Pioglitazone Blocks Ethanol Induction of Microglial Activation and Immune Responses in the Hippocampus, Cerebellum, and Cerebral Cortex in a Mouse Model of Fetal Alcohol Spectrum Disorders

Abstract

Background Fetal alcohol spectrum disorders (FASD) result from fetal exposure to alcohol and are the leading cause of mental retardation in the United States. There is currently no effective treatment that targets the causes of these disorders. Thus, novel therapies are critically needed to limit the neurodevelopmental and neurodegenerative pathologies associated with FASD. Methods A neonatal mouse FASD model was used to examine the role of the neuroimmune system in ethanol-induced neuropathology. Neonatal C57BL/6 mice were treated with ethanol, with or without pioglitazone, on postnatal days 4 through 9 and tissue was harvested one day post-treatment. Pioglitazone is a peroxisome proliferator-activated receptor (PPAR)-γ agonist that exhibits anti-inflammatory activity and is neuroprotective. We compared the effects of ethanol with or without pioglitazone on cytokine and chemokine expression and microglial morphology in the hippocampus, cerebellum, and cerebral cortex. Results In ethanol-treated animals compared to controls, cytokines IL-1β and TNF-α mRNA levels were increased significantly in the hippocampus, cerebellum, and cerebral cortex. Chemokine CCL2 mRNA was increased significantly in the hippocampus and cerebellum. Pioglitazone effectively blocked the ethanol-induced increase in the cytokines and chemokine in all tissues to the level expressed in handled-only and vehicle-treated control animals. Ethanol also produced a change in microglial morphology in all brain regions that was indicative of microglial activation, and pioglitazone blocked this ethanol-induced morphological change. Conclusions These studies indicate that ethanol activates microglia to a pro-inflammatory stage and also increases the expression of neuroinflammatory cytokines and chemokines in diverse regions of the developing brain. Further, the anti-inflammatory and neuroprotective PPAR-γ agonist pioglitazone blocked these effects. It is proposed that microglial activation and inflammatory molecules expressed as a result of ethanol treatment during brain development contribute to the sequelae associated with FASD. Thus, pioglitazone, and anti-inflammatory pharmaceuticals more broadly, have potential as novel therapeutics for FASD.

Differential Effects of Pioglitazone in the Hippocampal CA1 Region Following Transient Forebrain Ischemia in Low- and High-Fat Diet-Fed Gerbils

“…The administration of PGZ reduces ischemia-induced morphological changes in microglia of LFD-fed group, not in the HFD-fed group. This result was supported by a previous study that PGZ blocks ethanol-induced morphological changes in microglia in all brain regions that was indicative of microglial activation [48]. In this study, we also observed TNF-a and IL-1b levels in the hippocampus at 6 h after ischemia/reperfusion because these levels are peaked in the hippocampus at this time point [31].…”

Neonatal binge alcohol exposure increases microglial activation in the developing rat hippocampus

“…As an indirect estimate of microglia morphology and activation state, total cell area or “territory” (Drew et al, 2015) was measured for the cell layers of CA1, CA3, and DG using NeuroLucida software (MBF BioScience, Williston, VT). For each section, the area encompassed by an individual microglia was measured by tracing a contour from the tip of each microglial process to the next.…”

“…Recent work has suggested alcohol-induced neuroinflammation, as measured by increased activation of microglia and levels of associated cytokines, as a potential secondary source of damage in various models of alcohol exposure, both during development and in adulthood (McClain et al, 2011; Saito et al, 2010; Kane et al, 2011; Tiwari & Chopra, 2011; Marshall et al, 2013; Drew et al, 2015; Topper et al, 2015). Alcohol exposure during the third trimester-equivalent induces waves of apoptosis in the hippocampus, possibly triggering microglial migration to the damaged tissue and activation of the resident microglia to phagocytose dying cells and debris (Miller, 1998; Ikonomidou et al, 2000; Smith et al, 2015).…”

Function of Thymosin Beta-4 in Ethanol-Induced Microglial Activation

“…We used a synthetic copy of the naturally-occurring 43-amino acid Tβ4 peptide (RegeneRx, Bethesda, MD, USA). The animal experiment was performed as described previously [17,18]. In brief, 4 g/kg/d ethanol or control was administered daily by intraesophageal gavage on postnatal days 4 through 9.…”

“…Animals were anesthetized with isoflurane and perfused transcardially with a brief heparinized PBS flush followed by phosphate-buffered (PB) 4% paraformaldehyde fixative [18]. Double immunofluorescence was performed as previously described [12].…”