Paternal Exposure to the Neonicotinoid Pesticide Clothianidin Alters Sperm MicroRNA Profiles in Mice and Intergenerationally Reduces Locomotor Activity in Male Offspring

Accumulating evidence suggests that paternal environmental factors have epigenetic effects on sperm and influence offspring. Neonicotinoid pesticides (NNs), which are widely used around the world, are known to affect offspring phenotypes through maternal exposure in mice, but the effect of the paternal exposure remains unknown. The main purpose of this study was to examine the neurobehavioral effects on offspring mice resulting from paternal exposure to an NN, clothianidin (CLO). We also investigated microRNA (miRNA) expression in sperm-which plays an important role in epigenetics-to explore the involvement of sperm miRNAs in the paternal effects of CLO. Male mice were exposed to a no-observed-adverse-effect level dose of CLO (50 mg·kg^-1·day^-1) for 6 weeks. Male offspring of CLO-exposed fathers showed decreased levels of spontaneous locomotor activity, accompanied by decreased dopamine and histamine levels in the striatum. Hippocampal RNA sequencing analysis showed that paternal CLO exposure downregulated the expression of genes involved in neurogenesis processes, including the development of neural cells and synaptogenesis. The miRNAs, such as miR-149-5p and miR-295-3p, which were upregulated in paternal sperm, were found to target genes associated with the development of neurons, neuritogenesis, and axon guidance. These results suggest that these upregulated sperm miRNAs altered by CLO exposure reduced the expression of genes that regulate early embryogenesis and may lead to persistent effects on neural development in the offspring. This study provides novel insights into the impact of paternal exposure to NNs on offspring via sperm.