Do Modulators of the Mitochondrial KATP Channel Change the Function of Mitochondria in Situ?

Abstract

Pharmacological opening of mitochondrial cardiac ATP-sensitive potassium (K ATP ) channels has the chance to be a promising but still controversial cardioprotective mechanism. Physiological roles of mitochondrial K ATP channels in the myocardium remain unclear. We studied the effects of diazoxide, a specific opener of these channels, on the function of rat mitochondria in situ in saponin-permeabilized fibers using an ionic medium that mimics the cytosol. In the presence of NADHproducing substrates (malate ؉ glutamate), neither 100 M diazoxide nor 100 M glibenclamide (a K ATP channel blocker) changed the mitochondrial respiration in the absence or presence of ADP. Because the K ATP channel function could be modified by changes in adenine nucleotide concentrations near the mitochondria, we studied the effects of diazoxide and glibenclamide on the functional activity of mitochondrial kinases. Both diazoxide and glibenclamide did not change the in situ ADP sensitivity in the presence or absence of creatine (apparent K m values for ADP were, respectively, 59 ؎ 9 and 379 ؎ 45 M). Similarly, stimulation of the mitochondrial respiration with AMP in the presence of ATP due to adenylate kinase activity was not affected by the modulators of K ATP channels. However, when succinate was used as substrate, diazoxide significantly inhibited basal respiration by 22% and maximal respiration by 24%. Thus, at a cardioprotective dose, the main functional effect of diazoxide depends on respiratory substrates and seems not to be related to K ATP channel activity.

The Mitochondrial Potassium Cycle

“…Although several laboratories have reported results that seem to indicate uncoupling by mitoK ATP opening in isolated mitochondria (62-64), we have shown directly that this is not the case (65). Similarly, the suggestion that diazoxide uncouples in vivo (57) seems to be refuted by measurements of uncoupling in vivo (66)(67)(68).…”

Distribution of Kir6.0 and SUR2 ATP-sensitive potassium channel subunits in isolated ventricular myocytes

“…2,3 These observations favor an involvement of the mitoK ATP channel in the preconditioning response, although an independent effect of K ATP channel modulators on mitochondrial metabolism cannot be ruled out. [12][13][14] To understand the different roles of sarcK ATP and mitoK ATP channels in cardiac myocytes, the molecular composition of each needs to be resolved. In this study a panel of isoform specific, site-directed antibodies against K ATP channel subunit peptides was produced to investigate the cellular localization of K ATP channel subunit isoforms in isolated ventricular myocytes by (i) confocal microscopy and (ii) subcellular fractionation and Western blotting.…”

“…This condition was associated with defects in vascular sarcolemmal K ATP channels, rather than their cardiac counterparts. 30 In view of these results and reservations concerning the use of flavoprotein fluorescence to monitor mitoK ATP channel activity [12][13][14] , the confirmation of the presence and potential rôle of a cardiac mitoK ATP channel remains elusive.…”

“…6 Moreover, the insensitivity of Kir6.1/SUR2A channels to either diazoxide or 5HD argues against this combination in mitoK ATP channel structure. 6 In view of some reservations over the validity of the association of potassium channel modulator-induced changes in flavoprotein fluorescence with mitoK ATP channel activity, [12][13][14] care is required in attributing a pharmacological profile to the mitoK ATP channel and, therefore, all three of the K ATP channel subunits identified in mitochondria herein may contribute to its structure.…”

Plasmalemmal KATPchannels shape triggered calcium transients in metabolically impaired rat atrial myocytes

“…Ischemic preconditioning is a condition where a prior ischemic episode protects the heart against subsequent ischemic damage (9). Third, the mitochondrial K ATP channel blocker 5-hydroxydecanoate (5-HD) can abolish ischemic preconditioning in some studies (27), although not in others (35), whereas mitochondrial K ATP channel openers such as pinacidil and diazoxide (13) can mimic ischemic preconditioning (2). These arguments for a role of mitochondrial K ATP channels are challenged by the possibility that diazoxide may not target mitochondrial K ATP channels (27,28) but inhibit mitochondrial respiration or oxidant stress at reoxygenation.…”