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1.
A model is proposed for Ca2+ stationary exchange in the myometrium cells in the absence of effects activating calcium channels of the plasmic membrane. The results of the model analysis point to an important role of the calcium pump (but not on Na(+)-Ca2+ exchanger) of the sarcolemma in providing the long-term regulation of physiologically significant concentration of ionized calcium in the smooth muscle cells. Ability of the calcium pump to efficiently compensate Ca2+ basal current continuously entering the myocytes at rest is proved. It is suggested that the stationary transsarcolemmal exchange of calcium (the system "basal calcium current--ATP-dependent transfer of Ca2+") underlies the control mechanism of the myometrium basal tonus, while a disturbance of the stationary state (with the pump inhibition) provides activation of the smooth muscle tonic contraction.  相似文献   

2.
Ca2+ concentration has been estimated in isolated myometrium cells using Ca2(+)-sensitive quin-2 fluorescent probe. Two components of Ca permeability of the plasmatic membrane have been determined, a potential-independent one (activated by K+ depolarization and nitrendipine-sensitive), and a basal one (not sensitive to nitrendipine). Smooth muscle cells could maintain intracellular Ca2+ concentration at the physiological level. In the presence of nitrendipine, orthovanadate, an inhibitor of sarcolemma Ca pump, induced the increase in the basal tonus depending on the presence of the Ca2+ in the medium. This suggests that in conditions of the blockage of electrically controlled Ca channels and Ca pump of the plasmatic membrane, the noncompensated basal Ca2+ influx activates the tonic contraction of smooth muscles.  相似文献   

3.
Oxytocin and sigetin were studied for their effect on the active and passive transport of Ca2+ in the fraction of myometrium sarcolemma in women. Oxytocin (5.10(-7) M) introduced into the sarcolemma vesicles and sigetin (5.10(-3) M) added into the incubation medium inhibit Mg2+, ATP-dependent accumulation of Ca2+ in these structures. The both agents in the mentioned concentration do not affect the passive release of cation from vesicles. A conclusion is drawn that inhibition of the calcium pump of myometrium cell plasma membranes underlies the physiological action of oxytocin and sigetin as stimulators of the contractile activity of the myometrium.  相似文献   

4.
It is proved that in the fraction of inverted vesicles of the myometrium sarcolemma there are two components of calcium metabolism which depend on the proton concentration in the incubation medium. The first component, a static one, identified under alkalization of the incubation medium from pH 6.0 up to pH 8.0 under equilibrium conditions (Ca2+ concentration inside and outside vesicles is the same) is manifested as an increase of the calcium capacity of vesicles at the expense of Ca2+-binding centres of the inner surface of membrane vesicles. The second component, a dynamic one, is represented as a passive transmembrane flow of Ca2+ outflowing from the vesicles induced by alkalization of the extravesicle space. Alkalization-stimulated Ca2+ release from vesicles is analyzed kinetically. Possible functional role of two components of pH-dependent metabolism of Ca2+ in providing the electrical and pharmacological-mechanical conjugation in the smooth-muscular tissue is under discussion.  相似文献   

5.
Some peculiarities of Ca2+ exchange in the vesiculate fraction of myometrium sarcolemma during separate and combined functioning of the Ca-pump and Na(+)-Ca2+ antiporter in the presence of initial physiologically significant transmembrane gradients of Ca2+ and Na+ were studied. The effect of synergistic activation of the transfer substrate accumulation inside the vesicles was demonstrated. This effect was observed both in the presence of inside-out directed Ca2+ gradient and in its absence. At Ca2+ concentrations in the extravesicular space equimolar to those in contracted myocytes (5 x 10(-6)-10(-5) M), the co-functioning of the cationic antiporter and Ca-pump provided for effective translocation of the transfer substrate to the vesicles which fully prevented the dissipation of the initial oppositely directed Ca2+ gradient. The synergism of energy-dependent calcium fluxes seemed to be unrelated to changes in the chemical composition of the ATP-containing incubation medium responsible for the induction of Mg2+, ATP- and Na(+)-dependent Ca2+ transfer (addition to the medium of Mg2+ and isotonic replacement of Na+ for choline+, respectively). It is concluded that the observed synergism is due to the stimulating effect of the Na+ gradient on the turnover number of the myometrium sarcolemma Ca-pump.  相似文献   

6.
Studies with sarcolemma from cattle myometrium containing inside-out cytoplasmic vesicles, using Ca2+-EGTA buffer, showed that the affinity of ionized Ca2+ for the Mg2+- or ATP-dependent transport is higher than that for the Na+-Ca2+ exchange system (Kd = 3,2 X 10(-6) and (4.3-5.3) X 10(-5) M), respectively. The Km values for MgATP are 2.15 mM. Oxytocin added to the homogenization medium containing rabbit and cattle myometrium cells, i.e. during the formation of closed sarcolemmal fragments, resulted in inhibition of Mg2+, ATP-dependent accumulation of 45Ca2+ by plasma membranes. However, an addition of oxytocin to the incubation medium did not affect the kinetics of active accumulation of Ca2+. It was assumed that the system of non-electrogenic Na+-Ca2+ exchange in the myometrium possessing a low affinity for Ca2+ provides for the maintenance of ionized Ca2+ concentration in the myocytes at 10(-5) M. Therefore, this system cannot induce relaxation of mechanical tension of the uterus. Further decrease of Ca2+ in the myoplasm from 10(-5) to 10(-7) M and, correspondingly, the relaxation of myometrium is provided for by the Mg2+, ATP-dependent efflux of Ca2+ from the myocytes having a high affinity for this cation. The decrease of the activity of ATP-dependent Ca2+-pump by oxytocin is the cause of Ca2+ elevation in the myoplasm and, consequently, of myometrium contraction.  相似文献   

7.
It has been found that Ca-pump of the smooth muscle sarcolemma has much greater affinity to Ca2+ (Km = 0.5 M) than the system Na-Ca2+ of the exchanger (Km = 40-60 M). The maximal rate of Mg2+, ATP-dependent translocation of Ca2+ is 2-3 times higher than that of Na-dependent. The results of kinetic analysis show that Ca-pump of the smooth muscle sarcolemma is able to compensate the basal diffusion flow of this cation entering into unexcited cells of smooth muscle (5 x 10(-15) mol Ca2+ per 1 cm2 for 1 sec). It can also stationary support the value of Ca2+ concentration in relaxed myocytes on a physiologically significant level (10(-7)-10(-6) M).  相似文献   

8.
Using atomic absorption spectroscopy, it was shown that the amount of firmly bound Ca2+ in cattle mitochondria and myometrium sarcolemma is 160 +/- 10 and 30 +/- 10 mumol/kg of wet tissue, respectively. The Ca2+ 1 accumulating capacity of mitochondria (350 nmol per mg of protein) markedly exceeds that of sarcolemmal vesicles (30 nmol per mg of protein). Using a Ca2+-EGTA buffer, it was found that the affinity of ionized Ca for the mitochondrial transport system (Km = 5.69 microM) is higher than that for the Na+-Ca2+ system of sarcolemma exchange (Km = 30 microM), but is markedly lower than that for the Mg2+, ATP-dependent Ca2+ efflux (Km = 0.35 microM). A kinetic analysis demonstrated that the sarcolemmal Ca2+ pump is incapable of causing complete relaxation of the smooth muscle within the physiologically significant time, whereas the Ca2+ transport system of mitochondria evokes this process within 21 s. However, the contribution of the Ca2+ pump to the regulation of the Ca2+ content in myocytes is paralleled with the accumulation of Ca2+ in mitochondria and is realized at low concentrations of this cation in the myoplasm, i.e., at late steps of relaxation. A mechanism of Ca2+ control over myometrium relaxation is proposed. The system of non-electrogenic Na+-Ca2+ exchange maintains Ca2+ concentration in the myoplasm as high as 10(-5) M. Mitochondria which accumulate the bulk of Ca2+ rapidly decrease its concentration in the cytoplasm down to 10(-6)-10(-7) M; at these values, the activity of the sarcolemmal Ca2+ pump with a high affinity for the transfer substrate is manifested. In this way, the Ca2+ pump accomplishes fine regulation of Ca2+ concentration in the myocytes.  相似文献   

9.
The apparent values of intravesicular volume (45 microliter/mg of protein), maximal capacity of adsorbed calcium binding on the inner surface of the vesicles (4.5 nmol/mg of protein) and dissociation constants for the Ca2+-binding site complexes (36 microM) were determined from the analysis of peculiarities of passive transport of 45Ca2+ into cow myometrium sarcolemmal vesicles. The kinetics of passive efflux of ionized Ca2+ from the vesicles is described by a two-phase exponential curve. Dilution of the vesicles with a dilution medium is associated with a rapid efflux of ionized Ca2+ from the intravesicular space resulting in dissociation of the Ca2+-binding site complexes on the inner surface of the vesicles and, correspondingly, in the passage from a rapid to the slow phase of Ca2+ efflux from the vesicles which is limited by the dissociation of the Ca2+-binding site complexes. The values of the apparent rate constants for the transmembrane transfer of Ca2+ and dissociation of the Ca2+-binding site complexes (0.73 and 0.02 min-1, respectively) and the permeability of sarcolemmal vesicles for the cation (10(-15) mol of Ca2+/cm2.s) were determined. Alkalinization of the dilution medium stimulates 45Ca2+ release from the vesicles. The blockers of passive Co2+ and Mn2+ transport injected into the vesicles inhibit the efflux of 45Ca2+ from the vesicles. The data obtained were used to analyze the role of sarcolemma in the Ca2+ control of myometrium contraction.  相似文献   

10.
Vanadate (10(-4)-10(-3) M) effectively blocks Mg2+, ATP-dependent Ca2+ transport in sarcolemmal vesicles and induces a slowly tonic contraction of the smooth muscle. This contraction was observed both with and without nifedipine (10(-5) M) evoking complete inhibition of hyperpotassium contracture, the Ca2+ removal from the solution washing the muscular preparation stimulating the tone decrease. There is a close correlation between the dose-dependent effects of vanadate on the Ca pump activity and tension. It is concluded that in smooth muscles, at least in myometrium, the sarcolemmal Ca-pump is involved into the control of the tonic tension.  相似文献   

11.
The ways and mechanisms of the Ca2+ concentration regulation in myometrium cells are analyzed. The plasma membrane is thoroughly studied for its role in the calcium control provision for the contractile activity of the uterus. The systems of Mg2+-ATP-dependent transport of Ca2+, sodium-calcium metabolism as well as regularities of the Ca2+ passive transfer in the sarcolemma vesicles are considered. The systems of the Mg2+-ATP- and N+-dependent transport of calcium are discussed for their contribution into regulation of calcium concentration in the myoplasm. Oxytocin and ions of bivalent metals (stimulators of the contractile activity of the uterus) are studied for their effect on the activity of the sarcolemma calcium pump.  相似文献   

12.
The published data on the mechanisms and regulation of active and passive Ca2+ transport in the myometrium have been analyzed. Particular attention is paid to the cGMP-dependent and independent pathways of action of nitric oxide or its derivatives on intracellular Ca2+ homeostasis of uterine smooth muscle and its contractile activity. Information on the effect of nitric oxide on Ca2+ -transport systems of other types of smooth muscles is provided in a comparative aspect. Based on own experimental results and literature data a scheme of NO action in the myometrium is suggested in which nitric oxide or its derivatives cause Ca2+ -dependent polarization of the sarcolemma. In accordance with our results, this effect may be based on the increase of sarcolemma Ca2+ permeability under the influence of NO or its derivatives and the stimulation of at least the initial passive transport of the cation in the myocytes mediated by dihydropyridine-sensitive channels. Additional factors that contribute to the polarization of the membrane are the increase of protons transport from the muscle cells and stimulation of Na+, K+ -ATPase. Acting on the sarcoplasmic reticulum, nitrosactive compounds activate the inclusion of calcium in this compartment and inhibit Ca2+ -induced release of the cation. The latter effects are able to provide compensation for NO-induced Ca2+ increase in myocytes and supress the electromechanical coupling at Ca2+ release from the reticulum. NO-derivates also inhibit a key link in the smooth muscle contractile act--the formation of the Ca2+ -calmodulin complex.  相似文献   

13.
While using myometrium sarcolemma vesicles the action of sodium nitroprusside, NO2-, NO3- and H+ on delta pH-dependent Ca(2+)-transport and passive permeability for H+ vesicles sarcolemma was estimated in the wide concentration range (10(-10)-10(-3) M) of the substances tested. In order of studying calcium transport 45Ca2+ was used, while for H+ translocation registrating via sarcolemma delta pH-indicator 14C-methylamine was applied. Sodium nitroprusside was displayed as weakly effective, while nitrite-anions essentially increased delta pH-dependent Ca2+ transport in the physiologically significant nanomolar concentration region, however in the micromolar region these substances effect failed to differ from the control and restored its intensity starting at 10(-4) M and more. Under the experiment sodium nitroprusside produces considerable quantities of NO2-. Effectory action of NO3- was similar as of NO2-. In the micromolar region the compounds estimated increased considerably sarcolemma passive permeability for H+. Hydrogen peroxide decreased delta pH-dependent Ca(2+)-transport by 10(-8) M and 10(-3) M while at the concentration equal to 10(-3) M increased the sarcolemma passive permeability for proton. Sodium nitroprusside and NO2(-)-effect on the vesicles passive permeability for proton failed to be prevented by dithiotriitol, while H2O2 action was completely removed. The conclusion about the complex concentration-dependent character of the active oxygen metabolities to the sarcolemma transport processes was made, and it's noticeable that the important role in vivo, probably could be played by NO (NO2-) stable nitric metabolities.  相似文献   

14.
The effect of the fraction (1-10 kDa) obtained from the brain of cold-adapted animal (Yakut horse) on Ca2+ transport in sarcolemma vesicles of cardiomyocytes was investigated. It was shown that during insertion of Yakut horse brain fraction into incubation medium at the concentration from 10(-9) M to 3.10(-5) M at Ca2+ transport substrate concentration from 0.1 mM to 1.0 mM, the rate of Ca2+ passive penetration into vesicles slightly increased and at Ca2+ transport substrate concentration 3 mM, which is physiologic, a decrease of rate values was established for all concentrations of the fraction (1-10 kDa) of Yakut horse. While studying the kinetics of an active Ca2+ transport for all investigated concentrations of the fraction (1-10 kDa) of Yakut horse brain from 10(-9) M to 3.10(-5) M at Ca2+ concentration in incubation medium from 10(-7) to 3.10(-6) M, calcium accumulation rates by vesicles exceeded control values. So we can suppose that application of brain fraction (1-10 kDa) of genotypically cold-adapted animal, results in a decrease of intracellular Ca2+ concentration.  相似文献   

15.
Studies on the vesicular fraction of myometrium sarcolemma showed that in the absence of initial Ca2+ gradient the vesicles activity accumulate Ca2+ by utilizing the energy of the antiport-directed Na+ gradient. Monensin (50 microM) suppresses practically completely the Ca2+ transport. The amount of Ca2+ entering the vesicles against the concentration gradient diminishes with a decrease in the oppositely directed Na+ gradient. Cd2+ (5 mM) causes a complete inhibition of active Ca2+ transport, whereas Mn2+ and Mg2+ inhibit this process by 85% and 35%, respectively; amiloride (500 microM) is fairly ineffective. In the absence of initial Ca2+ and Na+ gradients valinomycin (0.05-1 microM) does not affect the changes in Ca2+ concentration in the intravesicular volume both with and without K+ gradient. Under conditions of initial equilibrium for Ca2+ and Na+ the magnitude and sign of the membrane potential for the K(+)-valinomycin system have no effect on Ca2+ transport regardless of value of absolute Na+ concentration inside and outside the vesicles. Depolarization of membrane vesicles does not interfere with the Na(+)-driven active Ca2+ transport into the sarcolemma which is dependent on the energy of the Na+ gradient. Using calibration curves, it was shown that the physiologically significant (6-fold) Na+ gradient increases Ca2+ concentration in the intravesicular volume from 100 to 160-170 microM. Ac active potential-independent Ca2+ transport through the smooth muscle sarcolemma requires about one third (0.3 kcal/mol) of the Na+ gradient; energy the remainder is dissipated. It is concluded that in smooth muscles the Na+ gradient can provide the active transsarcolemmal transport of Ca2+.  相似文献   

16.
Kinetics of Ca2+ energy-dependent transport in sarcolemma and mitochondrion fractions of myometrium was studied. On the basis of the results obtained the mechanism of calcium control of smooth muscle relaxation was analysed. In terms of this mechanism kinetic curves of myometrium relaxation were calculated. It follows from their pattern that the mitochondria play the role of the main intercellular depo of Ca2+, while the calcium pump of the sarcolemma carries out fine regulation of this process making its contribution to relaxation at its later stage.  相似文献   

17.
The effect of fatty acid and acylcarnitine on Ca2+ and Na+ transporting enzymes and carriers was studied in sealed cardiac sarcolemma vesicles of mixed polarity. Palmitoylcarnitine markedly reduced the Na+ gradient-induced Ca2+ uptake. Half-maximal reduction was obtained at 15 microM of the carnitine derivative. In a same concentration range palmitoylcarnitine caused a rapid release of accumulated Ca2+ when added to Ca2+-filled vesicles, which suggests that palmitoylcarnitine increases the permeability of the sarcolemma vesicles to Ca2+. A rapid release of Ca2+ was also observed if Ca2+ was taken up by action of the Ca2+ pump. The (Ca2+ + Mg2+)-ATPase, which most likely drives this active Ca2+ uptake, was 90% increased by 50 microM palmitoylcarnitine and evidence was presented that the acylcarnitine effect again was linked to an alteration of Ca2+ permeability of the vesicles. At the same concentration acylcarnitine was not able to unmask the latent protein kinase, so that probably the sarcolemma ATP permeability was not affected. Palmitoylcarnitine at 25 microM did not affect the ouabain-sensitive (Na+ + K+) -ATPase in native sarcolemma vesicles, however, it inhibited markedly if the enzyme was measured in SDS-treated vesicles. The effect of increased free fatty acid concentration on some of the sarcolemma transporting properties was tested by adding oleate-albumin complexes with different molar ratios to the sarcolemma vesicles. In contrast to molar ratios 1 and 5, the ratio of 7 was able to induce a rapid Ca2+ release and to inhibit (Na+ + K+)-ATPase in either native or SDS-treated vesicles markedly. 22Na release from 22Na-preloaded sarcolemma vesicles was shown to be stimulated by either palmitoylcarnitine (50 microM) or oleate-albumin complex (with a molar ratio of 7). The possible significance of the observed effects of lipid intermediates on ion permeability and (Na+ + K+)-ATPase activity in isolated sarcolemma vesicles for the derangement of cardiac cell function in ischemia is discussed.  相似文献   

18.
Oxytocin (10(-7) M) administered inside the myometrium sarcolemma vesicles closed outward by the cytoplasmic side is shown to inhibit Mg2+, ATP-dependent Ca2+ accumulation in these structures having no effect on the passive release of cation out of them. According to these results and to the data available in literature on the inhibitory action of the peptide hormone on Mg2+, Ca2+-ATPase of myometrium sarcolemma a conclusion is drawn that oxytocin inhibits the Ca pump activity in plasma membranes of the myometrium cells.  相似文献   

19.
Calmodulin-dependent Ca2+-pump ATPase of human smooth muscle sarcolemma   总被引:1,自引:0,他引:1  
L M Popescu  P Ignat 《Cell calcium》1983,4(4):219-235
An enzymatically active Ca2+-stimulated ATPase has been isolated from the sarcolemmal sheets of human smooth muscle (myometrium). Ca2+-ATPase activity was quantitated in an assay medium which simulated the characteristic free ionic concentrations of the cytosol. New computer programs for calculating the composition of solutions containing metals (Ca, Mg, Na, K) and ligands (EGTA, ATP), based on the updated stability constants, were used. In detergent-soluble form the enzyme has a high Ca2+-affinity expressed by an apparent Km (Ca2+) of 0.25 +/- 0.04 microM. The maximum specific activity (about 20 nmol of Pi/mg protein/min) was found in the micromolar domain of free-Ca2+ concentrations, the same levels required for normal maximal contractions in smooth muscle. The variation of free-Ca2+ concentration in the assay medium over 4 orders of magnitude (pCa 9 to pCa 5) resulted in a sigmoidal dependence of enzymatic activity, with a Hill coefficient of 1.4, which suggested the regulation of Ca2+-ATPase by allosteric effectors. The presence and the activator role of endogenous calmodulin in smooth muscle sarcolemma was proved by calmodulin-depletion experiments and by using suitable anticalmodulinic concentrations of trifluoperazine. The addition of exogenous calmodulin restored the enzyme activity. Apparently, the concentration of calmodulin in isolated smooth muscle sarcolemma is about 0.1% of sarcolemmal proteins, as deduced from the comparison of calmodulin-depletion and calmodulin-readdition experiments. Calmodulin increased significantly the enzyme Ca2+-affinity and Vmax (by a factor of about 10). At variance with the sarcoplasmic reticulum Ca2+-ATPase, the sarcolemmal Ca2+-ATPase is extremely sensitive to orthovanadate, half-maximal inhibition being observed at 0.8 microM vanadate. In conclusion, the Ca2+-ATPase isolated from smooth muscle sarcolemma appears very similar to the well-known Ca2+-pump ATPases of erythrocyte membrane, heart sarcolemma or axolemma. We suggest that this high-affinity Ca2+-ATPase represents the calmodulin-regulated Ca2+-extrusion pump of the smooth muscle sarcolemma.  相似文献   

20.
A plasma membrane-enriched fraction from rat myometrium shows ATP-Mg2+-dependent active calcium uptake which is independent of the presence of oxalate and is abolished by the Ca2+ ionophore A23187. Ca2+ loaded into vesicles via the ATP-dependent Ca2+ uptake was released by extravesicular Na+. This showed that the Na+/Ca2+ exchange and the Ca2+ uptake were both occurring in plasma membrane vesicles. In a medium containing KCl, vanadate readily inhibited the Ca2+ uptake (K1/2 5 microM); when sucrose replaced KCl, 400 microM-vanadate was required for half inhibition. Only a slight stimulation of the calcium pump by calmodulin was observed in untreated membrane vesicles. Extraction of endogenous calmodulin from the membranes by EGTA decreased the activity and Ca2+ affinity of the calcium pump; both activity and affinity were fully restored by adding back calmodulin or by limited proteolysis. A monoclonal antibody (JA3) directed against the human erythrocyte Ca2+ pump reacted with the 140 kDa Ca2+-pump protein of the myometrial plasma membrane. The Ca2+-ATPase activity of these membranes is not specific for ATP, and is not inhibited by mercurial agents, whereas Ca2+ uptake has the opposite properties. Ca2+-ATPase activity is also over 100 times that of calcium transport; it appears that the ATPase responsible for transport is largely masked by the presence of another Ca2+-ATPase of unknown function. Measurements of total Ca2+-ATPase activity are, therefore, probably not directly relevant to the question of intracellular Ca2+ control.  相似文献   

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