AJP - Cell Physiology, Vol 255, Issue 1 C51-C59, Copyright © 1988 by American Physiological Society
Changes in expression of a functional Gi protein in cultured rat heart cells
I. S. Allen, S. T. Gaa and T. B. Rogers
Department of Biological Chemistry, University of Maryland School of Medicine, Baltimore 21201.
The muscarinic cholinergic agonist, carbachol, and pertussis toxin were
used to examine the functional status of the guanine nucleotide-binding
protein that inhibits adenylate cyclase (Gi) in cultured neonatal rat heart
myocytes. The isoproterenol stimulation of adenylate cyclase activity in
myocyte membranes and adenosine 3',5'-cyclic monophosphate (cAMP)
accumulation in intact cells (4 days in culture) were insensitive to
carbachol (0.1 mM). However, in cells cultured for 11 days, carbachol (0.1
mM) inhibited isoproterenol-stimulated cAMP accumulation by 30%.
Angiotensin II (ANG II) was also found to inhibit isoproterenol-stimulated
cAMP accumulation in day 11 cells in a dose-dependent manner. Pertussis
toxin treatment reversed the inhibitory effects of both ANG II and
carbachol, suggesting a role for Gi in the process. Carbachol binding to
membranes from day 4 cells was relatively insensitive to guanine
nucleotides when compared with binding to membranes from day 11 or adult
cells. Furthermore, pertussis toxin-mediated 32P incorporation into a 39-
to 41-kDa substrate in day 11 membranes was increased 3.2-fold over that
measured in day 4 membranes. These findings support the view that, although
Gi is expressed, it is nonfunctional in 4-day-old cultured neonatal rat
heart myocytes and acquisition of functional Gi is dependent on culture
conditions. Furthermore, the ANG II receptor can couple to Gi in heart.
