Gating and permeation mechanisms of ion channels will be studied. Voltage-gated ion channels open or close in a fraction of a millisecond, controlling the ion flows that generate electrical signals. These channels are present in most cells of the body, and are vital to many processes, including thought, movement, timing of the heartbeat, and control of secretion of hormones e.g. insulin. In nature ion channels are separated into two component parts. A permeation module or pore is found in isolation in bacteria and elsewhere. To this is added a gating module to form the voltage-gated channel found, for example, in nerve and muscle fibers. The mechanism of the gating module is understood on in part. It is proposed here to investigate the molecule events that link the gating module to the pore. Gating occurs in steps, beginning with movements in the four individual subunits that constitute a channel, and culminating in a concerted step that opens the pore. The last step will be examined in potassium channel mutants in which the concerted step is conveniently isolated from the earlier steps. These channels will be prepared by site-directed-mutagenesis, expressed in a cell line, and examined in patch-clamp experiments. Particular attention will be given to the effects of pore occupancy on the gating apparatus, and to the hypothesis that the concerted step that opens the pore involves a change in occupancy. Sodium channel experiments to examine occupancy effects will be performed primarily on voltage-clamped internally perfused giant axons of the squid, the best preparation for measuring the gating currents that signal the conformation changes in the gating module.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
2R01NS012547-24
Application #
2854327
Study Section
Special Emphasis Panel (ZRG1-MDCN-3 (01))
Program Officer
Talley, Edmund M
Project Start
1977-01-01
Project End
2003-02-28
Budget Start
1999-03-01
Budget End
2000-02-29
Support Year
24
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Physiology
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Gomez-Lagunas, Froylan; Melishchuk, Alexey; Armstrong, Clay M (2003) Block of Shaker potassium channels by external calcium ions. Proc Natl Acad Sci U S A 100:347-51
Armstrong, Clay M (2003) The Na/K pump, Cl ion, and osmotic stabilization of cells. Proc Natl Acad Sci U S A 100:6257-62
Armstrong, C M; Cota, G (1999) Calcium block of Na+ channels and its effect on closing rate. Proc Natl Acad Sci U S A 96:4154-7
Armstrong, C M (1999) Distinguishing surface effects of calcium ion from pore-occupancy effects in Na+ channels. Proc Natl Acad Sci U S A 96:4158-63
Armstrong, C M; Hille, B (1998) Voltage-gated ion channels and electrical excitability. Neuron 20:371-80
Khodakhah, K; Melishchuk, A; Armstrong, C M (1998) Charge immobilization caused by modification of internal cysteines in squid Na channels. Biophys J 75:2821-9
Melishchuk, A; Loboda, A; Armstrong, C M (1998) Loss of shaker K channel conductance in 0 K+ solutions: role of the voltage sensor. Biophys J 75:1828-35
Khodakhah, K; Melishchuk, A; Armstrong, C M (1997) Killing K channels with TEA+. Proc Natl Acad Sci U S A 94:13335-8
Mathes, C; Rosenthal, J J; Armstrong, G M et al. (1997) Fast inactivation of delayed rectifier K conductance in squid giant axon and its cell bodies. J Gen Physiol 109:435-48
Khodakhah, K; Armstrong, C M (1997) Inositol trisphosphate and ryanodine receptors share a common functional Ca2+ pool in cerebellar Purkinje neurons. Biophys J 73:3349-57

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