Juvenile manifestation of ultrasound communication deficits in the neuroligin-4 null mutant mouse model of autism

• Nlgn4−/− mice are a construct valid autism model, now confirmed in pups and juveniles.
• USV in pups on brief maternal separation revealed remarkable gender and mild genotype effects.
• USV in juveniles on exposure to an anesthetized female was reduced in Nlgn4−/− mice.
• Neonatal development from PND4 to 21 did not yield further differences between genotypes.

Neuroligin-4 (Nlgn4) is a member of the neuroligin family of postsynaptic cell adhesion molecules. Loss-of-function mutations of NLGN4 are among the most frequent, known genetic causes of heritable autism. Adult Nlgn4 null mutant (Nlgn4−/−) mice are a construct valid model of human autism, with both genders displaying a remarkable autistic phenotype, including deficits in social interaction and communication as well as restricted and repetitive behaviors. In contrast to adults, autism-related abnormalities in neonatal and juvenile Nlgn4−/− mice have not been reported yet. The present study has been designed to systematically investigate in male and female Nlgn4−/− pups versus wildtype littermates (WT, Nlgn4+/+) developmental milestones and stimulus-induced ultrasound vocalization (USV). Neonatal development, followed daily from postnatal days (PND) 4 to 21, including physical development, neurological reflexes and neuromotor coordination, did not yield any differences between Nlgn4−/− and their WT littermates. USV in pups (PND8–9) in response to brief separation from their mothers revealed remarkable gender effects, and a genotype influence in females regarding latency to first call. In juveniles (PND22–23), USV monitoring upon exposure to an anesthetized female intruder mouse uncovered a clear genotype effect with reduced USV in Nlgn4−/− mice, and again a more prominent phenotype in females. Together, these data support an early manifestation of communication deficits in Nlgn4−/− mice that appear more pronounced in immature females with their overall stronger USV as compared to males.