Neurotoxicity Research

, Volume 35, Issue 4, pp 848–859 | Cite as

Eugenol Attenuates Scopolamine-Induced Hippocampal Cholinergic, Glutamatergic, and Mitochondrial Toxicity in Experimental Rats

  • Debapriya GarabaduEmail author
  • Mahima Sharma
Original Article


Eugenol is one of the essential chemical constituents of several functional food plants including Eugenia caryophyllata Thunb. (Family: Myrtaceae). Eugenol exhibits neuroprotective and anti-stress activities through multimodal mechanisms of action. Further, eugenol exerts anti-amnesic activity in Alzheimer’s disease (AD)–like animals perhaps through anti-oxidant mechanism to date. Hence, the present study was designed to elaborate the anti-amnesic activity of eugenol in scopolamine-challenged rodents. Scopolamine (3 mg/kg/day, i.p.) and eugenol (12.5, 25.0, and 50.0 mg/kg) were administered to male rats for 14 consecutive days of the experimental schedule at a time lag of 30 min. Eugenol (25.0 and 50.0 mg/kg) attenuated scopolamine-induced loss in learning ability in terms of increased escape latency at day-4 (D-4) and memory function in terms of decreased time spent in target quadrant at D-5 of Morris water maze test protocol. Moreover, eugenol attenuated scopolamine-induced loss in spatial memory in terms of decreased percentage of spontaneous alteration behavior in Y-maze test. Additionally, eugenol attenuated scopolamine-induced hippocampal cholinergic dysfunction (decrease in acetylcholine level, increase in acetylcholinesterase activity, and decrease in density and affinity of M1 and total muscarinic receptor), glutamate neurotoxicity (increase in levels of glutamate, calcium, calcium-dependent calpain-2, and brain-derived neurotropic factor), and mitochondrial dysfunction (decrease in formazan produced, membrane potential, and oxidative stress) in rats. Thus, it could be considered as an alternate candidate in the management of AD. Moreover, inclusion of functional foods containing eugenol could be a better option to manage memory formation in neurological disorders.


Eugenol Memory Hippocampus Acetylcholine Glutamate Mitochondria 


Funding Information

This study was financially assisted by GLA University, Mathura, Uttar Pradesh, India.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interest.


  1. Armstrong RA (2006) Measuring the spatial arrangement patterns of pathological lesions in histological sections of brain tissue. Folia Neuropathol 44:229–237PubMedGoogle Scholar
  2. Balaban H, Nazıroglu M, Demirci K, Övey IS (2016) The protective role of selenium on scopolamine-induced memory impairment, oxidative stress, and apoptosis in aged rats: the involvement of TRPM2 and TRPV1 channels. Mol Neurobiol 54:2852–2868CrossRefPubMedGoogle Scholar
  3. Beers JRF, Sizer IW (1952) A spectrophotometric method for measuring the breakdown of hydrogen peroxide by catalase. J Biol Chem 195:133–140Google Scholar
  4. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254CrossRefGoogle Scholar
  5. Cho JS, Kim TH, Lim JM, Song JH (2008) Effects of eugenol on Na+ currents in rat dorsal root ganglion neurons. Brain Res 1243:53–62CrossRefPubMedGoogle Scholar
  6. Choi SH, Park CH, Koo JW, Seo JH, Kim HS, Jeong SJ et al (2001) Memory impairment and cholinergic dysfunction by centrally administered Abeta and carboxyl-terminal fragment of Alzheimer’s APP in mice. FASEB J 15:1816–1818CrossRefPubMedGoogle Scholar
  7. Cummings JL (2004) Treatment of Alzheimer’s disease: current and future therapeutic approaches. Rev Neurol Dis 1:60–69PubMedGoogle Scholar
  8. Fischer IU, Dengler HJ (1990) Sensitive high-performance liquid chromatographic assay for the determination of eugenol in body fluids. J Chromatogr 525:369–377CrossRefPubMedGoogle Scholar
  9. Garabadu D, Shah A, Ahmad A, Joshi VB, Saxena B, Palit G, Krishnamurthy S (2011) Eugenol as an anti-stress agent: modulation of hypothalamic-pituitary-adrenal axis and brain monoaminergic systems in a rat model of stress. Stress 14:145–155CrossRefPubMedGoogle Scholar
  10. Garabadu D, Shah A, Singh S, Krishnamurthy S (2015) Protective effect of eugenol against restraint stress-induced gastrointestinal dysfunction: potential use in irritable bowel syndrome. Pharm Biol 53:968–974CrossRefPubMedGoogle Scholar
  11. Green LC, Wagner DA, Glogowski J, Skipper PL, Wishnok JS, Tannenbaum SR (1982) Analysis of nitrate, nitrite, and [15N] nitrate in biological fluids. Anal Biochem 126:131–138CrossRefPubMedPubMedCentralGoogle Scholar
  12. Grynkiewicz G, Poenie M, Tsien RY (1985) A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem 260:3440–3450Google Scholar
  13. Guénette SA, Ross A, Marier JF, Beaudry F, Vachon P (2007) Pharmacokinetics of eugenol and its effects on thermal hypersensitivity in rats. Eur J Pharmacol 562:60–67CrossRefPubMedGoogle Scholar
  14. Guo C, Shen J, Meng Z, Yang X, Li F (2016) Neuroprotective effects of polygalacic acid on scopolamine-induced memory deficits in mice. Phytomedicine 23:149–155CrossRefPubMedGoogle Scholar
  15. Hersch SM, Levey AI (1995) Diverse pre- and post-synaptic expression of m1–m4 muscarinic receptor proteins in neurons and afferents in the rat neostriatum. Life Sci 56:931–938CrossRefPubMedGoogle Scholar
  16. Huang SG (2002) Development of a high throughput screening assay for mitochondrial membrane potential in living cells. J Biomol Screen 7:383–389CrossRefPubMedGoogle Scholar
  17. James BP Jr, Brian MC (1997) Release of endogenous glutamate and g-amino butyric acid from rat striatal tissue slices measured by an improved method of high performance liquid chromatography with electrochemical detection. J Neurosci 75:207–214Google Scholar
  18. Jeronimo-Santos A, Vaz SH, Parreira S, Rapaz-Lerias S, Caetano AP, Buee-Scherrer V et al (2015) Dysregulation of TrkB receptors and BDNF function by amyloid-β peptide is mediated by calpain. Cereb Cortex 25:3107–3121CrossRefPubMedGoogle Scholar
  19. Joshi A, Bondada V, Geddes JW (2009) Mitochondrial micro-calpain is not involved in the processing of apoptosis-inducing factor. Exp Neurol 218:221–227CrossRefPubMedPubMedCentralGoogle Scholar
  20. Kakkar P, Das B, Viswanathan PN (1984) A modified spectrophotometric assay of superoxide dismutase. Indian J Biochem Biophys 21:130–132PubMedGoogle Scholar
  21. Kamboj SS, Kumar V, Kamboj A, Sandhir R (2008) Mitochondrial oxidative stress and dysfunction in rat brain induced by carbofuran exposure. Cell Mol Neurobiol 28:961–969CrossRefPubMedGoogle Scholar
  22. Krishnaswamy K, Raghuramulu N (1998) Bioactive phytochemicals with emphasis on dietary practices. Indian J Med Res 108:167–181PubMedGoogle Scholar
  23. Lau A, Tymianski M (2010) Glutamate receptors, neurotoxicity and neurodegeneration. Pflugers Arch 460:525–542CrossRefPubMedGoogle Scholar
  24. Liu Z, Niu W, Yang X, Wang Y (2013) Effects of combined acupuncture and eugenol on learning-memory ability and antioxidation system of hippocampus in Alzheimer disease rats via olfactory system stimulation. J Tradit Chin Med 33:399–402CrossRefPubMedGoogle Scholar
  25. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275Google Scholar
  26. Morris R (1984) Developments of a water-maze procedure for studying spatial learning in the rat. J Neurosci Methods 11:47–60CrossRefPubMedGoogle Scholar
  27. Mount C, Downton C (2006) Alzheimer disease: progress or profit? Nat Med 12:780–784CrossRefPubMedGoogle Scholar
  28. Mouri A, Noda Y, Hara H, Mizoguchi H, Tabira T, Nabeshima T (2007) Oral vaccination with a viral vector containing Abeta cDNA attenuates age-related Abeta accumulation and memory deficits without causing inflammation in a mouse Alzheimer model. FASEB J 21:2135–2148CrossRefPubMedGoogle Scholar
  29. Nade VS, Kawale LA, Valte KD, Shendye NV (2015) Cognitive enhancing effect of angiotensin-converting enzyme inhibitors and angiotensin receptor blockers on learning and memory. Indian J Pharmacol 47:263–269CrossRefPubMedPubMedCentralGoogle Scholar
  30. Nangle MR, Gibson TM, Cotter MA, Cameron NE (2006) Effects of eugenol on nerve and vascular dysfunction in streptozotocin-diabetic rats. Planta Med 72:494–500CrossRefPubMedGoogle Scholar
  31. Nathan PJ, Watson J, Lund J, Davies CH, Peters G, Dodds CM, Swirski B, Lawrence P, Bentley GD, O’Neill BV, Robertson J, Watson S, Jones GA, Maruff P, Croft RJ, Laruelle M, Bullmore ET (2013) The potent M1 receptor allosteric agonist GSK1034702 improves episodic memory in humans in the nicotine abstinence model of cognitive dysfunction. Int J Neuropsychopharmacol 16:721–731CrossRefPubMedGoogle Scholar
  32. National Research Council (US) Committee for the Update of the Guide for the Care and Use of Laboratory Animals (2011) Guide for the care and use of laboratory animals, 8th edn. National Academies Press (US), Washington, DCGoogle Scholar
  33. Ohkawa H, Ohishi N, Yagi K (1979) Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Anal Biochem 95:351–358CrossRefGoogle Scholar
  34. Palkovits M, Brownstein MJ (1988) Maps and guide to micro dissection of the rat brain. Elsevier, New York, NY, USAGoogle Scholar
  35. Pandareesh MD, Anand T, Khanum F (2016) Cognition enhancing and neuromodulatory propensity of Bacopa monnieri extract against scopolamine induced cognitive impairments in rat hippocampus. Neurochem Res 41:985–999CrossRefPubMedGoogle Scholar
  36. Pedersen PL, Greenawalt JW, Reynafarje B, Hullihen J, Decker GL, Soper JW et al (1978) Preparation and characterization of mitochondria and submitochondrial particles of rat liver and liver-derived tissues. Methods Cell Biol 20:411–481CrossRefPubMedGoogle Scholar
  37. Portelius E, Zetterberg H, Andreasson U, Brinkmalm G, Andreasen N, Wallin A et al (2006) An Alzheimer’s disease-specific beta-amyloid fragment signature in cerebrospinal fluid. Neurosci Lett 419:215–219CrossRefGoogle Scholar
  38. Schliebs R, Arendt T (2006) The significance of the cholinergic system in the brain during aging and in Alzheimer’s disease. J Neural Transm (Vienna) 113:1625–1644CrossRefGoogle Scholar
  39. Tota S, Nath C, Najmi AK, Shukla R, Hanif K (2012) Inhibition of central angiotensin converting enzyme ameliorates scopolamine induced memory impairment in mice: role of cholinergic neurotransmission, cerebral blood flow and brain energy metabolism. Behav Brain Res 232:66–76CrossRefPubMedGoogle Scholar
  40. Varel VH, Miller DN (2004) Plant oils thymol and eugenol affect cattle and swine waste emissions differently. Water Sci Technol 50:207–213CrossRefPubMedGoogle Scholar
  41. Wang H, Joseph JA (1999) Quantifying cellular oxidative stress by dichlorofluorescein assay using microplate reader. Free Rad Biol Med 27:612–616CrossRefPubMedGoogle Scholar
  42. Wu CL, Hwang CS, Chen SD, Yin JH, Yang DI (2010) Neuroprotective mechanisms of brain-derived neurotrophic factor against 3-nitropropionic acid toxicity: therapeutic implications for Huntington’s disease. Ann N Y Acad Sci 1201:8–12CrossRefPubMedGoogle Scholar
  43. Yamamura HI, Snyder SH (1974) Muscarinic cholinergic binding in rat brain. Natl Acad Sci U S A 71:1725–1729CrossRefGoogle Scholar
  44. Yankner BA (1996) Mechanisms of neuronal degeneration in Alzheimer’s disease. Neuron 16:921–932CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Division of PharmacologyInstitute of Pharmaceutical Research, GLA UniversityMathuraIndia

Personalised recommendations