Nitric oxide (NO) and B-type natriuretic peptide (BNP) are protective against 
ischemia-reperfusion injury as they increase intracellular cGMP level via 
activation of soluble (sGC) or particulate guanylate cyclases (pGC), 
respectively. The aim of the present study was to examine if the cGMP-elevating 
mediators, NO and BNP, share a common downstream signaling pathway via 
cGMP-dependent protein kinase (PKG) in cardiac cytoprotection. Neonatal rat 
cardiac myocytes in vitro were subjected to 2.5 h simulated ischemia (SI) 
followed by 2 h reoxygenation. Cell viability was tested by trypan blue 
exclusion assay. PKG activity of cardiac myocytes was assessed by phospholamban 
(PLB) phosphorylation determined by western blot. Cell death was 34 +/- 2% after 
SI/reoxygenation injury in the control group. cGMP-inducing agents significantly 
decreased irreversible cell injury: the cGMP analog 8-bromo-cGMP (8-Br-cGMP, 10 
nM) decreased it to 13 +/- 1% (p < 0.001), the direct NO-donor 
S-nitroso-N-acetylpenicillamine (SNAP, 1 microM) to 18 +/- 6% (p < 0.05) and BNP 
(10 nM) to 12 +/- 2% (p < 0.001), respectively. This protective effect was 
abolished by the selective PKG inhibitor KT-5823 (600 nM) in each case. As PLB 
is not a unique reporter for PKG activity since it is also phosphorylated by 
protein kinase A (PKA), we examined PLB phosphorylation in the presence of the 
PKA inhibitor KT-5720 (1 microM). The ratio of pPLB/PLB significantly increased 
after administration of both BNP and 8-Br-cGMP under ischemic conditions, which 
was abolished by the PKG inhibitor. This is the first demonstration that 
elevated cGMP produced either by the sGC activator SNAP or the pGC activator BNP 
exerts cytoprotective effects via a common downstream signaling pathway 
involving PKG activation.