scholarly journals Molecular identification of SqKv1A. A candidate for the delayed rectifier K channel in squid giant axon.

1996 ◽  
Vol 108 (3) ◽  
pp. 207-219 ◽  
Author(s):  
J J Rosenthal ◽  
R G Vickery ◽  
W F Gilly
Keyword(s):  
Time Course ◽  
Xenopus Oocytes ◽  
Giant Axon ◽  
K Channel ◽  
Delayed Rectifier ◽  
Giant Fiber ◽  
Western Blots ◽  
Internal Solution ◽  
Giant Axons ◽  
K Currents

We have cloned the cDNA for a squid Kvl potassium channel (SqKv1A). SqKv1A mRNA is selectively expressed in giant fiber lobe (GFL) neurons, the somata of the giant axons. Western blots detect two forms of SqKv1A in both GFL neuron and giant axon samples. Functional properties of SqKv1A currents expressed in Xenopus oocytes are very similar to macroscopic currents in GFL neurons and giant axons. Macroscopic K currents in GFL neuron cell bodies, giant axons, and in Xenopus oocytes expressing SqKv1A, activate rapidly and inactivate incompletely over a time course of several hundred ms. Oocytes injected with SqKv1A cRNA express channels of two conductance classes, estimated to be 13 and 20 pS in an internal solution containing 470 mM K. SqKv1A is thus a good candidate for the "20 pS" K channel that accounts for the majority of rapidly activating K conductance in both GFL neuron cell bodies and the giant axon.

scholarly journals Potassium conductance of the squid giant axon. Single-channel studies.

1988 ◽  
Vol 92 (2) ◽  
pp. 179-196 ◽  
Author(s):  
I Llano ◽  
C K Webb ◽  
F Bezanilla
Keyword(s):  
Time Course ◽  
Single Channel ◽  
Potassium Conductance ◽  
Giant Axon ◽  
Squid Giant Axon ◽  
K Channel ◽  
Delayed Rectifier ◽  
Fluctuation Analysis ◽  
Patch Clamp Technique ◽  
Relative Contribution

The patch-clamp technique was implemented in the cut-open squid giant axon and used to record single K channels. We present evidence for the existence of three distinct types of channel activities. In patches that contained three to eight channels, ensemble fluctuation analysis was performed to obtain an estimate of 17.4 pS for the single-channel conductance. Averaged currents obtained from these multichannel patches had a time course of activation similar to that of macroscopic K currents recorded from perfused squid giant axons. In patches where single events could be recorded, it was possible to find channels with conductances of 10, 20, and 40 pS. The channel most frequently encountered was the 20-pS channel; for a pulse to 50 mV, this channel had a probability of being open of 0.9. In other single-channel patches, a channel with a conductance of 40 pS was present. The activity of this channel varied from patch to patch. In some patches, it showed a very low probability of being open (0.16 for a pulse to 50 mV) and had a pronounced lag in its activation time course. In other patches, the 40-pS channel had a much higher probability of being open (0.75 at a holding potential of 50 mV). The 40-pS channel was found to be quite selective for K over Na. In some experiments, the cut-open axon was exposed to a solution containing no K for several minutes. A channel with a conductance of 10 pS was more frequently observed after this treatment. Our study shows that the macroscopic K conductance is a composite of several K channel types, but the relative contribution of each type is not yet clear. The time course of activation of the 20-pS channel and the ability to render it refractory to activation only by holding the membrane potential at a positive potential for several seconds makes it likely that it is the predominant channel contributing to the delayed rectifier conductance.

scholarly journals Fast Inactivation of Delayed Rectifier K Conductance in Squid Giant Axon and Its Cell Bodies

1997 ◽  
Vol 109 (4) ◽  
pp. 435-448 ◽  
Author(s):  
Chris Mathes ◽  
Joshua J.C. Rosenthal ◽  
Clay M. Armstrong ◽  
William F. Gilly
Keyword(s):  
Large Fraction ◽  
Giant Axon ◽  
Delayed Rectifier ◽  
Inactivation Kinetics ◽  
Temperature Sensitive ◽  
Fast Inactivation ◽  
Giant Fiber ◽  
Experimental Conditions ◽  
Giant Axons ◽  
External K

Inactivation of delayed rectifier K conductance (gK) was studied in squid giant axons and in the somata of giant fiber lobe (GFL) neurons. Axon measurements were made with an axial wire voltage clamp by pulsing to VK (∼−10 mV in 50–70 mM external K) for a variable time and then assaying available gK with a strong, brief test pulse. GFL cells were studied with whole-cell patch clamp using the same prepulse procedure as well as with long depolarizations. Under our experimental conditions (12–18°C, 4 mM internal MgATP) a large fraction of gK inactivates within 250 ms at −10 mV in both cell bodies and axons, although inactivation tends to be more complete in cell bodies. Inactivation in both preparations shows two kinetic components. The faster component is more temperature-sensitive and becomes very prominent above 12°C. Contribution of the fast component to inactivation shows a similar voltage dependence to that of gK, suggesting a strong coupling of this inactivation path to the open state. Omission of internal MgATP or application of internal protease reduces the amount of fast inactivation. High external K decreases the amount of rapidly inactivating IK but does not greatly alter inactivation kinetics. Neither external nor internal tetraethylammonium has a marked effect on inactivation kinetics. Squid delayed rectifier K channels in GFL cell bodies and giant axons thus share complex fast inactivation properties that do not closely resemble those associated with either C-type or N-type inactivation of cloned Kv1 channels studied in heterologous expression systems.

scholarly journals Potassium channel block by internal calcium and strontium.

1991 ◽  
Vol 97 (3) ◽  
pp. 627-638 ◽  
Author(s):  
C M Armstrong ◽  
Y Palti
Keyword(s):  
Time Course ◽  
External Solution ◽  
High Energy ◽  
K Channel ◽  
Giant Fiber ◽  
Distance Measurements ◽  
K Channels ◽  
High Energy Barrier ◽  
Intracellular Ca ◽  
A Site

We show that intracellular Ca blocks current flow through open K channels in squid giant fiber lobe neurons. The block has similarities to internal Sr block of K channels in squid axons, which we have reexamined. Both ions must cross a high energy barrier to enter the blocking site from the inside, and block occurs only with millimolar concentrations and with strong depolarization. With Sr (axon) or Ca (neuron) inside, IK is normal in time course for voltages less than about +50 mV; but for large steps, above +90 mV, there is a rapid time-dependent block or "inactivation." From roughly +70 to +90 mV (depending on concentration) the current has a complex time course that may be related to K accumulation near the membrane's outer surface. Block can be deepened by either increasing the concentration or the voltage. Electrical distance measurements suggest that the blocking ion moves to a site deep in the channel, possibly near the outer end. Block by internal Ca can be prevented by putting 10 mM Rb in the external solution. Recovery from block after a strong depolarization occurs quickly at +30 mV, with a time course that is about the same as that of normal K channel activation at this voltage. 20 mM Mg in neurons had no discernible blocking effect. The experiments raise questions regarding the relation of block to normal channel gating. It is speculated that when the channel is normally closed, the "blocking" site is occupied by a Ca ion that comes from the external medium.

scholarly journals Gating of IsK expressed in Xenopus oocytes depends on the amount of mRNA injected.

1994 ◽  
Vol 104 (1) ◽  
pp. 87-105 ◽  
Author(s):  
J Cui ◽  
R P Kline ◽  
P Pennefather ◽  
I S Cohen
Keyword(s):  
Xenopus Oocytes ◽  
Single Injection ◽  
Ion Current ◽  
K Channel ◽  
Delayed Rectifier ◽  
Activation Kinetics ◽  
Kinetic Schemes ◽  
Mrna Concentration ◽  
Kinetics Of

IsK is a K+ channel of the delayed rectifier type widely distributed throughout both excitable and nonexcitable cells. Its structure is different from other cloned K+ channels and molecular details of its gating remain obscure. Here we show that the activation kinetics of IsK expressed in Xenopus oocytes depend upon the amount of its mRNA injected, with larger amounts resulting in slower activation kinetics with a longer initial delay during activation. Similar changes in activation kinetics occur with time after a single injection of IsK mRNA. We present two kinetic schemes which illustrate how our experimental results could arise. Both imply an interaction among individual channel proteins during IsK activation. The dependence of channel gating on mRNA concentration provides a novel mechanism for long term regulation of ion current kinetics.

scholarly journals Slow components of potassium tail currents in rat skeletal muscle.

1983 ◽  
Vol 81 (4) ◽  
pp. 513-530 ◽  
Author(s):  
K G Beam ◽  
P L Donaldson
Keyword(s):  
Time Constant ◽  
Time Course ◽  
Outward Current ◽  
Giant Axon ◽  
K Channel ◽  
Tail Current ◽  
Anomalous Rectifier ◽  
Single Well ◽  
Current Flows ◽  
Slow Kinetic

The kinetics of potassium tail currents have been studied in the omohyoid muscle of the rat using the three-microelectrode voltage-clamp technique. The currents were elicited by a two-pulse protocol in which a conditioning pulse to open channels was followed by a test step to varying levels. The tail currents reversed at a single well-defined potential (VK). At hyperpolarized test potentials (-100 mV and below), tail currents were inward and exhibited two clearly distinguishable phases of decay, a fast tail with a time constant of 2-3 ms and a slow tail with a time constant of approximately 150 ms. At depolarized potentials (-60 mV and above), tail currents were outward and did not show two such easily separable phases of decay, although a slow kinetic component was present. The slow kinetic phase of outward tail currents appeared to be functionally distinct from the slow inward tail since the channels responsible for the latter did not allow significant outward current. Substitution of Rb for extracellular K abolished current through the anomalous (inward-going) rectifier and at the same time eliminated the slow inward tail, which suggests that the slow inward tail current flows through anomalous rectifier channels. The amplitude of the slow inward tail was increased and VK was shifted in the depolarizing direction by longer conditioning pulses. The shift in VK implies that during outward currents potassium accumulates in a restricted extracellular space, and it is suggested that this excess K causes the slow inward tail by increasing the inward current through the anomalous rectifier. By this hypothesis, the tail current slowly decays as K diffuses from the restricted space. Consistent with such a hypothesis, the decay of the slow inward tail was not strongly affected by changing temperature. It is concluded that a single delayed K channel is present in the omohyoid. Substitution of Rb for K has little effect on the magnitude or time course of outward current tails, but reduces the magnitude and slows the decay of the fast component of inward tails. Both effects are consistent with a mechanism proposed for squid giant axon (Swenson and Armstrong, 1981): that (a) the delayed potassium channel cannot close while Rb is inside it, and (b) that Rb remains in the channel longer than K.

Giant Axons and Synergic Contractions in Branchiomma Vesiculosum

1951 ◽  
Vol 28 (1) ◽  
pp. 22-31 ◽  
Author(s):  
J. A. COLIN NICOL
Keyword(s):  
Time Course ◽  
Body Wall ◽  
Giant Axon ◽  
The Body ◽  
Single Muscle ◽  
Graphic Method ◽  
Giant Axons ◽  
Maximal Tension ◽  
Contraction And Relaxation ◽  
Electrical Stimuli

Synergic contractions following giant axon stimulation in Branchiomma vesiculosum have been investigated by the graphic method of recording. Isotonic and isometric levers were used, and electrical stimuli from condenser discharges were applied to the surface of the animal. Single muscle twitches occur at stimulation frequencies up to 2 per sec., above which clonus, and finally tetanus result. At high rates of stimulation fatigue rapidly sets in; this fatigue is reversible. Data for the time course of contractions are presented. Maximal tension develops about 255 msec. after the beginning of contraction, and relaxation occupies about 1.8 sec. Maximal tension developed isometrically under stimulation at different frequencies (12 per min. to 13 per sec.) was measured. Maximal tension is developed initially, and there is no evidence for facilitation. Extension of the animal and of strips of the body wall under tension are described. The results are discussed in terms of the habits of the animal, and compared with similar studies of other invertebrates.

Single HERG delayed rectifier K+ channels expressed in Xenopus oocytes

1997 ◽  
Vol 272 (3) ◽  
pp. H1309-H1314 ◽  
Author(s):  
A. Zou ◽  
M. E. Curran ◽  
M. T. Keating ◽  
M. C. Sanguinetti
Keyword(s):  
Cardiac Myocytes ◽  
Xenopus Oocytes ◽  
Single Channel ◽  
K Channel ◽  
Delayed Rectifier ◽  
Class Iii ◽  
Channel Opening ◽  
The Mean ◽  
K Concentration ◽  
Channel Properties

HERG is a K+ channel with properties similar to the rapidly activating component (I(Kr)) of delayed rectifier K+ current, which is important for repolarization of human cardiac myocytes. In this study, we have characterized the single-channel properties of HERG expressed in Xenopus oocytes. Currents were measured in cell-attached patches with an extracellular K concentration of 120 mM. The single HERG channel conductance, determined at test potentials between -50 and -110 mV, was 12.1 +/- 0.6 pS. At positive test potentials (+40 to +80 mV), the probability of channel opening was low and slope conductance was 5.1 +/- 0.6 pS. The mean channel open times at -90 mV were 2.9 +/- 0.5 and 11.8 +/- 1.0 ms, and the mean channel closed times were 0.54 +/- 0.02 and 14.5 +/- 5.3 ms. Single HERG channels were blocked by MK-499, a class III antiarrhythmic agent that blocks I(Kr) in cardiac myocytes. The development of block was more rapid in inside-out patches than in cell-attached patches or in whole cell recordings, indicating that block occurs from the cytoplasmic side of the membrane. The single-channel properties of HERG are similar to I(Kr) channels of isolated cardiac myocytes, which provides further evidence that HERG proteins coassemble to form I(Kr) channels.

scholarly journals Modulation by protein kinase C activation of rat brain delayed-rectifier K+ channel expressed in Xenopus oocytes

FEBS Letters ◽  
1996 ◽  
Vol 381 (1-2) ◽  
pp. 71-76 ◽  
Author(s):  
Tuvia Peretz ◽  
Gal Levin ◽  
Ofira Moran ◽  
William B. Thornhill ◽  
Dodo Chikvashvili ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Rat Brain ◽  
Xenopus Oocytes ◽  
K Channel ◽  
Delayed Rectifier ◽  
Kinase C ◽  
Protein Kinase C Activation

Blockade of Na+and K+Currents by Local Anesthetics in the Dorsal Horn Neurons of the Spinal Cord 

1998 ◽  
Vol 88 (1) ◽  
pp. 172-179 ◽  
Author(s):  
Andrea Olschewski ◽  
Gunter Hempelmann ◽  
Werner Vogel ◽  
Boris V. Safronov
Keyword(s):  
Spinal Cord ◽  
Dorsal Horn ◽  
Epidural Anesthesia ◽  
Local Anesthetics ◽  
Molecular Mechanisms ◽  
K Channel ◽  
Delayed Rectifier ◽  
Patch Clamp Technique ◽  
Dorsal Horn Neurons ◽  
K Currents

Background The dorsal horn of the spinal cord is a pivotal point for transmission of neuronal pain. During spinal and epidural anesthesia, the neurons of the dorsal horn are exposed to local anesthetics. Unfortunately, little is known about the action of local anesthetics on the major ionic conductances in dorsal horn neurons. In this article, the authors describe the effects of bupivacaine, lidocaine, and mepivacaine on voltage-gated Na+ and K+ currents in the membranes of these neurons. Methods The patch-clamp technique was applied to intact dorsal horn neurons from laminae I-III identified in 200-microm slices of spinal cord from newborn rats. Under voltage-clamp conditions, the whole-cell Na+ and K+ currents activated by depolarization were recorded in the presence of different concentrations of local anesthetics. Results Externally applied bupivacaine, lidocaine, and mepivacaine produced tonic block of Na+ currents with different potencies. Half-maximum inhibiting concentrations (IC50) were 26, 112, and 324 microM, respectively. All local anesthetics investigated also showed a phasic, that is, a use-dependent, block of Na+ channels. Rapidly inactivating K+ currents (KA currents) also were sensitive to the blockers with IC50 values for tonic blocks of 109, 163, and 236 microM, respectively. The block of KA currents was not use dependent. In contrast to Na+ and KA currents, delayed-rectifier K+ currents were almost insensitive to the local anesthetics applied. Conclusions In clinically relevant concentrations, local anesthetics block Na+ and KA currents but not delayed-rectifier K+ currents in spinal dorsal horn neurons. The molecular mechanisms of Na+ and K+ channel block by local anesthetics seem to be different. Characterization of these mechanisms could be an important step in understanding the complexity of local anesthetic action during spinal and epidural anesthesia.

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