GENDER-SPECIFIC BRAIN REGIONAL VARIATION OF NEURONS, ENDOGENOUS ESTROGEN, NEUROINFLAMMATION AND GLIAL CELLS DURING ROTENONE-INDUCED MOUSE MODEL OF PARKINSON’S DISEASE S. MITRA, a N. CHAKRABARTI, b S. S. DUTTA, a S. RAY, a P. BHATTACHARYA, a P. SINHA b AND A. BHATTACHARYYA a * a Immunology Lab, Department of Zoology, University of Calcutta, 35, Ballygunge Circular Road, Kolkata 700019, India b Department of Physiology, University of Calcutta, 92, Acharya Prafulla Chandra Road, Kolkata 700009, India Abstract—Rotenone (RT) produces reactive oxygen species (ROS) by inhibiting the mitochondrial electron transport chain; causing dopaminergic (DA) cell death in the substan- tia nigra (SN) and simulates other models of induced Parkinson’s disease (PD). There is a sincere dearth of knowledge regarding the status of glial cells, neuroprotec- tive estrogen and the status of neuroinflammatory TNF-a in the different brain regions in either sex during healthy, as well as during PD conditions. In the present study of RT-induced mouse model of PD, we have selected the fron- tal cortex (FC), hippocampus (HC) and SN from either sex of Swiss albino mice as these are the major regions involved during PD pathogenesis. During non pathogenic condi- tions, the ROS-scavenging enzyme activity varied among the brain regions and also in between genders. The number of DOPA decarboxylase-positive cells, astrocytes and microglia was similar in the respective regions of the brain in both the sexes. The level of proinflammatory cytokine TNF-a was same in the respective FC and HC in either sex except that of SN. The expression level of estrogen and its receptors varied among the three brain regions. During RT treatment, ROS-scavenging enzyme activities increased, DOPA decarboxylase-positive neurons and fibers in DA as well as in norepinephrinergic (NE) systems become degen- erated, number of astrocytes decreased and microglial cells increased in those specific brain regions in either of the sex- es except in the SN region of males where astrocyte number remained unaltered and microglial cell percentage decreased. TNF-a increased in the FC and SN but remained unaltered in the HC of both sexes. Estradiol level decreased in the HC and SN but the level unevenly varied in the FC. Similarly, the estrogen bound and nuclear-cytosolic recep- tor a and b also varied differentially among the brain regions of the two sexes. Therefore our present study depicts that there exists a clear variation of neuronal and astroglial cell population, estrogen and its receptor levels in different brain regions of both the sexes during control and RT-treated pathogenic condition and these variations have major implication in PD pathogenesis and progression. Ó 2015 IBRO. Published by Elsevier Ltd. All rights reserved. Key words: rotenone, estrogen, frontal cortex, hippocampus, substantia nigra, glial cells. INTRODUCTION Parkinson’s disease (PD) is a multifactorial neurodegenerative disease caused by genetic and environmental factors. Of the several efforts for better understanding of the PD etiology, studies with animal model and human subject indicate that environmental toxins including rotenone (RT, a pesticide) might cause PD and the behavioral changes associated with the disease (Spivey, 2011; Tanner et al., 2011; Blesa et al., 2012). RT inhibits mitochondrial complex I and produces oxidative stress leading to dopaminergic (DA) cell death and also noradrenergic, serotonergic, cholinergic cell death in the brain, and reproduces the animal model of PD (Sherer et al., 2003; Yadava and Nicholls, 2007). DA neurons are uniquely vulnerable to reactive oxygen species (ROS) because of its reduced antioxidant capa- city, increased accumulations of iron, and the high con- centration of dopamine (Jenner, 1998; Greenamyre et al., 1999). Studies have indicated that ROS play key roles in cellular functions by altering several enzyme activities (Klann and Thiels, 1999). However, it is still debatable whether RT-induced ROS (Klintworth et al., 2009), or release of superoxides by activated microglia (Facchinetti et al., 1998) leads to neuronal cell death. The molecular anatomy of the different regions of the brain varies considerably when compared to each other during healthy condition. A recent study has demonstrated that there is a differential pattern of glial cells and DA neurotoxicity in the mid-brain and striatum http://dx.doi.org/10.1016/j.neuroscience.2014.12.052 0306-4522/Ó 2015 IBRO. Published by Elsevier Ltd. All rights reserved. * Corresponding author. Tel: +91-33-24615445x286; fax: +91-33- 24614849. E-mail address: arindam19@yahoo.com (A. Bhattacharyya). Abbreviations: BSA, bovine serum albumin; CDNB, 1 chloro-2,4- dinitrobenzene; DA, dopaminergic; ER-a, estrogen receptor-alpha; ER-b, estrogen receptor-beta; FC, frontal cortex; GFAP, glial fibrillary acidic protein; GST, glutathione S-transferase; HC, hippocampus; HRP, horseradish peroxidase; NBT/BCIP, nitroblue tetrazolium chloride/5- bromo-4-chloro-3-indolyl-phosphate; NE, norepinephrinergic; PBS, phosphate-buffered saline; PD, Parkinson’s disease; PMSF, phenylmethanesulfonylfluoride; ROS, reactive oxygen species; RT, rotenone; SC, subcutaneous; SN, substantia nigra; SOD, superoxide dismutase; TNF-a, tumor necrosis factor-alpha. Neuroscience 292 (2015) 46–70 46