Supplementary Material for: Changes in IP3 Receptor Expression and Function in Aortic Smooth Muscle of Atherosclerotic Mice

Peroxynitrite is an endothelium-independent vasodilator that induces
relaxation via membrane hyperpolarization. The activation of IP3
receptors triggers the opening of potassium channels and
hyperpolarization. Previously we found that relaxation to peroxynitrite
was maintained during the development of atherosclerosis due to changes
in the expression of calcium-regulatory proteins. In this study we
investigated: (1) the mechanism of peroxynitrite-induced relaxation in
the mouse aorta, (2) the effect of atherosclerosis on relaxation to
peroxynitrite and other vasodilators, and (3) the effect of
atherosclerosis on the expression and function of the IP3 receptor.
Aortic function was studied using wire myography, and atherosclerosis
was induced by fat-feeding ApoE-/- mice. The expression of
IP3 receptors was studied using Western blotting and
immunohistochemistry. Relaxation to peroxynitrite was attenuated by the
IP3 antagonists 2-APB and xestospongin C and also the Kv
channel blocker 4-aminopyridine (4-AP). Atherosclerosis attenuated
vasodilation to cromakalim and the AMPK activator A769662 but not
peroxynitrite. Relaxation was attenuated to a greater extent by 2-APB in
atherosclerotic aortae despite the reduced expression of IP3 receptors.
4-AP was less effective in ApoE-/- mice fat-fed for 4 months. Peroxynitrite relaxation involves an IP3-induced calcium release and KV
channel activation. This mechanism becomes less important as
atherosclerosis develops, and relaxation to peroxynitrite may be
maintained by increased calcium extrusion.