Effects of berberine hydrochloride on methamphetamine-induced anxiety behaviors and relapse in rats

Document Type : Original Article


1 Department of Addiction Studies, School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran

2 Addiction Research Center, Shahroud University of Medical Sciences, Shahroud, Iran

3 Student Research Committee, School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran

4 Department of Physiology, School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran

5 Department of Neuroscience, School of Advanced Technologies in Medicine, Mazandaran University of Medical Sciences, Sari, Iran


Objective(s): This research aimed at evaluating the effect of berberine hydrochloride on anxiety-related behaviors induced by methamphetamine (METH) in rats, assessing relapse and neuroprotective effects.
Materials and Methods: 27 male Wistar rats were randomly assigned into groups of Control, METH-withdrawal (METH addiction and subsequent withdrawal), and METH addiction with berberine hydrochloride oral treatment (100 mg/kg/per day) during the three weeks of withdrawal. Two groups received inhaled METH self-administration for two weeks (up to 10 mg/kg). The elevated plus maze (EPM) test and open field test (OFT) were carried out one day after the last berberine treatment and relapse was assessed by conditional place preference (CPP) test. TUNEL assay and immunofluorescence staining for NF-κB, TLR4, Sirt1, and α-actin expression in the hippocampus were tested.
Results: After 3 weeks withdrawal, berberine hydrochloride decreased locomotor activity and reduced anxiety-related behaviors in comparison with the METH-withdrawal group (p <0.001). The obtained results from CPP showed that berberine significantly reduced relapse (p <0.01). Significantly decrease in activation of TLR4, Sirt1, and α-actin in METH-withdrawal group was found and the percentage of TLR4, Sirt1, and α-actin improved in berberine-treated group (p <0.001). A significant activity rise of NF-κB of cells in the METH-withdrawal group was detected compared to berberine-treated and control groups (p <0.001).
Conclusion: Treatment with berberine hydrochloride via modulating neuroinflammation may be considered as a potential new medication for the treatment of METH addiction and relapse. The histological assays supported the neuroprotective effects of berberine in the hippocampus.


1. Yasaei R, Saadabadi A. Methamphetamine.  StatPearls [Internet]: StatPearls Publishing; 2020.
2. Moratalla R, Khairnar A, Simola N, Granado N, García-Montes JR, Porceddu PF, et al. Amphetamine-related drugs neurotoxicity in humans and in experimental animals: main mechanisms. Prog Neurobiol 2017; 155:149-170.
3. Krasnova IN, Cadet JL. Methamphetamine toxicity and messengers of death. Brain Res Rev 2009; 60:379-407.
4. Bannerman D, Rawlins J, McHugh S, Deacon R, Yee B, Bast T, et al. Regional dissociations within the hippocampus—memory and anxiety. Neurosci Biobehav Rev 2004; 28:273-283.
5. Felger JC. Imaging the role of inflammation in mood and anxiety-related disorders. Curr Neuropharmacol 2018; 16:533-558.
6. Coelho-Santos V, Leitao RA, Cardoso FL, Palmela I, Rito M, Barbosa M, et al. The TNF-alpha/NF-kappaB signaling pathway has a key role in methamphetamine-induced blood-brain barrier dysfunction. J Cereb Blood Flow Metab 2015; 35:1260-1271.
7. Koo JW, Russo SJ, Ferguson D, Nestler EJ, Duman RS. Nuclear factor-κB is a critical mediator of stress-impaired neurogenesis and depressive behavior. Proc Natl Acad Sci 2010; 107:2669-2674.
8. Femenia T, Qian Y, Arentsen T, Forssberg H, Heijtz RD. Toll-like receptor-4 regulates anxiety-like behavior and DARPP-32 phosphorylation. Brain Behav Immun 2018; 69:273-282.
9. Kawamoto EM, Cutler RG, Rothman SM, Mattson MP, Camandola S. TLR4-dependent metabolic changes are associated with cognitive impairment in an animal model of type 1 diabetes. Biochem Biophys Res Co 2014; 443:731-737.
10. Guerri C, Pascual M. Role of toll-like receptor 4 in alcohol-induced neuroinflammation and behavioral dysfunctions. In Neural-immune interactions in brain function and alcohol related disorders: Springer; 2013. p. 279-306.
11. Trotta T, Porro C, Calvello R, Panaro MA. Biological role of Toll-like receptor-4 in the brain. J Neuroimmunol 2014; 268:1-12.
12. Walter S, Letiembre M, Liu Y, Heine H, Penke B, Hao W, et al. Role of the toll-like receptor 4 in neuroinflammation in Alzheimer’s disease. Cell Physiol Biochem 2007; 20:947-956.
13. Wang X, Northcutt AL, Cochran TA, Zhang X, Fabisiak TJ, Haas ME, et al. Methamphetamine activates Toll-like receptor 4 to induce central immune signaling within the ventral tegmental area and contributes to extracellular dopamine increase in the nucleus accumbens shell. ACS Chem neurosci 2019; 10:3622-3634.
14. Chong ZZ, Shang YC, Wang S, Maiese K. SIRT1: new avenues of discovery for disorders of oxidative stress. Expert Opin Ther Targets 2012; 16:167-178.
15. Hisahara S, Chiba S, Matsumoto H, Tanno M, Yagi H, Shimohama S, et al. Histone deacetylase SIRT1 modulates neuronal differentiation by its nuclear translocation. Proc Natl Acad Sci 2008; 105:15599-15604.
16. Calvanese V, Lara E, Suárez-Álvarez B, Dawud RA, Vázquez-Chantada M, Martínez-Chantar ML, et al. Sirtuin 1 regulation of developmental genes during differentiation of stem cells. Proc Natl Acad Sci 2010; 107:13736-13741.
17. Gonfloni S, Iannizzotto V, Maiani E, Bellusci G, Ciccone S, Diederich M. P53 and Sirt1: routes of metabolism and genome stability. Biochem Pharmacol 2014; 92:149-156.
18. Shoba B, Lwin ZM, Ling LS, Bay BH, Yip GW, Kumar SD. Function of sirtuins in biological tissues. Anat Rec 2009; 292:536-543.
19. Lamprecht R. The roles of the actin cytoskeleton in fear memory formation. Front Behav Neurosci 2011; 5:39-49.
20. Young EJ, Briggs SB, Miller CA. The actin cytoskeleton as a therapeutic target for the prevention of relapse to methamphetamine use. CNS Neurol Disord Drug Targets 2015; 14:731-737.
21. Zou K, Li Z, Zhang Y, Zhang H-y, Li B, Zhu W-l, et al. Advances in the study of berberine and its derivatives: a focus on anti-inflammatory and anti-tumor effects in the digestive system. Acta Pharmacol Sin 2017; 38:157-167.
22. Sun Y, Xun K, Wang Y, Chen X. A systematic review of the anticancer properties of berberine, a natural product from Chinese herbs. Anti-cancer drugs 2009; 20:757-769.
23. Peng L, Kang S, Yin Z, Jia R, Song X, Li L, et al. Antibacterial activity and mechanism of berberine against Streptococcus agalactiae. Int J Clin Exp Pathol 2015; 8:5217.
24. Peng W-H, Wu C-R, Chen C-S, Chen C-F, Leu Z-C, Hsieh M-T. Anxiolytic effect of berberine on exploratory activity of the mouse in two experimental anxiety models: interaction with drugs acting at 5-HT receptors. Life Sci 2004; 75:2451-2462.
25. Mangrulkar SV, Selote RD, Chaple DR, Chourasia A. Antiamnesic effect of berberine in colchicines induced experimental alzheimer’s disease model. Int J Pharm Bio Sci 2013; 4:618-624.
26. Yoo J-H, Yang E-M, Cho J-H, Lee J-H, Jeong S, Nah S-Y, et al. Inhibitory effects of berberine against morphine-induced locomotor sensitization and analgesic tolerance in mice. Neurosci 2006; 142:953-961.
27. Kulkarni S, Dhir A. Berberine: A plant alkaloid with therapeutic potential for central nervous system disorders. Phytother Res 2010; 24:317-324.
28. Kulkarni SK, Dhir A. On the mechanism of antidepressant-like action of berberine chloride. Eur J Pharmacol 2008; 589:163-172.
29. Fan J, Zhang K, Jin Y, Li B, Gao S, Zhu J, et al. Pharmacological effects of berberine on mood disorders. J Cell Mol Med 2019; 23:21-28.
30. Shi M, Zhao C, Ge X, Yang H, Ge L, Zhu G, et al. Berberine prevents cognitive disorders induced by sepsis by regulating the inflammatory cytokines, oxidative stress and neuronal apoptosis in rat brain. Neuropsychiatry 2018; 8:17-26.
31. Chandirasegaran G, Elanchezhiyan C, Ghosh K, Sethupathy S. Berberine chloride ameliorates oxidative stress, inflammation and apoptosis in the pancreas of Streptozotocin induced diabetic rats. Biomed Pharmacother 2017; 95:175-185.
32. Karila L, Weinstein A, Aubin H-J, Benyamina A, Reynaud M, Batki SL. Pharmacological approaches to methamphetamine dependence: a focused review. Br J Clin Pharmacol 2010; 69:578-592.
33. Moghaddam HK, Baluchnejadmojarad T, Roghani M, Goshadrou F, Ronaghi A. Berberine chloride improved synaptic plasticity in STZ induced diabetic rats. Metab Brain Dis 2013; 28:421-428.
34. Rafaiee R, Ahmadiankia N, Mousavi SA, Rezaeian L, Niroumand Sarvandani M, Shekari A, et al. Inhalant self-administration of methamphetamine: the most similar model to human methamphetamine addiction. Iran J Psychiatry Behav Sci 2019; 13:e90561.
35. Belovicova K, Bogi E, Csatlosova K, Dubovicky M. Animal tests for anxiety-like and depression-like behavior in rats. Interdiscip Toxicol 2017; 10:40-43.
36. Walf AA, Frye CA. The use of the elevated plus maze as an assay of anxiety-related behavior in rodents. Nat Protoc 2007; 2:322-328.
37. Wellington D, Mikaelian I, Singer L. Comparison of ketamine–xylazine and ketamine–dexmedetomidine anesthesia and intraperitoneal tolerance in rats. J Am Assoc Lab Anim Sci 2013; 52:481-487.
38. Gage GJ, Kipke DR, Shain W. Whole animal perfusion fixation for rodents. J Vis Exp 2012:3564-3573.
39. Yu S, Zhu L, Shen Q, Bai X, Di X. Recent Advances in Methamphetamine Neurotoxicity Mechanisms and Its Molecular Pathophysiology. Behav Neurol 2015; 2015:103969-103980.
40. Zuloaga D, Jacobskind J, Raber J. Methamphetamine and the hypothalamic-pituitary-adrenal axis. Front Neurosci 2015; 9-17.
41. Zhou X-Q, Zeng X, Kong H, Sun X-L. Neuroprotective effects of berberine on stroke models in vitro and in vivo. Neurosci Lett 2008; 447:31-36.
42. Durairajan SSK, Liu L-F, Lu J-H, Chen L-L, Yuan Q, Chung SK, et al. Berberine ameliorates β-amyloid pathology, gliosis, and cognitive impairment in an Alzheimer’s disease transgenic mouse model. Neurobiol Aging 2012; 33:2903-2919.
43. Moneim AEA. The neuroprotective effect of berberine in mercury-induced neurotoxicity in rats. Metab Brain Dis 2015; 30:935-942.
44. Jiang W, Li S, Li X. Therapeutic potential of berberine against neurodegenerative diseases. China Life Sci 2015; 58:564-569.
45. Azam S, Jakaria M, Kim I-S, Kim J, Haque ME, Choi D-K. Regulation of Toll-Like Receptor (TLR) signaling pathway by polyphenols in the treatment of age-linked neurodegenerative diseases: focus on TLR4 signaling. Front Immunol 2019; 10-27.
46. He X-F, Zhang L, Zhang C-H, Zhao C-R, Li H, Zhang L-F, et al. Berberine alleviates oxidative stress in rats with osteoporosis through receptor activator of NF-kB/receptor activator of NF-kB ligand/osteoprotegerin (RANK/RANKL/OPG) pathway. Bosnian J Basic Med Sci 2017; 17:295-301.
47. Zhu L, Han J, Yuan R, Xue L, Pang W. Berberine ameliorates diabetic nephropathy by inhibiting TLR4/NF-κB pathway. Biol Res 2018; 51:9-21.
48. Li  X, Wu F, Xue L, Wang B, Li J, Chen Y, et al. Methamphetamine causes neurotoxicity by promoting polarization of macrophages and inflammatory response. Hum Exp Toxicol 2017; 37:486-495.
49. Kuo C-L, Chi C-W, Liu T-Y. The anti-inflammatory potential of berberine in vitro and in vivo. Cancer Lett 2004; 203:127-137.
50. Lee D-U, Kang YJ, Park MK, Lee YS, Seo HG, Kim TS, et al. Effects of 13-alkyl-substituted berberine alkaloids on the expression of COX-II, TNF-α, iNOS, and IL-12 production in LPS-stimulated macrophages. Life Sci 2003; 73:1401-1412.
51. Chen F, Yang Z, Liu Y, Li L, Liang W, Wang X, et al. Berberine inhibits the expression of TNFα, MCP-1, and IL-6 in AcLDL-stimulated macrophages through PPARγ pathway. Endocrine 2008; 33:331-337.
52. Yu L, Li Q, Yu B, Yang Y, Jin Z, Duan W, et al. Berberine attenuates myocardial ischemia/reperfusion injury by reducing oxidative stress and inflammation response: role of silent information regulator 1. Oxid Med Cell Longev 2016; 2016:1689602-1689618.
53.Chen D-L, Yang K-Y. Berberine Alleviates Oxidative Stress in Islets of Diabetic Mice by Inhibiting miR-106b Expression and Up-Regulating SIRT1. J Cell Biochem 2017; 118:4349-4357.
54. Li H-Y, Wang X-C, Xu Y-M, Luo N-C, Luo S, Hao X-Y, et al. Berberine improves diabetic encephalopathy through the SIRT1/ER stress pathway in db/db mice. Rejuvenation Res 2018; 21:200-209.
55. Zhu X, Guo X, Mao G, Gao Z, Wang H, He Q, et al. Hepatoprotection
of berberine against hydrogen peroxide‐induced apoptosis by upregulation of sirtuin 1. Phytother Res 2013; 27:417-421.
56. Cheng Y, Takeuchi H, Sonobe Y, Jin S, Wang Y, Horiuchi H, et al. Sirtuin 1 attenuates oxidative stress via upregulation of superoxide dismutase 2 and catalase in astrocytes. J Neuroimmunol 2014; 269:38-43.
57. Kim D, Nguyen MD, Dobbin MM, Fischer A, Sananbenesi F, Rodgers JT, et al. SIRT1 deacetylase protects against neurodegeneration in models for Alzheimer’s disease and amyotrophic lateral sclerosis. EMBO J 2007; 26:3169-3179.
58. Codocedo JF, Allard C, Godoy JA, Varela-Nallar L, Inestrosa NC. SIRT1 regulates dendritic development in hippocampal neurons. PLoS One 2012; 7:e47073-e47073.
59. Paraíso AF, Mendes KL, Santos SHS. Brain activation of SIRT1: role in neuropathology. Mol Neurobiol 2013; 48:681-689.
60. Haase R, Kirschning CJ, Sing A, Schrottner P, Fukase K, Kusumoto S, et al. A dominant role of Toll-like receptor 4 in the signaling of apoptosis in bacteria-faced macrophages. J Immunol 2003; 171:4294-4303.