Autism gene variant causes hyperserotonemia, serotonin receptor hypersensitivity, social impairment and repetitive behavior.

Fifty years ago, increased whole-blood serotonin levels, or hyperserotonemia, first linked disrupted 5-HT homeostasis to Autism Spectrum Disorders (ASDs). The 5-HT transporter (SERT) gene (SLC6A4) has been associated with whole blood 5-HT levels and ASD susceptibility. Previously, we identified multiple gain-of-function SERT coding variants in children with ASD. Here we establish that transgenic mice expressing the most common of these variants, SERT Ala56, exhibit elevated, p38 MAPK-dependent transporter phosphorylation, enhanced 5-HT clearance rates and hyperserotonemia. These effects are accompanied by altered basal firing of raphe 5-HT neurons, as well as 5HT(1A) and 5HT(2A) receptor hypersensitivity. Strikingly, SERT Ala56 mice display alterations in social function, communication, and repetitive behavior. Our efforts provide strong support for the hypothesis that altered 5-HT homeostasis can impact risk for ASD traits and provide a model with construct and face validity that can support further analysis of ASD mechanisms and potentially novel treatments.

Patterns of cell death in the perinatal mouse forebrain.

The importance of cell death in brain development has long been appreciated, but many basic questions remain, such as what initiates or terminates the cell death period. One obstacle has been the lack of quantitative data defining exactly when cell death occurs. We recently created a "cell death atlas," using the detection of activated caspase-3 (AC3) to quantify apoptosis in the postnatal mouse ventral forebrain and hypothalamus, and found that the highest rates of cell death were seen at the earliest postnatal ages in most regions. Here we have extended these analyses to prenatal ages and additional brain regions. We quantified cell death in 16 forebrain regions across nine perinatal ages from embryonic day (E) 17 to postnatal day (P) 11 and found that cell death peaks just after birth in most regions. We found greater cell death in several regions in offspring delivered vaginally on the day of parturition compared with those of the same postconception age but still in utero at the time of collection. We also found massive cell death in the oriens layer of the hippocampus on P1 and in regions surrounding the anterior crossing of the corpus callosum on E18 as well as the persistence of large numbers of cells in those regions in adult mice lacking the pro-death Bax gene. Together these findings suggest that birth may be an important trigger of neuronal cell death and identify transient cell groups that may undergo wholesale elimination perinatally. J. Comp. Neurol. 525:47-64, 2017. © 2016 Wiley Periodicals, Inc.

Letting a typical mouse judge whether mouse social interactions are atypical.

Diagnosis of an autism spectrum disorder (ASD) requires a qualitative assessment of social aptitude: one person judging whether another person interacts in a "typical" way. We hypothesized that mice could be used to make a similar judgment if they prefer "typical" over "atypical" social interactions with mouse models relevant to ASD. We used wild-type C57BL/6 (B6) mice as "judges" and evaluated their preference for a chamber containing a "typical" (B6 or 129S6) or an "atypical" mouse. For our atypical mouse stimuli, we chose two inbred strains with well-documented social phenotypes (BTBR and BALB/c), as well a mutant line with abnormal social behavior and seizures (Gabrb3 +/-). Overall, we observed a stimulus by time interaction (P < 0.0001), with B6 mice preferring the typical mouse chamber during the last 10 min of the 30-min test. For two of the individual stimulus pairings, we observed a similar chamber by time interaction (BALB/c vs. 129S6, P = 0.0007; Gabrb3 +/- vs. 129S6, P = 0.033). For the third stimulus pairing, we found a trend for preference of the typical mouse across time (BTBR vs. B6, P = 0.051). We repeated the experiments using 129S6 mice as judges and found a significant overall interaction (P = 0.034), but only one stimulus pairing reached significance on its own (BALB/c vs. 129S6, P = 0.0021). These data suggest that a characteristic pattern of exploration in B6 mice can distinguish some socially atypical animals from controls.

Genetic background modulates phenotypes of serotonin transporter Ala56 knock-in mice.

BACKGROUND: Previously, we identified multiple, rare serotonin (5-HT) transporter (SERT) variants in children with autism spectrum disorder (ASD). Although in our study the SERT Ala56 variant was over-transmitted to ASD probands, it was also seen in some unaffected individuals, suggesting that associated ASD risk is influenced by the epistatic effects of other genetic variation. Subsequently, we established that mice expressing the SERT Ala56 variant on a 129S6/S4 genetic background display multiple biochemical, physiological and behavioral changes, including hyperserotonemia, altered 5-HT receptor sensitivity, and altered social, communication, and repetitive behavior. Here we explore the effects of genetic background on SERT Ala56 knock-in phenotypes. METHODS: To explore the effects of genetic background, we backcrossed SERT Ala56 mice on the 129 background into a C57BL/6 (B6) background to achieve congenic B6 SERT Ala56 mice, and assessed autism-relevant behavior, including sociability, ultrasonic vocalizations, and repetitive behavior in the home cage, as well as serotonergic phenotypes, including whole blood serotonin levels and serotonin receptor sensitivity. RESULTS: One consistent phenotype between the two strains was performance in the tube test for dominance, where mutant mice displayed a greater tendency to withdraw from a social encounter in a narrow tube as compared to wildtype littermate controls. On the B6 background, mutant pup ultrasonic vocalizations were significantly increased, in contrast to decreased vocalizations seen previously on the 129 background. Several phenotypes seen on the 129 background were reduced or absent when the mutation was placed on the B6 background, including hyperserotonemia, 5-HT receptor hypersensivity, and repetitive behavior. CONCLUSIONS: Our findings provide a cogent example of how epistatic interactions can modulate the impact of functional genetic variation and suggest that some aspects of social behavior may be especially sensitive to changes in SERT function. Finally, these results provide a platform for the identification of genes that may modulate the risk of ASD in humans.

Absence of preference for social novelty and increased grooming in integrin ß3 knockout mice: initial studies and future directions.

Elevated whole blood serotonin 5-HT, or hyperserotonemia, is a common biomarker in autism spectrum disorder (ASD). The integrin ß3 receptor subunit gene (ITGB3) is a quantitative trait locus for whole blood 5-HT levels. Recent work shows that integrin ß3 interacts with the serotonin transporter (SERT) in both platelets and in the midbrain. Furthermore, multiple studies have now reported gene-gene interaction between the integrin ß3 and SERT genes in association with ASD. Given the lack of previous data on the impact of integrin ß3 on brain or behavioral phenotypes, we sought to compare mice with decreased or absent expression of the integrin ß3 receptor subunit (Itgb3 +/- and -/-) with wildtype littermate controls in behavioral tasks relevant to ASD. These mice did not show deficits in activity level in the open field or anxiety-like behavior on the elevated plus maze, two potential confounds in the evaluation of mouse social behavior. In the three-chamber social test, mice lacking integrin ß3 were shown to have normal sociability but did not show a preference for social novelty. Importantly, the absence of integrin ß3 did not impair olfaction or the ability to recall familiar social odors. Additionally, mice lacking integrin ß3 showed increased grooming behavior in novel environments. These preliminary studies reveal altered social and repetitive behavior in these mice, which suggests that the integrin ß3 subunit may be involved in brain systems relevant to ASD. Further work is needed to fully characterize these behavioral changes and the underlying brain mechanisms.