Putative Structural and Functional Coupling of the Mitochondrial BKCa Channel to the Respiratory Chain

Abstract

Potassium channels have been found in the inner mitochondrial membranes of various cells. These channels regulate the mitochondrial membrane potential, the matrix volume and respiration. The activation of these channels is cytoprotective. In our study, the single-channel activity of a large-conductance Ca2+-regulated potassium channel (mitoBKCa channel) was measured by patch-clamping mitoplasts isolated from the human astrocytoma (glioblastoma) U-87 MG cell line. A potassium-selective current was recorded with a mean conductance of 290 pS in symmetrical 150 mM KCl solution. The channel was activated by Ca2+ at micromolar concentrations and by the potassium channel opener NS1619. The channel was inhibited by paxilline and iberiotoxin, known inhibitors of BKCa channels. Western blot analysis, immuno-gold electron microscopy, high-resolution immunofluorescence assays and polymerase chain reaction demonstrated the presence of the BKCa channel β4 subunit in the inner mitochondrial membrane of the human astrocytoma cells. We showed that substrates of the respiratory chain, such as NADH, succinate, and glutamate/malate, decrease the activity of the channel at positive voltages. This effect was abolished by rotenone, antimycin and cyanide, inhibitors of the respiratory chain. The putative interaction of the β4 subunit of mitoBKCa with cytochrome c oxidase was demonstrated using blue native electrophoresis. Our findings indicate possible structural and functional coupling of the mitoBKCa channel with the mitochondrial respiratory chain in human astrocytoma U-87 MG cells.

Brain mitochondrial ATP-insensitive large conductance Ca+2-activated K+ channel properties are altered in a rat model of amyloid-β neurotoxicity

“…Other studies performed at higher Ca 2+ concentrations (200 μM) led, as expected, to V 1/2 (−20 mV) more negative compared to the value estimated for the control channel in the present case (11 mV) (Bednarczyk et al, 2013). More importantly is the observation that channels from our Aβ rat model showed a left shift in V 1/2 of~6 mV relative to controls, suggesting an intrinsic change in the channel gating mechanism.…”

“…In fact, interactions between BK Ca α/β4 subunits may involve other protein constituents not present when channels are incorporated in a lipid bilayer. Notably a recent study on the mitoBK Ca channel in astrocytoma U-87 MG cells suggests a structural and functional coupling between the mitoBK Ca channel with the mitochondrial respiratory chain that could include β4 (Bednarczyk et al, 2013). A modification of the β4 expression level could have under these conditions an important impact on the mitochondrial functionality.…”

Effectors of large-conductance calcium-activated potassium channel modulate glutamate excitotoxicity in organotypic hippocampal slice cultures

“…It remains unclear whether the mitochondrial or the plasma membrane BK Ca channel is the major mediator of pharmacological preconditioning. Both blockers are affect mitochondrial channel activity (Bednarczyk et al 2013a(Bednarczyk et al , 2013b, but organotipic hippocampal slice culture (applied in this manuscript) may differentiate access of these modulators to plasma membrane or mitochondrial BK Ca channels.…”

Mitochondrial mechanisms of endothelial dysfunction

“…With antibodies raised against the plasma membrane BK Ca channels, we have identified a pore-forming a subunit and regulatory b2 subunit of the channel in the endothelial inner mitochondrial membrane. Our findings indicate a possible structural and functional coupling of the mitoBK Ca channel with the mammalian mitochondrial respiratory chain [55]. In addition, we have shown that hemin inhibits mitoBK Ca and partially abolishes some of the cytoprotective properties of potassium channel opening [56].…”