DLK1-DIO3 IMPRINTED LOCUS SETS INPUT-SIDE DEPENDENT LEFT-RIGHT ASYMMETRY OF MOUSE HIPPOCAMPAL CA3-CA1 SYNAPSES

Left–Right asymmetries of the CNS are a well-documented feature across vertebrate species, offering evolutionary advantages in data processing speed and energy efficiency. In the mouse hippocampus, CA3-CA1 synapses show distinct input-side dependent asymmetry. Right CA3 inputs lead to larger, GluA1-dense synapses, whereas left CA3 inputs form smaller, GluN2B-dense synapses and exhibit higher capacity for t-LTP induction. Mice lacking this asymmetry show deficits in spatial navigation and working memory. Despite its significance, the molecular basis governing this input-side dependent asymmetry remains elusive. To investigate the genetic drivers of hippocampal asymmetry, we used inversus viscerum (iv) mice, which carry a spontaneous mutation at the Dnah11 locus affecting ciliary function and exhibit the right CA3-input phenotype bilaterally (right isomerism). Interestingly, the hippocampal phenotype in iv mice shifted during the backcrossing from SI/Col to C57Bl/6J background. During N5 rounds of backcrossing, iv5 mice maintained the initial right isomerism. However, after N10 rounds, iv10 animals showed a switch from right to left isomerism. These phenotypes were validated using TEM synaptic ultrastructure and patch-clamp recordings. Whole-genome sequence analysis of F2 pups from iv5 and iv10 crosses revealed an insertion mutation in a DLK1-DIO3 locus imprinted gene. Homozygous F2 pups for the insertion showed a left isomerism phenotype, while heterozygous pups followed imprinted inheritance. Subsequent genotyping confirmed the insertion in all left isomerism iv10 animals. Parental inheritance was further confirmed in reciprocal iv5/iv10 crossing litters. Ongoing experiments utilize CRISPR/Cas9 to knock-in the insertion into the DLK1-DIO3 locus in iv5 and wild-type backgrounds to confirm sufficiency.