Polyunsaturated dietary fats change the properties of calcium sparks in adult rat atrial myocytes

Abstract Background Atrial fibrillation has been associated with an increase in ryanodine receptor (RyR2) phosphorylation and local calcium release (calcium sparks). Carvedilol, a nonselective beta-adrenergic receptor blocker also inhibits the cardiac ryanodine receptor (RyR2), but it has been suggested that the enantiomer R-carvedilol only inhibits RyR2 activity and hence has the potential to inhibit calcium sparks without affecting RyR2 phosphorylation. Purpose This study aimed to determine the ability of the enantiomers R- and S-carvedilol to reverse RyR2 phosphorylation at s2808 and calcium sparks induced by the β2-adrenergic agonist fenoterol, in order to determine the relationship between RyR2 phosphorylation at s2808 and calcium spark frequency, and to assess the efficacy of R- and S-carvedilol. Methods Human right atrial myocytes were isolated and subjected to immunofluorescent labelling of total and s2808 phosphorylated RyR2, or loaded with fluo-4 and subjected to confocal calcium imaging. Beta-adrenergic receptors were first activated with 3μM fenoterol and then inhibited by different concentrations of carvedilol R- or S-enantiomers. Results Incubation of myocytes with fenoterol increased the s2808/RyR2 ratio from 0.32±0.03 to 0.66±0.05 (n=18, p<0.001). Incubation with 0.1, 0.3, 1 or 3μM R-carvedilol in the presence of fenoterol changed the s2808/RyR2 ratio to 0.64±0.05, 0.44±0.04, 0.34±0.07 and 0.28±0.05 (p<0.01) respectively. For comparison 3μM S-carvedilol reduced the s2808/RyR2 ratio to 0.23±0.06 in myocytes from 5 patients (p<0.01). Confocal calcium imaging revealed that fenoterol increased the spark density from 0.28±0.04 to 1.24±0.25 events/s/1000μm2 (n=9, p<0.01) and addition of 0.1, 0.3, or 1μM R-carvedilol changed the frequency to 1.32±0.52, 0.38±0.05, and 0.15±0.05 events/s/1000μm2 (p<0.01) respectively. Analysis of atrial myocytes from patients without atrial fibrillation revealed that the s2808/RyR2 ratio was similar in 25 patients treated with beta-blockers (0.39±0.04) and 57 that did not receive beta-blockers (0.44±0.03, p=0.33) while the s2808/RyR2 ratio was significantly smaller in 16 patients with atrial fibrillation receiving beta-blockers (0.43±0.08) than in 5 patients that did not (0.80±0.19, p<0.05). Conclusions R-carvedilol reverses the effects of beta-adrenergic stimulation on s2808 phosphorylation and calcium sparks in human atrial myocytes, and treatment with beta-blockers reduces excessive RyR2 phosphorylation at s2808 in patients with atrial fibrillation to levels observed in those without the arrhythmia, pointing to beta-adrenergic receptors as a target for controlling RyR2 phophorylation and activity in atrial fibrillation. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): Spanish Ministry of Science and Innovation & Spanish Ministry of Health and Consume

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Two functionally distinct 4-aminopyridine-sensitive outward K+ currents in rat atrial myocytes.

The Journal of General Physiology ◽

10.1085/jgp.100.6.1041 ◽

1992 ◽

Vol 100 (6) ◽

pp. 1041-1067 ◽

Cited By ~ 58

Author(s):

W A Boyle ◽

J M Nerbonne

Keyword(s):

Experimental Testing ◽

Potential Range ◽

Kinetic Properties ◽

Atrial Myocytes ◽

Adult Rat ◽

Outward Currents ◽

Recovery From Inactivation ◽

Functional Inactivation ◽

The Individual ◽

K Currents

In the experiments here, the detailed kinetic properties of the Ca(2+)-independent, depolarization-activated outward currents (Iout) in enzymatically dispersed adult rat atrial myocytes were studied. Although there is only slight attenuation of peak Iout during brief (100 ms) voltage steps, substantial decay is evident during long (10 s) depolarizations. The analyses here reveal that current inactivation is best described by the sum of two exponential components, which we have termed IKf and IKs to denote the fast and slow components, respectively, of Iout decay. At all test potentials, IKf inactivates approximately 20-fold more rapidly than IKs. Neither the decay time constants nor the fraction of Iout remaining at the end of 10-s depolarizations varies over the potential range of 0 to +50 mV, indicating that the rates of inactivation and recovery from inactivation are voltage independent. IKf recovers from inactivation completely, independent of the recovery of IKs, and IKf recovers approximately 20 times faster than IKs. The pharmacological properties of IKf and IKs are similar: both components are sensitive to 4-aminopyridine (1-5 mM) and both are relatively resistant to externally applied tetraethylammonium (50 mM). Taken together, these findings suggest that IKf and IKs correspond to two functionally distinct K+ currents with similar voltage-dependent properties and pharmacologic sensitivities, but with markedly different rates of inactivation and recovery from inactivation. From the experimental data, several gating models were developed in which voltage-independent inactivation is coupled either to channel opening or to the activation of the individual channel subunits. Experimental testing of predictions of these models suggests that voltage-independent inactivation is coupled to activation, and that inactivation of only a single subunit is required to result in functional inactivation of the channels. This model closely approximates the properties of IKf and IKs, as well as the composite outward currents, measured in adult rat atrial myocytes.

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Angiotensin II-induced changes of calcium sparks and ionic currents in human atrial myocytes: Potential role for early remodeling in atrial fibrillation

Cell Calcium ◽

10.1016/j.ceca.2005.10.008 ◽

2006 ◽

Vol 39 (2) ◽

pp. 175-186 ◽

Cited By ~ 60

Author(s):

Natig Gassanov ◽

Mathias C. Brandt ◽

Guido Michels ◽

Michael Lindner ◽

Fikret Er ◽

...

Keyword(s):

Atrial Fibrillation ◽

Angiotensin Ii ◽

Potential Role ◽

Ionic Currents ◽

Atrial Myocytes ◽

Calcium Sparks ◽

Human Atrial Myocytes ◽

Induced Changes

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Regulation of ANP secretion by endothelin-1 in cultured atrial myocytes: desensitization and receptor subtype

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1994.267.6.h2193 ◽

1994 ◽

Vol 267 (6) ◽

pp. H2193-H2203 ◽

Cited By ~ 4

Author(s):

M. F. Leite ◽

E. Page ◽

S. K. Ambler

Keyword(s):

Rate Constant ◽

Primary Cultures ◽

Receptor Activation ◽

Secretory Response ◽

Receptor Subtype ◽

Atrial Myocytes ◽

Experimental Conditions ◽

Endothelin 1 ◽

Adult Rat ◽

Eta Receptor

We examined endothelin-1 (ET-1) binding and ET-1-regulated atrial natriuretic peptide (ANP) secretion in primary cultures of adult rat atrial myocytes. ET-1 binding was analyzed as a reversible bimolecular reaction, with bimolecular association rate constant = 1.9 x 10(9) M-1.h-1, dissociation rate constant = 0.028 h-1, and a dissociation constant, calculated from these values = 0.015 nM. ET-1 increased ANP secretion with a one-half effective concentration (EC50) of 0.62 nM, which correlated with EC50 receptor occupancy under equivalent experimental conditions (0.75 nM). The secretory response rapidly desensitized (half-time = 10 min at 10 nM ET-1). The time courses for ET-1 binding, ET-1-stimulated secretion, and desensitization were all comparable. Recovery from desensitization was slow and paralleled the recovery of 125I-labeled ET-1 binding. The ETA receptor subtype-selective antagonist, BQ-123, inhibited 125I-ET-1 binding and ET-1-activated ANP secretion with high affinity, whereas the ETB-selective agonists, endothelin-3 and sarafotoxin S6c, inhibited binding with low affinity and did not effectively stimulate ANP secretion. We conclude that 1) ET-1 can stimulate ANP secretion by direct action on the atrial myocytes; 2) primary cultures of adult rat atrial myocytes express only the ETA receptor subtype; 3) the ANP secretory response to ET-1 desensitizes rapidly but recovers slowly; and 4) occupation of the ETA receptors by ET-1 initiates the unidirectional sequence of receptor activation, signal transduction, ANP secretion, and finally, desensitization.

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Effects of ANP receptor antagonists on ANP secretion from adult rat cultured atrial myocytes

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1995.268.3.e428 ◽

1995 ◽

Vol 268 (3) ◽

pp. E428-E432 ◽

Cited By ~ 12

Author(s):

S. Nachshon ◽

O. Zamir ◽

Y. Matsuda ◽

N. Zamir

Keyword(s):

Atrial Natriuretic Peptide ◽

Natriuretic Peptide ◽

Guanylyl Cyclase ◽

Second Messenger ◽

Target Cells ◽

Atrial Myocytes ◽

Receptor Antagonists ◽

Adult Rat ◽

Cgmp Formation ◽

Autoregulatory Mechanism

Atrial natriuretic peptide (ANP) is a hormone-secreted predominantly by atrial myocytes. ANP exerts many of its actions via activation of the particulate guanylyl cyclase receptor ANPR-A and the formation of guanosine 3',5'-cyclic monophosphate (cGMP), which serves as a second messenger in the target cells. Using membrane-permeable cGMP analogues (8-bromo-cGMP and dibutyryl- cGMP), we first tested the hypothesis that ANP secretion by adult rat cultured atrial myocytes can be modulated through the second messenger cGMP. Second, we examined the effects of two competitive ANPR-A receptor antagonists, namely HS-142-1 and anantin, on cGMP formation and ANP secretion from cultured atrial myocytes. Cultured atrial myocytes secreted large quantities of immunoreactive (ir) ANP under basal conditions. We found that cGMP analogues inhibited basal irANP secretion from cultured atrial myocytes, whereas HS-142-1 and anantin had stimulating effects. HS-142-1 and anantin reduced cGMP formation in cultured atrial myocytes at basal conditions. These results suggest an autoregulatory mechanism of ANP secretion by atrial myocytes in an autocrine/paracrine fashion.

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Cloning and expression of a new inositol 1,4,5-trisphosphate receptor type 1 splice variant in adult rat atrial myocytes

Cellular & Molecular Biology Letters ◽

10.2478/s11658-011-0043-7 ◽

2012 ◽

Vol 17 (1) ◽

Cited By ~ 2

Author(s):

Krishna Subedi ◽

Thoudam Singh ◽

Joon-Chul Kim ◽

Sun-Hee Woo

Keyword(s):

Amino Acid ◽

Rat Brain ◽

Cdna Clone ◽

Splice Variant ◽

Atrial Myocytes ◽

Adult Rat ◽

Inositol 1,4,5 Trisphosphate ◽

Trisphosphate Receptor ◽

Atrial Cell

AbstractInositol 1,4,5-trisphosphate receptor type 1 (IP3R1) is already known to be highly expressed in the brain, and is found in many other tissues, including the atrium of the heart. Although the complete primary structure of IP3R1 in the rat brain has been reported, the complete sequence of an IP3R1 clone from atrial myocytes has not been reported. We isolated an IP3R1 complementary DNA (cDNA) clone from isolated adult rat atrial myocytes, and found a new splice variant of IP3R1 that was different from a previously reported IP3R1 cDNA clone obtained from a rat brain (NCBI GenBank accession number: NM_001007235). Our clone had 99% similarity with the rat brain IP3R1 sequence; the exceptions were 39 amino acid deletions at the position of 1693–1731, and the deletion of phenylalanine at position 1372 that lay in the regulatory region. Compared with the rat brain IP3R1, in our clone proline was replaced with serine at residue 2439, and alanine was substituted for valine at residue 2445. These changes lie adjacent to or within the fifth transmembrane domain (2440–2462). Although such changes in the amino acid sequences were different from the rat brain IP3R1 clone, they were conserved in human or mouse IP3R1. We produced a plasmid construct expressing the atrial IP3R1 together with green fluorescent protein (GFP), and successfully overexpressed the atrial IP3R1 in the adult atrial cell line HL-1. Further investigation is needed on the physiological significance of the new splice variant in atrial cell function.

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