Introduction:
Endothelial dysfunction plays a key role in the pathogenesis of diabetic vascular disease, which predisposes to ischemic cardiovascular events. Small conductance calcium-activated-potassium (SK) channels are largely responsible for coronary arteriolar relaxation mediated by endothelium-dependent hyperpolarizing factors. Diabetic inactivation/inhibition of endothelial SK channels contributes to endothelial dysfunction. Endothelial dysfunction during diabetes (DM) is also associated with increases in metabolites NADH, and PKC. The pyridine nucleotide NADH has been recently found to inactivate endothelial SK channels. The overexpressed and activated PKC has been shown to play an important role in diabetes-induced endothelial dysfunction. However, it is undefined if PKC is involved in the metabolite NADH dysregulation of endothelial SK channels.
Hypothesis:
We hypothesized that PKC in a signaling cascade whereby NADH dysregulates endothelial SK channels.
Methods:
SK channels currents of human coronary artery endothelial cells were measured by whole cell patch clamp method in the presence or absence of NADH, and/or PKC activator PMA, PKC inhibitors or endothelial PKC
α
/PKCβ knock-down by using short interfering RNA.
Results:
NADH (30-300μM, n=7-9) or PKC activator PMA (30-300μM, n=6-12) reduced endothelial SK current density (p<0.05 vs. control (n=15), Fig. A-D), whereas the selective PKC
α
inhibitor LY333531 (50nM, n=12) significantly reversed the NADH-induced SK channel inhibition (p>0.05 vs. control (n=15), Fig. E). PKC
α
knock-down failed to affect NADH (n=13) and PMA (n=10) inhibition of endothelial SK currents (Fig. F). In contrast, PKCβ knock-down significantly prevented NADH (n=12) and PMA (n=6)-induced SK inhibition (p>0.05, vs. control, Fig. G).
Conclusions:
The metabolite NADH dysregulation of endothelial SK channels was via PKCβ activation.
Paradoxical Excitatory Impact of SK Channels on Dendritic Excitability
