Effects of Regional Deletion of Rab3A-interacting Molecule and PTEN on Brain Function




Haws, Michael

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This dissertation describes experiments designed to delete/knockdown molecules from targeted neuronal populations to study brain region-specific behavioral functions. To this end, I utilized two different conditional knockdown techniques to study the role of the presynaptic active zone molecule Rab3A-interacting Molecule (RIM1) and the phosphatase and tensin homologue on chromosome 10 (PTEN). The cre-lox system was used to eliminate RIM1 from the hippocampal dentate gyrus and area CA3, while adeno-associated virus expressing PTEN-directed interference RNA was injected into the basolateral amygdala to knockdown PTEN in local pyramidal neurons. In the case of RIM1, I hypothesized that deletion of RIM1 from the dentate gyrus or from area CA3 would replicate a subset of the learning and memory deficits found in RIM1α-/- mice. Though the conditional RIM1 knockout mice were not completely selective for the dentate gyrus or for area CA3, both conditional knockouts induced a different behavioral abnormality present in RIM1α-/- mice. My results help narrow the potential brain regions involved in key RIM1α-/- mice behavioral aberrations. In the case of PTEN, I hypothesized that deletion of PTEN specifically in the basolateral amygdala (BLA) would cause increased anxiety and neuronal hypertrophy. Knockdown of PTEN in the BLA did not induce anxiogenesis though it did increase soma volume, dendritic caliber, spine size, mushroom:thin spine ratio, and the frequency of spontaneous miniature excitatory post-synaptic currents. These findings are in contrast to previous findings of increased spine density with PTEN knockdown. This difference likely represents the more sensitive techniques employed in the present studies to ascertain dendritic spine type and density. Though PTEN knockdown had synaptic effects I did not observe any behavioral effects. However, limitations in viral knockdown of PTEN transcripts or viral infection rate may be responsible for the lack of effect. Indeed, limitations exist for both the transgenic and viral approaches used which proved to be challenging obstacles to designing experiments, interpreting data and coming to more extensive concrete conclusions. Transgene expression is often not as selective as desired. Virus injections may not localize to target region or may not infect enough neurons. Understanding and characterizing these and other limitations is vital.

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