Relevant to our research interests in animal models of neuropsychiatric diseases, our Laboratory of Behavioral Neuroscience investigates the behavioral phenotypes of transgenic and knockout mice with mutations in genes expressed in brain pathways involved in neuropsychiatric disorders. We developed and refined a multi-tiered strategy for mouse behavioral phenotyping that is widely used by the international research community. We collaborate with a large number of molecular genetics laboratories, e.g. behavioral testing of COMT knockout mice with Danny Weinberger, NIMH, and oxytocin knockout mice with Scott Young, NIMH in FY2008.? ? Dr. Crawley began a new initiative in 2002 to develop mouse behavioral tasks relevant to the symptoms of autism. She learned about the defining behavioral features of autism through a collaboration with Dr. Joseph Piven at the University of North Carolina Center for Autism Research, through interactions with Dr. Susan Swedo's autism research program at NIMH, through participation in international autism conferences and workshops, and from observing autistic children in classrooms and in videotaped interviews. During 2003-2007, Dr. Crawley contributed to the UNC STAART project at UNC, helping Dr. Piven and Research Assistant Professor Dr. Sheryl Moy to set up a mouse behavioral phenotyping laboratory, and participating in a comprehensive mouse strain distribution study. One particularly interesting strain that emerged from these expeirments is BTBR T+tf/J (BTBR), which displayed low sociability in our automated three-chambered social approach task.? ? During FY2008, our Laboratory of Behavioral Neuroscience pursued the phenotypes of BTBR and the biological mechanisms underlying these phenotypes. Lack of sociability was confirmed in multiple independently bred cohorts, on multiple social tasks including juvenile play, social approach, adult reciprocal social interactions, and social transmission of food preference (McFarlane et al., 2008), relevant to the first diagnostic symptom of autism, abnormal social interactions. High levels of repetitive self-grooming was detected (Yang et al., 2007; McFarlane et al., 2008), relevant to the third diagnostic symptom of autism, repetitive behaviors. These phenotypes were identical when mice were raised on a conventional circadian cycle and tested in the light, versus on a reverse light cycle and tested in the dark phase of the circadian cycle, when mouse social behaviors are maximal (McFarlane et al., 2008; Yang et al., in press). Postdoctoral fellow Mu Yang confirmed that the social deficit in BTBR mice was not caused by maternal care factors, using cross-fostering between BTBR and C57BL/6J (Yang et al., 2007). Dr. Yang and student intern Andrew Clarke tested the hypothesis that the corpus callosum deficit in BTBR is responsible for their phenotypes (Yang et al., submitted). Postdoctoral fellow Maria Luisa Scattoni and postbaccalaurate Shruti Gandhy detected unusual pattterns of vocalizations in BTBR as compared to C57BL/6J, FVB/NJ, and 129X1/SvJ inbred strains (Scattoni et al., in press). ? ? Current experiments are addressing the genes and biological mechanisms responsible for the autism-like phenotypes in BTBR. A new collaboration was initiated with Dr. Elliott Sherr at the University of California San Francisco to conduct a quantitative trait loci linkage analysis of the BTBR x B6 cross, to discover genes in the BTBR background that correlate with autism-like phenotypes. F2 mice are being tested behaviorally by Dr. Yang, new postbaccalaureate Mike Weber, and new student intern Kayla Perry. Drs. Mike Tyszka and Ralph Adolphs have joined this collaboration to conduct tractography of the neuroanatomy of BTBR. Collaborators Dr. Valerie Bolivar and graduate student Gretchen Kusek at the Wadsworth Center for Genome Research in Troy, NY had queried the JAX database for unusual single nucleotide polymorphisms (SNPs) in the coding regions of genes in the BTBR background. Three unique SNPs were detected in nonsynonymous coding region of the gene for Kmo (McFarlane et al., 2008), kynurenine-3-hydroxylase, an enzyme that regulate the metabolism of kynurenic acid in the brain. Kynurenic acid mediates neuronal survival and dendritic spine formation. These findings provide a proof of principle for using social deficits in inbred strains of mice to generate new hypotheses about unpredicted genes that could mediate symptoms of autism. We are pursuing the biological significance of the Kmo polymorphism in BTBR with collaborator Dr. Robert Schwarcz at the Maryland Psychiatric Research Center (MPRC). Dr. Schwarcz is an expert on metabolic enzymes for kynurenic acid, and the role of kynurenic acid in psychiatric disorders. Stress-related hormones that might cause repetitive self-grooming are being assayed in BTBR by Dr. Jim Koenig at MPRC. Oxytocin receptors and peptide are being assayed by Dr. Scott Young and his postdoctoral fellow Abbe Macbeth, and by Dr. Koenig. Our Senior Laboratory Manager Dr. Jill Silverman is conducting parts of these experiments and coordinating the collaborations with Drs. Schwarz, Koenig, and Young. ? ? A second approach toward identifying the genes mediating the diagnostic symptoms of autism is to evaluate the behaviors of lines of mice with experimentally targeted mutations in candidate genes for autism. Lines of mice with mutations in two candidate genes for autism were tested this year. Neuroligins, neurexins, and shanks are synaptic development genes for which mutations have been detected in a small number of autistic individuals. Postdoctoral fellow Kathy Chadman and student intern Sarah Boltuck conducted comprehensive behavioral phenotyping of neuroligin-3 knockin mice generated by collaborator Nat Heintz at Rockefeller University (Chadman et al., in press). Dr. Silverman, new postbaccalaureate Charlotte Barkan, and new student intern Seda Tolu are conducting comprehensive behavioral phenotyping of shank-1 knockout mice generated by collaborator Morgan Sheng at Massachusetts Institute of Technology. Mice with mutations in other synaptic candidate genes for autism are under discussion for importation in FY2009.? ? The third component of our mouse models of autism project is the translational evaluation of proposed treatments. The robust and highly replicated social deficits and repetitive self-grooming in BTBR provide a good model system for testing the ability of drugs and behavioral treatments to reverse and prevent autism-like symptoms. Dr. Silverman, Charlotte Barkan, and Seda Tolu are completing a dose-response curve for risperidone. Risperdal is the first drug approved by the FDA for autism, primarily for the irritability symptom. Drugs planned for the next BTBR challenges include fluoxetine and oxytocin, which have been reported to reverse some symptoms of autism. Compounds that enhance synaptic functions are planned for subsequent challenges, including the mGluR5 antagonist MPEP and the ampakine CX546, both of which reversed phenotypes in Fragile X mice. Comorbidity between Fragile X and autism is high, and may involve some common biological mechanisms. Behavioral interventions are being tested by Dr. Yang, Mike Weber, and Kayla Perry, including a """"""""sibling"""""""" intervention involving home cage social interactions between BTBR juveniles and B6 juveniles.? ? Taken together, these multidisciplinary analyses of multiple mouse models of autism are likely to move the field forward, by testing hypotheses about the causes of autism, and identifying useful treatments.
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