MicroRNAs (miRNA) are a recently recognised class of small, non-coding RNAs involved in the post-transcriptional regulation of gene expression and with crucial implication for mammalian development. In particular, they play key roles in neuronal development, from early neurogenesis to neuronal differentiation and synaptic development, and also in in vitro systems. The detection of embryotoxic hazards in the preclinical phase is still a challenge, often due to species-species variations. In this study we analysed whether miRNA expression profiles in a human pluripotent cell model can be a helpful tool for a more mechanistic approach to pharmacology and toxicology. Differentiating human pluripotent cells were repeatedly treated with non-cytotoxic doses of methylmercury chloride (MeHgCl), a well known brain developmental toxicant. The expression of proteins, mRNA and miRNAs were used to monitor successful neural differentiation. Significant changes in the expression of 12 miRNAs were detected. By using available bioinformatics tools, we obtained validated and predicted targets for the identified miRNAs, on which we performed functional clustering analysis. Through this approach, we identified several terms and functional clusters associated with neural development, together with indicators of general toxic effect, such as apoptosis or stress response-related genes. Interestingly, our results also suggest a previously undiscovered association between MeHgCl and the ubiquitin-proteasome protein degradation pathway. Although further investigations are needed, our results suggest that miRNA expression analysis is a powerful tool in pathway-oriented toxicity and could improve early-phase hazard assessments.