Stephen J. Tucker

Adenosine triphosphate (ATP)-sensitive potassium (KATP) channels couple electrical activity to cellular metabolism through their inhibition by intracellular ATP. ATP inhibition of KATP channels varies among tissues and is affected by the metabolic and regulatory state of individual cells, suggesting involvement of endogenous factors. It is reported here that phosphatidylinositol-4, 5-bisphosphate (PIP2) and phosphatidylinositol-4-phosphate (PIP) controlled ATP inhibition of cloned KATP channels (Kir6.2 and SUR1). These phospholipids acted on the Kir6.2 subunit and shifted ATP sensitivity by several orders of magnitude. Receptor-mediated activation of phospholipase C resulted in inhibition of KATP-mediated currents. These results represent a mechanism for control of excitability through phospholipids.

TREK-2 (KCNK10/K2P10), a two-pore domain potassium (K2P) channel, is gated by multiple stimuli such as stretch, fatty acids, and pH and by several drugs. However, the mechanisms that control channel gating are unclear. Here we present crystal structures of the human TREK-2 channel (up to 3.4 angstrom resolution) in two conformations and in complex with norfluoxetine, the active metabolite of fluoxetine (Prozac) and a state-dependent blocker of TREK channels. Norfluoxetine binds within intramembrane fenestrations found in only one of these two conformations. Channel activation by arachidonic acid and mechanical stretch involves conversion between these states through movement of the pore-lining helices. These results provide an explanation for TREK channel mechanosensitivity, regulation by diverse stimuli, and possible off-target effects of the serotonin reuptake inhibitor Prozac.

Biological ion channels are nanoscale transmembrane pores. When water and ions are enclosed within the narrow confines of a sub-nanometer hydrophobic pore, they exhibit behavior not evident from macroscopic descriptions. At this nanoscopic level, the unfavorable interaction between the lining of a hydrophobic pore and water may lead to stochastic liquid-vapor transitions. These transient vapor states are "dewetted", i.e. effectively devoid of water molecules within all or part of the pore, thus leading to an energetic barrier to ion conduction. This process, termed "hydrophobic gating", was first observed in molecular dynamics simulations of model nanopores, where the principles underlying hydrophobic gating (i.e., changes in diameter, polarity, or transmembrane voltage) have now been extensively validated. Computational, structural, and functional studies now indicate that biological ion channels may also exploit hydrophobic gating to regulate ion flow within their pores. Here we review the evidence for this process and propose that this unusual behavior of water represents an increasingly important element in understanding the relationship between ion channel structure and function.

There has been considerable debate as to whether adenosine triphosphate (ATP) is compartmentalized within cells and, in particular, whether the ATP concentration directly beneath the plasma membrane, experienced by membrane proteins, is the same as that of the bulk cytoplasm. This issue has been difficult to address because there is no indicator of cytosolic ATP, such as those available for Ca(2+), capable of resolving the submembrane ATP concentration ([ATP](sm)) in real time within a single cell. We show here that mutant ATP-sensitive K(+) channels can be used to measure [ATP](sm) by comparing the increase in current amplitude on patch excision with the ATP dose-response curve. In Xenopus oocytes, [ATP](sm) was 4.6 +/- 0.3 mm (n = 29) under resting conditions, slightly higher than that measured for the bulk cytoplasm (2.3 mm). In mammalian (COSm6) cells, [ATP](sm) was slightly lower and averaged 1.4 +/- 0.1 mm (n = 66). Metabolic poisoning (10 min of 3 mm azide) produced a significant fall in [ATP](sm) in both types of cells: to 1.2 +/- 0.1 mm (n = 24) in oocytes and 0.8 +/- 0.11 mm for COSm6 cells. We conclude that [ATP](sm) lies in the low millimolar range and that there is no gradient between bulk cytosolic and submembrane [ATP].