Original Article ROLE OF NITRIC OXIDE (NO) IN CAPSAICIN MEDIATED ANTI-PLATELET ACTIVITY IN IN VITRO, IN VIVO, EX-VIVO MODEL OF PLATELET AGGREGATION ASSAY AND ARTERIAL THROMBOSIS IN RAT: POTENTIAL THERAPEUTIC TARGET? MIHIR K. PATEL 1 , KIRANJ K. CHAUDAGAR 1 , ANITA A. MEHTA 1* Department of Pharmacology, L. M. College of Pharmacy, Navarangpura, Ahmedabad Email: dranitalmcp@gmail.com Received: 06 Feb 2017 Revised and Accepted: 24 Feb 2018 ABSTRACT Objective: Although recent advances in the treatment of congestive heart disease, mortality among patients’ remains a questionable remark. Therefore, we evaluated the role of capsaicin on in vitro and ex vivo platelet aggregation induced by Adenosine Di-Phosphate (ADP) as well as in in vivo thrombosis models and role of NO, KATP was also identified in the capsaicin-induced anti-platelet animal model as well as in vivo model of arterial thrombosis. Methods: According to body weight wistar rats were divided into five groups. Group I and Group II was treated with saline and capsaicin (3 mg/kg, i. v), while animals from Group III were treated with N(ω)-nitro-L-arginine methyl ester (L-NAME) (30 mg/kg, i. v) 30 min before administration of capsaicin (3 mg/kg, i. v). Group IV animals were treated with glibenclamide (10 mg/kg,i. v) 30 min before administration of capsaicin (3 mg/kg, i. v). Group V was considered as a positive control and administered clopidogrel (30 mg/kg, p. o). Animals were subjected for in vitro, ex-vivo platelet aggregation assay. ADP (30µM) was utilized as an aggregating agent in these experiments. After these assays; animals of each group were subjected for subaqueous tail bleeding time in a rat model and FeCl3-induced arterial thrombosis model in rats. Results: In ADP-induced in vitro platelet aggregation, a significant reduction in % platelet aggregation was observed at 50µM (64.35±4.641) and 100µM (52.72±4.192) concentration of capsaicin as compared to vehicle control (85.82±3.716). Capsaicin (3 mg/kg, i. v) also showed a significant reduction (49.53±4.075) in ex-vivo ADP-induced platelet aggregation as compared to vehicle control (89.38±2.057). In FeCl3 induced arterial thrombosis model, Capsaicin (3 mg/kg, i. v) exhibited an increase in time to occlusion in this rodent model and presence of the L-NAME and glibenclamide had inhibited the activity of capsaicin. Conclusion: In our study, capsaicin (50 µM, 100µM) exhibited potent anti-platelet activity in ADP-induced platelet aggregation, similarly capsaicin exhibited significant anti-platelet action in the ex-vivo study. Moreover, the presence of L-NAME and glibenclamide inhibited the anti-thrombotic and anti-platelet action of capsaicin. Therefore, it was concluded that NO and KATP may be involved in the anti-thrombotic action of capsaicin. Keywords: Capsaicin, Anti-platelet activity, Glibenclamide, Nitric Oxide, TRPV1 channel © 2018 The Authors. Published by Innovare Academic Sciences Pvt Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/) DOI: http://dx.doi.org/10.22159/ijpps.2018v10i4.22474 INTRODUCTION Heart attacks and strokes are the most common causes of mortality and morbidity across the world and both represent clinical manifestations of acute arterial thrombosis [1]. Among them, atherosclerosis presents a greater risk to cardiovascular and peripheral vascular system causing mortality in cardiovascular disease patients [2]. Thrombus formation is a key mediator in the development of atherosclerosis [3]. The platelet is believed to play a pivotal role in pathogenesis and progression of atherosclerosis [4]. Previously it was believed that the platelet is having a minor role in this process; however, it is recognized that the platelet places itself as a critical link between thrombus formation, inflammation, and atherosclerosis [5]. At the site of vascular injury, platelets come into contact with subendothelial components and form a plug-like structure to avoid future damage to the endothelium. However, if the injury continues to happen, it will activate the cascade of signalling molecules which will form thrombus at the site of injury. This will lead to life-threatening disease states such as myocardial infarction, atherosclerosis or ischemic stroke [6]. Upon platelet formation at the site of vascular injury a series of cascade initiates which mainly involves three phases. Phase I, Phase II and Phase III name as the initial phase, extension phase and stabilization phase respectively. The initial phase involves attachment of platelets to the exposed sub-endothelial layer following vascular injury and a monolayer of activated cells is formed. This activated monolayer further recruits more additional platelets to construct another strong layer during extension phase [7]. In order to restrict the recently formed thrombus to the initial injury site, regulation of platelet aggregation is the process to modulate a balance between activation and inhibition of signalling pathway of platelet [8]. Any defect in regulation of platelet activation or aggregation can cause arterial thrombosis, the major manifestation of atherosclerosis which triggers myocardial infarction and stroke. Nitric oxide (NO) is an endogenous gas present in endothelial cell of vascular endothelium, which is believed to play a predominant role in the regulation of platelet aggregation [9]. Endothelial NO responsible for maintenance of basal vascular tone and blood flow, and thereby regulation of blood pressure due to vasodilatory action [10]. A substantial amount of research has been carried out to elucidate the role of NO in platelet aggregation and found that NO can inhibit platelet activation or aggregation in vitro and in vivo [11]. The mechanism by which inhibition of platelet aggregation turns out by NO is caused by the cGMP-dependent pathway. Soluble guanylcyclase (sGC) is an enzyme responsible for the production of cyclic guanosine monophosphate (cGMP), and NO is involved in activation of sGC which further lead to activation of cGMP- dependent protein kinase (PKG) via a cGMP-dependent pathway. This activated PKG caused inhibition of platelet activation via phosphorylating TxTA2 receptors and thus inhibits its aggregating action on platelet. Further details include inhibition of influx of Ca 2+ and other positive ions via activating sarcoplasmic reticulum ATPase (SERCA) [12]. PKG is also involved in blockage of the release of Ca 2+ cation from the sarcoplasmic reticulum via inhibition of International Journal of Pharmacy and Pharmaceutical Sciences ISSN- 0975-1491 Vol 10, Issue 4, 2018