1471-2210-11-S1-O151471-2210 Oral presentation <p>Characterization of mice with a deletion of protein kinase G type I in cardiomyocytes and the effect on cardioprotection through either postconditioning or mitochondria-targeted <it>S</it>-nitrosothiol</p> MethnerCarmen LukowskiRobert HofmannFranz MurphyMichael KriegThomastk382@medschl.cam.ac.uk

Clinical Pharmacology Unit, University of Cambridge, UK

Pharmakologie, klinische Pharmazie und Toxikologie, Universität Tübingen, Germany

Forschergruppe 923, Technische Universität München, Germany

MRC Mitochondrial Biology Unit, Cambridge, UK

BMC Pharmacology <p>5th International Conference on cGMP: Generators, Effectors and Therapeutic Implications</p>John Burnett Jr, Franz Hofmann, Harald HHW Schmidt and Johannes-Peter StaschMeeting abstracts - A single PDF containing all abstracts in this supplement is available here.<p>5th International Conference on cGMP: Generators, Effectors and Therapeutic Implications</p>Halle, Germany24-26 June 2011http://www.cyclicgmp.net/1471-2210 2011 11 Suppl 1 O15 http://www.biomedcentral.com/1471-2210/11/S1/O15 10.1186/1471-2210-11-S1-O15
182011 2011Methner et al; licensee BioMed Central Ltd.This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Background

Protein kinase G type I (PKGI/cGMP kinase I) plays a critical role in survival signalling of pre- and postconditioning. However, it is unclear whether cGKI exerts its protective effects in the cardiomyocyte or if other cardiac cell types are involved, and whether nitric oxide (NO) has cGKI-independent effects on cardiomyocytes mitochondria.

Objective

We developed mice with a cardiomyocyte-specific ablation of the cGKI gene (CMG-KO) and tested whether protection against reperfusion injury by ischemic postconditioning (IPost), soluble guanylyl cyclase (sGC) activation, the adenosine A2B receptor (A2BAR), or the mitochondria-targeted S-nitrosothiol (MitoSNO) was affected. MitoSNO accumulates within mitochondria, driven by the membrane potential, where it generates NO and S-nitrosated thiol proteins 1 .

Methods and results

Conditional mice with floxed cGKI alleles were crossed to the MLC2a-Cre transgenic mice. Western Blot and immunohistochemistry confirmed that the Cre-mediated recombination produced the cGKI knock-out specifically in atrial and ventricular cardiomyocytes but not in other organs (Figure 1).

<p>Figure 1</p>

Representative Western Blots of A left ventricle and B cerebellum of CMG-KO and control mice from the same litters on a C57/Bl6N genetic background (CMG-CTR), clearly indicating the specific absence of cGKI in the heart. C Immunohistochemistry of cardiac tissue.

Representative Western Blots of A left ventricle and B cerebellum of CMG-KO and control mice from the same litters on a C57/Bl6N genetic background (CMG-CTR), clearly indicating the specific absence of cGKI in the heart. C Immunohistochemistry of cardiac tissue.

In situ hearts of underwent 30 min of regional ischemia followed by 2 h of reperfusion. As expected, in the control animals all interventions at early reperfusion lead to profound infarct size reduction: IPost (six cycles of 10 sec reperfusion and 10 sec of coronary occlusion), treatment with the specific sGC activator BAY 58-2667 (BAY58), the selective A2BAR agonist BAY 60-6583 (BAY60), as well as MitoSNO. In contrast, the hearts of CMG-KO animals were not protected by BAY58, whereas the protective effects of IPost, BAY60, and MitoSNO were unaffected by the lack of cGKI.

<p>Figure 2</p>

Results of open chest experiments. As expected, either IPost, activation of sGC with BAY58 or A2BAR activation with BAY60 resulted in a profound decrease of infarct size in CMG-CTR. While BAY58 failed to protect in the CMG-KO animals, IPost, BAY60, and MitoSNO still afforded protection, suggesting a signaling independent on cGKI in the cardiomyocyte. *p<0.05 vs. control.

Results of open chest experiments. As expected, either IPost, activation of sGC with BAY58 or A2BAR activation with BAY60 resulted in a profound decrease of infarct size in CMG-CTR. While BAY58 failed to protect in the CMG-KO animals, IPost, BAY60, and MitoSNO still afforded protection, suggesting a signaling independent on cGKI in the cardiomyocyte. *p<0.05 vs. control.

Conclusion

While cardiomyocyte cGKI is important for the protection afforded via cGMP-signalling, beneficial effects of IPost, activation of the A2BAR, as well as direct NO effects via mitochondrial S-nitrosylation does not depend on cGKI in cardiomyocytes.

<p>A mitochondria-targeted S-nitrosothiol modulates respiration, nitrosates thiols, and protects against ischemia-reperfusion injury</p>PrimeTABlaikieFHEvansCNadtochiySMJamesAMDahmCCVitturiDAPatelRPHileyCRAbakumovaIRequejoRChouchaniETHurdTRGarveyJFTaylorCTBrookesPSSmithRAMurphyMPProc Natl Acad Sci U S A2009106261076410769