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Journal of the Hellenic Veterinary Medical Society Journal of the Hellenic Veterinary Medical Society Vol. 71, 2020 Isobolographic analysis of analgesic interactions of silymarin with ketamine in mice NASER A.S. Department of physiology, biochemistry and pharmacology, Faculty of Veterinary Medicine, University of Mosul, Mosul, Iraq ALBADRANY Y. Department of physiology, biochemistry and pharmacology, Faculty of Veterinary Medicine, University of Mosul, Mosul, Iraq SHAABAN K.A. [email protected] https://doi.org/10.12681/jhvms.23653 Copyright © 2020 A.S. NASER, Y. ALBADRANY, K.A. SHAABAN To cite this article: NASER, A.S., ALBADRANY, Y., & SHAABAN, K.A. (2020). Isobolographic analysis of analgesic interactions of silymarin with ketamine in mice. Journal of the Hellenic Veterinary Medical Society, 71(2), 2171-2178. doi:https://doi.org/10.12681/jhvms.23653 http://epublishing.ekt.gr | e-Publisher: EKT | Downloaded at 23/09/2021 18:43:31 | J HELLENIC VET MED SOC 2020, 71(2): 2171-2178 Research article ΠΕΚΕ 2020, 71(2): 2171-2178 Ερευνητικό άρθρο Isobolographic analysis of analgesic interactions of silymarin with ketamine in mice A.S. Naser1*, Y. Albadrany2, K.A. Shaaban2 1Department of physiology, biochemistry and pharmacology, Faculty of Veterinary Medicine, University of Mosul, Mosul, Iraq 2Department of physiology, biochemistry and pharmacology, Faculty of Veterinary Medicine, University of Mosul, Mosul, Iraq ABSTRACT: The present study was undertaken to explore the analgesic effect of silymarin and ketamine alone or in combination in mice. Analgesia was measured by using a hot plate and the writhing test. The up-and-down method was used to determine the median effective analgesic dosages (ED50s) of silymarin and ketamine administered intraper- itoneally (ip) either alone or together. The ED50s of both drugs were analyzed isobolographically to determine the type of pharmacological interaction between them. The analgesic ED50s for silymarin and ketamine in mice were 57.22 and 1.96 mg/kg, ip, respectively. Concomitant administration of the silymarin and ketamine at fixed ration (0.5:0.5) of their individual ED50s was 38.4 mg/kg and 1.28 mg/kg, ip, respectively. Silymarin and ketamine at fixed ration (1:1) of their individual ED50s were 47.54 mg/kg and 1.58 mg/kg, ip, respectively . Depending on the isobolographic analysis and calculating Y value, the type of pharmacological interaction between silymarin and ketamine at a ratio of 0.5:0.5 and 1:1 of their analgesic ED50 values of each drug , was antagonistic .In the writhing test the concomitant administration of silymarin and ketamine at 120mg/kg and 4mg/kg , ip, respectively reduce significantly the numbers of writhing in com- pare with silymarin120 mg/kg,ip and ketamine 4mg/kg, ip separately. The results suggest that the co-administration of silymarin and ketamine was ineffective to reduce the central pain while the concomitant administration of silymarin and ketamine was effective to reduce the visceral pain. Keywords: silymarin, ketamine, analgesia, isobolographic, hotplate, mice. Corresponding Author: Date of initial submission: 21-07-2020 S. Senturk, Department of Internal Medicine, Faculty of Veterinary Medicine, Date of revised submission: 21-10-2019 Uludag University, 16059, Bursa, Turkey Date of acceptance: 02-01-2020 E-mail address: [email protected] http://epublishing.ekt.gr | e-Publisher: EKT | Downloaded at 23/09/2021 18:43:31 | 2172 A.S. NASER, Y. ALBADRANY, K.A. SHAABAN INTRODUCTION volvement of central and peripheral systems, respec- etamine is an anesthetic and pain killeragen- tively(Acharya et al., 2011; Barrot, 2012; de Campos Ktused in human and animals for anesthesia which Buzzi et al., 2010; Jain, Kulkarni, & Singh, 2001). characterized by hypnosis and good analgesia with muscle rigidity(Carter & Story, 2013; Persson, 2013; MATERIALS AND METHODS Tawfic, 2013). Ketamine inhibits the polysynaptic ac- 46 Male and female Swiss albino mice weighing tions of the excitatory neurotransmitter acetylcholine 28–33 g were housed at a temperature of 20 ± 2°C (Ach) and glutamate in the spinal cord and block the and 10/14 h light/dark cycle, with water and food ad N-methyl-D-aspartate receptor complex (Hsu, 2013). libitum. All experiments complied with institutional Ketamine used in human(Aiello & Mays, 1998)and regulations addressing animal use, and the mice re- veterinary clinic (Rang, Dale, Ritter, & Moore, 2003) ceived suitable attention and humane care. The Scien- as a general anesthetic agent usually in combination tific Committee of the physiology, biochemistry and with sedatives, tranquilizers and analgesics. Ket- pharmacology of the College of Veterinary Medicine amine combinations with sedatives, tranquilizers and at the University of Mosul has reviewed and approved analgesics are also used as balanced anesthetics and the protocol of this study. The required doses of The restraining agents in wild animals(Carter & Story, commercial powder of silymarin (175mg, 21ST Cen- 2013; Coetzee, 2013; Short, 1992). Further, ketamine tury HealthCare, Inc.) was dissolved in propylene have anticonvulsant activity in mice (Tricklebank, glycol(Sigma chemical CO. 99%) . Ketamine (10 % Singh, Oles, Preston, & Iversen, 1989) and chicks injectable solution, DOPHARMA Netherland.) was (Reder, Trapp, & Troutman, 1980) further diluted in saline solution (Pharmaceutical Solution Industry, Saudi Arabia) to obtain the desired Silymarin (milk thistle), a medical herb belong- concentrations of the drug. The volume of administra- ing to the Asteraceae family(Ottai& Abdel-Moniem, tion of each drug was at 5 ml/kg body weight given 2006), is home Southern Europe, Southern Russia, intraperitoneally (ip). Two experimenters simultane- Asia Minor and Northern Africa (Abenavoli, Capas- ously observed the responses of the mice during the so, Milic, & Capasso, 2010).Silymarin is mixture of experiments which were conducted between 9–12 silibinin A and B (silybin A and B), silydianin and a.m. silychristin. Other flavonolignans include isosilybin A and B, isosilychristin and taxifolin(Ottai & Ab- Experiments del-Moniem, 2006; Stolf, Cardoso, & Acco, 2017). The up-and-down method (Dixon, 1980)was used Silymarin was used for the management of nu- to determine the median effective analgesic dosag- merous liver disorders including cirrhosis, hepatitis es (ED50s) of silymarin and ketamine (administered (Herz, Heywood, Harborne, & Turner, 1977; Luper, either alone, or concomitantly – silymarin followed 1998) Except hepatoprotective effects, it has cardio- immediately with ketamine) in mice. The initial dose protective, neuroprotective, cytoprotective, anti-in- of silymarin was at 100 mg/kg, ipwhereas that of ket- flammatory , analgesic and anti-carcinogenic effects amine was at 2 mg/kg, ip. In the combination exper- (Manna, Mukhopadhyay, Van, & Aggarwal, 1999; iment, the initial dosages of silymarin and ketamine Naser & Amin, 2019; Škottová et al., 2003)A useful were 30 and 1 mg/kg, ip, respectively (50:50 of their practice is a combination of two drugs with the same individual ED50 values) and the initial dosages of sily- therapeutic effect where in each agent is administered marin and ketamine were 60 and 2 mg/kg, ip, respec- to obtain additive, synergistic or antagonism inter- tively (1:1 of their individual ED50 values) . We based action in a fixed ratio. If the combination resulted in our choices for silymarin and ketamine dosages on addition or synergism, the doses employed by each preliminary experiments in mice, as well as on previ- drug are reduced, then the side effects are decline; this ous studies (G. Jadhav & Upasani, 2009; Mohammad, type of study is calledisobolographic analysis (Raffa, Al-Baggou, & Naser, 2012; Sabiu, Sunmonu, Ajani, 2001). & Ajiboye, 2015; Vahdati Hassani et al., 2015) . An- algesia was measured by the thermal method using a The purpose of the presentstudy was to explore hot plate (Panlab, S.I.U., Cornella, Spain) held at a the analgesic effect of silymarin and ketamine com- temperature of 56°C. Mice were individually placed bination in mice. The models of pain induction used on the hot plate 15 min after the drug administration were the hot plateand chemical-induced writhing, and the latency time to the first hind paw removal/ which are predictive of acute pain responses with in- J HELLENIC VET MED SOC 2020, 71(2) ΠΕΚΕ 2020, 71(2) http://epublishing.ekt.gr | e-Publisher: EKT | Downloaded at 23/09/2021 18:43:31 | A.S. NASER, Y. ALBADRANY, K.A. SHAABAN 2173 licking and/or jumping response was measured (Bar- Inhibition % = Mean no of writhes (control) – mean rot, 2012; Le Bars, Gozariu, & Cadden, 2001)The no of writhes (test) / Mean no of writhes (control) × cutoff point latency for the induction of analgesia was 100. equal to or more than 6 s (i.e., positive analgesic re- sponse), and the maximum time allowed for the ani- Statistical Analysis mal to stay on the hot plate was 20 s to prevent tissue Data are expressed as mean + SEM. Statistical damage .. The ED50s of both drugs were subjected to analysis was done by using one way analysis of vari- isobolographic analysis to determine the type of inter- ance (ANOVA).P<0.05were considered significant. action involved in the administration of silymarin and ketamine concomitantly (Gonzalez, Zegpi, Noriega, RESULTS Prieto, & Miranda, 2011; Mohammad, Al-Zubaidy, & Alias, 2007; Tallarida, 1992)A straight line was drawn Hot plate test The ED for silymarin - and ketamine induced for the isobolographic analysis between isoeffective 50s analgesia in mice, as determined by the up
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