Lack of nitric oxide- and guanosine 3 ': 5 '-cyclic monophosphate-dependent regulation of alpha-thrombin-induced calcium transient in endothelial cells of spontaneously hypertensive rat hearts
P. Failli et al., Lack of nitric oxide- and guanosine 3 ': 5 '-cyclic monophosphate-dependent regulation of alpha-thrombin-induced calcium transient in endothelial cells of spontaneously hypertensive rat hearts, BR J PHARM, 130(7), 2000, pp. 1468-1476
1 While the expression and/or activity of endothelial nitric oxide synthase
CeNOS) has been characterized in spontaneously hypertensive (SHR) and norm
otensive Wistar Kyoto rat (WKY) hearts, in coronary endothelial cells (ECs)
from both strains, the effect of NO on intracellular calcium concentration
([Ca2+](i)) is still unknown.
2 Coronary microvascular ECs were isolated from SHR and WKY and characteriz
ed. Immunocytochemistry and Western blot analysis showed that eNOS was simi
larly expressed in ECs from both strains.
3 Measuring [Ca2+](i) by imaging analysis of fura-2-loaded cells, we demons
trated that a-thrombin (3-180 U l(-1)) induced a superimposable dose-depend
ent calcium transient in ECs from both strains.
4 In WKY ECs, S-nitroso-N-acetyl-DL-penicillamine (SNAP) dose-dependently (
10 - 100 mu M) and 0.1 mu M atrial natriuretic factor (ANF) reduced the max
imum and the decay time of alpha-thrombin-induced calcium transient. The in
hibitory effects of SNAP and ANF were prevented by blocking cyclic GMP-depe
ndent protein kinase. Non selective eNOS inhibitors prolonged the decay tim
e of alpha-thrombin-induced calcium transient, while the selective inducibl
e NOS inhibitor 1400W was ineffective. SNAP (100 mu M) and 0.1 mu M ANF inc
reased cyclic GMP content up to 22.9 and 42.3 fold respectively.
5 In SHR ECs, alpha-thrombin-induced calcium transient was not modified by
SNAP, ANF or eNOS inhibition. SNAP (100 mu M) and 0.1 mu M ANF increased cy
clic GMP content up to 9.3 and 51 fold respectively.
6 In WKY ECs, SNAP dose-dependently (10 - 100 mu M) reduced also bradykinin
-induced calcium transient, while in SHR ECs was ineffective.
7 We concluded that in SHR ECs, the cyclic GMP-dependent regulation of calc
ium transient is lost.