Paternal scrotal heat stress epigenetically impairs spatial memory in F1 male offspring in mice

Increasing evidence indicates that the maternal preconception environment and lifestyle affect offspring health; however, the effects of paternal preconception environment and lifestyle have rarely been examined. Therefore, we aimed to examine whether paternal infertility-related factors affect offspring produced via assisted reproductive technology in mice. Scrotal heat impairment, which can result from factors such as varicoceles, sauna use, physical inactivity, or occupations such as welding, is a common risk factor associated with reduced male fertility. To model these effects, we exposed the scrota of C57BL/6 J male mice to 39 °C water for 30 min once weekly for 5 consecutive weeks, establishing frequent and mild scrotal heat stress (fmSHS) that mimics human SHS. Sperm from these mice, along with untreated female oocytes, were used for in vitro fertilization with embryo transfer to produce first-generation (F1) offspring. Second-generation (F2) offspring were obtained by mating F1 males with untreated females. Reduced spatial memory was observed in F1 males derived from fmSHS-exposed fathers, whereas F2 fmSHS males showed no significant impairment in spatial-memory testing. In F1 fmSHS offspring, hippocampal histology and N-methyl-D-aspartate receptor expression did not differ from controls. However, genes involved in the long-term potentiation pathway were dysregulated in the hippocampi of F1 fmSHS males, and abnormal methylation of the Pi3kcg promoter was observed in both the sperm of fmSHS fathers and the hippocampi of F1 males. These findings highlight the adverse effects of paternal preconception scrotal heat stress on offspring health.