Oxygen deprivation inhibits a K+ channel independently of cytosolic factors in rat central neurons.

1. K+ channel modulation has been shown to be an integral and important cellular response to O2 deprivation. Although part of this modulation occurs as a result of changes in concentrations of several cytosolic factors such as ATP and Ca2+, it is unknown whether there are mechanisms other than those originating from the cytosol. To test the hypothesis that membrane-delimited mechanisms participate in the O2-sensing process and are involved in the modulation of K+ channel activity in central neurons, we performed experiments using patch-clamp techniques and dissociated cells from the rat neocortex and substantia nigra. 2. Whole-cell outward currents were studied in voltage-clamp mode using Na(+)-free or low-Na+ (5 mM, with 1 microM tetrodotoxin) extracellular medium plus 0.5 mM Co2+. O2 deprivation produced a biphasic response in current amplitude, i.e. an initial transient increase followed by a pronounced decrease in outward currents. The reduction in outward currents was a reversible process since perfusion with a medium of PO2 > 100 mmHg (1 mmHg = 133 Pa) led to a complete recovery. 3. In cell-free excised membrane patches, we found that a specific K+ current (large conductance, inhibited by micromolar concentrations of ATP and activated by Ca2+) was reversibly inhibited by lack of O2. This was characterized by a marked decrease in channel open-state probability and a slight reduction in unitary conductance. The magnitude of channel inhibition by O2 deprivation was closely dependent on O2 tension. The PO2 level for 50% channel inhibition was about 10 mmHg with little or no inhibition at PO2 > or = 20 mmHg. 4. Single-channel kinetic analysis showed that channel open times consisted of two components and closed times were composed of three. The hypoxia-induced inhibition of K+ channel activity was mediated by selective suppression of the longer time constant channel openings without significantly affecting closed time constants. This led to an increase in frequency of opening and closing and rapid channel flickerings. 5. Our data showed that O2 deprivation had no effect on another K+ current characterized by a much smaller conductance and Ca2+ independence. This provides evidence for the selective nature of the hypoxia-induced inhibition of some species of K+ channels. 6. These results therefore provide the first evidence for regulation of K+ channel activity by O2 deprivation in cell-free excised patches from central neurons.