Chapter 13 - Insights into the Pore of the Hair Cell Transducer Channel from Experiments with Permeant Blockers
Section snippets
OVERVIEW
This chapter considers recent experiments designed to infer properties of the ion‐conducting pore of the mechanoelectrical transducer channel of sensory hair cells using permeant blockers. By combining results from experiments with three classes of large cationic permeant blockers, the fluorescent dye FM1‐43, the aminoglycoside antibiotics, and the potassium‐sparing diuretic amiloride, information has been obtained on the free energy profile along the transducer channel's pore as sensed by
INTRODUCTION
The molecular identity of the hair cell transducer channel is, despite intensive research efforts, still in question at the time of writing (Corey, 2006). Nevertheless, recent pharmacological characterization is beginning to yield information about key properties of the channel that may help toward its eventual molecular identification by comparison with other known channel types. Historically, the pharmacology of the transducer channel has been studied to try and find the mode of action of
IONIC SELECTIVITY OF THE TRANSDUCER CHANNEL
The hair cell transducer channel is a nonselective cation channel with very similar permeabilities for the alkali cations and a much higher permeability for divalent metal ions, the highest for Ca2+ (Ohmori, 1985, Jorgensen and Kroese, 1995). The permeability sequence of the alkali metal ions corresponds to Eisenman sequence XI, pointing to a high negative charge density of the selectivity filter (Hille, 2001). The high affinity of the Ca2+ ions for the selectivity filter has the effect that Ca
Evidence for Permeation of FM1‐43 Through the Hair Cell Transducer Channel
FM1‐43 (Fig. 1) is a fluorescent styryl dye with a divalent cationic head group related to TEA and a long lipophilic tail which enables it to partition reversibly into the outer leaflet of the cell membrane when present in the extracellular solution (Betz et al., 1992). On incorporation into the membrane, its fluorescence increases by two orders of magnitude. It cannot cross the lipid bilayer so that when it becomes internalized into cells by endocytosis, it remains trapped in the inner leaflet
Evidence for Permeation of Aminoglycoside Antibiotics Through the Transducer Channel
The studies of Ohmori, 1985, Kroese et al., 1989 showed that extracellularly applied aminoglycoside antibiotics reversibly blocked transducer currents at negative but not at positive potentials. Kroese et al. (1989) also found that intracellularly applied aminoglycosides did not block the transducer currents even at concentrations of 500 μM, one to two orders of magnitude higher than the concentrations with which they achieved block from the extracellular side. The KD for extracellularly applied
Amiloride and Amiloride Derivatives as Permeant Transducer Channel Blockers: A Reinterpretation
The synthetic drug amiloride and related compounds find clinical application as potassium‐sparing diuretics, thanks to their high‐affinity blocking action (at submicromolar concentrations) on epithelial Na+ channels in the distal and collecting tubules of the kidney (Kleyman and Cragoe, 1988). Amiloride has been found to reversibly inhibit the hair cell transducer current, but at higher concentrations (KD around 50 μM; Jorgensen and Ohmori, 1988, Rüsch et al., 1994), which is probably why no
CONCLUSIONS
The results discussed in the preceding sections may be summarized by constructing a putative geometrical model of the transducer channel with a specific charge distribution lining the pore (Fig. 6). The interactions of the channel with the alkali metal cations, divalent cations (in particular Ca2+), the permeant cationic blocker molecules discussed in this chapter as well as other polycationic blocker molecules that have been investigated (Farris et al., 2004), all support the conclusion that
Acknowledgments
Supported by the Netherlands Organisation for Scientific Research (S.M.v.N) and the MRC (C.J.K.). The authors thank Dr. Cécil J. W. Meulenberg for his comments on an early version of this chapter and his help with the preparation of Fig. 1.
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