Heterozygous Disruption of Autism susceptibility candidate 2 Causes Impaired Emotional Control and Cognitive Memory

Abstract

Mutations in the Autism susceptibility candidate 2 gene (AUTS2) have been associated with a broad range of psychiatric illnesses including autism spectrum disorders, intellectual disability and schizophrenia. We previously demonstrated that the cytoplasmic AUTS2 acts as an upstream factor for the Rho family small GTPase Rac1 and Cdc42 that regulate the cytoskeletal rearrangements in neural cells. Moreover, genetic ablation of the Auts2 gene in mice has resulted in defects in neuronal migration and neuritogenesis in the developing cerebral cortex caused by inactivation of Rac1-signaling pathway, suggesting that AUTS2 is required for neural development. In this study, we conducted a battery of behavioral analyses on Auts2 heterozygous mutant mice to examine the involvement of Auts2 in adult cognitive brain functions. Auts2-deficient mice displayed a decrease in exploratory behavior as well as lower anxiety-like behaviors in the absence of any motor dysfunction. Furthermore, the capability for novel object recognition and cued associative memory were impaired in Auts2 mutant mice. Social behavior and sensory motor gating functions were, however, normal in the mutant mice as assessed by the three-chamber test and prepulse inhibition test, respectively. Together, our findings indicate that AUTS2 is critical for the acquisition of neurocognitive function.

Fig 1. Decrease of locomotor activity in Auts2 mutant mice during habituation.

(A) Spontaneous locomotor activity for habituation to the novel environment was measured every 5 min for 60 min. Auts2-deficient mice exhibited a decrease in exploratory behavior during the first 15 min. (B) Assessment of physical motor function and coordination of Auts2 mutant mice in a rotarod test. The latency of falling from an accelerating rotarod was measured for five consecutive days. Data are presented as the mean ± SEM (A: WT, n = 17, Auts2 ^neo/+ = 14 and B: WT, n = 10, Auts2 ^neo/+ = 9). *p<0.05; two-way ANOVA with repeated measures.

Fig 2. Abnormality of anxiety-like behaviors in Auts2 mutant mice.

(A) In open field tests, the time spent in the center field (left graph) and total distant traveled (right graph) were measured for 5 min to analyze mobility and exploration behavior. The ratio of distance traveled in an inner area scores as the percentage of total distance traveled (middle graph). Auts2 ^neo/+ mice spent significantly more time in the open environment than wild-type mice. (B) In elevated plus maze test, the duration of time spent in an arm (left graphs) and number of arm entries (right graphs) were measured for 5 min. The time (% Open arm time) and number of entries (% Open arm entry) in the open arms are presented by the percentage of total time (Total arm time) and number of total arm entries (Total arm entry), respectively. Auts2 ^neo/+ mice spent increased time in the unprotected open arm entry compared with wild-type mice. Data are presented as the mean ± SEM (WT, n = 17, Auts2 ^neo/+ = 14). *p<0.05, **p<0.01; Mann-Whitney U-test.

Fig 3. Normal social approach behaviors in Auts2-deficient mice.

(A) Sociability for contacting an unfamiliar mouse or empty chamber (middle graphs) and social novelty for a stranger mouse versus a familiar mouse (right graphs) of WT and Auts2 mutant mice were measured by a three-chamber test. Graphs show the time spent in the each chamber. Data are presented as the mean ± SEM (WT, n = 16, Auts2 ^neo/+ = 13), t test.

Fig 4. Impairment of cognitive memory functions in Auts2 mutant mice.

(A) Novel object recognition test. Graphs show the exploratory preference (left graph) and total exploration time (right graph) in training and retention sessions. The retention session was carried out 24 hrs after the training session. (B) Associative memory function was measured by the cued (Tone-dependent) and contextual (Context-dependent) fear-conditioning test 24hrs after the conditioning phase (Conditioning). Note that Auts2-deficient mice had defective auditory fear conditioning showing a decrease of freezing response while they displayed a higher response to lower nociceptive stimuli (foot shock) compared with the wild type mice (Nociceptive threshold). Data are presented as the mean ± SEM (WT, n = 17, Auts2 ^neo/+ = 14). *p<0.05, **p<0.01; Mann-Whitney U-test.

Fig 5. Normal behaviors of Auts2-deficient mice on PPI.

PPI (%) at four different prepulse intensities (69, 73, 77 and 81 dB) in PPI test (left graph) and acoustic startle response to two different pulses (middle and right graphs) as measured in trials without a prepulse. There were no significant differences between wild type and Auts2 mutant mice in the characterization of PPI whereas Auts2 mutant mice exhibited a higher acoustic startle response at 120 dB pulse compared to WT mice. Data are presented as the mean ± SEM (WT, n = 17, Auts2 ^neo/+ = 13). **p<0.0001; two-way ANOVA with repeated measures in PPI test and Mann-Whitney U-test in startle response.