Holly C. Dow

Assessment of NMDA receptor NR1 subunit hypofunction in mice as a model for schizophrenia.

N‐methyl‐D‐aspartate receptors (NMDARs) play a pivotal role in excitatory neurotransmission, synaptic plasticity and brain development. Clinical and experimental evidence suggests a dysregulation of NMDAR function and glutamatergic pathways in the pathophysiology of schizophrenia. We evaluated electrophysiological and behavioral properties of NMDAR deficiency utilizing mice that express only 5–10% of the normal level of NMDAR NR1 subunit. Auditory and visual event related potentials yielded significantly increased amplitudes for the P20 and N40 components in NMDAR deficient (NR1neo−/−) mice suggesting decreased inhibitory tone. Compared to wild types, NR1neo−/− mice spent less time in social interactions and showed reduced nest building. NR1neo−/− mice displayed a preference for open arms of a zero maze and central zone of an open field, possibly reflecting decreased anxiety‐related behavioral inhibition. However, locomotor activity did not differ between groups in either home cage environment or during behavioral testing. NR1neo−/− mice displayed hyperactivity only when placed in a large unfamiliar environment, suggesting that neither increased anxiety nor non‐specific motor activation accounts for differential behavioral patterns. Data suggest that NMDAR NR1 deficiency causes disinhibition in sensory processing as well as reduced behavioral inhibition and impaired social interactions. The behavioral signature in NR1neo−/− mice supports the impact of impaired NMDAR function in a mouse model with possible relevance to negative symptoms in schizophrenia.

Neuregulin 1 Transgenic Mice Display Reduced Mismatch Negativity, Contextual Fear Conditioning and Social Interactions

INTRODUCTION Neuregulin-1 (NRG1) is one of susceptibility genes for schizophrenia and plays critical roles in glutamatergic, dopaminergic and GABAergic signaling. Using mutant mice heterozygous for Nrg1 (Nrg1(+/-)) we studied the effects of Nrg1 signaling on behavioral and electrophysiological measures relevant to schizophrenia. EXPERIMENTAL PROCEDURE Behavior of Nrg1(+/-) mice and their wild type littermates was evaluated using pre-pulse inhibition, contextual fear conditioning, novel object recognition, locomotor, and social choice paradigms. Event-related potentials (ERPs) were recorded to assess auditory gating and novel stimulus detection. RESULTS Gating of ERPs was unaffected in Nrg1(+/-) mice, but mismatch negativity in response to novel stimuli was attenuated. The Nrg1(+/-) mice exhibited behavioral deficits in contextual fear conditioning and social interactions, while locomotor activity, pre-pulse inhibition and novel object recognition were not impaired. SUMMARY Nrg1(+/-) mice had impairments in a subset of behavioral and electrophysiological tasks relevant to the negative/cognitive symptom domains of schizophrenia that are thought to be influenced by glutamatergic and dopaminergic neurotransmission. These mice are a valuable tool for studying endophenotypes of schizophrenia, but highlight that single genes cannot account for the complex pathophysiology of the disorder.

Low sociability is associated with reduced size of the corpus callosum in the BALB/cJ inbred mouse strain.

The behavioral manifestations of autism, including reduced sociability (reduced tendency to seek social interaction), may be related to underdevelopment of the corpus callosum (CC). The BALB/cJ inbred mouse strain is a useful model system for testing the relationship between reduced sociability and CC underdevelopment. BALB/cJ mice show low levels of sociability, on average, but substantial intrastrain variability in sociability, as well as striking variability in CC development. This study tested the hypothesis that sociability is positively correlated with CC size within the BALB/cJ inbred strain. 30-day-old BALB/cJ and C57BL/6J mice were tested for sociability towards gonadectomized A/J stimulus mice in a social choice task. The size of the corpus callosum was measured histologically at the midsagittal plane. BALB/cJ mice showed a significant positive correlation between the tendency to sniff the stimulus mouse and size of the CC relative to brain weight. C57BL/6J mice showed consistently high levels of sociability and normal corpus callosum development. These results suggest that abnormal white matter structure is associated with deficits in sociability in BALB/cJ mice. Additional studies are warranted to elucidate the relationship between brain connectivity and sociability in this model system.

Development of home cage social behaviors in BALB/cJ vs. C57BL/6J mice

BALB/cJ and C57BL/6J inbred mouse strains have been proposed as useful models of low and high levels of sociability (tendency to seek social interaction), respectively, based primarily on behaviors of ∼30-day-old mice in the Social Approach Test (SAT). In the SAT, approach and sniffing behaviors of a test mouse toward an unfamiliar stimulus mouse are measured in a novel environment. However, it is unclear whether such results generalize to a familiar environment with a familiar social partner, such as with a littermate in a home cage environment. We hypothesized that C57BL/6J mice would show higher levels of social behaviors than BALB/cJ mice in the home cage environment, particularly at 30 days-of-age. We measured active and passive social behaviors in home cages by pairs of BALB/cJ or C57BL/6J littermates at ages 30, 41, and 69 days. The strains did not differ robustly in their active social behaviors. C57BL/6J mice were more passively social than BALB/cJ mice at 30 days, and C57BL/6J levels of passive social behaviors declined to BALB/cJ levels by 69 days. The differences in passive social behaviors at 30 days-of-age were primarily attributable to differences in huddling. These results indicate that different test conditions (SAT conditions vs. home cage conditions) elicit strain differences in distinct types of behaviors (approach/sniffing vs. huddling behaviors, respectively). Assessment of the more naturalistic social interactions in the familiar home cage environment with a familiar littermate will provide a useful component of a comprehensive assessment of social behaviors in mouse models relevant to autism.

GENETIC DISSECTION OF INTERMALE AGGRESSIVE BEHAVIOR IN BALB/cJ AND A/J MICE

Aggressive behaviors are disabling, treatment refractory, and sometimes lethal symptoms of several neuropsychiatric disorders. However, currently available treatments for patients are inadequate, and the underlying genetics and neurobiology of aggression is only beginning to be elucidated. Inbred mouse strains are useful for identifying genomic regions, and ultimately the relevant gene variants (alleles) in these regions, that affect mammalian aggressive behaviors, which, in turn, may help to identify neurobiological pathways that mediate aggression. The BALB/cJ inbred mouse strain exhibits relatively high levels of intermale aggressive behaviors and shows multiple brain and behavioral phenotypes relevant to neuropsychiatric syndromes associated with aggression. The A/J strain shows very low levels of aggression. We hypothesized that a cross between BALB/cJ and A/J inbred strains would reveal genomic loci that influence the tendency to initiate intermale aggressive behavior. To identify such loci, we conducted a genomewide scan in an F2 population of 660 male mice bred from BALB/cJ and A/J inbred mouse strains. Three significant loci on chromosomes 5, 10 and 15 that influence aggression were identified. The chromosome 5 and 15 loci are completely novel, and the chromosome 10 locus overlaps an aggression locus mapped in our previous study that used NZB/B1NJ and A/J as progenitor strains. Haplotype analysis of BALB/cJ, NZB/B1NJ and A/J strains showed three positional candidate genes in the chromosome 10 locus. Future studies involving fine genetic mapping of these loci as well as additional candidate gene analysis may lead to an improved biological understanding of mammalian aggressive behaviors.

Mouse Model of OPRM1 (A118G) Polymorphism Increases Sociability and Dominance and Confers Resilience to Social Defeat

A single nucleotide polymorphism (SNP) in the human μ-opioid receptor gene (OPRM1 A118G) has been widely studied for its association in drug addiction, pain sensitivity, and, more recently, social behavior. The endogenous opioid system has been shown to regulate social distress and reward in a variety of animal models. However, mechanisms underlying the associations between the OPRM1 A118G SNP and these behaviors have not been clarified. We used a mouse model possessing the human equivalent nucleotide/amino acid substitution to study social affiliation and social defeat behaviors. In mice with the Oprm1 A112G SNP, we demonstrate that the G allele is associated with an increase in home-cage dominance and increased motivation for nonaggressive social interactions, similar to what is reported in human populations. When challenged by a resident aggressor, G-allele carriers expressed less submissive behavior and exhibited resilience to social defeat, demonstrated by a lack of subsequent social avoidance and reductions in anhedonia as measured by intracranial self-stimulation. Protection from social defeat in G-allele carriers was associated with a greater induction of c-fos in a resilience circuit comprising the nucleus accumbens and periaqueductal gray. These findings led us to test the role of endogenous opioids in the A112G mice. We demonstrate that the increase in social affiliation in G carriers is blocked by pretreatment with naloxone. Together, these data suggest a mechanism involving altered hedonic state and neural activation as well as altered endogenous opioid tone in the differential response to aversive and rewarding social stimuli in G-allele carriers.

Sociability Deficits and Altered Amygdala Circuits in Mice Lacking Pcdh10, an Autism Associated Gene

BACKGROUND Behavioral symptoms in individuals with autism spectrum disorder (ASD) have been attributed to abnormal neuronal connectivity, but the molecular bases of these behavioral and brain phenotypes are largely unknown. Human genetic studies have implicated PCDH10, a member of the δ2 subfamily of nonclustered protocadherin genes, in ASD. PCDH10 expression is enriched in the basolateral amygdala, a brain region implicated in the social deficits of ASD. Previous reports indicate that Pcdh10 plays a role in axon outgrowth and glutamatergic synapse elimination, but its roles in social behaviors and amygdala neuronal connectivity are unknown. We hypothesized that haploinsufficiency of Pcdh10 would reduce social approach behavior and alter the structure and function of amygdala circuits. METHODS Mice lacking one copy of Pcdh10 (Pcdh10+/-) and wild-type littermates were assessed for social approach and other behaviors. The lateral/basolateral amygdala was assessed for dendritic spine number and morphology, and amygdala circuit function was studied using voltage-sensitive dye imaging. Expression of Pcdh10 and N-methyl-D-aspartate receptor (NMDAR) subunits was assessed in postsynaptic density fractions of the amygdala. RESULTS Male Pcdh10+/- mice have reduced social approach behavior, as well as impaired gamma synchronization, abnormal spine morphology, and reduced levels of NMDAR subunits in the amygdala. Social approach deficits in Pcdh10+/- male mice were rescued with acute treatment with the NMDAR partial agonist d-cycloserine. CONCLUSIONS Our studies reveal that male Pcdh10+/- mice have synaptic and behavioral deficits, and establish Pcdh10+/- mice as a novel genetic model for investigating neural circuitry and behavioral changes relevant to ASD.

Association between sociability and diffusion tensor imaging in BALB/cJ mice.

The purpose of this study was to use high‐resolution diffusion tensor imaging (DTI) to investigate the association between DTI metrics and sociability in BALB/c inbred mice. The sociability of prepubescent (30‐day‐old) BALB/cJ mice was operationally defined as the time that the mice spent sniffing a stimulus mouse in a social choice test. High‐resolution ex vivo DTI data on 12 BALB/cJ mouse brains were acquired using a 9.4‐T vertical‐bore magnet. Regression analysis was conducted to investigate the association between DTI metrics and sociability. Significant positive regression (p < 0.001) between social sniffing time and fractional anisotropy was found in 10 regions located in the thalamic nuclei, zona incerta/substantia nigra, visual/orbital/somatosensory cortices and entorhinal cortex. In addition, significant negative regression (p < 0.001) between social sniffing time and mean diffusivity was found in five areas located in the sensory cortex, motor cortex, external capsule and amygdaloid region. In all regions showing significant regression with either the mean diffusivity or fractional anisotropy, the tertiary eigenvalue correlated negatively with the social sniffing time. This study demonstrates the feasibility of using DTI to detect brain regions associated with sociability in a mouse model system. Copyright

Activation of basolateral amygdala in juvenile C57BL/6J mice during social approach behavior.

There is a strong need to better understand the neurobiology of juvenile sociability (tendency to seek social interaction), a phenotype of central relevance to autism spectrum disorders (ASD). Although numerous genetic mouse models of ASD showing reduced sociability have been reported, and certain brain regions, such as the amygdala, have been implicated in sociability, there has been little emphasis on delineating brain structures and circuits activated during social interactions in the critical juvenile period of the mouse strain that serves as the most common genetic background for these models-the highly sociable C57BL/6J (B6) strain. We measured expression of the immediate early genes Fos and Egr-1 to map activation of brain regions following the Social Approach Test (SAT) in juvenile male B6 mice. We hypothesized that juvenile B6 mice would show activation of the amygdala during social interactions. The basolateral amygdala (BLA) was activated by social exposure in highly sociable, 4-week-old B6 mice. In light of these data, and the many lines of evidence indicating alteration of amygdala circuits in human ASD, future studies are warranted to assess structural and functional alterations in the BLA, particularly at BLA synapses, in various mouse models of ASD.

Protocadherin 10 alters γ oscillations, amino acid levels, and their coupling; baclofen partially restores these oscillatory deficits

Approximately one in 45 children have been diagnosed with Autism Spectrum Disorder (ASD), which is characterized by social/communication impairments. Recent studies have linked a subset of familial ASD to mutations in the Protocadherin 10 (Pcdh10) gene. Additionally, Pcdh10s expression pattern, as well as its known role within protein networks, implicates the gene in ASD. Subsequently, the neurobiology of mice heterozygous for Pcdh10 (Pcdh10+/-) has been investigated as a proxy for ASD. Male Pcdh10+/- mice have demonstrated sex-specific deficits in social behavior, recapitulating the gender bias observed in ASD. Furthermore, in vitro slice preparations of these Pcdh10+/- mice demonstrate selective decreases to high frequency electrophysiological responses, mimicking clinical observations. The direct in vivo ramifications of such decreased in vitro high frequency responses are unclear. As such, Pcdh10+/- mice and their wild-type (WT) littermates underwent in vivo electrocorticography (ECoG), as well as ex vivo amino acid concentration quantification using High Performance Liquid Chromatography (HPLC). Similar to the previously observed reductions to in vitro high frequency electrophysiological responses in Pcdh10+/- mice, male Pcdh10+/- mice exhibited reduced gamma-band (30-80Hz), but not lower frequency (10 and 20Hz), auditory steady state responses (ASSR). In addition, male Pcdh10+/- mice exhibited decreased signal-to-noise-ratio (SNR) for high gamma-band (60-100Hz) activity. These gamma-band perturbations for both ASSR and SNR were not observed in females. Administration of a GABAB agonist remediated these electrophysiological alterations among male Pcdh10+/-mice. Pcdh10+/- mice demonstrated increased concentrations of GABA and glutamine. Of note, a correlation of auditory gamma-band responses with underlying GABA concentrations was observed in WT mice. This correlation was not present in Pcdh10+/- mice. This study demonstrates the role of Pcdh10 in the regulation of excitatory-inhibitory balance as a function of GABA in ASD.