Skip to main content
SpringerLink
Log in

Apoptosis of Purkinje and Granular Cells of the Cerebellum Following Chronic Ethanol Intake

  • Original Paper
  • Published:
The Cerebellum Aims and scope Submit manuscript

Abstract

Ethanol alters motricity, learning, cognition, and cellular metabolism in the cerebellum. We evaluated the effect of ethanol on apoptosis in Golgi, Purkinje, and granule cells of the cerebellum in adult rats. There were two groups of 20 rats: a control group that did not consume ethanol and an experimental group of UChA rats that consumed ethanol at 10 % (<2 g ethanol/kg body weight/day). At 120 days old, rats were anesthetized and decapitated, and their cerebella were collected and fixed. Cerebellar sections were subjected to immunohistochemistry for terminal deoxynucleotide transferase dUTP nick end labeling (TUNEL), caspase-3, X-linked inhibitor of apoptosis protein (XIAP), and insulin-like growth factor 1-receptor (IGF-1R); real-time PCR (RT-PCR) to determine caspase-3, XIAP, and IGF-1R gene expression; and transmission electron microscopy (TEM). We identified fragmentation of DNA and an increase in caspase-3 protein and XIAP in Purkinje cells, whereas granule cells exhibited increased caspase-3 and XIAP. IGF-1R expression was unchanged. There was no significant difference in gene expression of caspase-3, XIAP, and IGF-1R. There were an increase in lipid droplets, a reduction in the cellular cytoplasm in electron-dense nuclei, and changes in the myelin sheath in the cerebellar cortex. In conclusion, our data demonstrated that ethanol induced apoptosis in the Purkinje and granule cells of the cerebellum of adult UChA rats.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

References

  1. Edwards G, Gross MM. Alcohol dependence: provisional description of a clinical syndrome. BMJ. 1976;1:1058–61.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  2. World Health Organization. Global status report on alcohol and health. Geneva: WHO Press; 2011. p. 1–85.

    Google Scholar 

  3. Alfonso-Loeches S, Guerri C. Molecular and behavioral aspects of the actions of alcohol on the adult and developing brain. Crit Rev Clin Lab Sci. 2011;48:19–47.

    Article  CAS  PubMed  Google Scholar 

  4. De Smet HJ, Parquier PF, De Deyn PP, Marie P. The cerebellum and neurocognition: a review of clinical and neuroimaging studies. AJCN. 2010;4:1.

    Google Scholar 

  5. Maier SE, West JR. Regional differences in cell loss associate with binge-like alcohol exposure during the first two trimesters equivalent in the rat. Alcohol. 2001;23:49–57.

    Article  CAS  PubMed  Google Scholar 

  6. Sakata-Haga H, Sawada K, Hisano S, Fukui Y. Abnormalities of cerebellar foliation in rats prenatally exposed to ethanol. Acta Neuropathol. 2001;102(1):36–40.

    CAS  PubMed  Google Scholar 

  7. Apfel MI, Esberard CA, Rodrigues FK, Bahamad Jr FM, Sillero RO. Stereological study of the cerebellar Purkinje cells submitted to alcoholic intoxication in Wistar rats. Arq Neuropsiquiatr. 2002;60:258–63.

    Article  PubMed  Google Scholar 

  8. Ito M. The cerebellum and neural control. New York: Raven; 1984. p. 121–30.

    Google Scholar 

  9. Huang JJ, Yen CT, Tsao HW, Tsai ML, Huang C. Neuronal oscillations in Golgi cells and purkinje cells are accompanied by decreases in Shannon information entropy. Cerebellum. 2013;13:523–6.

    Google Scholar 

  10. Jaatinen P, Rintala J. Mechanisms of ethanol-induced degeneration in the developing, mature, and aging cerebellum. Cerebellum. 2008;7:332–47.

    Article  CAS  PubMed  Google Scholar 

  11. Luo J. Mechanisms of ethanol-induced death of cerebellar granule cells. Cerebellum. 2012;11:145–54.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  12. Sarna JR, Hawkes R. Patterned Purkinje cell death in the cerebellum. Prog Neurobiol. 2003;70:473–507.

    Article  CAS  PubMed  Google Scholar 

  13. Young C, Klocke BJ, Tenkova T, Choi J, Labruyere J, Qin Y-Q, et al. Ethanol-induced neuronal apoptosis in vivo requires BAX in the developing mouse brain. Cell Death Differ. 2003;10:1148–55.

    Article  CAS  PubMed  Google Scholar 

  14. Olney JW, Tenkova T, Dikranian K, Labruyere J, Qin YQ, Ikonomidou C. Ethanol-induced apoptotic neurodegeneration in the developing C57BL/6 mouse brain. Dev Brain Res. 2002;133:115–26.

    Article  CAS  Google Scholar 

  15. De La Monte SM, Longato L, Tong M, DeNucci S, Wands JR. The liver-brain axis of alcohol-mediated neurodegeneration: role of toxic lipids. Int J Environ Res Public Health. 2009;6:2055–75.

    Article  PubMed Central  PubMed  Google Scholar 

  16. Grivicich I, Regner A, Da Rocha AB. Morte celular por apoptose apoptosis: programmed cell death. Rev Bras Cancerol. 2007;53:335–43.

    Google Scholar 

  17. Quintanilla ME, Israel Y, Sapag A, Tampier L. The UChA and UChB rat lines: metabolic and genetic differences influencing ethanol intake. Addict Biol. 2006;11:310–23.

    Article  CAS  PubMed  Google Scholar 

  18. Chuffa LG, Amorim JP, Teixeira GR, Mendes LO, Fioruci BA, Pinheiro PF, et al. Long-term exogenous melatonin treatment modulates overall feed efficiency and protects ovarian tissue against injuries caused by ethanol-induced oxidative stress in adult UChB rats. Alcohol Clin Exp Res. 2011;35:1498–508.

    CAS  PubMed  Google Scholar 

  19. Chuffa LG, Seiva FR, Fávaro WJ, Amorim JP, Teixeira GR, Mendes LO, et al. Melatonin and ethanol intake exert opposite effects on circulating estradiol and progesterone and differentially regulate sex steroid receptors in the ovaries, oviducts, and uteri of adult rats. Reprod Toxicol. 2013;39:40–9.

    Article  CAS  PubMed  Google Scholar 

  20. Mardones J, Segovia-Riquelme N. Thirty-two years of selection of rats by ethanol preference: UChA and UChB strains. Neurobehav Toxicol Teratol. 1983;5:171–8.

    CAS  PubMed  Google Scholar 

  21. Ewenczyk A, Ziplow J, Tong M, De La Monte SM. Sustained impairments in brain insulin/IGF signaling in adolescent rats subjected to binge alcohol exposures during development. J Clin Exp Pathol. 2012;2:106. doi:10.4172/2161-0681.1000106.

    Article  Google Scholar 

  22. Maycotte P, Blancas S, Morán J. Role of inhibitor of apoptosis proteins and Smac/DIABLO in staurosporine-induced cerebellar granule neurons death. Neurochem Res. 2008;33(8):1534–40.

    Article  CAS  PubMed  Google Scholar 

  23. Oliveira SA, Fontanelli BAF, Stefanini MA, Chuffa LGA, Teixeira GR, Lizarte FSN, et al. Interaction of maternal separation on the UCh rat cerebellum. Microsc Res Tech. 2014;77:44–51.

    Article  CAS  PubMed  Google Scholar 

  24. Lewandowska E, Stepien T, Wierzba-Bobrowicz T, Felczak P, Szpak GM, Pasennik E. Alcohol-induced changes in the developing cerebellum. Ultrastructural and quantitative analysis of neurons in the cerebellar cortex. Folia Neuropathol. 2012;50(4):397–406.

    Article  PubMed  Google Scholar 

  25. Dlugos CA, Pentney RJ. Effects of chronic ethanol consumption on SER of Purkinje neurons in old F344 rats. Alcohol. 2000;20:125–32.

    Article  CAS  PubMed  Google Scholar 

  26. Pentney RJ. Measurements of dendritic path lengths provide evidence that ethanol-induced lengthening of terminal dendritic segments may result from dendritic regression. Alcohol Alcohol. 1995;30:87–96.

    CAS  PubMed  Google Scholar 

  27. Pentney RJ, Dlugos CA. Cerebellar Purkinje neurons with altered terminal dendritic segments are present in all lobules of the cerebellar vermis of ageing, ethanol-treated F344 rats. Alcohol Alcohol. 2000;35:35–43.

    Article  CAS  PubMed  Google Scholar 

  28. Pentney RJ, Quackenbush LJ. Dendritic hypertrophy in Purkinje neurons of old Fischer 344 rats after long-term ethanol treatment. Alcohol Clin Exp Res. 1990;14:878–86.

    Article  CAS  PubMed  Google Scholar 

  29. Pentney RJ, Quackenbush LJ. Effects of long durations of ethanol treatment during aging on dendritic plasticity in Fischer 344 rats. Alcohol Clin Exp Res. 1991;15:1024–30.

    Article  CAS  PubMed  Google Scholar 

  30. Hitomi J, Katayama T, Taniguchi M, Honda A, Imaizumi K, Tohyama M. Apoptosis induced by endoplasmic reticulum stress depends on activation of caspase-3 via caspase-12. Neurosci Lett. 2003;357:127–30.

    Article  Google Scholar 

  31. Shi Y. Mechanical aspects of apoptosome assembly. Curr Opin Cell Biol. 2006;18:677–84.

    Article  CAS  PubMed  Google Scholar 

  32. Taranukhin AG, Taranukhina EY, Saransaari P, Podkletnova IM, Pelto-Huikko M, Oja SS. Neuroprotection by taurine in ethanol-induced apoptosis in the developing cerebellum. J Biomed Sci 2010; 17. doi: 10.1186/1423-0127-17-S1-S12.

  33. Lee JH, Tajuddin NF, Druse AJ. Effects of ethanol and ipsapirone on the expression of genes encoding anti-apoptotic proteins and an antioxidant enzyme in ethanol-treated neurons. Brain Res. 2009;1249:54–60.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  34. Guo R, Zhong L, Ren J. Overexpression of aldehyde dehydrogenase-2 attenuates chronic alcohol exposure-induced apoptosis, change in Akt and Pim signalling in liver. Clin Exp Pharmacol Physiol. 2009;36(5–6):463–8.

    Article  CAS  PubMed  Google Scholar 

  35. Antonio AM, Gillespie RA, Druse-Manteuffel MJ. Effects of lipoic acid on antiapoptotic genes in control and ethanol-treated fetal rhombencephalic neurons. Brain Res. 2011;1383:13–21.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  36. Marks JL, Porte Jr D, Baskin DG. Localization of type I insulin-like growth factor receptor messenger RNA in the adult rat brain by in situ hybridization. Mol Endocrinol. 1991;5(8):1158–68.

    Article  CAS  PubMed  Google Scholar 

  37. Bondy CA. Transient IGF-I gene expression during the maturation of functionally related central projection neurons. J Neurosci. 1991;11:3442–55.

    CAS  PubMed  Google Scholar 

  38. Liu J, Lewohl JM, Harris RA, Iyer VR, Dodd PR, Randall PK, et al. Patterns of gene expression in the frontal cortex discriminate alcoholic from nonalcoholic individuals. Neuropsychopharmacology. 2006;31:1574–82.

    Article  CAS  PubMed  Google Scholar 

  39. Pignataro L, Varodayan FP, Tannenholz LE, Harrison NL. The regulation of neuronal gene expression by alcohol. Pharmacol Ther. 2009;124:324–35.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  40. Cassini C, Linden R. Prenatal exposure to ethanol: toxicity, biomarkers and detection methods. Rev Psiq Clin. 2011;38:116–21.

    Article  Google Scholar 

  41. Dlugos CA, Pentney RJ. Morphometric evidence that the total number of synapses on Purkinje neurons of old F344 rats is reduced after long-term ethanol treatment and restored to control levels after recovery. Alcohol Alcohol. 1997;32:161–72.

    Article  CAS  PubMed  Google Scholar 

  42. Botta P, Souza FMS, Sangrey T, Schutter E, Valenzuela F. Alcohol excites cerebellar Golgi cells by inhibiting the Na+/K+ ATPase. Neuropsychopharmacology. 2010;35:1984–96.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  43. Botta P, Souza FMS, Sangrey T, Schutter E, Valenzuela F. Excitation of rat cerebellar Golgi cells by ethanol: further characterization of the mechanism alcohol. Clin Exp Res. 2012;36(4):616–24.

    Article  CAS  Google Scholar 

  44. Botta P, Zucca A, Valenzuela F. Acute ethanol exposure inhibits silencing of cerebellar Golgi cell firing induced by granule cell axon input. Front Integr Neurosci. 2014;8:10. doi:10.3389/fnint.2014.00010.

    Article  PubMed Central  PubMed  Google Scholar 

  45. Wang GH, Jiang ZL, Li YC, Li X, Shi H, Gao YQ, et al. Free-radical scavenger edaravone treatment confers neuroprotection against traumatic brain injury in rats. J Neurotrauma. 2011;28(10):2123–34.

    Article  PubMed Central  PubMed  Google Scholar 

  46. Oomen CA, Farkas E, Roman V, Van Der Beek EM, Luiten PGM. Resveratrol preserves cerebrovascular density and cognitive function in aging mice. Front Aging Neurosci. 2009;1:4. doi:10.3389/neuro.24.004.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  47. Shirpoor A, Minassian S, Salami S, Khadem-Ansari MH, Ghaderi-Pakdel F, Yeghiazaryan M. Vitamin E protects developing rat hippocampus and cerebellum against ethanol-induced oxidative stress and apoptosis. Food Chem. 2009;113:115–20.

    Article  CAS  Google Scholar 

Download references

Acknowledgments

We would like to thank Mr. Wanderley Thiago da Silva for animal care,  Mr. Gelson Rodrigues for technical support and the funding agency (FAPESP 2011/50466-0).

Conflict of Interest

The authors declare no conflicts of interest.

Author information

Authors and Affiliations

  1. Graduate Program in General and Applied Biology, Institute of Bioscience, Univ. Estadual Paulista (UNESP), Botucatu, SP, 18618-970, Brazil

    Suelen A. Oliveira

  2. Department of Anatomy, Institute of Biosciences, Univ. Estadual Paulista (UNESP), P.O. Box 510, Rubião Júnior, s/n, Botucatu, SP, 18618-970, Brazil

    Luiz Gustavo A. Chuffa, Beatriz Aparecida Fioruci-Fontanelli & Francisco Eduardo Martinez

  3. Department of Surgery and Anatomy, Faculty of Medicine, Univ. São Paulo (USP), Ribeirão Preto, SP, 14049-900, Brazil

    Fermino Sanches Lizarte Neto, Paulo Cezar Novais & Luiz Fernando Tirapelli

  4. Department of Physiology, Universidade Federal de São Carlos, São Carlos, SP, 13565-905, Brazil

    Jorge Camargo Oishi

  5. Department of Morphology and Pathology, Universidade Federal de São Carlos, São Carlos, SP, 13565-905, Brazil

    Luiz Fernando Takase, Maira Aparecida Stefanini & Marcelo Martinez

Authors
  1. Suelen A. Oliveira
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Luiz Gustavo A. Chuffa
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Beatriz Aparecida Fioruci-Fontanelli
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Fermino Sanches Lizarte Neto
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Paulo Cezar Novais
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Luiz Fernando Tirapelli
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Jorge Camargo Oishi
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Luiz Fernando Takase
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Maira Aparecida Stefanini
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Marcelo Martinez
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Francisco Eduardo Martinez
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Francisco Eduardo Martinez.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Oliveira, S.A., Chuffa, L.G.A., Fioruci-Fontanelli, B.A. et al. Apoptosis of Purkinje and Granular Cells of the Cerebellum Following Chronic Ethanol Intake. Cerebellum 13, 728–738 (2014). https://doi.org/10.1007/s12311-014-0591-2

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12311-014-0591-2

Keywords

Search

Navigation