Single Na' channels from rat skeletal muscle plasma membrane vesicles were inserted into plan... more Single Na' channels from rat skeletal muscle plasma membrane vesicles were inserted into planar lipid bilayers formed from neutral phospho- lipids and were observed in the presence of batrachotoxin . The batrachotoxin- modified channel activates in the voltage range -120 to -80 mV and remains open almost all the time at voltages positive to -60 mV . Low levels of
Single Na' channels from rat skeletal muscle plasma membrane vesicles were inserted into plan... more Single Na' channels from rat skeletal muscle plasma membrane vesicles were inserted into planar lipid bilayers formed from neutral phospho- lipids and were observed in the presence of batrachotoxin . The batrachotoxin- modified channel activates in the voltage range -120 to -80 mV and remains open almost all the time at voltages positive to -60 mV . Low levels of
We have examined whether F-actin integrity is involved in activation of a volume-regulated Cl- cu... more We have examined whether F-actin integrity is involved in activation of a volume-regulated Cl- current (VRChlC) in B-lymphocytes. VRChlC activation was initiated in response to establishing a whole cell recording in the presence of a hyposmotic gradient. Parallel confocal microscopy experiments using Rhodamine-Phalloidin (R-P) as a specific marker of F-actin showed that the submembrane actin ring is reversibly disrupted in response to an hyposmotic gradient. Disruptions of cortical F-actin integrity by 50 microM cytochalasin B (CB) does not trigger activation of VRChlC under isosmotic conditions or potentiate the rate of activation when the osmolarity of the extracellular solution was decreased by 75%. However, incubation with CB increased the rate of VRChlC activation in response to a 90% hyposmotic gradient. Phalloidin, a stabilizer of F-actin, decreases the rate of VRChlC activation in response to a 90% gradient, but has no effect in response to a 75% gradient. These observations suggest that disassembly of cortical F-actin is not critical for VRChlC activation in B-lymphocytes. The integrity of cortical F-actin, however, can exert a modulatory effect on the rate of VRChlC activation in the presence of a hyposmotic gradient.
Many mammalian cells regulate their volume by the osmotic movement of water directed by anion and... more Many mammalian cells regulate their volume by the osmotic movement of water directed by anion and cation flux. Ubiquitous volume-dependent anion currents permit cells to recover volume after swelling in response to a hypotonic environment. This study addressed competition between glutamate (Glu) and Cl(-) permeation in volume-activated anion currents in order to provide insight into the ionic requirements for volume regulation, volume-dependent anion channel activity and to the architecture of the channel pore. The effect of changing the intracellular molar fraction (MF) of Glu and Cl(-) on conductance and relative anion permeability was evaluated as a function of the extracellular permeant anion and/or the ionic strength. Relative permeability of Glu to Cl(-) was determined by measuring reversal potentials under defined ionic conditions. Under conditions with high (150 m M) or low (50 m M) ionic strength solutions on both sides of the membrane, Cl(-) was always more permeable than Glu. When a transmembrane ionic strength gradient (150 m M extracellular: 50 m M intracellular) was set to drive water into the cell, and in the presence of extracellular Cl(-), Glu became up to 16-fold more permeable than Cl(-). Replacement of extracellular Cl(-) with Glu abolished this effect. These results indicate that it is possible for Glu to move into the extracellular environment during volume-regulatory events and they support the emerging role of glutamate as a modulator of anion channel activity.
Volume-regulated anion channels (VRAC) are ubiquitous ion channels that are normally nonconductin... more Volume-regulated anion channels (VRAC) are ubiquitous ion channels that are normally nonconducting, but can be opened upon cell swelling. Described initially in T lymphocytes, VRAC are characterized by activation in response to a hypotonic challenge, outward rectification of the current-voltage relationship, and a selectivity of I– > NO–3 > Br- > C1- > propionate > gluconate. Currents carried by these
Single Na' channels from rat skeletal muscle plasma membrane vesicles were inserted into plan... more Single Na' channels from rat skeletal muscle plasma membrane vesicles were inserted into planar lipid bilayers formed from neutral phospho- lipids and were observed in the presence of batrachotoxin . The batrachotoxin- modified channel activates in the voltage range -120 to -80 mV and remains open almost all the time at voltages positive to -60 mV . Low levels of
Single Na' channels from rat skeletal muscle plasma membrane vesicles were inserted into plan... more Single Na' channels from rat skeletal muscle plasma membrane vesicles were inserted into planar lipid bilayers formed from neutral phospho- lipids and were observed in the presence of batrachotoxin . The batrachotoxin- modified channel activates in the voltage range -120 to -80 mV and remains open almost all the time at voltages positive to -60 mV . Low levels of
We have examined whether F-actin integrity is involved in activation of a volume-regulated Cl- cu... more We have examined whether F-actin integrity is involved in activation of a volume-regulated Cl- current (VRChlC) in B-lymphocytes. VRChlC activation was initiated in response to establishing a whole cell recording in the presence of a hyposmotic gradient. Parallel confocal microscopy experiments using Rhodamine-Phalloidin (R-P) as a specific marker of F-actin showed that the submembrane actin ring is reversibly disrupted in response to an hyposmotic gradient. Disruptions of cortical F-actin integrity by 50 microM cytochalasin B (CB) does not trigger activation of VRChlC under isosmotic conditions or potentiate the rate of activation when the osmolarity of the extracellular solution was decreased by 75%. However, incubation with CB increased the rate of VRChlC activation in response to a 90% hyposmotic gradient. Phalloidin, a stabilizer of F-actin, decreases the rate of VRChlC activation in response to a 90% gradient, but has no effect in response to a 75% gradient. These observations suggest that disassembly of cortical F-actin is not critical for VRChlC activation in B-lymphocytes. The integrity of cortical F-actin, however, can exert a modulatory effect on the rate of VRChlC activation in the presence of a hyposmotic gradient.
Many mammalian cells regulate their volume by the osmotic movement of water directed by anion and... more Many mammalian cells regulate their volume by the osmotic movement of water directed by anion and cation flux. Ubiquitous volume-dependent anion currents permit cells to recover volume after swelling in response to a hypotonic environment. This study addressed competition between glutamate (Glu) and Cl(-) permeation in volume-activated anion currents in order to provide insight into the ionic requirements for volume regulation, volume-dependent anion channel activity and to the architecture of the channel pore. The effect of changing the intracellular molar fraction (MF) of Glu and Cl(-) on conductance and relative anion permeability was evaluated as a function of the extracellular permeant anion and/or the ionic strength. Relative permeability of Glu to Cl(-) was determined by measuring reversal potentials under defined ionic conditions. Under conditions with high (150 m M) or low (50 m M) ionic strength solutions on both sides of the membrane, Cl(-) was always more permeable than Glu. When a transmembrane ionic strength gradient (150 m M extracellular: 50 m M intracellular) was set to drive water into the cell, and in the presence of extracellular Cl(-), Glu became up to 16-fold more permeable than Cl(-). Replacement of extracellular Cl(-) with Glu abolished this effect. These results indicate that it is possible for Glu to move into the extracellular environment during volume-regulatory events and they support the emerging role of glutamate as a modulator of anion channel activity.
Volume-regulated anion channels (VRAC) are ubiquitous ion channels that are normally nonconductin... more Volume-regulated anion channels (VRAC) are ubiquitous ion channels that are normally nonconducting, but can be opened upon cell swelling. Described initially in T lymphocytes, VRAC are characterized by activation in response to a hypotonic challenge, outward rectification of the current-voltage relationship, and a selectivity of I– > NO–3 > Br- > C1- > propionate > gluconate. Currents carried by these
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