S-nitrosation of mitochondrial connexin 43 regulates mitochondrial function

Abstract

S-nitrosation (SNO) of connexin 43 (Cx43)-formed channels modifies dye uptake in astrocytes and gap junctional communication in endothelial cells. Apart from forming channels at the plasma membrane of several cell types, Cx43 is also located at the inner membrane of myocardial subsarcolemmal mitochondria (SSM), but not in interfibrillar mitochondria (IFM). The absence or pharmacological blockade of mitochondrial Cx43 (mtCx43) reduces dye and potassium uptake. Lack of mtCx43 is associated with loss of endogenous cardioprotection by ischemic preconditioning (IPC), which is mediated by formation of reactive oxygen species (ROS). Whether or not mitochondrial Lucifer Yellow (LY), ion uptake, or ROS generation are affected by SNO of mtCx43 and whether or not cardioprotective interventions affect SNO of mtCx43 remains unknown. In SSM from rat hearts, application of NO donors (48 nmol to 1 mmol) increased LY uptake (0.5 mmol SNAP 38.4 ± 7.1 %, p < 0.05; 1 mmol GSNO 28.1 ± 7.4 %, p < 0.05) and the refilling rate of potassium (SNAP 227.9 ± 30.1 %, p < 0.05; GSNO 122.6 ± 28.1 %, p < 0.05). These effects were absent following blockade of Cx43 hemichannels by carbenoxolone as well as in IFM lacking Cx43. Unlike potassium, the sodium permeability was not affected by application of NO. Furthermore, mitochondrial ROS formation was increased following NO application compared to control SSM (0.5 mmol SNAP 22.9 ± 1.8 %, p < 0.05; 1 mmol GSNO 40.6 ± 7.1 %, p < 0.05), but decreased in NO treated IFM compared to control (0.5 mmol SNAP 14.4 ± 4 %, p < 0.05; 1 mmol GSNO 13.8 ± 4 %, p < 0.05). NO donor administration to isolated SSM increased SNO of mtCx43 by 109.2 ± 15.8 %. Nitrite application (48 nmol) to mice was also associated with elevated SNO of mtCx43 by 59.3 ± 18.2 % (p < 0.05). IPC by four cycles of 5 min of ischemia and 5 min of reperfusion increased SNO of mtCx43 by 41.6 ± 1.7 % (p < 0.05) when compared to control perfused rat hearts. These data suggest that SNO of mtCx43 increases mitochondrial permeability, especially for potassium and leads to increased ROS formation. The increased amount of SNO mtCx43 by IPC or nitrite administration may link NO and Cx43 in the signal transduction cascade of cardioprotective interventions.Electronic supplementary materialThe online version of this article (doi:10.1007/s00395-014-0433-x) contains supplementary material, which is available to authorized users.

Interaction of Risk Factors, Comorbidities, and Comedications with Ischemia/Reperfusion Injury and Cardioprotection by Preconditioning, Postconditioning, and Remote Conditioning

“…The mechanism through which the signaling ROS are generated in response to the ischemic preconditioning stimulus is not clear, but one suggestion has implicated the activation of the mitochondrial ATP-sensitive potassium channel (K ATP ), which is related to mitochondrial connexin 43 (Cx43) (Heinzel et al, 2005) and appears to be mediated via protein kinase G (PKG) and mitochondrial protein kinase C (PKC)« (Costa and Garlid, 2008). The K + influx into mitochondria is believed to induce matrix alkalinization that then results in the production of superoxide from complex I of the electron transport chain (Soetkamp et al, 2014).…”

“…Cx43 is believed to form hemichannels in the inner mitochondrial membrane, thereby facilitating complex I function and the influx of K + into mitochondria in response to the ischemic preconditioning stimulus (Boengler et al, 2012(Boengler et al, , 2013aSoetkamp et al, 2014). The activation of the RISK or SAFE pathways is not involved in the protective function of Cx43 in ischemic preconditioning (Sanchez et al, 2013).…”

Molecular Basis of Cardioprotection

“…366 Mitochondrial Cx 43 is also a target of nitrosation. 367 The loss of IPC's protection with aging is associated with loss of Cx 43 in both sarcolemma and mitochondria. 368 Of note, although Cx 43 is important for protection by IPC, 358,369 it is not for that by POC.…”

“…215,388 Mitochondrial Cx 43 is a target of nitrosation during IPC. 367 Nitrosation of complex I along with reduced respiration and ROS formation contributes to RIPC.…”

Specific Mechanisms Underlying Right Heart Failure: The Missing Upregulation of Superoxide Dismutase-2 and Its Decisive Role in Antioxidative Defense

“…NO interacts far more ubiquitously at cellular and subcellular levels, in part, by nitrosation of cystein residues within signaling pathways of the renin-angiotensin-aldosterone system (RAAS), the sympathetic nervous system, and the mitochondria, and thus directly and indirectly influences the function and structure of the myocardium (21,39,45).…”