Voltage-sensitive calcium flux promoted by vesicles in an isolated cardiac sarcolemma preparation

Schilling, William P.; Lindenmayer, George E.

doi:10.1007/bf01872120

Cited by 21 publications

(8 citation statements)

References 53 publications

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“…In fact, ouabain is able to bind to its site under rather unfavorable conditions, that is, when no specific ligands are added to assay solutions (Forbush, 1983). Furthermore, our results are in agreement with previous observations of significant binding of labeled ouabain to sarcolemmal vesicles isolated from dog heart in the absence of supporting ligands (Wellsmith & Lindenmayer, 1980;Schilling & Drewe, 1986). Thus, the ouabain-sensitive S6Rb+ influx observed probably reflects slow passive Rb+-K + exchange fluxes through the sodium pump, first described for purified enzyme reconstituted in phospholipid vesicles (Karlish & Stein, 1982a), and subsequently identified in resealed erythrocyte ghosts (Kenney & Kaplan, 1986;Sachs, 1986).…”

Section: Role Of the Sodium Pump In Potassium Transportsupporting

confidence: 82%

Role of the sodium pump and the background K+ channel in passive K+(Rb+) uptake by isolated cardiac sarcolemmal vesicles

Otero

Szabó

1988

J. Membrain Biol.

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A simple procedure was developed for the isolation of a sarcolemma-enriched membrane preparation from homogenates of bullfrog (Rana catesbeiana) heart. Crude microsomes obtained by differential centrifugation were fractionated in Hypaque density gradients. The fraction enriched in surface membrane markers consisted of 87% tightly sealed vesicles. The uptake of 86Rb+ by the preparation was measured in the presence of an opposing K+ gradient using a rapid ion exchange technique. At low extravesicular Rb+ concentrations, at least 50% of the uptake was blocked by addition of 1 mM ouabain to the assay medium. Orthovanadate (50 microM), ADP (2.5 mM) or Mg (1 mM) were also partial inhibitors of Rb+ uptake under these conditions, and produced a complete block of Rb+ influx in the presence of 1 mM ouabain. When 86Rb+ was used as a tracer of extravesicular K+ (Rb+0 less than or equal to 40 microM, K+0 = 0.1-5 mM) a distinct uptake pathway emerged, as detected by its inhibition by 1 mM Ba2+ (K0.5 = 20 microM). At a constant internal K+ concentration (K+in = 50 mM), the magnitude of the Ba2+-sensitive K+ uptake was found to depend on K+0 in a manner that closely resembles the K+ concentration dependence of the background K+ conductance (IK1) observed electrophysiologically in intact cardiac cells. We conclude that K+ permeates passively this preparation through two distinct pathways, the sodium pump and a system identifiable as the background potassium channel.

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Section: Role Of the Sodium Pump In Potassium Transportsupporting

confidence: 82%

Role of the sodium pump and the background K+ channel in passive K+(Rb+) uptake by isolated cardiac sarcolemmal vesicles

Otero

Szabó

1988

J. Membrain Biol.

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“…At depolarized holding potentials the unbinding rate could be so small due to slowing of the conformational change. The binding of nitrendipine in the open state and in the inactivated state at depolarized holding potentials is supported by the radio-binding studies, which have revealed that the affinity of the receptor for 1,4-dihydropyridine is enhanced intensely by depolarization (Schilling & Drewe, 1986;Kokubun, Prod'hom, Becker, Porzig & Reuter, 1986).…”

Section: Existence Of Multiple Unavailable Statesmentioning

confidence: 93%

Voltage‐dependent decrease in the availability of single calcium channels by nitrendipine in guinea‐pig ventricular cells.

Kawashima

Ochi

1988

The Journal of Physiology

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SUMMARY1. The mechanism of Ca2+ channel block by nitrendipine was studied by recording single-channel activity from cell-attached patches on guinea-pig ventricular cells using patch pipettes containing 50 mM-Ba2+. Test depolarization pulses to around 10 mV with a duration of 100 ms were applied repetitively at 2 Hz.2. The percentage of non-blank sweeps was maximal (about 40%) at a holding potential between -65 and -130 mV and decreased sigmoidally with its depolarization. Nitrendipine shifted the availability-voltage relationship in a hyperpolarizing direction.3. From the number of consecutive non-blank sweeps and that of blank sweeps, the duration of the available state and that of the unavailable state were estimated.4. The histogram of the duration of the available state showed a singleexponential distribution. Its mean duration was about 1-5 s and was shortened by nitrendipine. Correspondingly, the decay of the mean current during the depolarization step was accelerated by nitrendipine.5. In the presence of 100 nM-nitrendipine the histogram of the duration of the unavailable state at large negative holding potentials was simulated as the sum of two exponential components, one with a time constant similar to that in the control and the other with a time constant of 6-7 s.6. The histogram of the duration of the unavailable state at depolarized holding potentials was simulated by a double-exponential curve also in the control. The duration of the slow component was prolonged by nitrendipine.7. The prolongation of the unavailable states initiated by drug binding during depolarization steps and maintained during depolarized holding potentials is the mechanism of the blockade. The rate constants of the state transitions between an available state and two unavailable states were estimated.

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“…concentrations that affect properties such as calcium flux responses or non-linear charge movement (Dunn, 1989;Ohkusa et al, 1991;Rfos & Pizarro, 1991). One possible explanation for this discrepancy is the regulation of DHP-binding affinity by different factors such as membrane potential or divalent-cation concentration (Schilling & Drewe, 1986;Hosey & Lanzdunski, 1988). One alternative possibility for the discrepancy between high-affinity binding and low-affinity pharmacology is the existence of multiple DHP-binding sites, having different affinities and leading to different responses.…”

Section: Discussionmentioning

confidence: 99%

“…In studies with frog skeletal muscle T-tubules, canine cardiac sarcolemma and chick heart membranes, comparable temperature effects on [3H]-nitrendipine binding affinity have been described (Jaimovich et al, 1986;Schilling & Drewe, 1986;Maan & Hosey, 1987 , 1984). It has been proposed that the decreased affinity for DHP observed at 37 ° C may be related with the more rapid dissociation which occurs at elevated temperatures (Hosey & Lanzdunski, 1988).…”

Section: Temperaturementioning

confidence: 99%

Dihydropyridine receptors in transverse tubules from normal and dystrophic chicken skeletal muscle

Moro

Saborido

Delgado

et al. 1995

J Muscle Res Cell Motil

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Calcium overload is a fundamental pathogenic event associated with chronic muscle degeneration in muscular dystrophies. The possibility that L-type voltage-dependent calcium channels were involved in the etiology of chicken muscular dystrophy was investigated by studying the dihydropyridine receptors in transverse tubule membranes isolated from skeletal muscle of normal (line 412) and dystrophic (line 413) chickens. The yield of T-tubular protein from dystrophic muscle was considerably increased compared with that from normal muscle (2.51 +/- 0.18 vs 1.04 +/- 0.31 mg protein x 100 g muscle-1). The binding of the calcium channel antagonist (+) [3H]PN200-110 to the dihydropyridine receptor in transverse tubule preparations was relatively slow, markedly affected by temperature and required divalent cations. (+) [3H]PN200-110 equilibrium binding assays revealed a single class of high-affinity sites and showed that maximum binding capacity (Bmax) (3.17 +/- 0.47 for normal and 3.51 +/- 0.52 pmol x mg protein-1 for dystrophic transverse tubules) and dissociation constant (Kd) (0.32 +/- 0.07 and 0.26 +/- 0.09 nM, respectively) were not significantly different in normal and dystrophic membranes. Kinetic studies indicated that normal and dystrophic transverse tubules did not differ significantly in association (2.54 x 10(6) and 2.27 x 10(6) M(-1)s(-1), respectively) and dissociation (8.5 x 10(-4) and 9.3 x 10(-4)s(-1), respectively) rate constants. Since dissociation kinetics for both preparations were monoexponential under all the experimental conditions employed, no low-affinity binding sites for (+) [3H]PN200-110 could be detected in chicken transverse tubules membranes. However, immunoblot assay, using a monoclonal antibody, revealed that dystrophic transverse tubules as compared with normal membranes were enriched twofold with the alpha 1-subunit of the dihydropyridine receptor. Therefore, although dihydropyridine-binding sites were not altered in transverse tubule membranes from dystrophic chicken skeletal muscle, both the increased yield in T-tubule vesicles and the enhanced immunodetection of the alpha 1-subunit of the dihydropyridine receptor, suggest that total content in dihydropyridine receptor is higher in dystrophic than in normal muscle.

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Voltage-sensitive calcium flux promoted by vesicles in an isolated cardiac sarcolemma preparation

Cited by 21 publications

References 53 publications

Role of the sodium pump and the background K+ channel in passive K+(Rb+) uptake by isolated cardiac sarcolemmal vesicles

Role of the sodium pump and the background K+ channel in passive K+(Rb+) uptake by isolated cardiac sarcolemmal vesicles

Voltage‐dependent decrease in the availability of single calcium channels by nitrendipine in guinea‐pig ventricular cells.

Dihydropyridine receptors in transverse tubules from normal and dystrophic chicken skeletal muscle

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