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The TRP superfamily represents a highly diverse group of cation-permeable ion channels related to the product of the (ransient eceptor otential) gene. The cloning and characterization of members of this cation channel family has experienced a remarkable growth during the last decade, uncovering a wealth of information concerning the role of TRP channels in a variety of cell types, tissues, and species. Initially, TRP channels were mainly considered as phospholipase C (PLC)-dependent and/or store-operated Ca-permeable cation channels. More recent research has highlighted the sensitivity of TRP channels to a broad array of chemical and physical stimuli, allowing them to function as dedicated biological sensors involved in processes ranging from vision to taste, tactile sensation, and hearing. Moreover, the tailored selectivity of certain TRP channels enables them to play key roles in the cellular uptake and/or transepithelial transport of Ca, Mg, and trace metal ions. In this chapter we give a brief overview of the TRP channel superfamily followed by a survey of current knowledge concerning their structure and activation mechanisms.

Ion channels in epithelial cells serve to move ions, and in some cases fluid, between compartments of the body. This function of the transfer of is fundamentally different from that of the transfer of , which is the main job of most channels in excitable cells. Nevertheless the basic construction of the channels is similar in many respects in the two tissue types. This chapter reviews the nature of channels in epithelia and discusses how their functions have evolved to accomplish the basic tasks for which they are responsible. I will focus on three channel types: epithelial Na channels, inward-rectifier K channels, and CFTR Cl channels.

The kinetics of an assembly of charged particles such as electrons, ions, or colloids, particularly when subjected to externally applied electric fields, has been of interest for many years and in many disciplines. In applied physics and electrical engineering, the motion of electrons and holes through semiconductor materials under the influence of an applied voltage plays an essential role in the function of modern electronic components such as transistors, diodes, and infrared lasers (Peyghambarian et al., 1993). Electrochemistry deals in large part with the motion of simple inorganic ions (e.g., Na, Cl) in electrolytic solutions and how this motion is influenced when electrodes are employed to generate an electric potential drop across the solution or a membrane interface (Bockris and Reddy, 1998). Larger macroions such as charged polystyrene spheres (radius 0.1–1 micron) can also be manipulated using applied electric fields (Ise and Yoshida, 1996). Many processes in molecular biology, from self-assembly of DNA strands into bundles (Wissenburg et al., 1995) to enzymeligand docking (Gilson et al., 1994), are steered by electrostatic forces between biological macroions which are mediated by the response of simple salt ions in the solution.

Understanding how physiological ion channels simultaneously exhibit the apparently contradictory properties of high throughput and great discrimination is a long-standing theoretical problem. These nanodevices all operate on the same basic principle: ions, solvated by bulk water, lose a significant part of their hydration shell as they pass through a constriction where a chemical selection process occurs (Hille, 2001). High throughput requires that the chosen ion faces no significant energy barrier, which would forbid its entry. On first blush, it seems that falling into a deep well is also forbidden, since that would apparently trap it in the channel and block further passage. While generally true, some channels function in multi-ion mode, so that they are permanently ion-occupied; permeation then occurs with the entry of a second (or third) ion, repelling the prior occupant and leading to conduction. In all instances, high selectivity requires that there is a mechanism by which all other physiologically prevalent ions face significant energetic discrimination.

All living cells are surrounded by a thin membrane, composed of two layers of phospholipid molecules, called the lipid bilayer. This thin membrane effectively confines some ions and molecules inside and exchanges others with outside and acts as a hydrophobic, low dielectric barrier to hydrophilic molecules. Because of a large difference between the dielectric constants of the membrane and electrolyte solutions, no charged particles, such as Na, K, and Cl ions, can jump across the membrane. The amount of energy needed to transport one monovalent ion, in either direction across the membrane, known as the , is enormously high. For a living cell to function, however, the proper ionic gradient has to be maintained, and ions at times must move across the membrane to maintain the potential difference across the membrane and to generate synaptic and action potentials. The delicate tasks of regulating the transport of ions across the membrane are carried out by biological nanotubes called “ion channels,” water-filled conduits inserted across the cell membrane through which ions can freely move in and out when the gates are open. These ion channels can be viewed as biological sub-nanotubes, the typical pore diameters of which are ~10 m or 10 Å.

Ion channels are proteins that form pores of nanoscopic dimensions in cell membranes. As a consequence of advance in protein crystallography we now know the three-dimensional structures of a number of ion channels. However, X-ray diffraction techniques yield an essentially static (time- and space-averaged) structure of an ion channel, in an environment often somewhat distantly related to that which the protein experiences when in a cell membrane. Thus, additional techniques are required to fully understand the relationship between channel structure and function. Potassium (K) channels (Yellen, 2002) provide an opportunity to explore the relationship between membrane protein structure, , and function. Furthermore, K channels are of considerable physiological and biomedical interest. They regulate K ion flux across cell membranes. K channel regulation is accomplished by a conformational change that allows the protein to switch between two alternative (closed vs. open) conformations, a process known as . Gating is an inherently dynamic process that cannot be fully characterized by static structures alone.

The electrical activity of living cells can be monitored in various ways, but for the study of ion channels and the drugs that affect them, the patch-clamp techniques are the most sensitive. In this chapter the principles of patch-clamp recording are reviewed, and recent developments in microfabricated patch-clamp electrodes are described.Technical challenges and prospects for the future are discussed.

A biosensor device based on the ion channel gramicidin A (gA) incorporated into a bilayer membrane is described. This generic immunosensing device utilizes gA coupled to an antibody and assembled in a lipid membrane. The membrane is chemically tethered to a gold electrode, which reports on changes in the ionic conduction of the lipid bilayer. Binding of a target molecule in the bathing solution to the antibody causes the gramicidin channels to switch from predominantly conducting dimers to predominantly nonconducting monomers. Conventional a.c. impedance spectroscopy between the gold and a counter electrode in the bathing solution is used to measure changes in the ionic conductivity of the membrane. This approach permits the quantitative detection of a range of target species, including bacteria, proteins, toxins, DNA sequences, and drug molecules.

The measurement of ionic currents flowing through single channels in cell membranes has been made possible by the giga-seal patch-clamp technique (Neher and Sakmann, 1976; Hamill et al., 1981). A tight seal between the rim of the electrode tip and the cell membrane drastically reduces the leakage current and extraneous background noise, enabling the resolution of the discrete changes in conductance that occur when single channels open or close. Although the noise from a small patch is much less than that from a whole-cell membrane, signals of interest are often obscured by the noise. Even if the signal frequently emerges from the noise, low-amplitude events such as small subconductance states can remain below the noise level and there may be little evidence of their presence. It is desirable, therefore, to have a method to measure and characterize not only relatively large ionic currents but also much smaller current fluctuations that are obscured by noise.