Na+-dependent alkaline earth metal uptake in cardiac sarcolemmal vesicles

doi:10.1016/0005-2736(85)90035-5

Biochimica et Biophysica Acta (BBA) - Biomembranes

Volume 817, Issue 2, 25 July 1985, Pages 327-332

https://doi.org/10.1016/0005-2736(85)90035-5 Get rights and content

Abstract

The ability of alkaline earth metals (M²⁺) to substitute for Ca²⁺ in Na⁺-Ca²⁺ exchange was examined in sarcolemmal vesicles isolated from the canine heart. ⁸⁵Sr²⁺ and ¹³³Ba²⁺, in addition to ⁴⁵Ca²⁺, were used to determine the characteristics of Na⁺-M²⁺ exchange. The Na_i⁺-dependent M²⁺ uptake was measured as a function of time, with $t$ ranging from 0.5 to 360 s, $[Na^{+}]_{i} = 140 mM$ and $[M^{2+}]_{o} = 40 μM$ . This function was linear for Ca²⁺ and Sr²⁺ uptake for approx. 6 s and for Ba²⁺ for about 60 s. Plateau levels were achieved within 120 s for Ca²⁺ and Sr²⁺ but Ba²⁺ took considerably longer. The $K_{m}$ values for Na⁺-M²⁺ exchange, derived from Eadie-Hofstee plots, were 30, 58, and 73 μM for Ca²⁺, Sr²⁺ and Ba²⁺, respectively. The Na_i⁺-dependent uptake of all three ions was stimulated in the presence of 0.36 μM valinomycin. Na⁺-Ca²⁺ exchange was also measured in the presence of either 20 μM Sr²⁺ or 100 μM Ba²⁺. Both of these ions behaved (at these concentrations) as competitive inhibitors of Na⁺-Ca²⁺ exchange with the $K_{I}$ being 32 μM for Sr²⁺ and 92 μM for Ba²⁺. Passive efflux was determined by first allowing Na⁺-M²⁺ exchange to continue to plateau values and then diluting the loaded vesicles in the presence of EGTA. The rate constants for the passive efflux were 8.4, 6.3 and 4.4 min⁻¹ for Ca²⁺, Sr²⁺ and Ba²⁺, respectively.

References (35)

K.D. Philipson et al.
J. Biol. Chem.
(1980)
H. Miyamoto et al.
J. Biol. Chem.
(1980)
B.J.R. Pitts
J. Biol. Chem.
(1979)
J.P. Reeves et al.
J. Biol. Chem.
(1984)
K.D. Philipson et al.
J. Biol. Chem.
(1984)
K.D. Philipson
J. Biol. Chem.
(1984)
K.D. Philipson et al.
J. Biol. Chem.
(1981)
K.D. Philipson et al.
J. Biol. Chem.
(1982)
O.H. Lowry et al.
J. Biol. Chem.
(1951)
T.L. Trosper et al.
Biochim. Biophys. Acta
(1983)

C. Bernard et al.

Bioelectrochem. Bioenergetics

(1974)

J.M. Vanderkooi et al.

Arch. Biochem. Biophys.

(1971)

S. Ringer

J. Physiol. (Lond.)

(1883)

G.A. Langer

Annu. Rev. Physiol.

(1982)

H. Reuter

Annu. Rev. Physiol.

(1979)

P. Caroni et al.

Nature

(1980)

L.J. Mullins

Am. J. Physiol.

(1979)

Cited by (15)

Pre-steady-state kinetics of Ba-Ca exchange reveals a second electrogenic step involved in Ca2+ translocation by the Na-Ca exchanger
2009, Biophysical Journal
Citation Excerpt :
It has been shown in previous studies that Sr2+ and Ba2+ induce Na+ transport by the exchanger, although at different rates. Sr-Na exchange is as fast as Na-Ca exchange, whereas Ba-Na exchange is much slower (6–9). In the experiments described here, Ba2+ or Sr2+ were applied on the extracellular side and a Ca2+ concentration jump was performed on the cytoplasmic side.
Kinetic properties of the Na-Ca exchanger (guinea pig NCX1) expressed in Xenopus oocytes were investigated with excised membrane patches in the inside-out configuration and photolytic Ca²⁺ concentration jumps with either 5 mM extracellular Sr²⁺ or Ba²⁺. After a Ca²⁺ concentration jump on the cytoplasmic side, the exchanger performed Sr-Ca or Ba-Ca exchange. In the Sr-Ca mode, currents are transient and decay in a monoexponential manner similar to that of currents in the Ca-Ca exchange mode described before. Currents recorded in the Ba-Ca mode are also transient, but the decay is biphasic. In the Sr-Ca mode the amount of charge translocated increases at negative potentials in agreement with experiments performed in the Ca-Ca mode. In the Ba-Ca mode the total amount of charge translocated after a Ca²⁺ concentration jump is ∼4 to 5 times that in Ca-Ca or Sr-Ca mode. In the Ba-Ca mode the voltage dependence of charge translocation depends on the Ca²⁺ concentration on the cytosolic side before the Ca²⁺ concentration jump. At low initial Ca²⁺ levels (∼0.5 μM), charge translocation is voltage independent. At a higher initial concentration (1 μM Ca²⁺), the amount of charge translocated increases at positive potentials. Biphasic relaxation of the current was also observed in the Ca-Ca mode if the external Ca²⁺ concentration was reduced to ≤0.5 mM. The results reported here and in previous publications can be described by using a 6-state model with two voltage-dependent conformational transitions.
Activation of human TRPC6 channels by receptor stimulation
2004, Journal of Biological Chemistry
Citation Excerpt :
They propose that Na+ entry via TRPC3 in the absence of extracellular Ca2+ produces a significant alteration in the Na+ gradient, which upon readdition of Ca2+ to the bath, fuels Ca2+ entry via the exchanger. It is well established that the Na+-Ca2+ exchanger will also transport Ba2+ and Sr2+ (29–31). Furthermore, PKC appears to be part of the Na+-Ca2+ exchanger macromolecular complex (32) and exchange activity may be activated (33) or inhibited (34) by a PKC-dependent phosphorylation event.
The human TRPC6 channel was expressed in human embryonic kidney (HEK) cells, and activity was monitored using the giga-seal technique. Whole cell membrane currents with distinctive inward and outward rectification were activated by carbachol (CCh) in TRPC6-expressing cells, but not in lacZ-transfected controls. The effect of CCh was steeply dose-dependent with a K_0.5 of ∼10 μm and a Hill coefficient of 3–4. A steep concentration-response relationship was also observed when TRPC6 activity was measured using a fluorescence-based imaging plate reader (FLIPR) assay for membrane depolarization. Ionomycin, thapsigargin, and dialysis of the cell with inositol 1,4,5-trisphosphate via the patch pipette had no effect on TRPC6 currents, but exogenous application of 1-oleoyl acetyl-sn-glycerol (OAG, 30–300 μm) produced a slow increase in channel activity. The PKC activator, phorbol 12-myristate 13-acetate (PMA, 0.5 μm) had no significant acute effect on TRPC6, or on the subsequent response to OAG. In contrast, the response to CCh was blocked >90% by PMA pretreatment. To further explore the role of DAG in receptor stimulation, TRPC6 currents were monitored following the sequential addition of CCh and OAG. Surprisingly, concentrations of CCh that produced little or no response in the absence of OAG, produced increases in TRPC6 currents in the presence of OAG that were larger than the sum of either agent alone. Likewise, the response to OAG was superadditive following prior stimulation of the cells with near threshold concentrations of CCh. Overall, these results suggest that generation of DAG alone may not fully account for activation of TRPC6, and that other receptor-mediated events act synergistically with DAG to stimulate channel activity. This synergy may explain, at least in part, the steep dose-response relationship observed for CCh-induced TRPC6 currents expressed in HEK cells.
Effects of active oxygen generated by DTT Fe2+ on cardiac Na+ Ca2+ exchange and membrane permeability to Ca2+
1989, Journal of Molecular and Cellular Cardiology
Sarcolemmal vesicles isolated from bovine heart were preincubated at 37°C with an oxygen radical generating system consisting of 1 mm dithiothreitol (DTT) and 50 μm FeSO₄. Exposure of the vesicles for 1 to 40 mins stimulated $Na^{+} Ca^{2+}$ exchange about 2.5-fold. The $DTT Fe^{2+}$ treatment decreased the apparent K_m for Ca²⁺ of Na_i⁺-dependent Ca²⁺ uptake by 80% (from 63 to 13 μm). The effect on V_max was much smaller however. The resulting stimulation of exchange activity was diminished by the presence of desferrioxamine (95%) or catalase (60%). In contrast, superoxide dismutase and sodium formate did not prevent the effects of $DTT Fe^{2+}$ on the exchanger. Neither Zn²⁺ nor Ga³⁺ could replace Fe²⁺ in the stimulation of $Na^{+} Ca^{2+}$ exchange. Passive Ca²⁺ efflux was determined by first allowing $Na^{+} Ca^{2+}$ exchange to continue to plateau values and then diluting the loaded vesicles in the presence of EGTA. Ca²⁺ leakage from the vesicles was slightly but significantly (P < 0.05) increased by the action of $DTT Fe^{2+}$ , the rate constants for the passive Ca²⁺ efflux being 0.22 and 0.26/min in control and treated groups, respectively. The calcium loading observed in myocytes in ischemia/reperfusion injury suggests that the stimulation of $Na^{+} Ca^{2+}$ exchange by active oxygen may moderate the myocardial response to oxygen mediated injuries including ischemia/reperfusion injury. However, the clinical relevance of these phenomena is far from clear as the stimulation depends in part on the K_m for Ca²⁺ prior to treatment.
Are Ba2+ and Sr2+ ions transported by the Na+Ca2+ exchanger in frog atrial cells?
1989, Journal of Molecular and Cellular Cardiology
Ba²⁺ and Sr²⁺ ions are widely used to replace Ca²⁺ ions for the study of Ca²⁺ channel currents in electrophysiological experiments. Using the double sucrose gap technique, we investigated the effects of Sr²⁺ and Ba²⁺ ions on the Na⁺Ca²⁺ exchange activity in frog atrial fibres where it is the major relaxation mechanism. With either Sr²⁺ or Ba²⁺ ions instead of Ca²⁺ in the extracellular bath, Na-free contractures reversibly developed but with different kinetics. Voltage clamp experiments showed that the tonic tension recorded in the presence of Sr²⁺ or Ba²⁺ was markedly increased following the addition of monensin, a Na⁺ ionophore known to increase the intracellular Na⁺ activity. In Na-free solutions (Li-substituted), it was possible to induce contractures by substituting Sr²⁺ or Ba²⁺ ions for extracellular Ca²⁺. These contractures could be relaxed by reintroducing Na⁺ or Ca²⁺ ions in the extracellular medium. Taken together, these results suggest that Sr²⁺ and Ba²⁺ ions can interact with the Na⁺Ca²⁺ exchange mechanism and potentially participate not only in Na⁺-cation but also in Ca²⁺-cation exchanges on either side of the sarcolemmal membrane.
Can strontium replace calcium as an activator of internal calcium release in cardiac muscles?: Study on a dual action of A23187
1987, Journal of Molecular and Cellular Cardiology
The positive inotropic effect of A23187 has previously been indicated to be attributed to both of the enhancement of the slow Ca inward current and the facilitation of Ca release from internal stores. The electromechanical effects of replacing external Ca by Sr on this dual action of A23187 were examined on guinea-pig papillary muscles driven at 0.2 Hz. Replacing most of external Ca by Sr (Sr 2.5 mm plus Ca 0.2 mm) (Sr-medium) increased the developed tension. This procedure simultaneously prolonged the time to peak developed tension (tPT) and the action potential duration at all repolarization phases (APD 10, APD 30 and APD 80). In such preparations, A23187 (10⁻⁶ m) caused a marked positive inotropic effect (938±180% of control) accompanied by a prolongation of APD at an early repolarization phase (APD 10), but did not affect tPT. Thereafter, returning superfusate to normal Ca medium (Ca 1.2 mm) in the presence of A23187 also caused a positive inotropic effect, but markedly shortened tPT. The electrical and mechanical responses to A23187 in Sr-medium were completely blocked by nifedipine (2×10⁻⁶ m), Ca-channel-blocker, while the positive inotropic effect in normal Ca medium was not blocked by nifedipine. Ryanodine, an inhibitor of internal Ca release, did not affect the positive inotropic effect of A23187 in Sr-medium. These results suggest that in Sr-medium, an internal Ca pool of the myocardium, most likely sarcoplasmic reticulum, does not play a role in the activation of contraction.
Ca2+ transport capacity of sarcolemmal Na+-Ca2+ exchange.Extrapolation of vesicle data to in vivo conditions
1986, Journal of Molecular and Cellular Cardiology
Na⁺−Ca²⁺ exchange activity is high in cardiac sarcolemmal vesicles suggesting an important physiologic role. Vesicular Na⁺−Ca²⁺ exchange, however, is usually measured under conditions which are far from physiologic. Using sarcolemmal vesicles, we have estimated the possible significance of both Ca²⁺ influx and efflux mediated by Na⁺−Ca²⁺ exchange under approximate in vivo ionic conditions. In this situation, Na⁺−Ca²⁺ exchange activity is far from maximal with intracellular Mg²⁺ causing significant inhibition. The capacity of the Na⁺−Ca²⁺ exchange system to extrude intracellular Ca²⁺ (at [Ca²⁺]=6.0 μm) is about 1.2 μmol Ca²⁺/kg wet weight/s and approximately equals the capacity of the sarcolemmal ATP-dependent Ca²⁺ pump. The capacity of the sarcoplasmic reticular Ca²⁺ pump to remove cytoplasmic Ca²⁺ is much larger. Significant Ca²⁺ influx through the exchanger is unlikely to occur in normal mammalian myocardium and would require reduced extracellular Na⁺ or elevated intracellular Na⁺.

View all citing articles on Scopus

View full text

Article preview

Biochimica et Biophysica Acta (BBA) - Biomembranes

Abstract

References (35)

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

Biochim. Biophys. Acta

Bioelectrochem. Bioenergetics

Arch. Biochem. Biophys.

J. Physiol. (Lond.)

Annu. Rev. Physiol.

Annu. Rev. Physiol.

Nature

Am. J. Physiol.

Cited by (15)

Pre-steady-state kinetics of Ba-Ca exchange reveals a second electrogenic step involved in Ca<sup>2+</sup> translocation by the Na-Ca exchanger

Activation of human TRPC6 channels by receptor stimulation

Effects of active oxygen generated by DTT Fe<sup>2+</sup> on cardiac Na<sup>+</sup> Ca<sup>2+</sup> exchange and membrane permeability to Ca<sup>2+</sup>

Are Ba<sup>2+</sup> and Sr<sup>2+</sup> ions transported by the Na<sup>+</sup>Ca<sup>2+</sup> exchanger in frog atrial cells?

Can strontium replace calcium as an activator of internal calcium release in cardiac muscles?: Study on a dual action of A23187

Ca<sup>2+</sup> transport capacity of sarcolemmal Na<sup>+</sup>-Ca<sup>2+</sup> exchange.Extrapolation of vesicle data to in vivo conditions

Regular paperNa+-dependent alkaline earth metal uptake in cardiac sarcolemmal vesicles

Abstract

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

Biochim. Biophys. Acta

Bioelectrochem. Bioenergetics

Arch. Biochem. Biophys.

J. Physiol. (Lond.)

Annu. Rev. Physiol.

Annu. Rev. Physiol.

Nature

Am. J. Physiol.

Regular paper
Na⁺-dependent alkaline earth metal uptake in cardiac sarcolemmal vesicles