Sex and age differences in social and cognitive function in offspring exposed to late gestational hypoxia

Abstract

Background: Gestational sleep apnea affects 8-26% of pregnancies and can increase the risk for autism spectrum disorder (ASD) in offspring. ASD is a neurodevelopmental disorder associated with social dysfunction, repetitive behaviors, anxiety, and cognitive impairment. To examine the relationship between gestational sleep apnea and ASD-associated behaviors, we used a chronic intermittent hypoxia (CIH) protocol between gestational days (GD) 15-19 in pregnant rats to model late gestational sleep apnea. We hypothesized that late gestational CIH would produce sex- and age-specific social, mood, and cognitive impairments in offspring.

Methods: Timed pregnant Long-Evans rats were exposed to CIH or room air normoxia from GD 15-19. Behavioral testing of offspring occurred during either puberty or young adulthood. To examine ASD-associated phenotypes, we quantified ASD-associated behaviors (social function, repetitive behaviors, anxiety-like behaviors, and spatial memory and learning), hippocampal activity (glutamatergic NMDA receptors, dopamine transporter, monoamine oxidase-A, EGR-1, and doublecortin), and circulating hormones in offspring.

Results: Late gestational CIH induced sex- and age-specific differences in social, repetitive and memory functions in offspring. These effects were mostly transient and present during puberty. In female pubertal offspring, CIH impaired social function, increased repetitive behaviors, and increased circulating corticosterone levels, but did not impact memory. In contrast, CIH transiently induced spatial memory dysfunction in pubertal male offspring but did not impact social or repetitive functions. Long-term effects of gestational CIH were only observed in female offspring, wherein CIH induced social disengagement and suppression of circulating corticosterone levels in young adulthood. No effects of gestational CIH were observed on anxiety-like behaviors, hippocampal activity, circulating testosterone levels, or circulating estradiol levels, regardless of sex or age of offspring.

Conclusions: Our results indicate that hypoxia-associated pregnancy complications during late gestation can increase the risk for ASD-associated behavioral and physiological outcomes, such as pubertal social dysfunction, corticosterone dysregulation, and memory impairments.

Keywords: Morris water maze; autism spectrum disorder; chronic intermittent hypoxia; hippocampus; marble burying behaviors; open field; prenatal programming; sex differences; social behaviors.

Figure 1. Social and exploratory behaviors.

Gestational CIH decreased social behaviors in female rats during puberty (A) but not during young adulthood (B). No effect of gestational CIH was observed on male rats during puberty (A) or young adulthood (B). No effect of sex was observed on social behaviors (A, B). No effect of gestational CIH or sex was observed in exploratory behavior (C, D). Analyzed by Two-way ANOVA with Fisher’s LSD multiple comparisons tests. ANOVA significance indicated by: ** = CIH; Post-hoc significance indicated by: # versus normoxic female; p≤0.05

Figure 2. Social disengagement.

No effect of gestational CIH or sex was observed on social disengagement in pubertal rats (A). Gestational CIH only increased percentage of young adult female rats that exhibited social disengagement (B). Social withdrawal was significantly associated with gestational CIH only in pubertal females (C). Social disengagement analyzed by Two-way ANOVA with Fisher’s LSD multiple comparisons tests (A, B). Relationship between social engagement and CIH measured by Fisher’s exact test (C). Significance indicated by: ** = CIH, *** = interaction; p≤0.05

Figure 3. Repetitive behaviors.

Gestational CIH increased marble burying in pubertal females (A) and young adult females (B). Males buried more marbles than females, regardless of age (A, B). Normalized by square-root transformation (A, B). Analyzed by Two-way ANOVA with Fisher’s LSD multiple comparisons tests. ANOVA significance indicated by: * = sex, ** = CIH; Post-hoc significance indicated by: # versus normoxic female, p≤0.05

Figure 5. Spatial learning and memory.

Pubertal males had longer pathlength to target during Morris water maze probe trial (A), with the sex difference observed primarily in gestational CIH exposed pubertal males. No effect of sex or gestational CIH was observed on pathlength in young adult rats (B). No effect of gestational CIH on latency to target was observed in females regardless of age (C). Age improved latency to target during day 2 of learning in males with no effect of age observed on other days (D). Analyzed by Two-way ANOVA with Fisher’s LSD multiple comparisons tests. ANOVA significance indicated by: * = sex; ** = age; Post-hoc significance indicated by: ## = versus CIH female; p≤0.05; P = Puberty, YA = Young Adult