Paternal nicotine exposure defines different behavior in subsequent generation via hyper-methylation of mmu-miR-15b.

The neurobehavioral effects of paternal smoking and nicotine use have not been widely reported. In the present study, nicotine exposure induced depression in the paternal generation, but reduced depression and promoted hyperactivity in F1 offspring. While this intergenerational effect was not passed down to the F2 generation. Further studies revealed that nicotine induced the down-regulation of mmu-miR-15b expression due to hyper-methylation in the CpG island shore region of mmu-miR-15b in both the spermatozoa of F0 mice and the brains of F1 mice. As the target gene of mmu-miR-15b, Wnt4 expression was elevated in the thalamus of F1 mice due to the inheritance of DNA methylation patterns from the paternal generation. Furthermore, the increased expression of Wnt4 elevated the phosphorylation level of its downstream protein GSK-3 through the canonical WNT4 pathway which involved in the behavioral alterations observed in F1 mice. Moreover, in vivo stereotaxic brain injections were used to induce the overexpression of mmu-miR-15b and WNT4 and confirm the neurobehavioral effects in vitro. The behavioral phenotype of the F1 mice resulting from paternal nicotine exposure could be attenuated by viral manipulation of mmu-miR-15b in the thalamus.