Sodium current and membrane potential in EDL muscle fibers from normal and dystrophic (mdx) mice

1991 ◽  
Vol 261 (4) ◽  
pp. C718-C725 ◽  
Author(s):  
C. Mathes ◽  
F. Bezanilla ◽  
R. E. Weiss
Keyword(s):  
Membrane Potential ◽  
Sodium Channels ◽  
Single Channel ◽  
Muscle Fibers ◽  
Mdx Mice ◽  
Apparent Difference ◽  
Sodium Currents ◽  
Edl Muscle ◽  
Single Channel Currents ◽  
Channel Properties

The macroscopic and single-channel properties of sodium currents and membrane potential were studied in intact extensor digitorum longus (EDL) muscle fibers from mdx (C57BL/10ScSn-mdx) and normal (C57BL/10SnJ) mice. The voltage dependence of activation and inactivation were determined and the associated gating charges were calculated to determine if the lack of dystrophin associated with the mdx condition has any influence on sodium channels either directly or by effects on the membrane environment of the channel. Sodium currents were recorded from cell-attached patches on EDL muscle fibers isolated by collagenase treatment and manual dissection. Both macroscopic and single-channel currents were studied. We found no apparent difference in the sodium channel properties from the two types of muscle. In addition, microelectrode measurements in both mdx and normal muscle fibers indicated similar resting membrane potentials (Vm around -95 mV), which suggests that the normal behavior of sodium channels in the muscle sarcolemma is unaffected by the X-linked gene defect.

Effects of dihydropyridine calcium channel modulators on cardiac sodium channels

1988 ◽  
Vol 254 (1) ◽  
pp. H140-H147 ◽  
Author(s):  
A. Yatani ◽  
D. L. Kunze ◽  
A. M. Brown
Keyword(s):  
Calcium Channels ◽  
Sodium Channels ◽  
Single Channel ◽  
Bay K 8644 ◽  
Sodium Currents ◽  
Whole Cell ◽  
Blocking Effects ◽  
Voltage Dependent ◽  
Cardiac Sodium Channels ◽  
Single Channel Currents

To investigate whether cardiac sodium channels have dihydropyridine (DHP) receptors we studied the effects of the optically pure (greater than 95%) enantiomers of the DHPs PN200–110 and BAY-K 8644 and the racemic DHP nitrendipine (NTD). Whole cell and single-channel sodium currents were recorded from cultured ventricular cells of neonatal rats using the patch-clamp method. NTD reduced cardiac sodium currents in a voltage-dependent manner. Inhibitory effects were due to an increase in traces without activity. The unit conductance remained unchanged. At negative holding potentials, NTD transiently increased the probability of channel opening. Both (+) and (-) PN 200–110 blocked sodium channels, although the (-) isomer was about one order of magnitude less effective. The blocking effects were voltage dependent. (+) BAY-K 8644 had similar blocking effects. (-) BAY-K 8644 produced an increase in sodium currents due to an increased frequency of channel openings and a marked prolongation of open time without any significant change in unit conductance. The DHPs have effects on cardiac sodium whole cell and single-channel currents that appear identical to and are as stereospecific as their effects on cardiac calcium currents, although the concentrations required are larger. In contrast the inwardly rectifying potassium channel (IK1) is unaffected by these DHPs. We conclude that functionally equivalent DHP receptors are present in cardiac sodium and calcium channels but not potassium channels and take this as evidence of the homology between sodium and calcium channels.

Effects of New World scorpion toxins on single-channel and whole cell cardiac sodium currents

1988 ◽  
Vol 254 (3) ◽  
pp. H443-H451 ◽  
Author(s):  
A. Yatani ◽  
G. E. Kirsch ◽  
L. D. Possani ◽  
A. M. Brown
Keyword(s):  
Sodium Channel ◽  
Sodium Channels ◽  
New World ◽  
Single Channel ◽  
Sodium Current ◽  
Sodium Currents ◽  
Scorpion Toxins ◽  
Whole Cell ◽  
Cardiac Sodium Channels ◽  
Single Channel Currents

Purified toxins from a North American scorpion, Centruroides noxius (Cn II-10), and a South American scorpion, Tityus serrulatus (Ts-gamma), were tested on cardiac sodium channels using patch-clamp methods to record whole cell and single-channel currents. The two toxins produced similar effects on sodium currents; potassium and calcium currents were not affected. Macroscopic sodium current amplitudes, measured at test potentials greater than -20 mV where the opening probability was high, decreased in a concentration-dependent manner with a half maximum inhibitory concentration of 6 X 10(-8) M. Block was unchanged by repetitive depolarizing pulses. In the presence of scorpion toxin, the currents were rapidly blocked by tetrodotoxin (3 X 10(-5) M). Both toxins shifted the voltage dependence of sodium channel inactivation to more negative potentials. At test potentials between -50 and -70 mV, where the sodium channel opening probability is normally low, both toxins produced an increase in sodium current and slowed the rates of activation and inactivation. At intermediate potentials between -50 and -20 mV the currents in the presence of toxins crossed over the control currents. At a test potential of -20 mV, the toxins decreased single-channel activity and increased the latency to first opening. At a test potential of -60 mV, the toxins significantly prolonged channel open time. The unitary current amplitudes were unchanged at either potential. We conclude that New World scorpion toxins produce apparently complex effects on whole cell currents primarily by retarding activation gating of cardiac sodium channels.

scholarly journals Renal sodium channels: Regulation and single channel properties

1995 ◽  
Vol 48 (4) ◽  
pp. 941-949 ◽  
Author(s):  
Douglas C. Eaton ◽  
Andrea Becchetti ◽  
Heping Ma ◽  
Brian N. Ling
Keyword(s):  
Sodium Channels ◽  
Single Channel ◽  
Channel Properties

scholarly journals Purified and unpurified sodium channels from eel electroplax in planar lipid bilayers.

1987 ◽  
Vol 90 (3) ◽  
pp. 375-395 ◽  
Author(s):  
E Recio-Pinto ◽  
D S Duch ◽  
S R Levinson ◽  
B W Urban
Keyword(s):  
Sodium Channel ◽  
Lipid Bilayers ◽  
Sodium Channels ◽  
Single Channel ◽  
Planar Bilayers ◽  
Electric Eel ◽  
Voltage Dependent ◽  
Single Channel Activity ◽  
Single Channel Currents ◽  
Channel Currents

Highly purified sodium channel protein from the electric eel, Electrophorus electricus, was reconstituted into liposomes and incorporated into planar bilayers made from neutral phospholipids dissolved in decane. The purest sodium channel preparations consisted of only the large, 260-kD tetrodotoxin (TTX)-binding polypeptide. For all preparations, batrachotoxin (BTX) induced long-lived single-channel currents (25 pS at 500 mM NaCl) that showed voltage-dependent activation and were blocked by TTX. This block was also voltage dependent, with negative potentials increasing block. The permeability ratios were 4.7 for Na+:K+ and 1.6 for Na+:Li+. The midpoint for steady state activation occurred around -70 mV and did not shift significantly when the NaCl concentration was increased from 50 to 1,000 mM. Veratridine-induced single-channel currents were about half the size of those activated by BTX. Unpurified, nonsolubilized sodium channels from E. electricus membrane fragments were also incorporated into planar bilayers. There were no detectable differences in the characteristics of unpurified and purified sodium channels, although membrane stability was considerably higher when purified material was used. Thus, in the eel, the large, 260-kD polypeptide alone is sufficient to demonstrate single-channel activity like that observed for mammalian sodium channel preparations in which smaller subunits have been found.

scholarly journals Single Channel Properties of P2X2 Purinoceptors

1999 ◽  
Vol 113 (5) ◽  
pp. 695-720 ◽  
Author(s):  
Shinghua Ding ◽  
Frederick Sachs
Keyword(s):  
Conformational Changes ◽  
Single Channel ◽  
Excess Noise ◽  
Hill Coefficient ◽  
Inward Rectification ◽  
Channel Current ◽  
Single Channel Current ◽  
The Mean ◽  
Single Channel Currents ◽  
Channel Properties

The single channel properties of cloned P2X2 purinoceptors expressed in human embryonic kidney (HEK) 293 cells and Xenopus oocytes were studied in outside-out patches. The mean single channel current–voltage relationship exhibited inward rectification in symmetric solutions with a chord conductance of ∼30 pS at −100 mV in 145 mM NaCl. The channel open state exhibited fast flickering with significant power beyond 10 kHz. Conformational changes, not ionic blockade, appeared responsible for the flickering. The equilibrium constant of Na+ binding in the pore was ∼150 mM at 0 mV and voltage dependent. The binding site appeared to be ∼0.2 of the electrical distance from the extracellular surface. The mean channel current and the excess noise had the selectivity: K+ > Rb+ > Cs+ > Na+ > Li+. ATP increased the probability of being open (Po) to a maximum of 0.6 with an EC50 of 11.2 μM and a Hill coefficient of 2.3. Lowering extracellular pH enhanced the apparent affinity of the channel for ATP with a pKa of ∼7.9, but did not cause a proton block of the open channel. High pH slowed the rise time to steps of ATP without affecting the fall time. The mean single channel amplitude was independent of pH, but the excess noise increased with decreasing pH. Kinetic analysis showed that ATP shortened the mean closed time but did not affect the mean open time. Maximum likelihood kinetic fitting of idealized single channel currents at different ATP concentrations produced a model with four sequential closed states (three binding steps) branching to two open states that converged on a final closed state. The ATP association rates increased with the sequential binding of ATP showing that the binding sites are not independent, but positively cooperative. Partially liganded channels do not appear to open. The predicted Po vs. ATP concentration closely matches the single channel current dose–response curve.

scholarly journals Single-channel recordings from cultured human retinal pigment epithelial cells.

1988 ◽  
Vol 91 (2) ◽  
pp. 193-222 ◽  
Author(s):  
J A Fox ◽  
B A Pfeffer ◽  
G L Fain
Keyword(s):  
Membrane Potential ◽  
Epithelial Cells ◽  
Retinal Pigment Epithelial ◽  
Single Channel ◽  
Retinal Pigment ◽  
Retinal Pigment Epithelial Cells ◽  
Open Probability ◽  
Pigment Epithelial ◽  
Single Channel Currents ◽  
Pigment Epithelial Cells

We have applied patch-clamp techniques to on-cell and excised-membrane patches from human retinal pigment epithelial cells in tissue culture. Single-channel currents from at least four ion channel types were observed: three or more potassium-selective channels with single-channel slope conductances near 100, 45, and 25 pS as measured in on-cell patches with physiological saline in the pipette, and a relatively nonselective channel with subconductance states, which has a main-state conductance of approximately 300 pS at physiological ion concentrations. The permeability ratios, PK/PNa, measured in excised patches were 21 for the 100-pS channels, 3 for the 25-pS channels, and 0.8 for the 300-pS nonselective channel. The 45-pS channels appeared to be of at least two types, with PK/PNa's of approximately 41 for one type and 3 for the other. The potassium-selective channels were spontaneously active at all potentials examined. The average open time for these channels ranged from a few milliseconds to many tens of milliseconds. No consistent trend relating potassium-selective channel kinetics to membrane potential was apparent, which suggests that channel activity was not regulated by the membrane potential. In contrast to the potassium-selective channels, the activity of the nonselective channel was voltage dependent: the open probability of this channel declined to low values at large positive or negative membrane potentials and was maximal near zero. Single-channel conductances observed at several symmetrical KCl concentrations have been fitted with Michaelis-Menten curves in order to estimate maximum channel conductances and ion-binding constants for the different channel types. The channels we have recorded are probably responsible for the previously observed potassium permeability of the retinal pigment epithelium apical membrane.

Evidence for a Ca-activated inwardly rectifying K channel in human macrophages

1989 ◽  
Vol 257 (1) ◽  
pp. C77-C85 ◽  
Author(s):  
E. K. Gallin
Keyword(s):  
Membrane Potential ◽  
Single Channel ◽  
Reversal Potential ◽  
K Channel ◽  
Resting Membrane Potential ◽  
Base Line ◽  
Channel Activity ◽  
Human Macrophages ◽  
Excised Patches ◽  
Single Channel Currents

Cell-attached patch studies of cultured human macrophages demonstrate that exposure to ionomycin induces inward-rectifying single-channel currents that differ from the voltage-dependent 28 pS inward-rectifying K currents previously described in these cells (J. Membr. Biol. 103: 55-66, 1988). With 150 mM KCl in the electrode and NaCl Hanks' solution in the bath, the ionomycin-induced single-channel conductance for inward currents was 37 pS, and the reversal potential was 57 mV. Channel activity was often associated with a shift in the base-line current level indicating that the cell membrane potential hyperpolarized. The ability of ionomycin to induce channel activity depended on extracellular [Ca] supporting the view that the channels were gated by calcium. Ionomycin-induced channels were permeable to K, relatively impermeable to Cl or Na, exhibited bursting kinetics, and had no apparent voltage dependence. Barium (3 mM in the patch electrode) did not significantly block the ionomycin-induced channel at rest but blocked channel activity when the patch was hyperpolarized beyond the resting membrane potential. Exposure of macrophages to platelet-activating factor, which is known to increase intracellular [Ca] [( Ca]i) (J. Cell Biol. 103: 439-450, 1986), also transiently induced channel activity. In excised patches with 3 microM [Ca]i bursting inward-rectifying channels with a 41 pS conductance were noted that probably correspond to the ionomycin-induced channels present in cell-attached patches. Increasing [Ca]i from 10(-8) to 3 x 10(-6) M induced inward-rectifying channel activity in previously quiescent excised patches.(ABSTRACT TRUNCATED AT 250 WORDS)

Blockade by cAMP of native sodium channels of adult rat skeletal muscle fibers

1998 ◽  
Vol 275 (6) ◽  
pp. C1465-C1472 ◽  
Author(s):  
Jean-François Desaphy ◽  
Annamaria De Luca ◽  
Diana Conte Camerino
Keyword(s):  
Skeletal Muscle ◽  
Sodium Channels ◽  
Single Channel ◽  
Muscle Fibers ◽  
Rat Skeletal Muscle ◽  
Patch Clamp Technique ◽  
Cyclic Monophosphate ◽  
Adult Rat ◽  
Skeletal Muscle Fibers ◽  
Steady State Inactivation

Although the skeletal muscle sodium channel is a good substrate for cAMP-dependent protein kinase (PKA), no functional consequence was observed for this channel expressed in heterologous systems. Therefore, we investigated the effect of 8-(4-chlorophenylthio)adenosine 3′,5′-cyclic monophosphate (CPT-cAMP), a membrane-permeable cAMP analog, on the native sodium channels of freshly dissociated rat skeletal muscle fibers by means of the cell-attached patch-clamp technique. Externally applied CPT-cAMP (0.5 mM) reduced peak ensemble average currents by ∼75% with no change in kinetics. Single-channel conductance and normalized activation curves were unchanged by CPT-cAMP. In contrast, steady-state inactivation curves showed a reduction of the maximal available current and a negative shift of the half-inactivation potential. Similar effects were observed with dibutyryl adenosine 3′,5′-cyclic monophosphate but not with cAMP, which does not easily permeate the cell membrane. Incubation of fibers for 1 h with 10 μM H-89, a PKA inhibitor, did not prevent the effect of CPT-cAMP. Finally, the β-adrenoreceptor agonist isoproterenol mimicked CPT-cAMP when applied at 0.5 mM but had no effect at 0.1 mM. These results indicate that cAMP inhibits native skeletal muscle sodium channels by acting within the fiber, independently of PKA activation.

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