The role of endothelial connexin40 in the regulation of arterial function and blood pressure

Abstract

Coordination of cellular activity is essential for cardiovascular functions such as the heart beat and hyperaemia. This is facilitated by gap junctions that connect the cytoplasmic contents of adjacent cells. Human carriers of mutations in the gap junction subunit protein, connexin40 (Cx40), are predisposed to lone atrial fibrillation. However, in addition to the atria of the heart, Cx40 is highly expressed in the vascular endothelium, where it is necessary for the coordination of arterial responses. It was therefore hypothesised, that the existence of a dysfunctional Cx40, in the vascular endothelium, would impair arterial function and regulation of blood pressure. To this end, a mutant Cx40 was created containing a threonine to serine missense mutation at the highly conserved amino acid in position 202 (Cx40T202S). A transgenic mouse strain (Cx40T202STg) was created to specifically express Cx40T202S in the vascular endothelium using the endothelium specific promoter Tie2. The aim of this thesis was to determine whether expression of Cx40T202S in the vascular endothelium of mice impaired arterial function and regulation of blood pressure. The ability of Cx40T202S to form electrically patent channels was assessed in single and paired Xenopus oocytes and its ability to form chemically patent channels was assessed in mouse coronary endothelial cells. The effect of Cx40T202S expression in the vascular endothelium was assessed in third order mesenteric arteries that were isolated and mounted in a pressure myograph. The effect of Cx40T202S expression on blood pressure regulation was assessed using radiotelemetric implants. Gap junctions composed of Cx40T202S were found to be electrically but not chemically patent. Arteries from Cx40T202Tg mice were found to reliably and specifically express the Cx40T202S transgene in the vascular endothelium. Myogenic constriction was found to occur at lower pressures in arteries from Cx40T202STg mice and this was due to an absence of basally activated endothelium derived hyperpolarisation. Cx40T202S expression was also found to impair vasodilation by calcitonin gene related peptide and reduce arterial distensibility. However, no effect was seen on vasoconstriction caused by activation of smooth muscle alpha-1-adrenergic, or purinergic P2X1 receptors in third order mesenteric arteries pressurised to 70mmHg. When compared to wild type, the changes to arterial function seen in Cx40T202STg mice caused blood pressure to be significantly elevated during inactivity, and this effect was augmented by locomotor activity at night. However, pressor responses to L-NAME or phenylephrine were unaffected in Cx40T202STg mice. It was therefore concluded that expression of a dysfunctional Cx40, in the vascular endothelium, enhances arterial constriction, increases arterial stiffness, and elevates blood pressure. Show more