TREK-1: Identification of a physiological role and identification of novel binding partners.

Abstract

The emergence of two pore potassium channels as a highly evolved and involved potassium conductance in a large variety of tissues and cell types has provided a new family of targets for ion channel research. The TREK-1 channel has been extensively researched and is implicated in a number of physiological roles throughout the nervous system, the heart, and many smooth muscle tissues. In this thesis we describe the emergence of TREK-1 as a channel which may be involved in the generation of the nitrergic inhibitory junction potential (IJP), a physiological event in the gastrointestinal tract, of which the underlying ionic conductance has long been a point of controversy. Using a series of Ca2+ store releasing compounds, we suggest TREK-1 as a new ionic conductance candidate involved in generation of the nitrergic IJP in murine colon, and that the nitrergic IJPs, may not be as dependent on Ca2+ stores as previously suggested. The re-emergence of a K+ conductance as a candidate for the nitrergic IJP revisits an original paradigm which was investigated previously. Original thought pointed to activation of a K+ conductance as being fundamentally responsible for the hyperpolarization response during nitrergic IJPs; however the findings that the nitrergic IJPs were insensitive to "classical" channel blockers pioneered the idea that other channel conductances may be involved. The relatively novel emergence of TREK-1 in the K+ channel family, and the discovery that TREK-1 is a K+ conductance which is insensitive to classical channel blockers provided us with a new K+ channel candidate with which to revisit the underlying conductance responsible for the nitrergic IJP. This thesis also reveals a number of intracellular proteins which may be involved in TREK-1 binding and modulation. Dynein and Filamin A are intracellular proteins which have both been previously reported to bind to and modulate the trafficking and or surface expression levels of ion channels. We discovered these proteins binding to the C terminus of TREK-1 through a yeast 2-hybrid screen and investigated if these proteins bind to TREK-1 in vitro.