Na+/Ca2+ exchange current and contractions measured under Cl(-)-free conditions in developing rabbit hearts

Previous studies suggesting a greater functional role of cardiac Na+/Ca2+ exchange at birth were performed using tightly buffered free cytosolic Ca2+ concentration ([Ca2+]i). Because Na+/Ca2+ exchange current (INaCa) is influenced by physiological fluctuations in [Ca2+]i, we used conditions of minimally buffered [Ca2+]i to simultaneously record INaCa and cell contractions in single ventricular myocytes isolated from 1 to 27-day-old and adult rabbits. With conventional Cl(-)-containing solutions. Ni(2+)-sensitive outward and inward charge movements were unbalanced, suggesting the presence of a contaminating current (presumably the Ca(2+)-activated Cl- current). Removing Cl- abolished this discrepancy in all age groups and allowed for the accurate quantitation of INaCa. Under Cl(-)-free conditions, outward and inward charge movements were high at birth (4 days: 0.42 +/- 0.03 and -0.38 +/- 0.04 pC/pF, respectively) and decreased postnatally (adult: 0.08 +/- 0.01 and -0.07 +/- 0.01 pC/pF, respectively). Newborn but not adult myocytes contracted during depolarizations in the presence of nifedipine, ryanodine, and thapsigargin. The magnitudes of outward charge movement (Ca2+ influx) and cell shortening exhibited similar voltage dependence, consistent with INaCa-mediated contractions. These results indicate that INaCa can directly support contraction in newborn rabbit ventricular myocytes.

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β2-Adrenergic receptor agonists stimulate L-type calcium current independent of PKA in newborn rabbit ventricular myocytes

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00101.2007 ◽

2007 ◽

Vol 293 (5) ◽

pp. H2826-H2835 ◽

Cited By ~ 10

Author(s):

Leon P. Collis ◽

Shekhar Srivastava ◽

William A. Coetzee ◽

Michael Artman

Keyword(s):

Ventricular Myocytes ◽

Ventricular Myocardium ◽

Adult Rabbit ◽

Patch Clamp Technique ◽

Postnatal Maturation ◽

Newborn Rabbit ◽

Age Related ◽

Rabbit Ventricular Myocardium ◽

Rabbit Ventricular Myocytes ◽

Stimulation Of

Selective stimulation of β2-adrenergic receptors (ARs) in newborn rabbit ventricular myocardium invokes a positive inotropic effect that is lost during postnatal maturation. The underlying mechanisms for this age-related stimulatory response remain unresolved. We examined the effects of β2-AR stimulation on L-type Ca2+ current ( ICa,L) during postnatal development. ICa,L was measured (37°C; either Ca2+ or Ba2+ as the charge carrier) using the whole-cell patch-clamp technique in newborn (1 to 5 days old) and adult rabbit ventricular myocytes. Ca2+ transients were measured concomitantly by dialyzing the cell with indo-1. Activation of β2-ARs (with either 100 nM zinterol or 1 μM isoproterenol in the presence of the β1-AR antagonist, CGP20712A) stimulated ICa,L twofold in newborns but not in adults. The β2-AR-mediated increase in Ca2+ transient amplitude in newborns was due exclusively to the augmentation of ICa,L. Zinterol increased the rate of inactivation of ICa,L and increased the Ca2+ flux integral. The β2-AR inverse agonist, ICI-118551 (500 nM), but not the β1-AR antagonist, CGP20712A (500 nM), blocked the response to zinterol. Unexpectedly, the PKA blockers, H-89 (10 μM), PKI 6-22 amide (10 μM), and Rp-cAMP (100 μM), all failed to prevent the response to zinterol but completely blocked responses to selective β1-AR stimulation of ICa,L in newborns. Our results demonstrate that in addition to the conventional β1-AR/cAMP/PKA pathway, newborn rabbit myocardium exhibits a novel β2-AR-mediated, PKA-insensitive pathway that stimulates ICa,L. This striking developmental difference plays a major role in the age-related differences in inotropic responses to β2-AR agonists.

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Two classes of gating current from L-type Ca channels in guinea pig ventricular myocytes.

The Journal of General Physiology ◽

10.1085/jgp.99.6.863 ◽

1992 ◽

Vol 99 (6) ◽

pp. 863-895 ◽

Cited By ~ 42

Author(s):

R Shirokov ◽

R Levis ◽

N Shirokova ◽

E Ríos

Keyword(s):

Guinea Pig ◽

Time Constant ◽

Voltage Dependence ◽

Ventricular Myocytes ◽

Slow Component ◽

The Other ◽

Charge Movement ◽

Ca Channels ◽

Voltage Distribution ◽

Polarized Cells

Intramembrane charge movement was recorded in guinea pig ventricular myocytes at 19-22 degrees C using the whole-cell patch clamp technique. From a holding potential of -110 mV, the dependence of intramembrane charge moved on test voltage (Q(V)) followed the sum of two Boltzmann components. One component had a transition voltage (V) of -48 mV and a total charge (Qmax) of congruent to 3 nC/microF. The other had a V of -18 mV and a Qmax of 11 nC/microF. Ba2+ currents through Ca channels began to activate at -45 mV and peaked at congruent to -15 mV. Na+ current peaked at -35 to -30 mV. Availability of charge (in pulses from -70 to +10 mV) depended on the voltage of conditioning depolarizations as two Boltzmann terms plus a constant. One term had a V of -88 mV and a Qmax of 2.5 nC/microF; the other had a V of -29 mV and a Qmax of 6.3 nC/microF. From the Q(V) dependence, the voltage dependence of the ionic currents, and the voltage dependence of the availability of charge, the low voltage term of Q(V) and availability was identified as Na gating charge, at a total of 3.5 nC/microF. The remainder, 11 nC/microF, was attributed to Ca channels. After pulses to -40 mV and above, the OFF charge movement had a slow exponentially decaying component. Its time constant had a bell-shaped dependence on OFF voltage peaking at 11 ms near -100 mV. Conditioning depolarizations above -40 mV increased the slow component exponentially with the conditioning duration (tau approximately equal to 480 ms). Its magnitude was reduced as the separation between conditioning and test pulses increased (tau approximately equal to 160 ms). The voltage distribution of the slow component of charge was measured after long (5 s) depolarizations. Its V was -100 mV, a shift of -80 mV from the value in normally polarized cells. This voltage was the same at which the time constant of the slow component peaked. Qmax and the steepness of the voltage distribution were unchanged by depolarization. This indicates that the same molecules that produce the charge movement in normally polarized cells also produce the slow component in depolarized cells. 100 microns D600 increased by 77% the slow charge movement after a 500-ms conditioning pulse. These results demonstrate two classes of charge movement associated with L-type Ca channels, with kinetics and voltage dependence similar to charge 1 and charge 2 of skeletal muscle.(ABSTRACT TRUNCATED AT 400 WORDS)

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Role of Na+/Ca2+exchange in endothelin-1-induced increases in Ca2+transient and contractility in rabbit ventricular myocytes: pharmacological analysis with KB-R7943

British Journal of Pharmacology ◽

10.1038/sj.bjp.0702454 ◽

1999 ◽

Vol 126 (8) ◽

pp. 1785-1795 ◽

Cited By ~ 46

Author(s):

Huang-Tian Yang ◽

Kiyoharu Sakurai ◽

Hiromi Sugawara ◽

Tomoo Watanabe ◽

Ikuo Norota ◽

...

Keyword(s):

Ventricular Myocytes ◽

Endothelin 1 ◽

Pharmacological Analysis ◽

Rabbit Ventricular Myocytes

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SR Ca2+ refilling upon depletion and SR Ca2+ uptake rates during development in rabbit ventricular myocytes

AJP Cell Physiology ◽

10.1152/ajpcell.00241.2007 ◽

2007 ◽

Vol 293 (6) ◽

pp. C1906-C1915 ◽

Cited By ~ 20

Author(s):

Jingbo Huang ◽

Leif Hove-Madsen ◽

Glen F. Tibbits

Keyword(s):

Ventricular Myocytes ◽

Age Groups ◽

Rabbit Heart ◽

Developmental Changes ◽

Reverse Mode ◽

Time Integral ◽

Younger Age ◽

Highly Sensitive ◽

Uptake Rates ◽

Rabbit Ventricular Myocytes

While it has been reported that a sparse sarcoplasmic reticulum (SR) and a low SR Ca2+ pump density exist at birth, we and others have recently shown that significant amounts of Ca2+ are stored in the neonatal rabbit heart SR. Here we try to determine developmental changes in SR Ca2+ loading mechanisms and Ca2+ pump efficacy in rabbit ventricular myocytes. SR Ca2+ loading (loadSR) and k0.5 (Ca2+ concentration at half-maximal SR Ca2+ uptake) were higher and lower, respectively, in younger age groups. Inhibition of the L-type calcium current ( ICa) with 15 μM nifedipine dramatically reduced loadSR in older but not in younger age groups. In contrast, subsequent inhibition of the Na+/Ca2+ exchanger (NCX) with 10 μM KB-R7943 strongly reduced loadSR in the younger but not the older age groups. Accordingly, the time integral of the inward NCX current (tail INCX) elicited on repolarization was highly sensitive to nifedipine in the older groups and sensitive to KB-R7943 in the younger groups. Interestingly, slow SR loading took place in the presence of both nifedipine and KB-R7943 in all age groups, although it was less prominent in the older groups. We conclude that the SR loading capacity at the earliest postnatal stages is at least as large as that of adult myocytes. However, reverse-mode NCX plays a prominent role in SR Ca2+ loading at early postnatal stages while ICa is the main source of SR Ca2+ loading at late postnatal and adult stages.

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CaM kinase augments cardiac L-type Ca2+ current: a cellular mechanism for long Q-T arrhythmias

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1999.276.6.h2168 ◽

1999 ◽

Vol 276 (6) ◽

pp. H2168-H2178 ◽

Cited By ~ 36

Author(s):

Yuejin Wu ◽

Leigh B. MacMillan ◽

R. Blair McNeill ◽

Roger J. Colbran ◽

Mark E. Anderson

Keyword(s):

Ventricular Myocytes ◽

Ryanodine Receptors ◽

Cam Kinase ◽

Early Afterdepolarizations ◽

Ultrastructural Studies ◽

Intracellular Ca ◽

Dependent Protein ◽

T Tubules ◽

Rabbit Ventricular Myocytes

Early afterdepolarizations (EAD) caused by L-type Ca2+ current ( I Ca,L) are thought to initiate long Q-T arrhythmias, but the role of intracellular Ca2+ in these arrhythmias is controversial. Rabbit ventricular myocytes were stimulated with a prolonged EAD-containing action potential-clamp waveform to investigate the role of Ca2+/calmodulin-dependent protein kinase II (CaM kinase) in I Ca,L during repolarization. I Ca,L was initially augmented, and augmentation was dependent on Ca2+ from the sarcoplasmic reticulum because the augmentation was prevented by ryanodine or thapsigargin. I Ca,Laugmentation was also dependent on CaM kinase, because it was prevented by dialysis with the inhibitor peptide AC3-I and reconstituted by exogenous constitutively active CaM kinase when Ba2+ was substituted for bath Ca2+. Ultrastructural studies confirmed that endogenous CaM kinase, L-type Ca2+ channels, and ryanodine receptors colocalized near T tubules. EAD induction was significantly reduced in current-clamped cells dialyzed with AC3-I (4/15) compared with cells dialyzed with an inactive control peptide (11/15, P = 0.013). These findings support the hypothesis that EADs are facilitated by CaM kinase.

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Faculty Opinions recommendation of Action potential duration restitution and alternans in rabbit ventricular myocytes: the key role of intracellular calcium cycling.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1097052.553023 ◽

2007 ◽

Author(s):

Godfrey Smith

Keyword(s):

Action Potential ◽

Intracellular Calcium ◽

Action Potential Duration ◽

Ventricular Myocytes ◽

Calcium Cycling ◽

Rabbit Ventricular Myocytes ◽

Action Potential Duration Restitution

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Paradoxical Effect of Dofetilide on Action Potential Duration and Calcium Transient Amplitude in Newborn Rabbit Ventricular Myocytes

Journal of Cardiovascular Pharmacology ◽

10.1097/01.fjc.0000151896.57637.66 ◽

2005 ◽

Vol 45 (2) ◽

pp. 165-174 ◽

Cited By ~ 5

Author(s):

Shekhar Srivastava ◽

Leon Collis ◽

Anita Go ◽

Salvatore Mancarella ◽

William A Coetzee ◽

...

Keyword(s):

Action Potential ◽

Action Potential Duration ◽

Ventricular Myocytes ◽

Calcium Transient ◽

Paradoxical Effect ◽

Newborn Rabbit ◽

Rabbit Ventricular Myocytes

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Ontogeny of Ca2+-induced Ca2+ release in rabbit ventricular myocytes

AJP Cell Physiology ◽

10.1152/ajpcell.00417.2007 ◽

2008 ◽

Vol 294 (2) ◽

pp. C516-C525 ◽

Cited By ~ 25

Author(s):

Jingbo Huang ◽

Leif Hove-Madsen ◽

Glen F. Tibbits

Keyword(s):

Developmental Stages ◽

Ventricular Myocytes ◽

Age Groups ◽

Inhibitory Effect ◽

Dominant Mode ◽

Older Age ◽

Lower Efficiency ◽

Younger Age ◽

Rabbit Ventricular Myocytes ◽

Subsequent Addition

It is commonly accepted that L-type Ca2+ channel-mediated Ca2+-induced Ca2+ release (CICR) is the dominant mode of excitation-contraction (E-C) coupling in the adult mammalian heart and that there is no appreciable CICR in neonates. However, we have observed that cell contraction in the neonatal heart was significantly decreased after sarcoplasmic reticulum (SR) Ca2+ depletion with caffeine. Therefore, the present study investigated the developmental changes of CICR in rabbit ventricular myocytes at 3, 10, 20, and 56 days of age. We found that the inhibitory effect of the L-type Ca2+ current ( ICa) inhibitor nifedipine (Nif; 15 μM) caused an increasingly larger reduction of Ca2+ transients on depolarization in older age groups [from ∼15% in 3-day-old (3d) myocytes to ∼90% in 56-day-old (56d) myocytes]. The remaining Ca2+ transient in the presence of Nif in younger age groups was eliminated by the inhibition of Na+/Ca2+ exchanger (NCX) with the subsequent addition of 10 μM KB-R7943 (KB-R). Furthermore, Ca2+ transients were significantly reduced in magnitude after the depletion of SR Ca2+ with caffeine in all age groups, although the effect was significantly greater in the older age groups (from ∼40% in 3d myocytes up to ∼70% in 56d myocytes). This SR Ca2+-sensitive Ca2+ transient in the earliest developmental stage was insensitive to Nif but was sensitive to the subsequent addition of KB-R, indicating the presence of NCX-mediated CICR that decreased significantly with age (from ∼37% in 3d myocytes to ∼0.5% in 56d myocytes). In contrast, the ICa-mediated CICR increased significantly with age (from ∼10% in 3d myocytes to ∼70% in 56d myocytes). The CICR gain as estimated by the integral of the CICR Ca2+ transient divided by the integral of its Ca2+ transient trigger was smaller when mediated by NCX (∼1.0 for 3d myocytes) than when mediated by ICa (∼3.0 for 56d myocytes). We conclude that the lower-efficiency NCX-mediated CICR is a predominant mode of CICR in the earliest developmental stages that gradually decreases as the more efficient L-type Ca2+ channel-mediated CICR increases in prominence with ontogeny.

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