Due to massive production, improper use and disposal of plastics, microplastics have become global environmental pollutants affecting both freshwater and marine ecosystems. Several studies have documented the uptake of microplastics in wild species and their biological effects, ranging from epithelial damage, inflammation, biochemical biomarkers response, and gene expression alteration. However, the biological effects of microplastics are not fully understood yet, especially the potential impact on immune functions. In this study, adult zebrafish have been exposed for twenty days to two concentrations of a mix of polystyrene and high-density polyethylene microplastics. Gene expression was evaluated in head-kidney, while biochemical biomarkers were evaluated in head and body homogenates. Acetylcholinesterase activity was slightly inhibited by microplastics exposure, while no significant effects on lactate dehydrogenase activity were observed. Microplastics exposure up-regulated genes involved in sterol biosynthesis, xenobiotics metabolism, and adaptive immunity. The results presented in this study support the hypothesis that microplastics exposure could somehow affect acetylcholinesterase functioning and induce immune and metabolic responses at transcriptional level.