Antioxidant and Anti-Apoptotic Properties of Allopregnanolone in Protecting Dopaminergic Neurons from 6-OHDA-Induced Injury

Document Type : Original Article

Authors

1 Department of Basic Sciences, Faculty of Veterinary Medicine, Shahid Bahonar University of Kerman, kerman, Iran

2 Department of Biology, Faculty of Basic Sciences, Shahid Bahonar University of Kerman. Kerman, Iran

3 Department of Biotechnology, Faculty of Biological Science and Technology, University of Isfahan, Isfahan, Iran

Abstract

Dopaminergic neurodegeneration is associated with oxidative stress, mitochondrial dysfunction, neuroinflammation, and cell death. Allopregnanolone, a multifunctional neurosteroid, is involved in various biological processes within the body. It has been demonstrated that allopregnanolone exhibits protective effects in neurodegenerative conditions. However, its cellular mechanisms in dopaminergic neurons remain incompletely understood. Cell toxicity was induced using 6-hydroxydopamine (6-OHDA), and Cell survival was assessed through the MTT assay. Intracellular reactive oxygen species (ROS) and mitochondrial membrane potential were evaluated using fluorescence probes. Additionally, immunoblotting was employed to measure the levels of apoptosis biomarkers in the cells. Treatment with 6-OHDA significantly decreased cell survival rate and exacerbated the loss of mitochondrial membrane potential. Moreover, there was a notable increase in intracellular ROS levels, the Bax/Bcl-2 ratio, caspase-3, and cytochrome c activity in 6-OHDA-treated cells. Pretreatment with allopregnanolone (250 µM) significantly mitigated these effects in cells exposed to 6-OHDA. Furthermore, the blockade of GABAA receptors by bicuculline significantly reduced the protective effect of allopregnanolone. The data indicate that allopregnanolone's protective effects are attributed to its antioxidant and anti-apoptotic properties, suggesting its therapeutic potential for preventing dopaminergic damage.

Keywords

References

  1. Jankovic J. Parkinson’s disease: clinical features and diagnosis. J Neurol Neurosurg psychiatry. 2008; 79 (4):368–76.
  2. Mercado G, Kaeufer C, Richter F, Peelaerts W. Infections in the etiology of Parkinson’s disease and synucleinopathies: A renewed perspective, mechanistic insights, and therapeutic implications. J Park Dis. 2024; 14 (7):1301–29.
  3. Delamarre A, Meissner WG. Epidemiology, environmental risk factors and genetics of Parkinson’s disease. Presse Med. 2017; 46 (2):175–81.
  4. Tolleson CM, Fang JY. Advances in the mechanisms of Parkinson’s disease. Discov Med. 2013; 15 (80):61–6.
  5. Hickey P, Stacy M. Available and emerging treatments for Parkinson’s disease: a review. Drug Des Devel Ther. 2011; 241–54.
  6. Sarkar S, Raymick J, Imam S. Neuroprotective and therapeutic strategies against Parkinson’s disease: recent perspectives. Int J Mol Sci. 2016; 17 (6):904.
  7. Esmaeili-Mahani S, Vazifekhah S, Pasban-Aliabadi H, Abbasnejad M, Sheibani V. Protective effect of orexin- A on 6-hydroxydopamine-induced neurotoxicity in SH-SY5Y human dopaminergic neuroblastoma cells. Neurochem Int. 2013; 63 (8):719–25.
  8. Lu X, Kim-Han JS, Harmon S, Sakiyama-Elbert SE, O’Malley KL. The Parkinsonian mimetic, 6-OHDA, impairs axonal transport in dopaminergic axons. Mol Neurodegener. 2014; 9:1–11.
  9. Han DY, Di XJ, Fu YL, Mu TW. Combining valosin-containing protein (VCP) inhibition and suberanilohydroxamic acid (SAHA) treatment additively enhances the folding, trafficking, and function of epilepsy-associated γ-aminobutyric acid, type A (GABAA) receptors. J Biol Chem. 2015; 290 (1):325–37.
  10. Alberio T, Bondi H, Colombo F, Alloggio I, Pieroni L, Urbani A, et al. Mitochondrial proteomics investigation of a cellular model of impaired dopamine homeostasis, an early step in Parkinson’s disease pathogenesis. Mol Biosyst. 2014; 10 (6):1332–44.
  11. Bassani TB, Bartolomeo CS, Oliveira RB, Ureshino RP. Progestogen-mediated neuroprotection in central nervous system disorders. Neuroendocrinology. 2023; 113 (1):14–35.
  12. Nezhadi A, Sheibani V, Esmaeilpour K, Shabani M, Esmaeili-Mahani S. Neurosteroid allopregnanolone attenuates cognitive dysfunctions in 6-OHDA-induced rat model of Parkinson’s disease. Behav Brain Res. 2016; 305: 258–64.
  13. Zhi-chi WT tong C. Allopregnanolone restores dopaminergic neurons and motor performance in Parkinson’s disease mice and its molecular mechanisms. Acta Anat Sin. 2020; 51 (4):473.
  14. Mehdizadeh A, Aali BS, Hajializadeh Z, Torkzadeh-Mahani S, Esmaeili-Mahani S. Neurosteroid dehydroepiandrosterone attenuates 6-hydroxydopamine-induced apoptosis in a cell model of Parkinson’s disease. Physiol Pharmacol. 2019; 23 (3):166–73.
  15. Patte-Mensah C, Kibaly C, Mensah-Nyagan AG. Substance P inhibits progesterone conversion to neuroactive metabolites in spinal sensory circuit: a potential component of nociception. Proc Natl Acad Sci. 2005; 102 (25): 9044–9.
  16. Mensah-Nyagan AG, Do-Rego JL, Beaujean D, Pelletier G, Vaudry H. Neurosteroids: expression of steroidogenic enzymes and regulation of steroid biosynthesis in the central nervous system. Pharmacol Rev. 1999; 51 (1):63–82.
  17. Bixo M, Andersson A, Winblad B, Purdy RH, Progesterone TB. 5alpha-pregnane-3, 20-dione and 3alpha-hydroxy-5alpha-pregnane-20-one in specific regions of the human female brain in different endocrine states, 1997; 764:451–5.
  18. Tong-Tong W, Xin YE, Wei B, Zhi-Chi C, Juan-Juan DU, Wei-Da FU, et al. Allopregnanolone protecting cell line SH-SY5Y against 6-hydroxydopamine induced lesion. Acta Anat Sin. 2021; 52(1):5.
  19. Diviccaro S, Cioffi L, Falvo E, Giatti S, Melcangi RC. Allopregnanolone: An overview on its synthesis and effects. J Neuroendocrinol. 2022; 34 (2):e12996.
  20. Sun C, Ou X, M Farley J, Stockmeier C, Bigler S, Diaz Brinton R, et al. Allopregnanolone increases the number of dopaminergic neurons in substantia nigra of a triple transgenic mouse model of Alzheimer’s disease. Curr Alzheimer Res. 2012; 9 (4):473–80.
  21. De Nicola AF, Meyer M, Garay L, Kruse MS, Schumacher M, Guennoun R, et al. Progesterone and allopregnanolone neuroprotective effects in the wobbler mouse model of amyotrophic lateral sclerosis. Cell Mol Neurobiol. 2022; 42 (1):23–40.
  22. Casas S, Giuliani F, Cremaschi F, Yunes R, Cabrera R. Neuromodulatory effect of progesterone on the dopaminergic, glutamatergic, and GABAergic activities in a male rat model of Parkinson’s disease. Neurol Res. 2013; 35 (7):719–25.
  23. Charalampopoulos I, Tsatsanis C, Dermitzaki E, Alexaki VI, Castanas E, Margioris AN, et al. Dehydroepiandrosterone and allopregnanolone protect sympathoadrenal medulla cells against apoptosis via antiapoptotic Bcl-2 proteins. Proc Natl Acad Sci. 2004; 101 (21):8209–14.
  24. Marx CE, Trost WT, Shampine LJ, Stevens RD, Hulette CM, Steffens DC, et al. The neurosteroid allopregnanolone is reduced in prefrontal cortex in Alzheimer’s disease. Biol Psychiatry. 2006; 60 (12):1287–94.
  25. Naylor JC, Kilts JD, Hulette CM, Steffens DC, Blazer DG, Ervin JF, et al. Allopregnanolone levels are reduced in temporal cortex in patients with Alzheimer’s disease compared to cognitively intact control subjects. Biochim Biophys Acta (BBA)-Molecular Cell Biol Lipids. 2010; 1801 (8):951–9.
  26. Di Michele F, Longone P, Romeo E, Lucchetti S, Brusa L, Pierantozzi M, et al. Decreased plasma and cerebrospinal fluid content of neuroactive steroids in Parkinson’s disease. Neurol Sci. 2003; 24:172–3.
  27. Majewska MD. Neurosteroids: endogenous bimodal modulators of the GABAA receptor mechanism of action and physiological significance. Prog Neurobiol. 1992; 38(4):379–94.
  28. Wang JM, Johnston PB, Ball BG, Brinton RD. The neurosteroid allopregnanolone promotes proliferation of rodent and human neural progenitor cells and regulates cell-cycle gene and protein expression. J Neurosci. 2005; 25(19):4706–18.
  29. Melcangi RC, Garcia-Segura LM, Mensah-Nyagan AG. Neuroactive steroids: state of the art and new perspectives. Cell Mol Life Sci. 2008; 65:777–97.
  30. Lejri I, Grimm A, Miesch M, Geoffroy P, Eckert A, Mensah-Nyagan AG. Allopregnanolone and its analog BR 297 rescue neuronal cells from oxidative stress-induced death through bioenergetic improvement. Biochim Biophys Acta (BBA)-Molecular Basis Dis. 2017; 1863 (3):631–42.
  31. Majewska MD, Harrison NL, Schwartz RD, Barker JL, Paul SM. Steroid hormone metabolites are barbiturate-like modulators of the GABA receptor. Science (80- ). 1986; 232 (4753):1004–7.
  32. Frye CA, Koonce CJ, Walf AA. Novel receptor targets for production and action of allopregnanolone in the central nervous system: a focus on pregnane xenobiotic receptor. Front Cell Neurosci. 2014; 8:106.
  33. Carver CM, Reddy DS. Neurosteroid interactions with synaptic and extrasynaptic GABA A receptors: regulation of subunit plasticity, phasic and tonic inhibition, and neuronal network excitability. Psychopharmacology (Berl). 2013; 230:151–88.
  34. Afrazi S, Esmaeili-Mahani S, Sheibani V, Abbasnejad M. Neurosteroid allopregnanolone attenuates high glucose-induced apoptosis and prevents experimental diabetic neuropathic pain: in vitro and in vivo studies. J Steroid Biochem Mol Biol. 2014; 139:98–103.
  35. Nezhadi A, Esmaeili-Mahani S, Sheibani V, Shabani M, Darvishzadeh F. Neurosteroid allopregnanolone attenuates motor disability and prevents the changes of neurexin 1 and postsynaptic density protein 95 expression in the striatum of 6-OHDA-induced rats’ model of Parkinson’s disease. Biomed Pharmacother. 2017; 88:1188–97.
  36. Qian X, Cao H, Ma Q, Wang Q, He W, Qin P, et al. Allopregnanolone attenuates Aβ25-35-induced neurotoxicity in PC12 cells by reducing oxidative stress. Int J Clin Exp Med. 2015; 8(8):13610.
  37. Latchoumycandane C, Anantharam V, Jin H, Kanthasamy A, Kanthasamy A. Dopaminergic neurotoxicant 6-OHDA induces oxidative damage through proteolytic activation of PKCδ in cell culture and animal models of Parkinson’s disease. Toxicol Appl Pharmacol. 2011; 256(3):314–23.
  38. Saito Y, Nishio K, Ogawa Y, Kinumi T, Yoshida Y, Masuo Y, et al. Molecular mechanisms of 6-hydroxydopamine-induced cytotoxicity in PC12 cells: involvement of hydrogen peroxide-dependent and-independent action. Free Radic Biol Med. 2007; 42(5):675–85.
  39. Wu CC, Bratton SB. Regulation of the intrinsic apoptosis pathway by reactive oxygen species. Antioxid Redox Signal. 2013; 19(6):546–58.
  40. Djebaili M, Guo Q, Pettus EH, Hoffman SW, Stein DG. The neurosteroids progesterone and allopregnanolone reduce cell death, gliosis, and functional deficits after traumatic brain injury in rats. J Neurotrauma. 2005; 22(1):106–18.
  41. Djebaili M, Hoffman SW, Stein DG. Allopregnanolone and progesterone decrease cell death and cognitive deficits after a contusion of the rat pre-frontal cortex. Neuroscience. 2004; 123(2):349–59.
  42. Wang M. Neurosteroids and GABA-A receptor function. Front Endocrinol (Lausanne). 2011; 2:44.
Veterinary and Comparative Biomedical Research
Volume 2, Issue 2
December 2025
Pages 50-59

Files

Share

How to cite

Statistics