Early-life ethanol exposure alters the splicing of microexons in the rat brain

Background Prenatal and early postnatal ethanol exposure can lead to fetal alcohol spectrum disorder (FASD), a neurodevelopmental condition characterized by impairments in cognitive functions such as attention, learning, and executive function. Despite its high prevalence, the molecular mechanisms underlying FASD are not well understood. One promising avenue of research involves the regulation of alternative splicing, particularly the inclusion of microexons. These short exonic sequences, which range from 3 to 51 nucleotides, are enriched in the brain and play critical roles in synaptic development and neuronal identity. While alterations in microexon splicing have been associated with other neurodevelopmental disorders, their role in FASD pathophysiology remains unexplored. Methods To investigate the impact of postnatal ethanol exposure on microexon splicing, we performed an analysis of rat brain tissue following acute and repeated ethanol exposure. We focused on two ethanol-sensitive microexons: E20 in the Intersectin 1 transcript and E8a in the lysine-specific demethylase 1 transcript. Splicing patterns were assessed in the hippocampus and prefrontal cortex at different time points using quantitative PCR and Western blotting to evaluate changes in splicing factor expression. Results Acute postnatal ethanol exposure resulted in a transient reduction in E20 inclusion in both the hippocampus and prefrontal cortex. In contrast, E8a inclusion was significantly reduced in the hippocampus at 24 h postexposure, coinciding with decreased expression of the splicing factor Srrm3. Repeated ethanol exposure did not affect E20 or E8a splicing in the hippocampus but increased E8a inclusion in the prefrontal cortex, correlating with elevated levels of the splicing factors Srrm3 and Srrm4. Conclusions These results demonstrate that postnatal ethanol exposure transiently disrupts the splicing of specific microexons in a region- and time-dependent manner. This finding suggests that altered microexon splicing may contribute to the molecular pathogenesis of FASD, highlighting alternative splicing as a dynamic regulatory mechanism with potential therapeutic implications for FASD.