Sex Differences in Social Behaviour in a Mouse Model of Aromatase Deficiency: An Autism Spectrum Disorder-Like Phenotype

2017-03-23T03:18:06Z (GMT) by Kristina Vacy
Deficits in social interaction, communication and repetitive behaviours are the core markers of the autism spectrum disorders (ASD), which are five times more likely to be diagnosed in boys than girls, reason unknown. Post-mortem studies showed that ASD brains expressed less brain aromatase, and 15% fewer neurons in the amygdala. Functional MRI showed a reduction in amygdala activity during social processing tasks and this same region expressed abundant levels of aromatase, the enzyme that catalyzes the conversion of androgens to oestrogens.
   
   Our lab demonstrated that mouse posterodorsal medial amygdala (PDMeA) contains high levels of aromatase, with males expressing more than females. We hypothesized that male, but not female aromatase knockout (ArKO) mouse, will exhibit behaviours consistent with a mouse model of ASD and that administration of carbetocin (an analogue of oxytocin) will rescue the social interaction deficit in the male ArKO.
   
   Our aim was to analyse the social interaction behavior of the oestrogen-deficient (ArKO) model and test whether carbetocin administration could reverse any social interaction deficit.
 
   The three chamber social interaction task was used to examine social approach in three week-old male and female ArKO and Wildtype mice littermates (WT). The effects of carbetocin treatment (2mg/kg and 20mg/kg i.p.) on social behaviour was investigated in both juvenile and adult male mice using this task. We found that male, but not female ArKO mice spent less time interacting with sex and aged matched unfamiliar mice (a “ stranger”) compared with WT of the same sex (n=6-10/group, p<0.05) during the social approach trial. In a later experiment, we injected six month-old male ArKO and WT intraperitoneally with 2mg/kg or 20mg/kg carbetocin but neither dose increased social approach. We also recorded and analysed the ultrasonic vocalizations (USV) made by male and female ArKO pups and WT littermates at post-natal day 9 during a 5 min separation from the litter. Male but not female ArKO pups made fewer and shorter USV (n=7-15/group, p<0.05).
   
   As seizure disorders are more common in ASD, we tested seizure susceptibility in 40-60 day old male ArKO and WT mice by subcutaneously administering 80mg/kg of the pro-convulsant drug pentylenetetrazol (PTZ). We then recorded time taken to reach the clonic and hind leg extension stages of the resulting seizure and found a trend for the ArKO to reach the hind leg extension stage more quickly compared to WT.
   
   To measure neural activation of the PDMeA during social interaction, we exposed 3 week-old male and female transgenic mice with a Enhanced Fluorescent Green Protein reporter linked to the aromatase to a stranger or a novel object. Their brains were harvested for immunohistochemical analysis two hours after testing, and were co-labeled with c-fos, a marker for neural activation, and GFP. We found greater c-fos immunoreactivity in mice exposed to the stranger compared with object, thus we also exposed 3 week-oldmale ArKO to a stranger. Stereology analysis revealed significantly more c-fos immunoreactivity in the ArKO compared with WT (N=4-5/group, p<0.05).
 
   Our results revealed behavioural sex differences between genotypes, with male (but not female) ArKO mice showing a behavioural phenotype consistent with ASD. These inferred that aromatase may play a role in regulating ASD-like behaviour in a sex specific manner. However the addition of the oxytocin analogue carbetocin did not rescue the social interaction deficit of the male ArKO.