Sudden cessation of fluoxetine before alcohol drinking reinstatement alters microglial morphology and TLR4/inflammatory neuroadaptation in the rat brain

Jesús Aranda,Juan Suárez,Pedro J Serrano-Castro,Francisco Alén,María Del Mar Fernández-Arjona,Patricia Rivera,Francisco Javier Pavón,Inés Smith-Fernández,Leticia Rubio,Fernando Rodríguez De Fonseca,Antonia Serrano

doi:10.1007/s00429-021-02321-9

Abstract

Preclinical studies on the effects of abrupt cessation of selective serotonin reuptake inhibitors (SSRIs), a medication often prescribed in alcohol use disorder (AUD) patients with depression, results in alcohol consumption escalation after resuming drinking. However, a potential neuroinflammatory component on this escalation remains unexplored despite the immunomodulatory role of serotonin. Here, we utilized a rat model of 14-daily administration of the SSRI fluoxetine (10 mg/kg/day) along alcohol self-administration deprivation to study the effects of fluoxetine cessation on neuroinflammation after resuming alcohol drinking. Microglial morphology and inflammatory gene expression were analyzed in prelimbic cortex, striatum, basolateral amygdala and dorsal hippocampus. Results indicated that alcohol drinking reinstatement increased microglial IBA1 immunoreactivity and altered morphometric features of activated microglia (fractal dimension, lacunarity, density, roughness, and cell area, perimeter and circularity). Despite alcohol reinstatement, fluoxetine cessation modified microglial morphology in a brain region-specific manner, resulting in hyper-ramified (spatial complexity of branching), reactive (lower heterogeneity and circularity)-like microglia. We also found that microglial cell area correlated with changes in mRNA expression of chemokines (Cx3cl1/fractalkine, Cxcl12/SDF1α, Ccl2/MCP1), cytokines (IL1β, IL6, IL10) and the innate immune toll-like receptor 4 (TLR4) in dorsal hippocampus. Specifically, TLR4 correlated with microglial spatial complexity assessed by fractal dimension in striatum, suggesting a role in process branching. These findings suggest that alcohol drinking reinstatement after fluoxetine treatment cessation disturbs microglial morphology and reactive phenotype associated with a TLR4/inflammatory response to alcohol in a brain region-specific manner, facts that might contribute to alcohol-induced damage through the promotion of escalation of alcohol drinking behavior.

Highlights

Alcohol is a psychoactive substance highly consumed in the general population worldwide (Organization for Economic Co-operation and Development 2020)
But not saccharine drinking, induced significant overall effects on the IBA-1 immunoreactivity, but not IBA-1 + cell number, in the prelimbic cortex (PrL) (F1,20 = 12.13, p = 0.0027), Str (F1,20 = 29.88, p < 0.0001) and basolateral amygdala (BLA) (F1,20 = 46.08, p < 0.0001), showing a higher intensity in both ethanolexposed rats and ethanol-exposed rats treated with fluoxetine
Drinking induced a significant overall effect on IBA-1 + cell number in the dorsal hippocampus (F1,20 = 15.03, p = 0.001), showing a specific increase in the dentate gyrus, but not CA3 and CA1 areas, of both ethanol-exposed rats and ethanol-exposed rats treated with fluoxetine

Summary

Introduction

Alcohol is a psychoactive substance highly consumed in the general population worldwide (Organization for Economic Co-operation and Development 2020). One-fourth of people with mental health problems have a co-occurring substance use disorder, most commonly, AUD This dual diagnosis is challenging since clinical outcomes of antidepressant treatment in co-morbid dual depression is poor and there is a need for analyzing antidepressant medications in a comprehensive, integrated approach. In this regard, pharmacological interventions using selective serotonin reuptake inhibitors (SSRIs), among others (Ballesta et al 2019, 2020), are able to reduce clinical symptoms of alcohol withdrawal including hyperlocomotion, anxiety and negative mood (Torrens et al 2005; Uzbay 2008; Simon O’Brien et al 2011; Bell et al 2017). We have suggested that dysregulation of glutamatergic receptor function and endocannabinoid signaling in the central amygdala likely underlies the contribution of SSRI treatment cessation in alcohol relapse (Suárez et al 2020)

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Conclusion