Elsevier

Neuroscience

Volume 345, 14 March 2017, Pages 155-165
Neuroscience

Touchscreen learning deficits and normal social approach behavior in the Shank3B model of Phelan–McDermid Syndrome and autism

https://doi.org/10.1016/j.neuroscience.2016.05.016Get rights and content

Highlights

  • Shank3B +/− mice exhibit deficits in higher order learning and memory.

  • Shank3B +/− mice exhibit normal social approach.

  • Shank3B +/− mice exhibit modestly increased repetitive self-grooming.

Abstract

SHANK3 is a synaptic scaffolding protein localized in the postsynaptic density and has a crucial role in synaptogenesis and neural physiology. Deletions and point mutations in SHANK3 cause Phelan–McDermid Syndrome (PMS), and have also been implicated in autism spectrum disorder (ASD) and intellectual disabilities, leading to the hypothesis that reduced SHANK3 expression impairs basic brain functions that are important for social communication and cognition. Several mouse models of Shank3 deletions have been generated, varying in the specific domain deleted. Here we report impairments in cognitive function in mice heterozygous for exon 13–16 (coding for the PDZ domain) deletion. The touchscreen pairwise discrimination task was chosen by virtue of its: (a) conceptual and technical similarities to the Cambridge Neuropsychological Test Automated Battery (CANTAB) and NIH Toolbox Cognition Battery used for testing cognitive functions in humans, (b) minimal demand on motor abilities, and (c) capability to measure many aspects of learning and memory and complex cognitive functions, including cognitive flexibility. The similarity between our mouse tasks and human cognitive assays means a high translational validity in future intervention studies using preclinical models. Our study revealed that Shank3B heterozygous mice (+/−) were slower to reach criterion in the pairwise visual discrimination task, and exhibited trends toward making more errors (first trial errors) and more correction errors than wildtype mice (+/+). Open field activity was normal in +/−, ruling out hypo- or hyperactivity as potential confounds in the touchscreen test. Sociability in the three chamber test was also normal in both +/+ and +/−. These results indicate a deficit in discrimination learning in the Shank3B model of PMS and ASD, suggesting that this mouse model is a useful preclinical tool for studying neurobiological mechanisms behind cognitive impairments in PMS and ASD. The current findings are the starting point for our future research in which we will investigate multiple domains of cognition and explore pharmacological interventions.

Introduction

SHANKs are scaffolding proteins enriched in the postsynaptic density. They are crucial for the formation and stabilization of synapses (Qualmann et al., 2004, Grabrucker et al., 2009). SHANK3 (also referred to as PROSAP2) encodes a structural component of excitatory synapses important for synaptic morphology and functions (Herbert, 2011, Harony-Nicolas et al., 2015). Consisting of five domains (ankyrin repeats, SH3, PDZ, proline-rich, and SAM) (Naisbitt et al., 1999, Sheng and Kim, 2000, Bourgeron, 2007, Buxbaum, 2009), SHANK3 can interact with multiple key synaptic components, including glutamate receptor complexes, anchoring proteins, and actin cytoskeleton (Bockers et al., 2001, Roussignol et al., 2005, Baron et al., 2006, Durand et al., 2008, Bertaso et al., 2010). Heterozygous deletions or point mutations of SHANK3 are thought to be the main cause of Phelan–McDermid Syndrome (PMS, also referred to as 22q13 Deletion Syndrome), a genetic disorder characterized by global developmental delays, delayed or absent speech, moderate to severe intellectual disability, autism, some dysmorphic features, neonatal hypotonia, and seizures (Bonaglia et al., 2001, Phelan, 2008, Phelan and McDermid, 2012, Harony-Nicolas et al., 2015). Haploinsufficiency of SHANK3 due to deletion or de novo mutations occurs in approximately 1% of autism spectrum disorder (ASD) cases, making SHANK3 abnormalities one of the most common genetic causes of autism (Durand et al., 2007, Moessner et al., 2007, Buxbaum, 2009, Betancur and Buxbaum, 2013, Boccuto et al., 2013).

In addition to impaired social communication and repetitive behaviors, a hallmark feature of autism is restricted interests, deficits in set shifting and behavioral inflexibility (D’Cruz et al., 2013, Dawson et al., 2002, Miller et al., 2015, de Vries et al., 2015). A crucial step toward understanding cognitive inflexibility in ASD is to characterize associative learning in this disorder.

Our present study aimed at evaluating associative learning in the Shank3B model of PMS and ASD. Four independent groups have generated mouse models of Shank3 deficiency or ablation (Bozdagi et al., 2010, Peca et al., 2011, Wang et al., 2011, Kouser et al., 2013), and impaired learning and memory have been reported in all models. The current study employed the PDZ domain deletion model originally generated in the Feng lab (Peca et al., 2011). This model has both construct validity (reduced expression of Shank3 mRNA and protein) and some face validity (Excessive/injurious repetitive self-grooming and altered sociability) (Peca et al., 2011). +/+ and +/− were used in the current study, because: (a) heterozygous deletion is translational and analogous to deletions found in clinical populations, (b) excessive/injurious self-grooming and low general locomotor activity in null mutants of this line could confound results of the touchscreen operant learning task. Since no previous studies have evaluated complex learning in any of the deletion models, we chose the automated touchscreen task for its conceptual and technical similarities to the Cambridge Neuropsychological Test Automated Battery (CANTAB), an automated computerized battery of cognitive assays commonly used to test cognitive function in humans. In order to rule out hypo- or hyperactivity as confounds in this cognitive assay, we conducted the open field test to measure general locomotor activity. In a previous study on the Shank3B model, altered sociability was found in null mutants, but no data were reported in heterozygous mutants (Peca et al., 2011). Given the relevance of heterozygous deletions to the human disease condition, we therefore also evaluated sociability in the three chamber test in the current study.

Section snippets

Subjects

All procedures were approved by the Institutional Animal Care and Use Committees (IACUC) of the University of California Davis, and followed the NIH Guide for the Care and Use of Laboratory Animals. The Shank3B line characterized by a mutation within the PDZ domain was originally generated by the Feng lab (Peca et al., 2011). A neo cassette replaced exons 13–16 of the Shank3 gene, resulting in a deficiency of isoforms Shank3α and Shank3β, and a reduction in expression of the Shank3γ isoform.

Poor performance of −/− during pre-training stages

Eight null mutants (−/−) were initially included in the study. Genotype differences were not statistically significant for numbers of trials completed on habituation day 1 (F2,26 = 1.40, NS) or habituation day 2 (F2,26 = 1.82, NS), although trends were observed for −/− to complete fewer trials than +/+ on both days. As shown in Table 1, significant genotype effects were found in a number of trials completed in Stage 2 (F2,26 = 3.63, p < .05) and days to reach criterion in Stage 3 (F2,26 = p < 5.29, p < .05).

Discussion

Mouse models are indispensable tools for studying neurobiological mechanisms behind cognitive impairments caused by genetic abnormalities. Cognitive functions have been studied in a number of Shank3 deletion models, using simple assays (Bozdagi et al., 2010, Peca et al., 2011, Wang et al., 2011, Kouser et al., 2013). One line of mice homozygous for the exon 4–9 (coding for ankryin domain) deletion exhibited impaired novel object recognition but normal fear conditioning and spatial learning (

Conclusion

We report for the first time that pairwise discrimination associative learning is disrupted in +/− Shank3B mice, opening a new pathway to study neurobiological mechanisms behind intellectual disabilities caused by deletions/mutations in SHANK3. The touchscreen task requires habit forming, rule following, and attending to specific sensory stimuli (auditory, visual, and olfactory). Resistance to change or cognitive inflexibility could manifest as deficits in reversal learning. Over or

Acknowledgments

This work was supported by the Joe P. Tupin Research Award in Psychiatry and the MIND Institute’s Intellectual and Developmental Disabilities Research Center (NICHD-IDDRC) HD079125-01 (JLS). The authors are grateful to Drs. Jonathan Brigman (University of New Mexico) and Stacey J. Sukoff Rizzo for their knowledgeable suggestions on our protocol designs.

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