Endothelin-1 Enhances Activities of Cav1.2 in Arterial Vascular Smooth Muscle Cells of Peripheral Artery Disease with Hindlimb Ischemia-Reperfusion

Abstract

Background and Objective: The ischemia-reperfusion (IR) injury is a main feature of pathophysiological process in peripheral artery disease (PAD). In PAD, an exaggerated blood pressure response increases cardiovascular and cerebrovascular risks for the disease. Considering that L-type voltage-gated calcium channel (Cav1.2) in arterial vascular smooth muscle cells (VSMCs) plays a crucial role in regulating blood pressure, the purpose of this study was to determine the activities of Cav1.2 following hindlimb IR as well as a contribution of endothelin-1 (ET-1), a potent vasoconstrictor, to the activities of Cav1.2. Hypothesis: We hypothesized that IR injury of PAD upregulates ET-1 signaling pathways in limb muscle and thereby amplifies the activities of Cav1.2 in VSMCs. Methods: An experimental PAD model with IR was induced by 6 hours of ischemia followed by 18 hours of reperfusion in rats (IR18h rats). ELISA and western blot analysis were used to examine the levels of ET-1 in the hindlimb muscle, and protein expression of Cav1.2 and ET-1 receptor ETA in VSM. Patch clamp was used to determine the Cav1.2 currents. Data are presented as mean ± standard deviation. Results: The hindlimb IR increased the levels of ET-1 in the hindlimb muscle, and expression of Cav1.2 and ET-1’s ETA in VSMCs. In particular, compared with the control group, there was a promising trend for the increasing Cav1.2 in VSM of the IR18h group (1.00±0.61 in control/n=4 vs. 8.77±8.04 in IR18h/n=5, P=0.09). Meanwhile, ETA expression was significantly higher in IR18h group than control group (1.00±1.12 in control/n=4 vs. 3.71±1.59 in IR18h/n=5, P<0.05). The hindlimb IR also increased amplitude of Cav1.2. In addition, ET-1 (1 μM) enhanced Cav1.2 current density in VSMCs of control rats from 0.47± 0.14 pA/pF to 0.68 ± 0.14 pA/pF (n=3; P<0.05) and from 0.54 ± 0.17 pA/pF to 0.74 ± 0.15 pA/pF (n=5; P<0.05) in the VSMCs of IR18h rats, respectively. Pretreatment of the VSMCs with an ETA antagonist, BQ-123 (1 μM), prevented the increasing effects of ET-1 on Cav1.2 current. In contrast, an ETB antagonist, BQ-788 (1 μM), had no distinct effects on the ET-1 increased-Cav1.2 current. Conclusions: ET-1 increases the activities of Cav1.2 in VSMCs via ETA receptor and upregulation of ET-1 signaling pathways in VSMCs of PAD is likely to lead to amplified blood pressure response via increasing the activities of Cav1.2. NIH R01 HL141198 & HL164571; and American Heart Association grant # 940567 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.