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Page 8 of 21 Bradshaw et al. Vessel Plus 2023;7:35 https://dx.doi.org/10.20517/2574-1209.2023.121

[100]
[103]
respiration . Diazoxide facilitated mitochondrial swelling in ischemia (whereas mitochondria otherwise
tend to contract in this setting) and reduced mitochondrial calcium accumulation, which are beneficial for
[100]
mitochondrial function . In animal myocytes undergoing ischemia and reperfusion, pharmacologic
opening of mitoK channels was associated with decreased matrix calcium overload because of
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mitochondrial membrane depolarization .
[104]
The mechanisms and effects of cellular swelling observed in ischemic myocytes secondary to stress have
been studied for several decades , and the prevention of swelling has been suggested to be a potential
[105]
mechanism of K cardioprotection. Studies in the 2010s elucidated relationships between cellular volume
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changes during stress and myocyte contractility. Stresses that caused cellular swelling (hypoosmotic stress;
exposure to hyperkalemic, hypothermic cardioplegia; or exposure to metabolic inhibition) resulted in
reduced contractility that was prevented with the addition of K channel opener diazoxide [77,82,106] .
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Hyperosmotic stress resulted in myocyte shrinkage and improved contractility, and this inverse relationship
[77]
between function and myocyte size suggested a cellular mechanism for myocardial stunning . Thus, a
proposed cellular model of myocyte stunning was created to test mechanisms of diazoxide
[77]
cardioprotection .

Interestingly, in myocytes, an open sK channel appeared necessary for the observed cellular swelling and
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reduced contractility after exposure to hypothermic, hyperkalemic cardioplegia . An interplay between
[61]
cellular volume and mitochondrial volume may be important in cardioprotection afforded by K channel
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openers.

Pharmacologic channel blockers have also been utilized to determine potential mechanisms of
cardioprotection and implicated channel subunits. The attenuation of both swelling and reduced
contractility by diazoxide was unchanged by sarcolemmal or mitochondrial K channel blockers,
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contradicting other work with channel blockers [106-108] . This led to the proposal that pharmacologic channel
blockers are nonspecific and only definitive methods such as genetic deletion could confirm channel
subunit involvement in cardioprotection.

While the mitoK channel became a focus of cardioprotection, there was still evidence suggesting a role
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for the sK channel. A study using the Langendorff model found that diazoxide’s cardioprotection was
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[109]
provided via sK channels rather than mitoK channels . However, work using whole-cell voltage
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clamping of ventricular myocytes determined that diazoxide does not open sarcolemmal K channels; this
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led the investigation of diazoxide cardioprotection away from the sK channel .
[41]
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Mitochondrial respiration
There are several potential K -independent mechanisms of diazoxide cardioprotection at the
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mitochondrial level including the inhibition of succinate dehydrogenase (SDH), increased reactive oxygen
species signaling, and protein kinase C activation [42,110-117] . While the majority of studies have implicated K
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channels, some have emphasized that diazoxide and other openers have important K channel-
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independent actions, and therefore cardioprotective mechanisms remain unclear [42,118] .
K channel opener diazoxide is a known inhibitor of mitochondrial enzyme complex II, SDH, a
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component of the electron transport chain [119,120] . Consistent with this idea, other SDH inhibitors have been
found to be cardioprotective, and reversal of SDH and succinate accumulation is the primary driver of
mitochondrial reactive oxygen species (ROS) production that underlies ischemia/reperfusion
injury [110,121-123] . The exact nature of the interaction between SDH and mitoK is unknown. In myocyte
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