Elucidation of auto-regulatory mechanisms in voltage-gated potassium channels
Abstract
Inactivation is the auto-regulatory process of A-type voltage-gated K + (Kv) channels. Kv channels may inactivate directly from the open state (OSI), or from an activated closed state (CSI). This work focuses on the neuronal A-type Kv4 channel, which regulates the firing and shape of the action potential. Kv4 channels might undergo CSI, but this has not been conclusively established; and the underlying molecular mechanism has remained elusive. It is often necessary to apply several electrophysiological, pharmacological and computational approaches to determine the inactivation pathways of ion channels. However, intrinsic hurdles preclude their standardized application. Here, we have conclusively answered the following questions: 1) what is the pathway of inactivation of Kv4 channels? 2) what are the molecular underpinnings of this pathway? To answer the first question, we implemented a simple method to analyze macroscopic currents and deduce the pathways of inactivation of native and recombinant A-type Kv4 channels - including auxiliary subunits Kv channel interacting protein (KChIP) and dipeptidyl peptidase-like protein (DPP). This approach applies two complementary pulse protocols: either a single pulse or a variable duration conditioning pulse followed by a test pulse. The results unambiguously determined that Kv4 channels undergo preferential CSI. To answer the second question, we hypothesized that the intracellular activation gate also acts as the inactivation gate. To test this hypothesis, we investigated heterologously expressed Kv4.1 ternary channels upon intracellular exposure to quaternary ammonium (QA) derivative blockers as the channels open and rapidly inactivate. The results show that the activation gate closes during inactivation, thereby trapping the QA derivative blocking the pore; subsequent activation (opening) of the channels results in use-dependent QA derivative escape. This is compelling evidence for the dual activation/inactivation role of the Kv4 intracellular gate. Finally, to show how CSI and CSI+OSI can determine spiking properties in the nervous system, we implemented a computational model of the hippocampal CA1 neuron. CSI and the hybrid CSI+OSI differentially affect impulse trains, back-propagating impulses and impulse properties. Taken together, this work has solved the pathway and molecular mechanism of CSI in ternary Kv4 channels. This knowledge will help develop new interventions that may help treat epilepsy, pain syndromes and other neurological disorders.
Subject Area
Neurosciences|Physiology|Biophysics
Recommended Citation
Fineberg, Jeffrey D, "Elucidation of auto-regulatory mechanisms in voltage-gated potassium channels" (2013). ProQuest ETD Collection - Thomas Jefferson University. AAI3605411.
https://jdc.jefferson.edu/dissertations/AAI3605411
