Membrane domain localization of pH-regulating transporters in frog skeletal muscle membrane vesicles

1996 ◽  
Vol 271 (4) ◽  
pp. C1367-C1379 ◽  
Author(s):  
R. W. Putnam ◽  
P. B. Douglas ◽  
N. A. Ritucci
Keyword(s):  
Skeletal Muscle ◽  
Atpase Activity ◽  
Surface Membrane ◽  
Membrane Vesicles ◽  
Muscle Membrane ◽  
Frog Skeletal Muscle ◽  
Membrane Domain ◽  
Ph Response ◽  
Kinase C ◽  
Inside Out

The distribution of pH-regulating transporters in surface and transverse (T) tubular membrane (TTM) domains of frog skeletal muscle was studied. 2',7'-Bis(carboxyethyl)-5(6)- carboxyfluorescein-loaded giant sarcolemmal vesicles, containing surface membrane, exhibited reversible Na+/H+ exchange. A microsomal vesicle fraction was shown to be enriched in TTM on the basis of high Na(+)-K(+)-ATPase and Mg(2+)-ATPase activity, high ouabain and nitrendipine binding, and low Ca(2+)-ATPase activity. TTM vesicles were well sealed and oriented inside out. Vesicles were loaded with the pH-sensitive dye pyranine. In response to an inwardly directed Na+ gradient, vesicles displayed virtually no alkalinization unless monensin was present. No pH response to an imposed Na+ gradient was seen regardless of the direction of the pH gradient across the vesicles, after phosphorylation of the vesicles with protein kinase C, or when exposed to guanosine 5'-O-(3-thiotriphosphate). In the presence of CO2, addition of Na+ or Cl- had no effect on vesicle pH. These data indicate that the TTM lacks functional pH-regulating transporters [Na+/H+ and (Na+ + HCO3-)/Cl- exchangers], suggesting that pH-regulating transporters are localized only to the surface membrane domain in frog muscle.

The Effect of Lanthanum on Excitation–Contraction Coupling in Frog Skeletal Muscle

1974 ◽  
Vol 52 (6) ◽  
pp. 1126-1135 ◽  
Author(s):  
D. J. Parry ◽  
A. Kover ◽  
G. B. Frank
Keyword(s):  
Skeletal Muscle ◽  
Action Potential ◽  
Muscle Membrane ◽  
Frog Skeletal Muscle ◽  
Tension Development ◽  
Excitation Contraction Coupling ◽  
Contraction Coupling ◽  
Reduced Rate ◽  
Caffeine Contractures

Exposure of frog toe muscles to 1 mM La3+ results in a decrease in amplitude and rate of tension development of potassium contractures and twitches. At this concentration La3+ also inhibits the uptake of calcium, both in the resting condition and during stimulation. Caffeine contractures are unaffected even after a 5-min pre-exposure to La3+. The depolarization induced by various concentrations of K+ is reduced by about 10 mV as is the amplitude of the action potential. The rate of rise of the action potential is reduced by about 40% after 1 min in La3+ Ringer. Neither the decreased amplitude nor the reduced rate of depolarization is considered to be sufficient to explain the inhibition of tension development. It is suggested that La3+ partially uncouples excitation from contraction by preventing the release of a trigger-Ca2+ fraction from some site on the muscle membrane. This fraction normally plays a role in excitation–contraction coupling, although some tension may still be developed in the absence of a trigger-Ca2+ influx.

Kinetics of glucose transport in rat skeletal muscle membrane vesicles: effects of insulin and contractions

1992 ◽  
Vol 262 (5) ◽  
pp. E700-E711 ◽  
Author(s):  
T. Ploug ◽  
H. Galbo ◽  
T. Ohkuwa ◽  
J. Tranum-Jensen ◽  
J. Vinten
Keyword(s):  
Skeletal Muscle ◽  
Plasma Membrane ◽  
Glucose Transport ◽  
Cytochalasin B ◽  
Equilibrium Distribution ◽  
Initial Rate ◽  
Membrane Vesicles ◽  
Membrane Preparation ◽  
Muscle Membrane ◽  
Distribution Space

To study the mechanism of acceleration of glucose transport in skeletal muscle after stimulation with insulin and contractions, we isolated a subcellular vesicular membrane fraction, highly enriched in the plasma membrane enzyme K(+)-stimulated p-nitrophenylphosphatase and also enriched in some intracellular membranes. Protein recovery, morphology, lipid content, marker enzyme activities, total intravesicular volume, Western blot quantitation of GLUT-1, and glucose-inhibitable cytochalasin B binding were identical in membrane fractions from control, insulin-stimulated, contraction-stimulated, and insulin- and contraction-stimulated muscle. Time course of D-[3H]glucose entry in membrane vesicles at equilibrium exchange conditions showed that initial rate of transport at 30 mM of glucose was increased 19-fold and that equilibrium distribution space was increased 4-fold in vesicles from maximum stimulated muscle. The effects of insulin and contractions on initial rate of transport as well as on equilibrium distribution space were additive, and stimulation increased the substrate saturability of glucose transport. Furthermore, cytochalasin B binding to membranes prepared by using less centrifugation time than usual showed that, after stimulation with insulin and contractions, at least 35% of the total number of glucose transporters were redistributed from one kind of vesicles to a more slowly sedimenting kind of vesicles, probably reflecting translocation within the membrane preparation from intracellular vesicles to the plasma membrane upon stimulation. In the present membrane preparation the effects of insulin and/or contractions on glucose transport resemble those seen in intact muscle, and the effects are thus not dependent on cellular integrity.(ABSTRACT TRUNCATED AT 250 WORDS)

High-affinity [3H]PN200-110 and [3H]ryanodine binding to rabbit and frog skeletal muscle

1994 ◽  
Vol 266 (2) ◽  
pp. C462-C466 ◽  
Author(s):  
K. Anderson ◽  
A. H. Cohn ◽  
G. Meissner
Keyword(s):  
Skeletal Muscle ◽  
Rabbit Skeletal Muscle ◽  
Muscle Membrane ◽  
Scatchard Analysis ◽  
Physical Interaction ◽  
Frog Skeletal Muscle ◽  
Release Channel ◽  
High Affinity ◽  
Voltage Dependent ◽  
Student’S T

In vertebrate skeletal muscle, the voltage-dependent mechanism of sarcoplasmic reticulum (SR) Ca2+ release, commonly referred to as excitation-contraction (E-C) coupling, is mediated by the voltage-sensing dihydropyridine receptor (DHPR), which is believed to affect SR Ca2+ release through a physical interaction with the SR ryanodine receptor (RYR)/Ca2+ release channel. Scatchard analysis of ligand binding of [3H]PN200-110 to the DHPR and [3H]ryanodine to the RYR indicated the presence of high-affinity sites in muscle homogenates, with maximum binding (Bmax) values of 72 +/- 26 and 76 +/- 30 pmol/g wet wt for rabbit skeletal muscle, and 27 +/- 14 and 44 +/- 13 pmol/g wet wt for frog skeletal muscle, respectively. The Bmax values corresponded to a PN200-110-to-ryanodine binding ratio of 0.98 +/- 0.26 and 0.61 +/- 0.24 for rabbit and frog skeletal muscle, respectively, and were found by Student's t test to be significantly different (P < 0.02, n = 7). These results are compared with measurements with isolated rabbit skeletal muscle membrane fractions and discussed in relation to our current understanding of the mechanism of E-C coupling in skeletal muscle.

scholarly journals Molecular architecture of membranes involved in excitation-contraction coupling of cardiac muscle.

1995 ◽  
Vol 129 (3) ◽  
pp. 659-671 ◽  
Author(s):  
X H Sun ◽  
F Protasi ◽  
M Takahashi ◽  
H Takeshima ◽  
D G Ferguson ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Cardiac Muscle ◽  
Calcium Release ◽  
Surface Membrane ◽  
Freeze Fracture ◽  
Muscle Membrane ◽  
Charge Movement ◽  
Molecular Architecture ◽  
Direct Role ◽  
Large Particles

Peripheral couplings are junctions between the sarcoplasmic reticulum (SR) and the surface membrane (SM). Feet occupy the SR/SM junctional gap and are identified as the SR calcium release channels, or ryanodine receptors (RyRs). In cardiac muscle, the activation of RyRs during excitation-contraction (e-c) coupling is initiated by surface membrane depolarization, followed by the opening of surface membrane calcium channels, the dihydropyridine receptors (DHPRs). We have studied the disposition of DHPRs and RyRs, and the structure of peripheral couplings in chick myocardium, a muscle that has no transverse tubules. Immunolabeling shows colocalization of RyRs and DHPRs in clusters at the fiber's periphery. The positions of DHPR and RyR clusters change coincidentally during development. Freeze-fracture of the surface membrane reveals the presence of domains (junctional domains) occupied by clusters of large particles. Junctional domains in the surface membrane and arrays of feet in the junctional gap have similar sizes and corresponding positions during development, suggesting that both are components of peripheral couplings. As opposed to skeletal muscle, membrane particles in junctional domains of cardiac muscle do not form tetrads. Thus, despite their proximity to the feet, they do not appear to be specifically associated with them. Two observations establish the identify of the structurally identified feet arrays/junctional domain complexes with the immunocytochemically defined RyRs/DHPRs coclusters: the concomitant changes during development and the identification of feet as the cytoplasmic domains of RyRs. We suggest that the large particles in junctional domains of the surface membrane represent DHPRs. These observations have two important functional consequences. First, the apposition of DHPRs and RyRs indicates that most of the inward calcium current flows into the restricted space where feet are located. Secondly, contrary to skeletal muscle, presumptive DHPRs do not show a specific association with the feet, which is consistent with a less direct role of charge movement in cardiac than in skeletal e-c coupling.

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