Autism is a developmental disorder and affected individuals have alterations in brain function disrupting their ability to perform particular motor and cognitive tasks. The main long term objective of this project is to determine in one major affected brain region, the cerebellum, how changes in connectivity at the synaptic level are disturbed based on errors in the early development of key afferent projections and target neurons. Glutamatergic mossy fibers, olivocerebellar climbing fibers and parallel fibers all synapse on key cerebellar cortical neurons, some of which show neuropathological changes in autism. The major objective of this study focuses on key glutamatergic transporters and receptors in two functionally distinct parts of the cerebellar cortex, the Crus II area in the posterior lobe recently implicated in cognitively-based functions and vermis lobule VI in the anterior lobe important for sensorimotor-based roles. Quantitative glutamatergic measures will be related to scores from the Autism Diagnostic Interview-Revised (ADI-R) for reciprocal social interaction, communication, and stereotyped and repetitive motor behavior domains. To accomplish these objectives, four specific aims are proposed in ten adult (age 16-30 years old) autistic post-mortem cases and ten age-, gender and postmortem interval-matched controls. The first specific aim, to be completed in the first year, uses histological ligand binding to determine the number and ligand binding affinity of key ionotropic (AMPAR, NMDAR) and metabotropic glutamate receptors (mGluR1) in the three laminae of Crus II and lobule VI, targets for cerebellar afferents. The second specific aim, to be completed in the first year, uses in situ hybridization histochemistry in the same cerebellar regions to quantify the mRNA levels of three types of postsynaptic glutamate receptor subtypes (AMPAR1, NMDAR1 and mGluR1). The third specific aim with trials in the first year and completed in the second year, quantifies the densities of key glutamate transporters at terminals ending on PCs (Le., excitatory amino acid transporter type 4, EAAT4;on membranes of Bergmann glia that predominately wrap around climbing synapses (EAA T1, L-glutamate/L-aspartate transporter, GLAST);and a glutamate receptor (GluRo2) that is important for induction of long-term depression (LTD) and is predominately localized to parallel fiber synapses on PCs. Co-localization of GluRo2 with PSD93 will determine the percentage of postsynaptic density and co-localization of EAAT4 with synaptophysin will determine the percentage of presynaptic density. The final specific aim, to be completed in the second year, takes a translational approach and evaluates the relationships of glutamate receptors, transporters and synaptic density to severity of social-communicative and repetitive behavior symptoms as measured by the Autism Diagnostic Interview - Revised (ADI-R).
This aim examines whether significant changes in key excitatory glutamate component(s) in the two functionally distinct cerebellar regions relate differentially to autism symptom type and severity in the clinical cases. Data from these studies over the two year project will help delineate core neural substrates in autistic individuals and should help guide geneticists in their search for possible candidate genes in the disorder. Furthermore, understanding the cellular and molecular mechanisms within this model region could contribute to our understanding of current pharmacotherapeutic treatments of autistic individuals and may suggest novel treatments targeted to neural substrates.
This project has high relevance to understanding the core neural substrates within the glutamate system that may contribute to the autism phenotype and how alterations at the synaptic level, within functionally different cerebellar areas, can be related to sensorimotor and/or cognitive behavior. Quantitative data from the proposed aims will be correlated with available data from the ADI/ADI-R to draw a more comprehensive translational link between the neuropathological findings with measures of social interaction, communication, and restricted and repetitive behaviors. Finally, by identifying specific glutamate receptor subtypes and underlying mRNA that are highly impacted in autism, the study should provide important information regarding improved drug treatment to target affected substrates instead of limiting treatment to behavioral symptoms.
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