PT  - JOURNAL ARTICLE
AU  - Mark S. Taylor
AU  - Chris Okwuchukwuasanya
AU  - Christian K. Nickl
AU  - Werner Tegge
AU  - Joseph E. Brayden
AU  - Wolfgang R. G. Dostmann
TI  - Inhibition of cGMP-Dependent Protein Kinase by the Cell-Permeable Peptide DT-2 Reveals a Novel Mechanism of Vasoregulation
AID  - 10.1124/mol.65.5.1111
DP  - 2004 May 01
TA  - Molecular Pharmacology
PG  - 1111--1119
VI  - 65
IP  - 5
4099  - http://molpharm.aspetjournals.org/content/65/5/1111.short
4100  - http://molpharm.aspetjournals.org/content/65/5/1111.full
SO  - Mol Pharmacol2004 May 01; 65
AB  - Cyclic GMP-dependent protein kinase (PKG) serves as an important physiological regulator of vascular reactivity and tone. However, available inhibitors of PKG have exhibited variable effects in intact tissue, hindering the elucidation of the functional role of PKG in blood vessels. In this study, we have determined the effects of our previously engineered potent and selective PKG Iα inhibitor DT-2 on basal and cGMP-stimulated purified recombinant PKG, and compared DT-2 with commonly used PKG inhibitors (8R,9S,11S)-(–)-9-methoxy-carbamyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a-trizadibenzo-(a,g)-cycloocta-(c,d,e)-trinden-1-one (KT-5823), Rp-8-(4-chlorophenylthio)-guanosine-3′,5′-cyclic monophosphorothioate (Rp-8-pCPT-cGMPS), and (β-phenyl-1,N2-etheno-8-bromoguanosine-3′,5′-cyclic monophosphorothioate, Rp-isomer (Rp-8-Br-PET-cGMPS). As expected, all inhibitors reduced cGMP-stimulated PKG activity. However, only DT-2 decreased cGMP-independent or basal PKG activity, whereas KT5823 showed no effect and the Rp-compounds actually had partial agonist activity. To evaluate the potential functional impact of this unique inhibition by DT-2 under physiologically relevant conditions, we analyzed the inhibitors in isolated pressurized cerebral arteries. KT-5823 and Rp-8-pCPT-cGMPS demonstrated marginal reversal of vasodilation induced by 8-Br-cGMP. By comparison, DT-2 completely reversed 8-Br-cGMP induced dilations with comparable potency to Rp-8-Br-PET-cGMPS. In fact, DT-2 constricted arteries beyond their starting (pre-8-Br-cGMP) diameters and caused constriction even in the absence of exogenous 8-Br-cGMP, an effect that was not observed with any other inhibitor. The direct constricting effect of DT-2 was essentially abolished in cultured arteries, where PKG expression was reduced by approximately 90%. These findings indicate that DT-2 not only effectively inhibits cGMP-stimulated PKG activity but also reduces basal PKG activity both in vitro and in vivo. Moreover, these distinctive inhibitory properties of DT-2 suggest an important role for constitutive PKG activity in the continuous regulation of cerebral artery tone.