scholarly journals Abnormal Na channel gating in murine cardiac myocytes deficient in myotonic dystrophy protein kinase

2003 ◽  
Vol 12 (2) ◽  
pp. 147-157 ◽  
Author(s):  
Hwa C. Lee ◽  
Manoj K. Patel ◽  
Dilawaar J. Mistry ◽  
Qingcai Wang ◽  
Sita Reddy ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Current Density ◽  
Channel Gating ◽  
Resting Membrane Potential ◽  
Na Channel ◽  
Na Channels ◽  
Wild Type ◽  
Membrane Excitability ◽  
Na Current ◽  
Deficient Mice

DMPK is a serine/threonine kinase implicated in the human disease myotonic muscular dystrophy (DM). Skeletal muscle Na channels exhibit late reopenings in Dmpk-deficient mice and peak current density is reduced, implicating DMPK in regulation of membrane excitability. Since complete heart block and sudden cardiac death occur in the disease, we tested the hypothesis that cardiac Na channels also exhibit abnormal gating in Dmpk-deficient mice. We made whole cell and cell-attached patch clamp recordings of ventricular cardiomyocytes enzymatically isolated from wild-type, Dmpk+/−, and Dmpk−/− mice. Recordings from membrane patches containing one or a few Na channels revealed multiple Na channel reopenings occurring after the macroscopic Na current had subsided in both Dmpk+/− and Dmpk−/− muscle, but only rare reopenings in wild-type muscle (>3-fold difference, P < 0.05). This resulted in a plateau of non-inactivating Na current in Dmpk-deficient muscle. The magnitude of this plateau current was independent on the magnitude of the test potential from −40 to 0 mV and was also independent of gene dose. Macroscopic Na current density was similar in wild-type and Dmpk-deficient muscle, as was steady-state Na channel gating. Decay of macroscopic currents was slowed in Dmpk−/− muscle, but not in Dmpk+/− or wild-type muscle. Entry into, and recovery from, inactivation were similar at multiple test potentials in wild-type and Dmpk-deficient muscle. Resting membrane potential was depolarized, and action potential duration was significantly prolonged in Dmpk-deficient muscle. Thus in cardiac muscle, Dmpk deficiency results in multiple late reopenings of Na channels similar to those seen in Dmpk-deficient skeletal muscle. This is reflected in a plateau of non-inactivating macroscopic Na current and prolongation of cardiac action potentials.

A novel mutation in transmembrane protein 168 causes fatal ventricular arrhythmogenesis in Brugada syndrome

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
H Ogita ◽  
D.P Zankov ◽  
A Shimizu
Keyword(s):  
Ventricular Tachycardia ◽  
Current Density ◽  
Brugada Syndrome ◽  
Transmembrane Protein ◽  
Left Ventricular ◽  
Public Institution ◽  
Na Channel ◽  
Funding Source ◽  
Wild Type ◽  
Na Current

Abstract   Brugada syndrome (BrS) is diagnosed by a typical electrocardiography (ECG) with ST-segment elevation in precordial leads and tends to induce sudden cardiac death (SCD) due to ventricular tachycardia/fibrillation. About 20% of SCDs in non-structural cardiac diseases are considered to be caused by BrS. In patients with BrS, loss of function mutations in the Na+ channel is often observed, but the causative gene mutation is not detected for about 70% of BrS patients. Here, we investigated a family with clinically diagnosed BrS, in which no known gene mutations related to BrS had not been found, by whole exome sequencing. Novel heterozygous variant (c. 1616G&gt;A, p. R539Q) in transmembrane protein 168 (TMEM168) was identified only in symptomatic family members. Similar to endogenous TMEM168, both wild-type and mutant TMEM168 localized at the nuclear membrane. Na+ current density in whole-cell patch-clamp recordings was significantly reduced in HL-1 cardiomyocytes transfected with TMEM168 R539Q mutant, compared with those with wild-type TMEM168. Next, heterozygous Tmem168 1616G&gt;A knock-in mice were generated by the CRISPR/Cas9 genome editing technology. Although the knock-in mice had no abnormalities in ECG at the physiological state, the treatment with ajmaline caused various arrhythmias including ventricular tachycardia/fibrillation in the knock-in mice, but not in wild-type mice. Na+ current density and the parameters of action potentials were remarkably impaired in the cardiomyocytes of the knock-in mice. Optical mapping analysis in the whole heart showed the reduced left ventricular conduction velocity in the knock-in mice. The expression of Nav1.5, an α-subunit of the cardiac Na+ channel, was significantly decreased in the mutant TMEM168-transfected HL-1 cells and the knock-in hearts. We found that the decrease was caused by the enhanced ubiquitination of Nav1.5, which was mediated by increased binding of Nedd4–2 E3 ubiquitin ligase to Nav1.5 in the knock-in hearts. Co-immunoprecipitation experiments demonstrated that overactivity of Nedd4–2 is a result of Tmem168 mutant-mediated sequestration of a chaperon protein αB-crystallin, a Nav1.5-binding molecule that interferes with the interaction of Nedd4–2 with Nav1.5. These findings reveal the molecular mechanism of TMEM168 R539Q mutation-induced fatal ventricular arrhythmias in BrS. Funding Acknowledgement Type of funding source: Public Institution(s). Main funding source(s): JSPS Grants-in-aid for Scientific Research

scholarly journals Na channel distribution in vertebrate skeletal muscle.

1986 ◽  
Vol 87 (6) ◽  
pp. 907-932 ◽  
Author(s):  
J H Caldwell ◽  
D T Campbell ◽  
K G Beam
Keyword(s):  
Skeletal Muscle ◽  
Current Density ◽  
Sharp Decrease ◽  
Na Channel ◽  
Na Current ◽  
Na Currents ◽  
Current Densities ◽  
Kinetics Of ◽  
A Current ◽  
Vertebrate Skeletal Muscle

The loose patch voltage clamp has been used to map Na current density along the length of snake and rat skeletal muscle fibers. Na currents have been recorded from (a) endplate membrane exposed by removal of the nerve terminal, (b) membrane near the endplate, (c) extrajunctional membrane far from both the endplate and the tendon, and (d) membrane near the tendon. Na current densities recorded directly on the endplate were extremely high, exceeding 400 mA/cm2 in some patches. The membrane adjacent to the endplate has a current density about fivefold lower than that of the endplate, but about fivefold higher than the membrane 100-200 micron from the endplate. Small local variations in Na current density are recorded in extrajunctional membrane. A sharp decrease in Na current density occurs over the last few hundred micrometers from the tendon. We tested the ability of tetrodotoxin to block Na current in regions close to and far from the endplate and found no evidence for toxin-resistant channels in either region. There was also no obvious difference in the kinetics of Na current in the two regions. On the basis of the Na current densities measured with the loose patch clamp, we conclude that Na channels are abundant in the endplate and near-endplate membrane and are sparse close to the tendon. The current density at the endplate is two to three orders of magnitude higher than at the tendon.

scholarly journals Cardiac sodium channels (hH1) are intrinsically more sensitive to block by lidocaine than are skeletal muscle (mu 1) channels.

1995 ◽  
Vol 106 (6) ◽  
pp. 1193-1209 ◽  
Author(s):  
H B Nuss ◽  
G F Tomaselli ◽  
E Marbán
Keyword(s):  
Skeletal Muscle ◽  
Local Anesthetics ◽  
Time Course ◽  
Large Fraction ◽  
Expression System ◽  
Na Channel ◽  
Na Channels ◽  
Na Current ◽  
Unexpected Findings ◽  
Cardiac Sodium Channels

When lidocaine is given systemically, cardiac Na channels are blocked preferentially over those in skeletal muscle and nerve. This apparent increased affinity is commonly assumed to arise solely from the fact that cardiac Na channels spend a large fraction of their time in the inactivated state, which exhibits a high affinity for local anesthetics. The oocyte expression system was used to compare systematically the sensitivities of skeletal (mu 1-beta 1) and cardiac (hH1-beta 1) Na channels to block by lidocaine, under conditions in which the only difference was the choice of alpha subunit. To check for differences in tonic block, Na currents were elicited after 3 min of exposure to various lidocaine concentrations at -100 mV, a potential at which both hH1-beta 1 and mu 1-beta 1 channels were fully reprimed. Surprisingly, hH1-beta 1 Na channels were threefold more sensitive to rested-state block by lidocaine (402 +/- 36 microM, n = 4-22) than were mu 1-beta 1 Na channels (1,168 +/- 34 microM, n = 7-19). In contrast, the inactivated state binding affinities determined at partially depolarized holding potentials (h infinity approximately 0.2) were similar (Kd = 16 +/- 1 microM, n = 3-9 for hH1-beta 1 and 12 +/- 2 microM, n = 4-11 for mu 1-beta 1). Lidocaine produced more use-dependent block of peak hH1-beta 1 Na current elicited by trains of short-(10 ms) or long- (1 s) duration step depolarizations (0.5 Hz, -20 mV) than of mu 1-beta 1 Na current. During exposure to lidocaine, hH1-beta 1 channels recover from inactivation at -100 mV after a prolonged delay (20 ms), while mu 1-beta 1 channels begin repriming immediately. The overall time course of recovery from inactivation in the presence of lidocaine is much slower in hH1-beta 1 than in mu 1-beta 1 channels. These unexpected findings suggest that structural differences in the alpha subunits impart intrinsically different lidocaine sensitivities to the two isoforms. The differences in steady state affinities and in repriming kinetics are both in the correct direction to help explain the increased potency of cardiac Na channel block by local anesthetics.

Mechanism of modulation of the voltage-gated skeletal and cardiac muscle sodium channels by fatty acids

1997 ◽  
Vol 272 (2) ◽  
pp. C592-C600 ◽  
Author(s):  
S. Bendahhou ◽  
T. R. Cummins ◽  
W. S. Agnew
Keyword(s):  
Skeletal Muscle ◽  
Fatty Acids ◽  
Arachidonic Acid ◽  
Unsaturated Fatty Acids ◽  
Inactivation Kinetics ◽  
Na Channel ◽  
Na Channels ◽  
Channel Conductance ◽  
Na Current ◽  
Voltage Gated

Voltage-gated rat skeletal muscle and cardiac Na+ channels are modulated by exogenous unsaturated fatty acids. Application of 1-10 microM arachidonic or oleic acids reversibly depressed Na+ channel conductance and shifted the inactivation curve to hyperpolarizing potentials. These effects were not prevented by inhibitors of lipoxygenase, cyclooxygenase, cytochrome P-450 epoxygenase, or protein kinase C. Neither palmitic acid nor methyl ester oleate had an effect on the inward Na+ current, suggesting that trivial variations in membrane fluidity are not responsible for the Na+ current depression or kinetic changes. Arachidonic acid altered fast Na+ inactivation without changing the slow inactivation kinetics. Moreover, skeletal muscle Na+ channel gating currents were markedly decreased by 2 microM arachidonic acid. Finally, nonstationary noise analysis indicated that both the number of channels and the open probability were slightly decreased without change in the single-channel conductance. These data suggest that unsaturated fatty acids such as arachidonic and oleic acids 1) specifically regulate voltage-gated Na+ channels and 2) interact directly with Na+ channels, perhaps at a fatty acid binding domain, by decreasing the total gating charge and altering fast-inactivation kinetics.

Skeletal muscle Na currents in mice heterozygous for Six5 deficiency

2001 ◽  
Vol 6 (3) ◽  
pp. 153-158 ◽  
Author(s):  
DILAAWAR J. MISTRY ◽  
J. RANDALL MOORMAN ◽  
SITA REDDY ◽  
J. PAUL MOUNSEY
Keyword(s):  
Skeletal Muscle ◽  
Protein Kinase ◽  
Myotonic Dystrophy ◽  
Trinucleotide Repeat ◽  
Channel Gating ◽  
Na Channel ◽  
Wild Type ◽  
Na Currents ◽  
Significant Difference ◽  
Murine Skeletal Muscle

Myotonic dystrophy results from a trinucleotide repeat expansion between the myotonic dystrophy protein kinase gene ( Dmpk), which encodes a serine-threonine protein kinase, and the Six5 gene, which encodes a homeodomain protein. The disease is characterized by late bursts of skeletal muscle Na channel openings, and this is recapitulated in Dmpk −/− and Dmpk +/− murine skeletal muscle. To test whether deficiency of the nearby Six5 gene also affected Na channel gating in murine skeletal muscle, we measured Na currents from cell-attached patches in Six5 +/− mice and age-matched wild-type and Dmpk +/− mice. Late bursts of Na channel activity were defined as an opening probability >10% measured from 10 to 110 ms after depolarization. There was no significant difference in the occurrence of late Na channel bursts in wild-type and Six5 +/− muscle, whereas in Dmpk +/− muscle there was greater than fivefold increase in late bursts ( P < 0.001). Compared with wild-type mice, Na current amplitude was unchanged in Six5 +/− muscle, whereas in Dmpk +/− muscle it was 36% reduced ( P < 0.05). Thus, since Six5 +/− mice do not exhibit the Na channel gating abnormality of Dmpk deficiency, we conclude that Six5 deficiency does not contribute to the Na channel gating abnormality seen in dystrophia myotonica patients.

Treadmill running causes significant fiber damage in skeletal muscle of KATP channel-deficient mice

2005 ◽  
Vol 22 (2) ◽  
pp. 204-212 ◽  
Author(s):  
M. Thabet ◽  
T. Miki ◽  
S. Seino ◽  
J.-M. Renaud
Keyword(s):  
Skeletal Muscle ◽  
Muscle Fibers ◽  
Treadmill Running ◽  
Wild Type ◽  
Connective Tissues ◽  
Fiber Damage ◽  
Hindlimb Muscles ◽  
Skeletal Muscle Fibers ◽  
And Function ◽  
Deficient Mice

Although it has been suggested that the ATP-sensitive K+ (KATP) channel protects muscle against function impairment, most studies have so far given little evidence for significant perturbation in the integrity and function of skeletal muscle fibers from inactive mice that lack KATP channel activity in their cell membrane. The objective was, therefore, to test the hypothesis that KATP channel-deficient skeletal muscle fibers become damaged when mice are subjected to stress. Wild-type and KATP channel-deficient mice (Kir6.2−/− mice) were subjected to 4–5 wk of treadmill running at either 20 m/min with 0° inclination or at 24 m/min with 20° uphill inclination. Muscles of all wild-type mice and of nonexercised Kir6.2−/− mice had very few fibers with internal nuclei. After 4–5 wk of treadmill running, there was little evidence for connective tissues and mononucleated cells in Kir6.2−/− hindlimb muscles, whereas the number of fibers with internal nuclei, which appear when damaged fibers are regenerated by satellite cells, was significantly higher in Kir6.2−/− than wild-type mice. Between 5% and 25% of the total number of fibers in Kir6.2−/− extensor digitum longus, plantaris, and tibialis muscles had internal nuclei, and most of such fibers were type IIB fibers. Contrary to hindlimb muscles, diaphragms of Kir6.2−/− mice that had run at 24 m/min had few fibers with internal nuclei, but mild to severe fiber damage was observed. In conclusion, the study provides for the first time evidence 1) that the KATP channels of skeletal muscle are essential to prevent fiber damage, and thus muscle dysfunction; and 2) that the extent of fiber damage is greater and the capacity of fiber regeneration is less in Kir6.2−/− diaphragm muscles compared with hindlimb muscles.

Na current density at and away from end plates on rat fast- and slow-twitch skeletal muscle fibers

1992 ◽  
Vol 262 (1) ◽  
pp. C229-C234 ◽  
Author(s):  
R. L. Ruff
Keyword(s):  
Skeletal Muscle ◽  
Current Density ◽  
Muscle Fibers ◽  
Postsynaptic Membrane ◽  
Na Current ◽  
End Plate ◽  
Skeletal Muscle Fibers ◽  
Slow Twitch ◽  
Slow Twitch Fibers ◽  
Fast Twitch

Na current density and membrane capacitance were studied with the loose patch voltage clamp technique on rat fast- and slow-twitch skeletal muscle fibers at three different regions on the fibers: 1) the end plate border, 2) greater than 200 microns from the end plate (extrajunctional), and 3) on the end plate postsynaptic membrane. Fibers were treated with collagenase to improve visualization of the end plate and to enzymatically remove the nerve terminal. The capacitance of membrane patches was similar on fast- and slow-twitch fibers and patches of membrane on the end plate had twice the capacitance of patches elsewhere. For fast- and slow-twitch fibers, the sizes of the Na current normalized to the area of the patch were as follows: end plate greater than end plate border greater than extrajunctional. For both types of fibers, the amplitudes of the Na current normalized to the capacitance of the membrane patch were as follows: end plate approximately end plate border greater than extrajunctional. At each of the three regions, the Na current densities were larger on fast-twitch fibers and fast-twitch fibers had a larger increase in Na current density at the end plate border compared with extrajunctional membrane.

Urokinase-dependent plasminogen activation is required for efficient skeletal muscle regeneration in vivo

Blood ◽  
2001 ◽  
Vol 97 (6) ◽  
pp. 1703-1711 ◽  
Author(s):  
Frederic Lluı́s ◽  
Josep Roma ◽  
Mònica Suelves ◽  
Maribel Parra ◽  
Gloria Aniorte ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Plasminogen Activator ◽  
Muscle Regeneration ◽  
Plasminogen Activators ◽  
Skeletal Muscle Regeneration ◽  
Plasminogen Activation ◽  
Wild Type ◽  
Type Plasminogen Activator ◽  
Deficient Mice

Plasminogen activators urokinase-type plasminogen activator (uPA) and tissue-type plasminogen activator (tPA) are extracellular proteases involved in various tissue remodeling processes. A requirement for uPA activity in skeletal myogenesis was recently demonstrated in vitro. The role of plasminogen activators in skeletal muscle regeneration in vivo in wild-type, uPA-deficient, and tPA-deficient mice is investigated here. Wild-type and tPA−/− mice completely repaired experimentally damaged skeletal muscle. In contrast, uPA−/− mice had a severe regeneration defect, with decreased recruitment of blood-derived monocytes to the site of injury and with persistent myotube degeneration. In addition, uPA-deficient mice accumulated fibrin in the degenerating muscle fibers; however, the defibrinogenation of uPA-deficient mice resulted in a correction of the muscle regeneration defect. A similar severe regeneration deficit with persistent fibrin deposition was also reproducible in plasminogen-deficient mice after injury, suggesting that fibrinolysis by uPA-mediated plasminogen activation plays a fundamental role in skeletal muscle regeneration. In conclusion, the uPA-plasmin system is identified as a critical component of the mammalian skeletal muscle regeneration process, possibly because it prevents intramuscular fibrin accumulation and contributes to the adequate inflammatory response after injury. These studies demonstrate the requirement of an extracellular proteolytic cascade during muscle regeneration in vivo.

scholarly journals Skeletal muscle reperfusion injury is mediated by neutrophils and the complement membrane attack complex

1999 ◽  
Vol 277 (6) ◽  
pp. C1263-C1268 ◽  
Author(s):  
Constantinos Kyriakides ◽  
William Austen ◽  
Yong Wang ◽  
Joanne Favuzza ◽  
Lester Kobzik ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Reperfusion Injury ◽  
Membrane Attack Complex ◽  
Wild Type ◽  
Complement Components ◽  
Neutrophil Activity ◽  
Deficient Mice ◽  
Terminal Complement

The relative inflammatory roles of neutrophils, selectins, and terminal complement components are investigated in this study of skeletal muscle reperfusion injury. Mice underwent 2 h of hindlimb ischemia followed by 3 h of reperfusion. The role of neutrophils was defined by immunodepletion, which reduced injury by 38%, as did anti-selectin therapy with recombinant soluble P-selectin glycoprotein ligand-immunoglobulin (Ig) fusion protein. Injury in C5-deficient and soluble complement receptor type 1-treated wild-type mice was 48% less than that of untreated wild-type animals. Injury was restored in C5-deficient mice reconstituted with wild-type serum, indicating the effector role of C5–9. Neutropenic C5-deficient animals showed additive reduction in injuries (71%), which was lower than C5-deficient neutrophil-replete mice, indicating neutrophil activity without C5a. Hindlimb histological injury was worse in ischemic wild-type and C5-deficient animals reconstituted with wild-type serum. In conclusion, the membrane attack complex and neutrophils act additively to mediate skeletal muscle reperfusion injury. Neutrophil activity is independent of C5a but is dependent on selectin-mediated adhesion.

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