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, 39 (9), 2049-60

Autistic-like Syndrome in Mu Opioid Receptor Null Mice Is Relieved by Facilitated mGluR4 Activity

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Autistic-like Syndrome in Mu Opioid Receptor Null Mice Is Relieved by Facilitated mGluR4 Activity

Jérôme A J Becker et al. Neuropsychopharmacology.

Abstract

The etiology of Autism Spectrum Disorders (ASDs) remains largely unknown. Identifying vulnerability genes for autism represents a major challenge in the field and allows the development of animal models for translational research. Mice lacking the mu opioid receptor gene (Oprm1(-/-)) were recently proposed as a monogenic mouse model of autism, based on severe deficits in social behavior and communication skills. We confirm this hypothesis by showing that adult Oprm1(-/-) animals recapitulate core and multiple comorbid behavioral symptoms of autism and also display anatomical, neurochemical, and genetic landmarks of the disease. Chronic facilitation of mGluR4 signaling, which we identified as a novel pharmacological target in ASDs in these mice, was more efficient in alleviating behavioral deficits than the reference molecule risperidone. Altogether, our data provide first evidence that disrupted mu opioid receptor signaling is sufficient to trigger a comprehensive autistic syndrome, maybe through blunted social reward processes, and this mouse model opens promising avenues for therapeutic innovation.

Figures

Figure 1
Figure 1
Oprm1−/− mice show core symptoms of autism. (a) In the direct social interaction test, Oprm1−/− animals show decreased time in close social contact, due to lower frequency and duration of nose and paw contacts. Following was decreased, whereas self-grooming, especially after social contact, was increased in mutants (n=8 per genotype). (b) In the three-chamber task, WT controls but not Oprm1−/− mice spend more time interacting with a mouse rather than a toy (interaction phase) or a novel rather than a familiar conspecific (recognition phase) (n=13–14 per genotype). (c) Male Oprm1−/− animals are more aggressive in the resident-intruder test, with increased numbers of attacks and episodes of ptosis (n=8 per genotype). (d) Oprm1−/− mice show impaired ability to build a nest (n=8 per genotype). (e) Oprm1−/− mice display motor stereotypies, with increased rearing, grooming, circling, and head shakes (n=8 per genotype). (f) Mutant mice show deficient patterns of exploration in a Y-maze (n=16 per genotype). Spontaneous alternations (SPA) are decreased in Oprm1−/− animals; alternate arm returns (AAR) and same arm returns (SAR) are increased. Data are presented as mean±SEM. Solid stars: genotype effect, open stars: stimulus effect. One star p<0.05, two stars p<0.01, three stars p<0.001. See also Supplementary Figure S1.
Figure 2
Figure 2
Oprm1−/− mice display multiple secondary symptoms of autism. (a) Defensive anxiety is higher in mutant animals, hiding more marbles than controls in the marble-burying test (n=10 per genotype). (b) Conflict anxiety is also increased, as shown by longer feeding latencies in the novelty-suppressed feeding (NSF) test (n=10 per genotype). (c, d) Increased anxiety of Oprm1−/− mice in NSF is associated with increased number of Fos positive (Fos+) nuclei in the central nucleus of the amygdala (CeA) and decreased Fos staining in the ventral tegmental area (VTA). (e) Principal component analysis on NSF data, including behavioral (red dots) and Fos (grey dots) responses, opposes food intake with latency to eat along the first principal component (PC1, 31.7% of variance). Fos expression in brain regions implicated in the regulation of anxiety-like behavior clusters with the latency parameter, and Fos-staining in neural circuits involved in the control of reward processes correlates with food intake (left panel: variables' space). Oprm1+/+ and Oprm1−/− individual mice are dissociated in the subjects' space (right panel). (f, g) Motor function is altered in mutant animals. Oprm1−/− mice show impaired motor performance in the accelerating rotarod task (n=7 per genotype) and higher variability of pace size (sigma) in the footprint test (n=15–16 per genotype). (h) Sensitivity to pentylenetetrazole-induced seizures (n=8 per genotype) is increased in mice lacking the mu opioid receptor. Data are presented as mean±SEM. Genotype effect: one star p<0.05, two stars p<0.01, three stars p<0.001. See Supplementary Figure S2 for more tests and parameters and complete list of abbreviations.
Figure 3
Figure 3
Oprm1−/− mice display transcriptional modifications and abnormal striatal synapses. (a) Transcription of multiple candidate genes for autism is modified in mutant animals across four brain regions (see details in Supplementary Table S3). (b) Oprm1−/− mice display abnormalities in the number of striatal symmetrical synapses (left panels, 120 frames per mouse at × 40 000 magnification). Striatal asymmetrical synapses show reduced surface of postsynaptic densities (right panel). (c) Representative microphotographs ( × 40 000) illustrate morphology of striatal asymmetrical synapses in Oprm1+/+ and Oprm1−/− mice. Scale bar: 200 nm. Asterisk: presynaptic element. (d) Clustering analysis of gene expression data classifies genes in four clusters (AD). Cluster (D) gathers genes mostly involved in glutamatergic signaling that all show lowered mRNA levels in the caudate putamen and nucleus accumbens of Oprm1−/− mice, among which Grm4 (framed), coding mGluR4 glutamate receptors, was detected only in these regions. Data in (a) and (b) are presented as mean±SEM. Genotype effect: one star p<0.05, two stars p<0.01, three stars p<0.001. CeA, central nucleus of the amygdala; CPu, caudate putamen; NAc, nucleus accumbens; PFC, prefrontal cortex. See also Supplementary Figure S4.
Figure 4
Figure 4
VU0155041, a positive allosteric modulator of mGluR4 glutamate receptors and not the antipsychotic risperidone, restores initiation of social interaction in Oprm1−/− mice. (a) When a naive wild-type social partner is used in the dyadic interaction test, chronic risperidone (0.2 mg/kg) restores normal frequency of nose and paw contacts as well as grooming episodes but fails to increase the time spent in close contact, the duration of nose and paw contacts, and the number of following episodes in mutant animals (n=14 per genotype and treatment). (b) Chronic treatment with VU0155041 (5 mg/kg) improves all these parameters except the number of following episodes and increases the occurrence of paw contacts in controls (9–10 per genotype and treatment). (c) Using a paradigm in which each experimental mouse encounters a genotype and treatment-matched animal confirms the efficiency of VU0155041 to normalize or even facilitate social interaction in mutant mice, including the frequency of following episodes (n=7–8 per genotype and treatment). (d) Chronic VU0155041 administration normalizes deficient C-fos expression following social interaction in the NAc, MeA, and VTA as well as deficient Egr1 expression in the VTA of Oprm−/− mice. Data are presented as mean±SEM. Open stars: treatment effect; black stars: genotype × treatment interaction (post-hoc: Newman–Keuls test); grey stars: comparison with saline-treated Oprm1+/+ mice. One star p<0.05, two stars p<0.01, three stars p<0.001.
Figure 5
Figure 5
VU0155041 alleviates secondary symptoms of autism in Oprm1−/− mice more efficiently than risperidone. (a) Both risperidone (left) and VU0155041 (right) treatments suppress motor stereotypies. (b) In the marble burying test, risperidone and VU0155041 normalize stereotyped burying in mutant mice; VU0155041, however, increases anxiety-induced burying in Oprm1+/+ mice. (c) Latencies to feed in the NSF do not significantly differ between Oprm1+/+ and Oprm1−/− mice under risperidone treatment, while VU0155041 reverses the high latency to eat of Oprm1−/− mice to control levels. Food intake remains unchanged. Data are presented as mean±SEM. Open stars: treatment effect; solid stars: genotype × treatment interaction (post-hoc: Newman-Keuls); One star p<0.05, two stars p<0.01, three stars p<0.001. See also Supplementary Figure S5.

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