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Activity-dependent regulation of -̆amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors in rat neocortex

University of British Columbia

The study of α-amino-3-hydroxy-5-methyl-isoxazole propionic acid (AMPA) receptors is of great interest given that these receptors mediate most fast excitatory synaptic neurotransmission in brain and are involved in neuroplastic phenomena such as long-term potentiation and long-term depression. Understanding the molecular mechanisms involved in the regulation of AMPA receptors may therefore provide insight into many aspects of neuronal function, including normal synaptic transmission and synaptic neuroplasticity. To assess activity-dependent regulation of cortical AMPA receptors, radioligand binding methods employing the competitive AMPA receptor antagonist[³H]-6-cyano-7- nitroquinoxaline-2,3-dione (CNQX) were used to study the effects of various regulatory stimuli on the agonist binding site of AMPA receptors in rat cortical slices. AMPA receptor regulation was studied in response to a variety of stimuli, including agonist (AM PA), pharmacological depolarization (veratridine + glutamate), as well as the phosphorylating enzymes calcium (Ca²+) /calmodulin-dependent kinase II (CaMKH) and protein kinase A (PKA). Treatment with AMPA or veratridine led to approximately 20% decreases in [³H]-CNQX binding. Similar decreases in[³H]-CNQX binding were seen following treatment with CaMKII (∼35%) and PKA (∼30%). The effects of AMPA and veratridine could be blocked by inhibitors of CaMKll and PKA, suggesting that phosphorylation reactions are involved in AMPA receptor regulation by AMPA and veratridine. Moreover, loperamide, a non specific inhibitor of voltage-gated Ca²⁺ channels was able to inhibit the AMPA and veratridine-induced regulation of AMPA receptors. These results suggested that AMPA and veratridine may result in the activation of voltage gated Ca²⁺ channels, in turn leading to changes in[³H]-CNQX binding through the activation of CaMKII and/or PKA. Ca²⁺ alone was able to decrease[³H]- CNQX binding over a concentration range of 0.1 to 1 mM, an effect which could be blocked by specific inhibitors of CaMKII or PKA. These data indicate that AMPA and veratridine, agents intended to mimic aspects of synaptic transmission, lead to the regulation of AMPA receptors via a Ca²⁺ influx through voltage-gated Ca²⁺ channels and the activation of specific phosphorylating enzymes. These results provide for a novel mechanism ot AMPA receptor regulation and may establish the framework for a clearer understanding of the modulation of synaptic activity in normal conditions, following modifications of synaptic strength, and in some forms of neuropathology.

Thesis/Dissertation

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2009-06-05

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http://hdl.handle.net/2429/8806

Neuroscience

University of British Columbia

UBCV

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