A Ba2+ chelator suppresses long shut events in fully activated high-conductance Ca(2+)-dependent K+ channels

Abstract

High-conductance Ca(2+)-activated K+ channels from rat skeletal muscle were incorporated into planar lipid bilayers, and the channel kinetics were studied with a high internal Ca2+ concentration (Cai). Raising the Cai is known to increase the channel open probability. This effect is due to an increases in openings frequency and duration, and saturates at a Cai around 100 microM. Raising the Cai also increases the occurrence of less frequent but very long (> 5 s) shut events. The mechanism underlying this slow kinetic process was studied. Raising Cai above 100 microM does not further increase the frequency of the long shut events. This was not consistent with the hypothesis that the long closures result from a classical channel-block mechanism induced by internal Ca2+. The transmembrane voltage and the presence of K+ ions in the external compartment both affect the slow kinetic process. A comparison of these effects with the properties of the channel block induced by Ba2+ ions added to the internal compartment strongly suggested that the long shut events are due to a contamination of the internal solutions by Ba2+. This was confirmed by showing that a crown-ether compound that strongly chelates Ba2+ completely suppresses the long shut events when added to the inner compartment.

Hypersensitivity of BK Ca to Ca 2+ Sparks Underlies Hyporeactivity of Arterial Smooth Muscle in Shock

“…To exclude the contamination of Ba 2ϩ , 50 mol/L crown-ether, (ϩ)-18-crown-6-tetra carboxylic acid (18C6TA) was added in the bath solution before the experiment. 34 The identity of BK Ca channel currents was confirmed by their ChTX (100 nmol/L) and tetraethylammonium sensitivity (200 mol/L; data not nor IbTX (1 g/kg) exhibited any significant effect on MAP in the sham group (nϭ5 to 7 rats for each group). D and E, ChTX (D) and IbTX (E) increased MAP and partially restored vasoreactivity 2 hours after HS.…”

Preserved BK Channel Function in Vasospastic Myocytes from a Dog Model of Subarachnoid Hemorrhage

“…Elevated Ca 2+ (>100 µ M ) can induce a low-activity mode with long closed periods in the BK channel [28] and contaminating Ba 2+ in electrophysiological solutions may similarly cause long shut intervals [15, 29]. Analyses were thus repeated after exclusion of shut intervals >5 s (which were rarely observed).…”

“…Ba 2+ contaminating stock chemicals used to make intracellular electrophysiology solutions has been reported to cause a slow block of BK channels at test potentials >100 mV [15]. This phenomenon was infrequently observed in some experiments as a late decline in current.…”

Ca2+-activated K+ channel inhibition by reactive oxygen species

“…For single-channel membranes, Po was obtained as the time spent in the fully open current level divided by the total time of the record, usually 60 s. Po values were calculated excluding channel closures lasting Ͼ200 ms as these events are due to ion channel blockage induced by the contaminant Ba 2ϩ (9,25). Oocyte isolation and RNA injection.…”