Molecular Dissection of IA Channels in Cortical Pyramidal Neurons

The rapidly activating and inactivating voltage-gated K<super>+</super>: Kv) current, I <SUB>A</SUB>, is broadly expressed in neurons and is a key regulator of action potential repolarization, repetitive firing, back propagation: into dendrites) of action potentials, and responses to synaptic inputs. Interestingly, results from previous studies on a number of neuronal cell types, including hippocampal, cortical and spinal neurons, suggest that macroscopic I _A is composed of multiple components and that each component is likely encoded by distinct Kv channel &#945;-subunits. The goals of the experiments presented here were to test this hypothesis and to determine the molecular identities of the Kv channel &#945;-subunits that generate I _A in cortical pyramidal neurons. Combining genetic disruption of individual Kv &#945;-subunit genes with pharmacological approaches to block Kv currents selectively, the experiments here revealed that Kv1.4, Kv4.2 and Kv4.3 &#945;-subunits encode distinct components of I _A that together underlie the macroscopic I _A in mouse cortical pyramidal neurons. Recordings from neurons lacking both Kv4.2 and Kv4.3: Kv4.2-/-/Kv4.3-/-) revealed that, although Kv1.4 encodes a minor component of I _A, the Kv1.4-encoded current was found in all the Kv4.2-/-/Kv4.3-/- cortical pyramidal neurons examined. Of the cortical pyramidal neurons lacking both Kv4.2 and Kv1.4, 90% expressed a Kv4.3-encoded I _A larger in amplitude than the Kv1.4-encoded component. The experimental findings also demonstrate that the targeted deletion of the individual Kv &#945;-subunits encoding components of I _A results in electrical remodeling that is Kv &#945;-subunit specific. Accumulating, evidence indicates that native neuronal Kv4 channels function in macromolecular protein complexes that contain accessory subunits and other regulatory molecules. The K<super>+</super> Channel Interacting Proteins: KChIP) are among the identified Kv4 channel accessory subunits and are thought to be important for the formation and function of Kv4 channel complexes in neurons. Molecular genetic, biochemical and electrophysiological experiments were undertaken to examine directly the roles of KChIPs in the formation and function of Kv4-encoded I _A channels. The results of the experiments presented here revealed that KChIP2, KChIP3 and KChIP4 are critical for the formation of functional Kv4-encoded channels in cortical pyramidal neurons. All three KChIPs were found to be expressed in and to co-immunoprecipitate with Kv4.2 from posterior: visual) cortex. Further, the mean density of I _A in cortical pyramidal neurons was reduced by genetic disruption of KChIP3 expression. In cortical samples from mice lacking KChIP2 or KChIP3 the levels of KChIP3 and 4 or KChIP2 and 4 were increased, respectively. In neurons expressing constructs to mediate RNA inference induced reductions in the expression of KChIP2, 3 and 4 I _A densities were markedly reduced and Kv current remodeling was evident.

Name thesis:degree_name

Doctor of Philosophy (PhD)

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Biology and Biomedical Sciences: Neurosciences

Year dc:date.available

2012

Author dc:creator

• Norris, Aaron

Contributors dc:contributor

• Jeanne Nerbonne

• Neurosciences

Language dc:language

English (en)

DOI

https://doi.org/10.7936/K7QR4V40

OAI identifier oai:identifier

oai:openscholarship.wustl.edu:etd-1626