Grand-paternal age and the development of autism-like symptoms in mice progeny

Abstract

Advanced paternal age (APA) contributes to the risk of autism spectrum disorders (ASDs) in children. In this study, we used a mouse model to investigate the effects of APA on behavioral features related to autistic syndromes (that is, social deficits, communication impairments and stereotypic/repetitive behaviors). We also examined whether such effects are transmitted across generations. To do this, males aged 15 months (APA) and 4 months (control) were bred with 4-month-old females, and the resulting offspring (F1) and their progeny (F2; conceived by 4-month-old parents) were tested for the presence and severity of ASD-like behaviors. Our results indicate that APA resulted in offspring that displayed distinctive symptoms of ASD. We found that both F1 conceived from old fathers and F2 derived from old grandfathers displayed increased ultrasound vocalization (USV) activity, decreased sociability, increased grooming activity and increased anxiety-like responses. Moreover, such abnormalities were partially transmitted to the second generation of mice, having APA grandfathers. In conclusion, our study suggests that the risk of ASD could develop over generations, consistent with heritable mutations and/or epigenetic alterations associated with APA.

Figure 1

Increased USV activity in APA pups upon isolation from their mothers (n=25 pups per generation per group). (a) Both generations of APA pups emitted an increased number of calls with respect to controls throughout postnatal development (*P<0.001). (b) The percentage of high intensity calls was greater in F1 and F2 progenies derived from old fathers versus controls, on P4 and P8 (*P<0.001). (c) Although APA pups emitted more calls when isolated in clean Petri dishes (*P<0.001), no significant difference between the groups was observed when pups were isolated in the presence of nest materials (NS: P=0.89). APA, advanced paternal age; CTR, control; NS, not significant; USV, ultrasound vocalization.

Figure 2

APA induced perturbations of social behaviors (n=15 mice per generation per group). (a) All mice showed significant preferences for spending time in the chamber containing the stranger mouse (*P<0.05, **P<0.01, ***P<0.001), although the magnitude of preference was lower in APA progenies. (b) The sociability score was significantly lower in both generations of APA mice with respect to controls (*P<0.05). (c) APA mice spent less time in social sniffing (*P<0.05, **P<0.01). (d) In the test for social novelty, both generations of mice derived from young fathers spent significantly more time in the chamber containing the unfamiliar mouse (*P<0.05, **P<0.01). Conversely, APA F1 failed to display such a preference (*P<0.05, NS: P=0.16). (e) The time spent sniffing the wire cage containing the unfamiliar mouse over the familiar one, was significantly less in APA progenies versus the controls (*P<0.05, **P<0.01). APA, advanced paternal age; CTR, control; NS, not significant.

Figure 3

Increased self-grooming activity (n=15 mice per generation per group). APA progenies spent significantly more time in self-grooming compared with controls (*P<0.05). Conversely, there were no significant differences in digging nor in the sum of all stereotyped behaviors analyzed. APA, advanced paternal age; CTR, control.

Figure 4

Effects of APA on anxiety-like behaviors (n=12 mice per generation per group). (a, b) F1 mice derived from aged fathers displayed less number of entries into the central area of the open field with respect to the controls. Moreover the time spent in the central area was lower in APA with respect to the controls (*P<0.05). (c) These mice also made significantly fewer stretch–attend postures in the open arms of the elevated plus maze than did the controls (*P<0.05). F2 progeny did not display changes in anxiety-like behaviors. APA, advanced paternal age; CTR, control.