Copyright © 2007, European Society of Cardiology
Pharmacological and physiological stimuli do not promote Ca2+-sensitive K+ channel activity in isolated heart mitochondria
Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP, Brazil
* Corresponding author. Av. Prof. Lineu Prestes, 748, Cidade Universitária, São Paulo, SP, 05508-900, Brazil. Fax: +55 11 38155579. Email address: alicia{at}iq.usp.br
Objective: Mitochondrial calcium-activated K+ (mitoKCa) channels have been described as channels that are activated by Ca2+, inner mitochondrial membrane depolarization and drugs such as NS-1619. NS-1619 is cardioprotective, leading to the assumption that this effect is related to the opening of mitoKCa channels. Here, we show several weaknesses in this hypothesis.
Methods: Isolated mitochondria from rat hearts were tested for evidence of mitoKCa activity by analyzing functional parameters in K+-rich and K+-free media.
Results: NS-1619 promoted mitochondrial depolarization both in K+-rich and K+-free media. Respiratory rate increments were also seen in the presence of NS-1619 for both media. In parallel, NS-1619 promoted respiratory inhibition, as evidenced by respiratory measurements in state 3. Mitochondrial volume measurements conducted using light scattering showed that NS-1619 led to swelling, in a manner unaltered by inhibitors of mitoKCa channels, antagonists of adenosine triphosphate-sensitive potassium channels or inhibitors of the permeability transition. Swelling was also maintained when K+ in the media was substituted with tetraethylammonium (TEA+), which is not transported by any known K+ carrier. Electron microscopy experiments gave support to the idea that NS-1619-induced mitochondrial swelling took place in the absence of K+. In addition to testing the pharmacological effects of NS-1619, we attempted, unsuccessfully, to promote mitoKCa activity by altering Ca2+ concentrations in the medium and inducing mitochondrial uncoupling.
Conclusion: Our data indicate that NS-1619 promotes non-selective permeabilization of the inner mitochondrial membrane to ions, in addition to partial respiratory inhibition. Furthermore, we found no specific K+ transport in isolated heart mitochondria compatible with mitoKCa opening, whether by pharmacological or physiological stimuli. Our results indicate that NS-1619 has extensive mitochondrial effects unrelated to mitoKCa and suggest that tissue protection mediated by NS-1619 may occur through mechanisms other than activation of these channels.
KEYWORDS Oxygen consumption; K+ channel; Ischemia; Mitochondria; Preconditioning
Abbreviations: NS-1619, 1,3-dihydro-1-[2-hydroxy-5-(trifluoromethyl)phenyl]-5-(trifluoromethyl)-2H-benzimidazol-2-one • 5-HD, 5-hydroxydecanoate • CCCP, carbonyl cyanide m-chloro phenyl hydrazone • DNP, dinitrophenol • CsA, cyclosporin A • EGTA, ethylene glycol bis(2-aminoethyl ether)-N,N,N'N'-tetraacetic acid • mitoKCa, mitochondrial ATP-sensitive K+ channels • mitoKCa, mitochondrial calcium-activated K+ channels • Pax, paxilline • TEA+, tetraethylammonium ion • S.E.M., standard error of the mean • TMPD, N,N,N,N-tetramethyl-p-phenylene-diamine
Time for primary review 25 days
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