Abstract
An increased flux of potassium ions into the mitochondrial matrix through the ATP-sensitive potassium channel (mitoKATP) has been shown to provide protection against ischemia-reperfusion injury. Recently, it was proposed that the mitochondrial-targeted isoform of the renal outer medullary potassium channel (ROMK) protein creates a pore-forming subunit of mitoKATP in heart mitochondria. Our research focuses on the properties of mitoKATP from heart-derived H9c2 cells. For the first time, we detected single-channel activity and describe the pharmacology of mitoKATP in the H9c2 heart-derived cells. The patch-clamping of mitoplasts from wild type (WT) and cells overexpressing ROMK2 revealed the existence of a potassium channel that exhibits the same basic properties previously attributed to mitoKATP. ROMK2 overexpression resulted in a significant increase of mitoKATP activity. The conductance of both channels in symmetric 150/150 mM KCl was around 97 ± 2 pS in WT cells and 94 ± 3 pS in cells overexpressing ROMK2. The channels were inhibited by 5-hydroxydecanoic acid (a mitoKATP inhibitor) and by Tertiapin Q (an inhibitor of both the ROMK-type channels and mitoKATP). Additionally, mitoKATP from cells overexpressing ROMK2 were inhibited by ATP/Mg2+ and activated by diazoxide. We used an assay based on proteinase K to examine the topology of the channel in the inner mitochondrial membrane and found that both termini of the protein localized to the mitochondrial matrix. We conclude that the observed activity of the channel formed by the ROMK protein corresponds to the electrophysiological and pharmacological properties of mitoKATP.
Highlights
IntroductionThe identification of ischemic preconditioning (IPC) as a cardioprotective mechanism has raised the possibility that stimulating the endogenous pathways may protect the heart from ischemic injury [1]
Ischemic heart disease remains a major worldwide health concern
The identification of ischemic preconditioning (IPC) as a cardioprotective mechanism has raised the possibility that stimulating the endogenous pathways may protect the heart from ischemic injury [1]
Summary
The identification of ischemic preconditioning (IPC) as a cardioprotective mechanism has raised the possibility that stimulating the endogenous pathways may protect the heart from ischemic injury [1]. It has been suggested that the maintenance of mitochondrial integrity is crucial in the preconditioning of heart [2] and brain tissue [3]. The ATP-sensitive potassium channel (mitoKATP) opening protects cardiac and neuronal tissue against ischemia-reperfusion injury [8,9]. MitoKATP was later found in other mammalian tissues including the heart [13,14,15], brain [16,17], skeletal muscle [18], skin fibroblasts [19] and renal tissue [20]. The presence of the channel has been reported in non-mammalians including Crithidia fasciculate euglenozoa [21], Caenorhabditis elegans nematode [22], Acanthamoeba castellanii amoebae [23], and in plants, including Triticum turgidum wheat and Pisum sativum peas [24]
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have