J BIOCHEM MOLECULAR TOXICOLOGY Volume 16, Number 6, 2002 Melatonin Inhibits the Contractile Effect of Vanadate in the Isolated Pulmonary Arterial Rings of Rats: Possible Role of Hydrogen Peroxide Mahmoud N. Nagi, Mahmoud A. Mansour, Othman A. Al-shabanah, and Hassan A. El-Kashef Department of Pharmacology, College of Pharmacy, King Saud University, P.O. Box 2457, Riyadh, 11451, Saudi Arabia; E-mail: mnagi@ksu.edu.sa Received 25 June 2002; revised 15 September 2002; accepted 24 September 2002 ABSTRACT: The effect and possible mechanism of ac- tion of vanadate on the isolated pulmonary arterial rings of normal rats were studied. Pulmonary arterial rings contracted in response to vanadate (0.1–1 mM) in a concentration-dependent manner. Preincubation of the pulmonary arterial rings with 1 mM melatonin significantly reduced the contractile effect of vanadate by more than 60%. Furthermore, addition of hydro- gen peroxide (50 M) or enzymatic generation of hy- drogen peroxide by the addition of glucose oxidase (10 U/mL) to the medium containing glucose produced remarkable increases in the pulmonary arterial tension, 46.2 ± 7.3 and 78.7 ± 9.7 g tension/g tissue, respec- tively. Similarly, incubation of the pulmonary arterial rings with 1 mM melatonin significantly reduced the contractile responses of the arterial rings to hydrogen peroxide and glucose/glucose oxidase to 25.7 ± 2.9 and 24.7 ± 4.4 g tension/g tissue, respectively. Vanadate, in vitro, significantly stimulated the oxidation of NADH by xanthine oxidase, and the rate of oxidation was in- creased by increasing either time or vanadate concen- tration. Similarly, addition of melatonin to a reaction mixture containing xanthine oxidase and vanadate sig- nificantly inhibited the rate of NADH oxidation in a concentration-dependent fashion. The results of the present study indicated that vanadate induced contrac- tion in the isolated pulmonary arterial rings, which was significantly reduced by melatonin. Furthermore, the contractile effect of vanadate on the pulmonary arterial rings may be attributed to the intracellular generation of hydrogen peroxide. C  2002 Wiley Periodicals, Inc. J Biochem Mol Toxicol 16:273–278, 2002; Published on- line in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.10049 KEYWORDS: Vanadate; Melatonin; Hydrogen Peroxide; Xanthine Oxidase; Pulmonary Arterial Rings; Rats Correspondence to: Mahmoud N. Nagi. c  2002 Wiley Periodicals, Inc. INTRODUCTION Vanadate is a commonly occurring form of the trace element vanadium. Vanadium is widely distributed in the environment and can exert potent toxic and car- cinogenic effects especially in workers in petrochemi- cal, steel, and mining industries [1]. It has been reported that vanadate contracts several smooth muscles includ- ing rat vas deference [2], vascular and intestinal smooth muscle [3], rat gall bladder [4], guinea-pig airway smooth muscle [5], and isolated rat aorta [6]. The ex- act mechanism(s) responsible for the vanadate-induced contractions in smooth muscles is not yet certain. How- ever, several mechanisms have been proposed, includ- ing inhibition of phosphotyrosine phosphatase activity and consequent increase in protein tyrosine phosphory- lation of several substrates [3,7], inhibition of Na + /K + ATPase [8], inhibition of Ca 2+ -dependent ATPase activ- ity [5], or even through the increase in phosphoinositide metabolism [6]. Several reports demonstrated that vanadate stim- ulates the oxidation of NAD(P)H by enzymatic and nonenzymatic sources of superoxide anion radical [9–12] via a sequence of free-radical chain reactions with the net production of hydrogen peroxide (H 2 O 2 ) [13]. Furthermore, several enzymes were known to generate cellular superoxide in a controlled way: one of these enzymes is a membrane-located NADPH ox- idase, which is present in the endothelium [14]. A second and well-known superoxide generating sys- tem consists of xanthine/xanthine oxidase, which is found membrane-associated on endothelial and other cells [14]. Recently, it has been shown that incuba- tion of mouse epidermal JB6 cells with vanadate in- duces generation of H 2 O 2 , which seems to play an essential role in the action of vanadate in apoptosis [1]. 273