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Excitation-contraction coupling in zebrafish ventricular myocardium is regulated by trans-sarcolemmal Ca2+ influx and sarcoplasmic reticulum Ca2+ release.

Haustein M, Hannes T, Trieschmann J, Verhaegh R, Köster A, Hescheler J, Brockmeier K, Adelmann R, Khalil M - PLoS ONE (2015)

Bottom Line: We found an overall negative force-frequency relationship (FFR).Inhibition of L-type Ca(2+)-channels by verapamil (1 μM) decreased force of contraction to 22 ± 7% compared to baseline (n=4, p<0.05).In conclusion, force development in adult zebrafish ventricular myocardium requires not only trans-sarcolemmal Ca2+-flux, but also intact sarcoplasmic reticulum Ca(2+)-cycling.

View Article: PubMed Central - PubMed

Affiliation: Department of Paediatric Cardiology, Cologne Heart Centre, Medical Faculty, University of Cologne, Cologne, North Rhine-Westphalia, Germany.

ABSTRACT
Zebrafish (Danio rerio) have become a popular model in cardiovascular research mainly due to identification of a large number of mutants with structural defects. In recent years, cardiomyopathies and other diseases influencing contractility of the heart have been studied in zebrafish mutants. However, little is known about the regulation of contractility of the zebrafish heart on a tissue level. The aim of the present study was to elucidate the role of trans-sarcolemmal Ca(2+)-flux and sarcoplasmic reticulum Ca(2+)-release in zebrafish myocardium. Using isometric force measurements of fresh heart slices, we characterised the effects of changes of the extracellular Ca(2+)-concentration, trans-sarcolemmal Ca(2+)-flux via L-type Ca(2+)-channels and Na(+)-Ca(2+)-exchanger, and Ca(2+)-release from the sarcoplasmic reticulum as well as beating frequency and β-adrenergic stimulation on contractility of adult zebrafish myocardium. We found an overall negative force-frequency relationship (FFR). Inhibition of L-type Ca(2+)-channels by verapamil (1 μM) decreased force of contraction to 22 ± 7% compared to baseline (n=4, p<0.05). Ni(2+) was the only substance to prolong relaxation (5 mM, time after peak to 50% relaxation: 73 ± 3 ms vs. 101 ± 8 ms, n=5, p<0.05). Surprisingly though, inhibition of the sarcoplasmic Ca(2+)-release decreased force development to 54 ± 3% in ventricular (n=13, p<0.05) and to 52 ± 8% in atrial myocardium (n=5, p<0.05) suggesting a substantial role of SR Ca(2+)-release in force generation. In line with this finding, we observed significant post pause potentiation after pauses of 5 s (169 ± 7% force compared to baseline, n=8, p<0.05) and 10 s (198 ± 9% force compared to baseline, n=5, p<0.05) and mildly positive lusitropy after β-adrenergic stimulation. In conclusion, force development in adult zebrafish ventricular myocardium requires not only trans-sarcolemmal Ca2+-flux, but also intact sarcoplasmic reticulum Ca(2+)-cycling. In contrast to mammals, FFR is strongly negative in the zebrafish heart. These aspects need to be considered when using zebrafish to model human diseases of myocardial contractility.

No MeSH data available.


Related in: MedlinePlus

Contribution of SR to developed force.(A) Post-pause potentiation behaviour of zebrafish ventricular slices (representative experiment, top: 5 seconds rest-interval, bottom: 10 seconds rest-interval). (B) The twitch amplitudes of the first contraction were increased after a rest interval. Prolonging of rest intervals resulted in higher amplitudes. Force of contraction was normalised to steady-state force before the pause. (C) Effect of Ry Blocking of RyR receptor by 1 μM Ry leads to a reduction of force of contraction in ventricular slices and whole atrium. Force of contraction was normalised to baseline. (D) Original averaged twitches before and after Ry application of a ventricular slice (upper panel) and an atrium (lower panel). (E) Abbreviated FFR protocol (2.0Hz, 1.0 Hz, 3.0 Hz) before and after application of Ry (1 μM). Force of contraction decreased stronger at higher stimulation frequencies after application of Ry. Data are expressed as mean ±SEM, asterisks indicate statistically significant differences (p < 0.05 vs. steady state or between stimulation frequencies).
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pone.0125654.g004: Contribution of SR to developed force.(A) Post-pause potentiation behaviour of zebrafish ventricular slices (representative experiment, top: 5 seconds rest-interval, bottom: 10 seconds rest-interval). (B) The twitch amplitudes of the first contraction were increased after a rest interval. Prolonging of rest intervals resulted in higher amplitudes. Force of contraction was normalised to steady-state force before the pause. (C) Effect of Ry Blocking of RyR receptor by 1 μM Ry leads to a reduction of force of contraction in ventricular slices and whole atrium. Force of contraction was normalised to baseline. (D) Original averaged twitches before and after Ry application of a ventricular slice (upper panel) and an atrium (lower panel). (E) Abbreviated FFR protocol (2.0Hz, 1.0 Hz, 3.0 Hz) before and after application of Ry (1 μM). Force of contraction decreased stronger at higher stimulation frequencies after application of Ry. Data are expressed as mean ±SEM, asterisks indicate statistically significant differences (p < 0.05 vs. steady state or between stimulation frequencies).

Mentions: A main feature of mammalian myocardium is an increase in contractility after rest which—in mammals—mainly depends on intact SR Ca2+ loading during rest. We studied post pause potentiation after 5 s and 10 s (Fig 4A and 4B) rest intervals. Force of contraction increased to 169 ± 7% after 5 s rest compared to steady state baseline (n = 8, p < 0.05) and 198 ± 9% after 10 s rest compared to steady state baseline (n = 5, p < 0.05). Thus, we observed a considerable degree of post pause potentiation.


Excitation-contraction coupling in zebrafish ventricular myocardium is regulated by trans-sarcolemmal Ca2+ influx and sarcoplasmic reticulum Ca2+ release.

Haustein M, Hannes T, Trieschmann J, Verhaegh R, Köster A, Hescheler J, Brockmeier K, Adelmann R, Khalil M - PLoS ONE (2015)

Contribution of SR to developed force.(A) Post-pause potentiation behaviour of zebrafish ventricular slices (representative experiment, top: 5 seconds rest-interval, bottom: 10 seconds rest-interval). (B) The twitch amplitudes of the first contraction were increased after a rest interval. Prolonging of rest intervals resulted in higher amplitudes. Force of contraction was normalised to steady-state force before the pause. (C) Effect of Ry Blocking of RyR receptor by 1 μM Ry leads to a reduction of force of contraction in ventricular slices and whole atrium. Force of contraction was normalised to baseline. (D) Original averaged twitches before and after Ry application of a ventricular slice (upper panel) and an atrium (lower panel). (E) Abbreviated FFR protocol (2.0Hz, 1.0 Hz, 3.0 Hz) before and after application of Ry (1 μM). Force of contraction decreased stronger at higher stimulation frequencies after application of Ry. Data are expressed as mean ±SEM, asterisks indicate statistically significant differences (p < 0.05 vs. steady state or between stimulation frequencies).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4418605&req=5

pone.0125654.g004: Contribution of SR to developed force.(A) Post-pause potentiation behaviour of zebrafish ventricular slices (representative experiment, top: 5 seconds rest-interval, bottom: 10 seconds rest-interval). (B) The twitch amplitudes of the first contraction were increased after a rest interval. Prolonging of rest intervals resulted in higher amplitudes. Force of contraction was normalised to steady-state force before the pause. (C) Effect of Ry Blocking of RyR receptor by 1 μM Ry leads to a reduction of force of contraction in ventricular slices and whole atrium. Force of contraction was normalised to baseline. (D) Original averaged twitches before and after Ry application of a ventricular slice (upper panel) and an atrium (lower panel). (E) Abbreviated FFR protocol (2.0Hz, 1.0 Hz, 3.0 Hz) before and after application of Ry (1 μM). Force of contraction decreased stronger at higher stimulation frequencies after application of Ry. Data are expressed as mean ±SEM, asterisks indicate statistically significant differences (p < 0.05 vs. steady state or between stimulation frequencies).
Mentions: A main feature of mammalian myocardium is an increase in contractility after rest which—in mammals—mainly depends on intact SR Ca2+ loading during rest. We studied post pause potentiation after 5 s and 10 s (Fig 4A and 4B) rest intervals. Force of contraction increased to 169 ± 7% after 5 s rest compared to steady state baseline (n = 8, p < 0.05) and 198 ± 9% after 10 s rest compared to steady state baseline (n = 5, p < 0.05). Thus, we observed a considerable degree of post pause potentiation.

Bottom Line: We found an overall negative force-frequency relationship (FFR).Inhibition of L-type Ca(2+)-channels by verapamil (1 μM) decreased force of contraction to 22 ± 7% compared to baseline (n=4, p<0.05).In conclusion, force development in adult zebrafish ventricular myocardium requires not only trans-sarcolemmal Ca2+-flux, but also intact sarcoplasmic reticulum Ca(2+)-cycling.

View Article: PubMed Central - PubMed

Affiliation: Department of Paediatric Cardiology, Cologne Heart Centre, Medical Faculty, University of Cologne, Cologne, North Rhine-Westphalia, Germany.

ABSTRACT
Zebrafish (Danio rerio) have become a popular model in cardiovascular research mainly due to identification of a large number of mutants with structural defects. In recent years, cardiomyopathies and other diseases influencing contractility of the heart have been studied in zebrafish mutants. However, little is known about the regulation of contractility of the zebrafish heart on a tissue level. The aim of the present study was to elucidate the role of trans-sarcolemmal Ca(2+)-flux and sarcoplasmic reticulum Ca(2+)-release in zebrafish myocardium. Using isometric force measurements of fresh heart slices, we characterised the effects of changes of the extracellular Ca(2+)-concentration, trans-sarcolemmal Ca(2+)-flux via L-type Ca(2+)-channels and Na(+)-Ca(2+)-exchanger, and Ca(2+)-release from the sarcoplasmic reticulum as well as beating frequency and β-adrenergic stimulation on contractility of adult zebrafish myocardium. We found an overall negative force-frequency relationship (FFR). Inhibition of L-type Ca(2+)-channels by verapamil (1 μM) decreased force of contraction to 22 ± 7% compared to baseline (n=4, p<0.05). Ni(2+) was the only substance to prolong relaxation (5 mM, time after peak to 50% relaxation: 73 ± 3 ms vs. 101 ± 8 ms, n=5, p<0.05). Surprisingly though, inhibition of the sarcoplasmic Ca(2+)-release decreased force development to 54 ± 3% in ventricular (n=13, p<0.05) and to 52 ± 8% in atrial myocardium (n=5, p<0.05) suggesting a substantial role of SR Ca(2+)-release in force generation. In line with this finding, we observed significant post pause potentiation after pauses of 5 s (169 ± 7% force compared to baseline, n=8, p<0.05) and 10 s (198 ± 9% force compared to baseline, n=5, p<0.05) and mildly positive lusitropy after β-adrenergic stimulation. In conclusion, force development in adult zebrafish ventricular myocardium requires not only trans-sarcolemmal Ca2+-flux, but also intact sarcoplasmic reticulum Ca(2+)-cycling. In contrast to mammals, FFR is strongly negative in the zebrafish heart. These aspects need to be considered when using zebrafish to model human diseases of myocardial contractility.

No MeSH data available.


Related in: MedlinePlus