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Highly co‐operative Ca2+ activation of intermediate‐conductance K+ channels in granulocytes from a human cell line.
Author(s) -
Varnai P,
Demaurex N,
Jaconi M,
Schlegel W,
Lew D P,
Krause K H
Publication year - 1993
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.1993.sp019952
Subject(s) - charybdotoxin , chemistry , apamin , ionomycin , biophysics , agonist , patch clamp , conductance , extracellular , ionophore , reversal potential , receptor , membrane potential , potassium channel , biochemistry , biology , in vitro , membrane , mathematics , combinatorics
1. To study Ca(2+)‐activated K+ currents in dimethyl sulphoxide (DMSO)‐differentiated HL‐60 cells (HL‐60 granulocytes), we have combined the patch clamp technique with microfluorimetric measurements of the cytosolic free Ca2+ concentration ([Ca2+]i). 2. Elevations of [Ca2+]i induced by the receptor agonist N‐formyl‐L‐methionyl‐L‐phenylalanine (f‐MLP), by cellular spreading or by the Ca2+ ionophore ionomycin, activated whole‐cell currents. The kinetics of the current elevations closely paralleled the kinetics of the elevations in [Ca2+]i. Cellular spreading induced oscillations in [Ca2+]i and parallel oscillatory changes in the amplitude of the recorded currents. 3. The reversal potential of the Ca(2+)‐activated current was a function of the extracellular K+ concentration (56.1 mV per log [K+]), demonstrating that the underlying conductance was selective for K+. 4. The current was blocked by charybdotoxin, but insensitive to apamin. 5. The whole‐cell current was inwardly rectifying. No time‐dependent activation or inactivation of the current could be observed within the range of voltages tested (‐100 to +100 mV). 6. The dependence of the current amplitude on the measured [Ca2+]i revealed a half‐maximal activation at approximately 350 nM [Ca2+]i, and a highly co‐operative activation by [Ca2+]i with an apparent Hill coefficient of approximately 8. Neither the half‐maximal activation by [Ca2+]i nor the apparent Hill coefficient depended on the voltage, and they were identical for Ca2+ elevations caused by the ionophore and the receptor agonist. 7. Analysis of Ca(2+)‐activated single‐channel events in cell‐attached recordings revealed an inwardly rectifying K+ channel with a slope conductance of 35 pS. Fluctuation analysis of the Ca(2+)‐activated whole‐cell current suggested an underlying single‐channel conductance of a similar size (28 pS). 8. In summary, we describe a charybdotoxin‐sensitive, intermediate‐conductance Ca(2+)‐activated K+ channel in HL‐60 granulocytes. The characteristics of the Ca2+ activation of this current (i.e. sensitivity to submicromolar [Ca2+]i, high co‐operativity and voltage independence) are similar to the Ca2+ activation of the apamin‐sensitive small‐conductance K+ channel. Our results also suggest that [Ca2+]i elevations are the predominant, if not the only, activators of this channel during physiological stimulation of HL‐60 granulocytes.