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Inhibition of ion channels and heart beat in Drosophila by selective COX-2 inhibitor SC-791.

Frolov RV, Singh S - PLoS ONE (2012)

Bottom Line: To better understand these unexpected effects, we have recently investigated the mechanism of inhibition of human K(v)2.1 channels by a highly selective COX-2 inhibitor SC-791.Using intracellular recordings in combination with a pharmacological approach and utilizing available Drosophila mutants, we found that SC-791 inhibited voltage-activated K(+) and L-type Ca(2+) channels in larval body-wall muscles and reduced heart rate in a concentration-dependent manner.The magnitude of heart rate reduction, when SC-791 and ryanodine were applied together, was smaller than expected for independent mechanisms, raising the possibility that SC-791 might be interfering with excitation-contraction coupling in Drosophila heart.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Toxicology, State University of New York at Buffalo, Buffalo, New York, United States of America. rvfrolov@gmail.com

ABSTRACT
Recent findings suggest that modulation of ion channels might be implicated in some of the clinical effects of coxibs, selective inhibitors of cyclooxygenase-2 (COX-2). Celecoxib and its inactive analog 2,5-dimethyl-celecoxib, but not rofecoxib, can suppress or augment ionic currents and alter functioning of neurons and myocytes. To better understand these unexpected effects, we have recently investigated the mechanism of inhibition of human K(v)2.1 channels by a highly selective COX-2 inhibitor SC-791. In this study we have further explored the SC-791 action on ion channels and heartbeat in Drosophila, which lacks cyclooxygenases and thus can serve as a convenient model to study COX-2-independent mechanisms of coxibs. Using intracellular recordings in combination with a pharmacological approach and utilizing available Drosophila mutants, we found that SC-791 inhibited voltage-activated K(+) and L-type Ca(2+) channels in larval body-wall muscles and reduced heart rate in a concentration-dependent manner. Unlike celecoxib and several other K(+) channel blockers, SC-791 did not induce arrhythmia. Instead, application of SC-791 resulted in a dramatic slowing of contractions and, at higher concentrations, in progressively weaker contractions with gradual cessation of heartbeat. Isradipine, a selective blocker of L-type Ca(2+) channels, showed a similar pattern of heart arrest, though no prolongation of contractions was observed. Ryanodine was the only channel modulating compound of those tested additionally that was capable of slowing contractions. Like SC-791, ryanodine reduced heart rate without arrhythmia. However, it could not stop heartbeat completely even at 500 µM, the highest concentration used. The magnitude of heart rate reduction, when SC-791 and ryanodine were applied together, was smaller than expected for independent mechanisms, raising the possibility that SC-791 might be interfering with excitation-contraction coupling in Drosophila heart.

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Effects of SC-791 on heartbeat.Traces represent the relative displacement of an edge of the larval heart in the experiments with SC-791 (A) or celecoxib (C); heartbeat was video recorded immediately before drug application, at 18–20 min after wash-in of 30 µM SC-791 or at 8–10 min after application of 10 μM celecoxib, and at 8–10 min after washing the drug out; (B) depicts heartbeat irregularity in the presence of 30 μM SC-791; the coefficient of variation (CV) was calculated as described previously [8]; the number of experiments was 8. (D) Average heart rate is shown for 6–10 larvae before drug application, after rapid wash-in of 3 μM, 10 μM, 30 μM, or 100 μM SC-791 (horizontal bar), and after the wash-out; treatment with 0.5% Me2SO, the maximum concentration of Me2SO used with SC-791, did not change the heart rate significantly. (E) A dose-response curve for the effect of SC-791 on heart rate is shown.
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pone-0038759-g003: Effects of SC-791 on heartbeat.Traces represent the relative displacement of an edge of the larval heart in the experiments with SC-791 (A) or celecoxib (C); heartbeat was video recorded immediately before drug application, at 18–20 min after wash-in of 30 µM SC-791 or at 8–10 min after application of 10 μM celecoxib, and at 8–10 min after washing the drug out; (B) depicts heartbeat irregularity in the presence of 30 μM SC-791; the coefficient of variation (CV) was calculated as described previously [8]; the number of experiments was 8. (D) Average heart rate is shown for 6–10 larvae before drug application, after rapid wash-in of 3 μM, 10 μM, 30 μM, or 100 μM SC-791 (horizontal bar), and after the wash-out; treatment with 0.5% Me2SO, the maximum concentration of Me2SO used with SC-791, did not change the heart rate significantly. (E) A dose-response curve for the effect of SC-791 on heart rate is shown.

Mentions: The larval heart has a pacemaker region and a non-pacemaking myocardium [25]. Electrical activity in the heart is driven by myogenic pacemaker located in the caudal-most segment of the heart [26]. Larval heart tube myocytes do not express sodium channels; instead, depolarization is provided by L-type Ca2+ channels as in the body-wall muscles [27], though L-type Ca2+ channels are apparently not involved in the pacemaker activity [27]. Among potassium channels, involvement of Shab, KCNQ and eag in Drosophila heartbeat has been documented previously [8], [27], [28], [29]. SC-791 inhibited heartbeat in dose-dependent manner with an EC50 (the effective concentration to alter a functional parameter by 50%) of 30 μM (see example in Fig. 3A); drug effects developed slowly and usually reached a relatively steady-state level around 20 min after a rapid wash-in (Fig. 3D). They included a reduced heart rate and heart wall displacement amplitude and prolonged contraction duration. During contraction, the maximal displacement of heart wall was observed at 160±10 ms after initiation of contraction in control and at 254±17 ms in the presence of 30 μM SC-791. The character of SC-791 effect on heartbeat was distinct from that of celecoxib. In the presence of celecoxib, irregularity of heartbeat with a significant variation in both the peak-to-peak intervals and the amplitude of heart wall displacement was observed, especially at concentrations above 3 μM [8] (Fig. 3C). In contrast, analysis of variation in peak-to-peak intervals and in amplitude of heart wall displacement indicated that SC-791 did not increase heartbeat irregularity significantly (Fig. 3B). In addition, the effects of SC-791 on heartbeat were largely irreversible.


Inhibition of ion channels and heart beat in Drosophila by selective COX-2 inhibitor SC-791.

Frolov RV, Singh S - PLoS ONE (2012)

Effects of SC-791 on heartbeat.Traces represent the relative displacement of an edge of the larval heart in the experiments with SC-791 (A) or celecoxib (C); heartbeat was video recorded immediately before drug application, at 18–20 min after wash-in of 30 µM SC-791 or at 8–10 min after application of 10 μM celecoxib, and at 8–10 min after washing the drug out; (B) depicts heartbeat irregularity in the presence of 30 μM SC-791; the coefficient of variation (CV) was calculated as described previously [8]; the number of experiments was 8. (D) Average heart rate is shown for 6–10 larvae before drug application, after rapid wash-in of 3 μM, 10 μM, 30 μM, or 100 μM SC-791 (horizontal bar), and after the wash-out; treatment with 0.5% Me2SO, the maximum concentration of Me2SO used with SC-791, did not change the heart rate significantly. (E) A dose-response curve for the effect of SC-791 on heart rate is shown.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3368905&req=5

pone-0038759-g003: Effects of SC-791 on heartbeat.Traces represent the relative displacement of an edge of the larval heart in the experiments with SC-791 (A) or celecoxib (C); heartbeat was video recorded immediately before drug application, at 18–20 min after wash-in of 30 µM SC-791 or at 8–10 min after application of 10 μM celecoxib, and at 8–10 min after washing the drug out; (B) depicts heartbeat irregularity in the presence of 30 μM SC-791; the coefficient of variation (CV) was calculated as described previously [8]; the number of experiments was 8. (D) Average heart rate is shown for 6–10 larvae before drug application, after rapid wash-in of 3 μM, 10 μM, 30 μM, or 100 μM SC-791 (horizontal bar), and after the wash-out; treatment with 0.5% Me2SO, the maximum concentration of Me2SO used with SC-791, did not change the heart rate significantly. (E) A dose-response curve for the effect of SC-791 on heart rate is shown.
Mentions: The larval heart has a pacemaker region and a non-pacemaking myocardium [25]. Electrical activity in the heart is driven by myogenic pacemaker located in the caudal-most segment of the heart [26]. Larval heart tube myocytes do not express sodium channels; instead, depolarization is provided by L-type Ca2+ channels as in the body-wall muscles [27], though L-type Ca2+ channels are apparently not involved in the pacemaker activity [27]. Among potassium channels, involvement of Shab, KCNQ and eag in Drosophila heartbeat has been documented previously [8], [27], [28], [29]. SC-791 inhibited heartbeat in dose-dependent manner with an EC50 (the effective concentration to alter a functional parameter by 50%) of 30 μM (see example in Fig. 3A); drug effects developed slowly and usually reached a relatively steady-state level around 20 min after a rapid wash-in (Fig. 3D). They included a reduced heart rate and heart wall displacement amplitude and prolonged contraction duration. During contraction, the maximal displacement of heart wall was observed at 160±10 ms after initiation of contraction in control and at 254±17 ms in the presence of 30 μM SC-791. The character of SC-791 effect on heartbeat was distinct from that of celecoxib. In the presence of celecoxib, irregularity of heartbeat with a significant variation in both the peak-to-peak intervals and the amplitude of heart wall displacement was observed, especially at concentrations above 3 μM [8] (Fig. 3C). In contrast, analysis of variation in peak-to-peak intervals and in amplitude of heart wall displacement indicated that SC-791 did not increase heartbeat irregularity significantly (Fig. 3B). In addition, the effects of SC-791 on heartbeat were largely irreversible.

Bottom Line: To better understand these unexpected effects, we have recently investigated the mechanism of inhibition of human K(v)2.1 channels by a highly selective COX-2 inhibitor SC-791.Using intracellular recordings in combination with a pharmacological approach and utilizing available Drosophila mutants, we found that SC-791 inhibited voltage-activated K(+) and L-type Ca(2+) channels in larval body-wall muscles and reduced heart rate in a concentration-dependent manner.The magnitude of heart rate reduction, when SC-791 and ryanodine were applied together, was smaller than expected for independent mechanisms, raising the possibility that SC-791 might be interfering with excitation-contraction coupling in Drosophila heart.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Toxicology, State University of New York at Buffalo, Buffalo, New York, United States of America. rvfrolov@gmail.com

ABSTRACT
Recent findings suggest that modulation of ion channels might be implicated in some of the clinical effects of coxibs, selective inhibitors of cyclooxygenase-2 (COX-2). Celecoxib and its inactive analog 2,5-dimethyl-celecoxib, but not rofecoxib, can suppress or augment ionic currents and alter functioning of neurons and myocytes. To better understand these unexpected effects, we have recently investigated the mechanism of inhibition of human K(v)2.1 channels by a highly selective COX-2 inhibitor SC-791. In this study we have further explored the SC-791 action on ion channels and heartbeat in Drosophila, which lacks cyclooxygenases and thus can serve as a convenient model to study COX-2-independent mechanisms of coxibs. Using intracellular recordings in combination with a pharmacological approach and utilizing available Drosophila mutants, we found that SC-791 inhibited voltage-activated K(+) and L-type Ca(2+) channels in larval body-wall muscles and reduced heart rate in a concentration-dependent manner. Unlike celecoxib and several other K(+) channel blockers, SC-791 did not induce arrhythmia. Instead, application of SC-791 resulted in a dramatic slowing of contractions and, at higher concentrations, in progressively weaker contractions with gradual cessation of heartbeat. Isradipine, a selective blocker of L-type Ca(2+) channels, showed a similar pattern of heart arrest, though no prolongation of contractions was observed. Ryanodine was the only channel modulating compound of those tested additionally that was capable of slowing contractions. Like SC-791, ryanodine reduced heart rate without arrhythmia. However, it could not stop heartbeat completely even at 500 µM, the highest concentration used. The magnitude of heart rate reduction, when SC-791 and ryanodine were applied together, was smaller than expected for independent mechanisms, raising the possibility that SC-791 might be interfering with excitation-contraction coupling in Drosophila heart.

Show MeSH
Related in: MedlinePlus