Background: Ketamine is a widely used anesthetic in animal research, but its use is strictly regulated in several countries, including Japan and China. As an alternative, the medetomidine-midazolam-butorphanol (MMB) combination is commonly used in Japan. However, medetomidine is a racemic mixture containing the inactive R-enantiomer, which may reduce anesthetic predictability and safety.
Objective: The aim of the study was to evaluate the efficacy and safety of a modified anesthetic combination (dMMB), in which dexmedetomidine replaces medetomidine, across three commonly used mouse strains (ICR, C57BL/6, BALB/c).
Methods: Male and female mice were administered either MMB or dMMB subcutaneously. Anesthetic depth, recovery profiles, heart rate, SpO2, body temperature, ocular opacity, and blood glucose levels were assessed. Atipamezole was used to reverse anesthesia, and thermoregulatory recovery was monitored postinjection.
Results: dMMB produced similar anesthetic depth to MMB, with faster and more consistent recovery, particularly in males. Body temperature recovery was significantly enhanced in dMMB-treated B6 males. No significant differences in side effects (ocular opacity or blood glucose levels) were observed between protocols, though strain-specific glucose elevations were noted in dMMB-treated males.
Conclusion: dMMB is a safe, effective, and ketamine-free injectable anesthetic protocol, offering advantages in recovery and thermoregulation. It may be a valuable alternative in research settings where ketamine is restricted and medetomidine may become unavailable.
| [1] |
Brown EN, Pavone KJ, Naranjo M. Multimodal general anesthesia: theory and practice. Anesth Analg. 2018;127:1246-1258.
|
| [2] |
Shim JH. Multimodal analgesia or balanced analgesia: the better choice? Korean J Anesthesiol. 2020;73:361-362.
|
| [3] |
Kawai S, Takagi Y, Kaneko S, Kurosawa T. Effect of three types of mixed anesthetic agents alternate to ketamine in mice. Exp Anim. 2011;60:481-487.
|
| [4] |
Tang Y, Yang K, Xu J, Qiu Y, Meng L, Zhao S. Toxicological assessment of ketamine in juvenile zebrafish (Danio rerio). Toxics. 2025;13(2):82.
|
| [5] |
Scheinin H, Virtanen R, MacDonald E, Lammintausta R, Scheinin M. Medetomidine—a novel alpha 2-adrenoceptor agonist: a review of its pharmacodynamic effects. Prog Neuro-Psychopharmacol Biol Psychiatry. 1989;13:635-651.
|
| [6] |
Sinclair MD. A review of the physiological effects of alpha2-agonists related to the clinical use of medetomidine in small animal practice. Can Vet J. 2003;44:885-897.
|
| [7] |
Schwinn DA, Correa-Sales C, Page SO, Maze M. Functional effects of activation of alpha-1 adrenoceptors by dexmedetomidine: in vivo and in vitro studies. J Pharmacol Exp Ther. 1991;259:1147-1152.
|
| [8] |
Guo TZ, Tinklenberg J, Oliker R, Maze M. Central alpha 1-adrenoceptor stimulation functionally antagonizes the hypnotic response to dexmedetomidine, an alpha 2-adrenoceptor agonist. Anesthesiology. 1991;75:252-256.
|
| [9] |
Giovannitti JA Jr, Thoms SM, Crawford JJ. Alpha-2 adrenergic receptor agonists: a review of current clinical applications. Anesth Prog. 2015;62:31-39.
|
| [10] |
Di Franco C, Evangelista F, Briganti A. Multiple uses of dexmedetomidine in small animals: a mini review. Front Vet Sci. 2023;10:1135124.
|
| [11] |
Morris TH. Use of medetomidine and atipamezole in laboratory animals. J Vet Anaesth. 1991;18:277-279.
|
| [12] |
Mekada K, Yoshiki A. Substrains matter in phenotyping of C57BL/6 mice. Exp Anim. 2021;70:145-160.
|
| [13] |
Shintaku T, Ohba T, Niwa H, et al. Effects of isoflurane inhalation anesthesia on mouse ECG. Hirosaki Med J. 2015;66:1-7.
|
| [14] |
Drysch M, Wallner C, Schmidt SV, et al. An optimized low-pressure tourniquet murine hind limb ischemia reperfusion model: inducing acute ischemia reperfusion injury in C57BL/6 wild type mice. PLoS One. 2019;14:e0210961.
|
| [15] |
Durst MS, Arras M, Palme R, Talbot SR, Jirkof P. Lidocaine and bupivacaine as part of multimodal pain management in a C57BL/6J laparotomy mouse model. Sci Rep. 2021;11:10918.
|
| [16] |
Watanabe H, Numata K, Ito T, Takagi K, Matsukawa A. Innate immune response in Th1- and Th2-dominant mouse strains. Shock. 2004;22:460-466.
|
| [17] |
Kirihara Y, Takechi M, Kurosaki K, Kobayashi Y, Kurosawa T. Anesthetic effects of a mixture of medetomidine, midazolam and butorphanol in two strains of mice. Exp Anim. 2013;62:173-180.
|
| [18] |
Tanaka T. Biological and environmental factors influencing reproductive performance in ICR mice, Mus musculus. Birth Defects Res. 2024;116:e2337.
|
| [19] |
Gergye CH, Zhao Y, Moore RH, Lee VK. A comparison of ketamine or etomidate combined with xylazine for intraperitoneal anesthesia in four mouse strains. J Am Assoc Lab Anim Sci. 2020;59:519-530.
|
| [20] |
Hwang DJ, Song HK, Kim KS, Jung YS, Hwang DY, Cho JY. Comparative analysis of basal locomotor activity-related metabolic phenotypes between C57BL/6 mice and ICR mouse substrains derived from three different sources. Lab Anim Res. 2017;33:140-149.
|
| [21] |
Obernier JA, Baldwin RL. Establishing an appropriate period of acclimatization following transportation of laboratory animals. ILAR J. 2006;47:364-369.
|
| [22] |
Shibuta H, Yamana R, Kashimoto J, Kamio K, Suda A. Comparison of the anesthetic effect by the injection route of mixed anesthesia (medetomidine, midazolam and butorphanol) and the effect of this anesthetic agent on the respiratory function. J Vet Med Sci. 2020;82:35-42.
|
| [23] |
Burnside WM, Flecknell PA, Cameron AI, Thomas AA. A comparison of medetomidine and its active enantiomer dexmedetomidine when administered with ketamine in mice. BMC Vet Res. 2013;9:48.
|
| [24] |
Tashiro M, Tohei A. Recommended doses of medetomidine-midazolam-butorphanol with atipamezole for preventing hypothermia in mice. J Vet Med Sci. 2022;84:445-453.
|
| [25] |
Watanabe M, Nikaido Y, Sasaki N. Validation of the anesthetic effect of a mixture of remimazolam, medetomidine, and butorphanol in three mouse strains. Exp Anim. 2024;73:223-232.
|
| [26] |
Tsukamoto Y, Yamada N, Miyoshi K, Yamashita K, Ohsugi T. Anesthetic effect of a mixture of alfaxalone, medetomidine, and butorphanol for inducing surgical anesthesia in ICR, BALB/c, and C57BL/6 mouse strains. J Vet Med Sci. 2019;81:937-945.
|
| [27] |
Gaines Das R, North D. Implications of experimental technique for analysis and interpretation of data from animal experiments: outliers and increased variability resulting from failure of intraperitoneal injection procedures. Lab Anim. 2007;41:312-320.
|
| [28] |
Laferriere CA, Leung VS, Pang DS. Evaluating intrahepatic and intraperitoneal sodium pentobarbital or ethanol for mouse euthanasia. J Am Assoc Lab Anim Sci. 2020;59:264-268.
|
| [29] |
Navarro KL, Huss M, Smith JC, Sharp P, Marx JO, Pacharinsak C. Mouse anesthesia: the art and science. ILAR J. 2021;62:238-273.
|
| [30] |
Buitrago S, Martin TE, Tetens-Woodring J, Belicha-Villanueva A, Wilding GE. Safety and efficacy of various combinations of injectable anesthetics in BALB/c mice. J Am Assoc Lab Anim Sci. 2008;47:11-17.
|
| [31] |
Constantinides C, Mean R, Janssen BJ. Effects of isoflurane anesthesia on the cardiovascular function of the C57BL/6 mouse. ILAR J. 2011;52:e21-e31.
|
| [32] |
Calderone L, Grimes P, Shalev M. Acute reversible cataract induced by xylazine and by ketamine-xylazine anesthesia in rats and mice. Exp Eye Res. 1986;42(4):331-337.
|
| [33] |
Kodera SY, Yoshida M, Dezaki K, et al. Inhibition of insulin secretion from rat pancreatic islets by dexmedetomidine and medetomidine, two sedatives frequently used in clinical settings. Endocr J. 2013;60(3):337-346.
|
| [34] |
Guedes AG, Rude EP. Effects of pre-operative administration of medetomidine on plasma insulin and glucose concentrations in healthy dogs and dogs with insulinoma. Vet Anaesth Analg. 2013;40(5):472-481.
|
| [35] |
Ansah OB, Raekallio M, Vainio O. Comparison of three doses of dexmedetomidine with medetomidine in cats following intramuscular administration. J Vet Pharmacol Ther. 1998;21:380-387.
|
| [36] |
Waxman DJ, Holloway MG. Sex differences in the expression of hepatic drug metabolizing enzymes. Mol Pharmacol. 2009;76:215-228.
|
| [37] |
Craft RM, Mogil JS, Aloisi AM. Sex differences in pain and analgesia: the role of gonadal hormones. Eur J Pain. 2004;8:397-411.
|
| [38] |
Wasilczuk AZ, Rinehart C, Aggarwal A, et al. Hormonal basis of sex differences in anesthetic sensitivity. Proc Natl Acad Sci USA. 2024;121(3):e2312913120.
|
| [39] |
Flecknell P. Laboratory Animal Anaesthesia and Analgesia. 5th ed. Elsevier; 2023.
|
| [40] |
Cruz JI, Loste JM, Burzaco OH. Observations on the use of medetomidine/ketamine and its reversal with atipamezole for chemical restraint in the mouse. Lab Anim. 1998;32(1):18-22.
|
| [41] |
Kuusela E, Raekallio M, Anttila M, Falck I, Mölsä S, Vainio O. Clinical effects and pharmacokinetics of medetomidine and its enantiomers in dogs. J Vet Pharmacol Ther. 2000;23:15-20.
|
| [42] |
Granholm M, McKusick BC, Westerholm FC, Aspegrén JC. Evaluation of the clinical efficacy and safety of dexmedetomidine or medetomidine in cats and their reversal with atipamezole. Vet Anaesth Analg. 2006;33:214-223.
|
| [43] |
Granholm M, McKusick BC, Westerholm FC, Aspegrén JC. Evaluation of the clinical efficacy and safety of intramuscular and intravenous doses of dexmedetomidine and medetomidine in dogs and their reversal with atipamezole. Vet Rec. 2007;160:891-897.
|
| [44] |
Gómez-Villamandos RJ, Palacios C, Benítez A, et al. Dexmedetomidine or medetomidine premedication before propofol-desflurane anaesthesia in dogs. J Vet Pharmacol Ther. 2006;29:157-163.
|
| [45] |
Sacks M, Ringer SK, Bischofberger AS, Berchtold SM, Bettschart-Wolfensberger R. Clinical comparison of dexmedetomidine and medetomidine for isoflurane balanced anaesthesia in horses. Vet Anaesth Analg. 2017;44:1128-1138.
|
| [46] |
König C, Plank AC, Kapp A, Timotius IK, von Hörsten S, Zimmermann K. Thirty mouse strain survey of voluntary physical activity and energy expenditure: influence of strain, sex and day-night variation. Front Neurosci. 2020;14:531.
|
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2025 The Author(s). Animal Models and Experimental Medicine published by John Wiley & Sons Australia, Ltd on behalf of The Chinese Association for Laboratory Animal Sciences.
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