Korean J Anesthesiol Introduction Reactive oxygen species (ROS) are normally generated at basal amounts, but ischemia or oxygen re-supply during reperfusion lead to abnormally high ROS levels that cannot be cleared by normal anti-oxidizing systems, causing tissue reperfusion injury [1]. An oxygen (O2) molecule generates the intermediate species, superoxide radical (O2.), hydrogen peroxide (H2O2), and hydroxyl radical (OH.), during the monovalent reduction reaction, and it is eventually converted to water (H2O) by catalase. In this process, the intermediates O2. and OH. are important free radicals that cause peroxidation (LOO.) of intracellular lipids and tissue damage [2]. In particular, OH., the most reactive ROS [1], is produced by the Fenton reaction (Fe2+ + HOOH → Fe3+ + OH. + OH-) and the Haber-Weiss reaction (O2.- + NO. → OONO- + H+ → OONOH → NOO. + OH.) [3]. ROS generated during reperfusion causes organ damage in the heart [3,4], brain [5,6], and blood vessels [7,8]. In 1980, Furchgott and Zawadzki [9] found an endothelium- derived relaxing factor (EDRF) later shown to be nitric oxide (NO) [10]. Other vasoactive substances, including endothelium- derived hyperpolarizing factor (EDHF) and the endothelium- derived contracting factor (EDCF), are released from the endothelium to regulate vascular tone. However, because the endothelium is composed of a cell monolayer, it is easily damaged by physical and chemical stimuli, including ROS during reperfusion. Inhalation anesthetics [1,4,11] and lidocaine, a local anesthetics, can prevent ischemia-reperfusion injury in the brain [12]. In this study, we tested the effects of lidocaine and procaine, two local anesthetics, on endothelial-dependent relaxation in the rabbit aorta to examine if their anti-oxidizing effects could suppress or reduce the endothelial injury by ROS. Materials and Methods Preparation of ring slices and recording All experiments conformed with the regulations of the Labo- ratory Animal Committee. Under enflurane inhalation anesthesia, ear intravenous injection of heparin 600 IU/kg was performed on rabbits, with exsanguinations 3 minutes later by cleaving the carotid artery. The abdominal aorta of the rabbits (2-2.5 kg, Male, n = 27) was extirpated and 3-4 mm long ring slices were prepared by separating the lipid tissue and connective tissue, not engaging any tension on them, in a petri dish with Krebs- Henseleit solution (K-H) solution (NaCl 120.0, NaHCO3 25.0, KCl 5.0, MgSO4 1.2, CaCl2 2.5, NaH2PO4 1.4, glucose 11.0 mM) bubbled with 95% oxygen and 5% carbon dioxide. Maintaining the temperature at 37 ± 0.5oC, one end of the aorta slice was www.ekja.org Lee, et al. fixed in the tissue bath with 5 ml of K-H solution. The opposite end was connected to the force displacement transducer (TSD 125CⓇ, Biopac Inc., USA) and the K-H solution was exchanged every 15 minutes during the 90-minute equilibrating period. The final resting tension was fixed at 1.5 g. The vascular smooth muscle tension was recorded through an amplifier (DA100CⓇ, Biopac Inc.) with a data acquisition system (MP100Ⓡ, Biopac Inc.) and a personal computer. Following the precontraction with phenylephrine (PE) 10-6 M, acetylcholine (ACh) 3 × 10-8, 10-7, 3 × 10-7, and 10-6 M were consecutively administered to observe the percent change (%) of the aortic tone. Changes of the aortic tone by ACh administration before ROS exposure (control) and after ROS exposure (experimental) were compared. Method of ROS exposure ACh-induced contractions were measured before and after electrolysis (EL) with an electric current (constant current, 15 mA) to the positive and negative electrodes in the K-H solution for 35 seconds to generate ROS and induce endothelial injury. More than 1 cm was maintained between the abdominal aorta ring slices and the positive and negative electrodes to avoid a direct stimulus of the electric field to the tissue. The K-H solution was then exchanged, followed by precontraction with PE and the second ACh treatment. Vascular relaxation in the ROS scavenger pretreatment To examine the action of the scavenger on ROS, 1,000 U/ ml of catalase (n = 8), the hydrogen peroxide scavenger, was pretreated for 15 minutes. The hydroxyl radical scavengers, deferoxamine (n = 9) 0.1 mM, sodium salicylate 1 mM (n = 8), and mannitol 5 × 10-3 M (n = 12) were also pretreated, each for 30 minutes. Then, ROS was generated by EL to induce endothelial injury. The lidocaine and procaine pretreatment groups Experimental group values were obtained by 15 minutes of reagent pretreatment followed by ROS exposure. The experimental groups included pretreatment with lidocaine (L : n = 13, 13, 12, 10, 13, 11) or procaine (P : n = 6, 15, 12, 8, 10, 12) at concentrations of 10-5, 3 × 10-5, 10-4, 3 × 10-4, 10-3, and 3 × 10-3 M. Effect of 3-amino-1,2,4-triazole (3AT) on endothelial injury by ROS The samples were pretreated with 3AT, a catalase inhibitor, for 30 minutes. After a 15-minute waiting period, 3 × 10-4 M of lidocaine 105