 1997-07, Vol.94 (15), p.7921-7926
• Main Authors: Sowell, Margaret O., Ye, Chianping, Ricupero, Dennis A., Hansen, Sherry, Quinn, Stephen J., Vassilev, Peter M., Mortensen, Richard M.
• Format: Article
• Language: English
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.94.15.7921

Cardiac muscarinic receptors activate an inwardly rectifying K + channel, I K + Ach , via pertussis toxin (PT)-sensitive heterotrimeric G proteins (in heart G i2 , G i3 , or G o ). We have used embryonic stem cell (ES cell)-derived cardiocytes with targeted inactivations of specific PT-sensitive α subunits to determine which G proteins are required for receptor-mediated regulation of I K + Ach in intact cells. The muscarinic agonist carbachol increased I K + Ach activity in ES cell-derived cardiocytes from wild-type cells, in cells lacking α o , and in cells lacking the PT-insensitive G protein α q . In cells with targeted inactivation of α i2 or α i3 , channel activation by both carbachol and adenosine was blocked. Carbachol-induced channel activation was restored in the α i2 - and α i3 -null cells by reexpressing the previously targeted gene and guanosine 5′-[γ-thio] triphosphate was able to fully activate I K + Ach in excised membranes patches from these mutants. In contrast, negative chronotropic responses to both carbachol and adenosine were preserved in cells lacking α i2 or α i3 . Our results show that expression of two specific PT-sensitive α subunits (α i2 and α i3 but not α o ) is required for normal agonist-dependent activation of I K + Ach and suggest that both α i2 - and α i3 -containing heterotrimeric G proteins may be involved in the signaling process. Also the generation of negative chronotropic responses to muscarinic or adenosine receptor agonists do not require activation of I K + Ach or the expression of α i2 or α i3 .