Binge alcohol alters exercise-driven neuroplasticity

Highlights

• Binge alcohol exerts a prolonged influence on cortical microglia morphology.

• Exercise increases cortical microglia.

• Binge alcohol suppresses this exercise-driven increase in microglia.

Abstract

Exercise is increasingly being used as a treatment for alcohol use disorders (AUD), but the interactive effects of alcohol and exercise on the brain remain largely unexplored. Alcohol damages the brain, in part by altering glial functioning. In contrast, exercise promotes glial health and plasticity. In the present study, we investigated whether binge alcohol would attenuate the effects of subsequent exercise on glia. We focused on the medial prefrontal cortex (mPFC), an alcohol-vulnerable region that also undergoes neuroplastic changes in response to exercise. Adult female Long-Evans rats were gavaged with ethanol (25% w/v) every 8 h for 4 days. Control animals received an isocaloric, non-alcohol diet. After 7 days of abstinence, rats remained sedentary or exercised for 4 weeks. Immunofluorescence was then used to label microglia, astrocytes, and neurons in serial tissue sections through the mPFC. Confocal microscope images were processed using FARSIGHT, a computational image analysis toolkit capable of automated analysis of cell number and morphology. We found that exercise increased the number of microglia in the mPFC in control animals. Binged animals that exercised, however, had significantly fewer microglia. Furthermore, computational arbor analytics revealed that the binged animals (regardless of exercise) had microglia with thicker, shorter arbors and significantly less branching, suggestive of partial activation. We found no changes in the number or morphology of mPFC astrocytes. We conclude that binge alcohol exerts a prolonged effect on morphology of mPFC microglia and limits the capacity of exercise to increase their numbers.

Introduction

Binge drinking damages corticolimbic brain regions important for memory, decision-making and behavioral control (Crews and Boettiger, 2009, Duka et al., 2011), and recent studies indicate that it results in detectable brain dysfunction (Maurage et al., 2012, Campanella et al., 2013). It also decreases hippocampal neurogenesis (Nixon and Crews, 2002, Nixon and Crews, 2004, Nixon et al., 2008) and disrupts glial function (de la Monte and Kril, 2014). In contrast, exercise benefits neural health through a variety of mechanisms that include enhanced neurogenesis (van Praag et al., 1999), gliogenesis (Li et al., 2005, Mandyam et al., 2007), angiogenesis (Black et al., 1990, Swain et al., 2003, Rhyu et al., 2010) and trophic factor upregulation (Gomez-Pinilla et al., 2001, Vaynman and Gomez-Pinilla, 2005). Exercise therefore has the potential to heal the alcohol-damaged brain and indeed has been shown to ameliorate the consequences of developmental alcohol exposure (Thomas et al., 2008, Helfer et al., 2009). However, the interactive effects of alcohol and exercise on the brain remain largely unexplored.

Exercise is increasingly being used as an adjunctive treatment for alcohol use disorders (AUD). Several recent reviews of a growing number of clinical trials indicate that exercise is feasible in those with AUD, and is effective at enhancing their cardiovascular health as well as treating co-morbid mental health problems, such as anxiety and depression (Giesen et al., 2015, Stoutenberg et al., 2016). The effect of exercise training on drinking behaviors is much less clear and there is a compelling need for carefully controlled trials (Stoutenberg et al., 2016). A better understanding of the interactive effects of alcohol and exercise on the brain will inform activity-based treatment strategies for AUD.

We have shown that exercise reverses binge-induced hippocampal damage in female rats (Maynard and Leasure, 2013), substantiating the idea that exercise can counter the damaging effects of binge alcohol. It remains unknown, however, whether alcohol influences the brain benefits of subsequent exercise. In the present report, we examined banked tissue from our prior study (Maynard and Leasure, 2013) in order to determine whether binge alcohol impacted exercise-induced cellular plasticity in the female brain. We focused on the medial prefrontal cortex (mPFC), a region that is both vulnerable to alcohol (Sullivan et al., 2000, Kubota et al., 2001, Sullivan and Pfefferbaum, 2005) and responsive to exercise (Mandyam et al., 2007, Brockett et al., 2015). In addition, the mPFC has connections to the hippocampus (Warburton and Brown, 2010, Varela et al., 2014), which we and others have shown to be damaged by binge alcohol (Nixon and Crews, 2002, Nixon and Crews, 2004, Nixon et al., 2008, Maynard and Leasure, 2013).

As neurogenesis does not occur in the mPFC, we focused on glial plasticity. As immunocompetent cells of the brain, microglia are constantly surveying the neural parenchyma, ready to respond to changes in their environment by taking on several stages of activation (Streit, 2002, Nimmerjahn et al., 2005). Previous research indicates that a single binge episode is able to prime microglia to respond to subsequent neuroimmune challenges (McClain et al., 2011, Marshall et al., 2016). This has been found in male rats, however, sex differences have been found in microglia priming in the mPFC in response to stress (Bollinger et al., 2016). Therefore, investigation into the effects of binge alcohol on microglia in the female brain is warranted.

We hypothesized that exercise would less effectively drive glial plasticity (including number and morphology of astrocytes and microglia) in the mPFC of binged rats, compared to controls. However, we anticipated that after 5 weeks of abstinence (including 4 weeks of exercise), binge effects on exercise-driven plasticity in the mPFC would be subtle. We therefore used computational image analysis, which can detect small morphological differences, and which generates a rich collection of quantitative measurements (Bjornsson et al., 2008, Al-Kofahi et al., 2010, Narayanaswamy et al., 2011), to analyze both number and morphology of mPFC astrocytes and microglia. These computational arbor analytics revealed that microglia in the binged animals continued to display an altered morphology following 5 weeks of abstinence. Additionally, the combination of binge exposure and exercise resulted in a drastic decrease in the number of surviving microglia. Our findings suggest that binge alcohol exerts a prolonged effect on microglia, suggestive of microglial priming, and alters the typical microglial response to exercise in the mPFC.