AMPA receptors (AMPAR) mediate the majority of fast excitatory synaptic transmission in the central nervous system. Trafficking of AMPARs in and out of synapses is a highly dynamic process and regulation of this trafficking plays a critical role in synaptic plasticity and learning and memory. However, the direct observation of dynamic AMPAR trafficking in live animals during synaptic plasticity has not been accomplished. To examine plasticity in vivo, we will investigate AMPAR dynamics in live animals undergoing various physiologically relevant sensory experiences using two-photon microscopy. We have been able to express pHluorin tagged AMPARs in layer ll/lll pyramidal neurons in the mouse barrel cortex using in utero electroporation. Following electroporation we make a cranial window over the barrel cortex region, map out barrel columns in the cortex using intrinsic optical imaging and then image pHluorin tagged AMPARs with two-photon microscopy. In preliminary studies we have investigated AMPAR dynamics under acute whisker deflection and chronic whisker trimming and regrowth conditions. We have found that both acute whisker deflection and whisker deprivation and regrowth lead to specific changes in AMPAR synaptic levels. To further understand how AMPARs behave during learning tasks, we will observe AMPAR dynamics in the mouse visual cortex during stimulus-specific response potentiation, a well-studied learning paradigm in the visual system and in other cortical regions. To study the molecular mechanisms underlying the induction and long-term maintenance of plasticity we will investigate the AMPAR subunit dependence and the structural regions of each subunit required for the regulation of AMPAR synaptic trafficking in vivo. In addition, we will examine AMPAR trafficking in vivo in our collection of transgenic and knock out mouse lines in which various key synaptic proteins, such as PSD95, SAP97, GRIP1/2, PICK1 and PKC zeta have been removed or altered. The findings from these experiments will help us identify the essential regulators of AMPAR trafficking in vivo under conditions that elicit synaptic plasticity and learning and elucidate the molecular mechanisms underlying synaptic plasticity in the brain in health and disease.
This research will elucidate basic molecular mechanisms that regulate synaptic transmission and plasticity in the brain but it also has broad relevance for many neurological and psychiatric diseases. Dysfunction of synaptic transmission and plasticity underlies many neurological and psychiatric disorders. This research may therefore reveal novel targets for the development of therapeutic treatments for several brain disorders including schizophrenia autism depression drug addiction and pain
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