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Noninvasive technique for measurement of heartbeat regularity in zebrafish (Danio rerio) embryos.

Chan PK, Lin CC, Cheng SH - BMC Biotechnol. (2009)

Bottom Line: A significant decrease in heart rate was found by our method in treated embryos (p < 0.01).Moreover, there was a significant increase of the rhythmicity index (p < 0.01), which was supported by an increase in beat-to-beat interval variability (p < 0.01) of treated embryos as shown by Poincare plot.This method is capable of measuring the heart rate and heartbeat regularity simultaneously via the analysis of caudal blood flow in zebrafish embryos.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology and Chemistry, City University of Hong Kong, 83 Tat Chee Avenue, HKSAR, PR China.

ABSTRACT

Background: Zebrafish (Danio rerio), due to its optical accessibility and similarity to human, has emerged as model organism for cardiac research. Although various methods have been developed to assess cardiac functions in zebrafish embryos, there lacks a method to assess heartbeat regularity in blood vessels. Heartbeat regularity is an important parameter for cardiac function and is associated with cardiotoxicity in human being. Using stereomicroscope and digital video camera, we have developed a simple, noninvasive method to measure the heart rate and heartbeat regularity in peripheral blood vessels. Anesthetized embryos were mounted laterally in agarose on a slide and the caudal blood circulation of zebrafish embryo was video-recorded under stereomicroscope and the data was analyzed by custom-made software. The heart rate was determined by digital motion analysis and power spectral analysis through extraction of frequency characteristics of the cardiac rhythm. The heartbeat regularity, defined as the rhythmicity index, was determined by short-time Fourier Transform analysis.

Results: The heart rate measured by this noninvasive method in zebrafish embryos at 52 hour post-fertilization was similar to that determined by direct visual counting of ventricle beating (p > 0.05). In addition, the method was validated by a known cardiotoxic drug, terfenadine, which affects heartbeat regularity in humans and induces bradycardia and atrioventricular blockage in zebrafish. A significant decrease in heart rate was found by our method in treated embryos (p < 0.01). Moreover, there was a significant increase of the rhythmicity index (p < 0.01), which was supported by an increase in beat-to-beat interval variability (p < 0.01) of treated embryos as shown by Poincare plot.

Conclusion: The data support and validate this rapid, simple, noninvasive method, which includes video image analysis and frequency analysis. This method is capable of measuring the heart rate and heartbeat regularity simultaneously via the analysis of caudal blood flow in zebrafish embryos. With the advantages of rapid sample preparation procedures, automatic image analysis and data analysis, this method can potentially be applied to cardiotoxicity screening assay.

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Related in: MedlinePlus

Effect of terfenadine on heart rate and heart rate variability. Bradycardia was induced by terfenadine treatment (A). Rhythmicity index was increased by terfenadine treatment (B), suggesting induction of irregularity of heart rate by terfenadine.
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Figure 6: Effect of terfenadine on heart rate and heart rate variability. Bradycardia was induced by terfenadine treatment (A). Rhythmicity index was increased by terfenadine treatment (B), suggesting induction of irregularity of heart rate by terfenadine.

Mentions: Apart from the poincare plot, the variation of the heartbeat was also demonstrated by the changes of fbasic time by time by STFT analysis by our program. Figure 5 illustrated the changes of frequency by analyzing the signal of dynamic pixels time by time from control and terfenadine-treated embryos. In control embryo (Fig. 5A), the fbasic (red colour) in each second analyzed was kept consistent along the examination period. However, in terfenadine-treated embryo with bradycardia (Fig. 5B), the fbasic was changed along time. In addition, some terfenadine-treated embryos had shown similar heart rate with control embryos (Fig. 5C), but the fbasic was also shifted time by time. The rhythmicity index, as defined by the coefficient of variation in STFT power spectra, in control embryos was 0.03421 ± 0.0018 (Fig. 6). The rhythmicity index of terfenadine-treated embryos (including both embryos with bradycardia and with normal heart rate) was 0.07011 ± 0.0047 which was statistically significant different to the rhythmicity index in control embryos (p < 0.05).


Noninvasive technique for measurement of heartbeat regularity in zebrafish (Danio rerio) embryos.

Chan PK, Lin CC, Cheng SH - BMC Biotechnol. (2009)

Effect of terfenadine on heart rate and heart rate variability. Bradycardia was induced by terfenadine treatment (A). Rhythmicity index was increased by terfenadine treatment (B), suggesting induction of irregularity of heart rate by terfenadine.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2664803&req=5

Figure 6: Effect of terfenadine on heart rate and heart rate variability. Bradycardia was induced by terfenadine treatment (A). Rhythmicity index was increased by terfenadine treatment (B), suggesting induction of irregularity of heart rate by terfenadine.
Mentions: Apart from the poincare plot, the variation of the heartbeat was also demonstrated by the changes of fbasic time by time by STFT analysis by our program. Figure 5 illustrated the changes of frequency by analyzing the signal of dynamic pixels time by time from control and terfenadine-treated embryos. In control embryo (Fig. 5A), the fbasic (red colour) in each second analyzed was kept consistent along the examination period. However, in terfenadine-treated embryo with bradycardia (Fig. 5B), the fbasic was changed along time. In addition, some terfenadine-treated embryos had shown similar heart rate with control embryos (Fig. 5C), but the fbasic was also shifted time by time. The rhythmicity index, as defined by the coefficient of variation in STFT power spectra, in control embryos was 0.03421 ± 0.0018 (Fig. 6). The rhythmicity index of terfenadine-treated embryos (including both embryos with bradycardia and with normal heart rate) was 0.07011 ± 0.0047 which was statistically significant different to the rhythmicity index in control embryos (p < 0.05).

Bottom Line: A significant decrease in heart rate was found by our method in treated embryos (p < 0.01).Moreover, there was a significant increase of the rhythmicity index (p < 0.01), which was supported by an increase in beat-to-beat interval variability (p < 0.01) of treated embryos as shown by Poincare plot.This method is capable of measuring the heart rate and heartbeat regularity simultaneously via the analysis of caudal blood flow in zebrafish embryos.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology and Chemistry, City University of Hong Kong, 83 Tat Chee Avenue, HKSAR, PR China.

ABSTRACT

Background: Zebrafish (Danio rerio), due to its optical accessibility and similarity to human, has emerged as model organism for cardiac research. Although various methods have been developed to assess cardiac functions in zebrafish embryos, there lacks a method to assess heartbeat regularity in blood vessels. Heartbeat regularity is an important parameter for cardiac function and is associated with cardiotoxicity in human being. Using stereomicroscope and digital video camera, we have developed a simple, noninvasive method to measure the heart rate and heartbeat regularity in peripheral blood vessels. Anesthetized embryos were mounted laterally in agarose on a slide and the caudal blood circulation of zebrafish embryo was video-recorded under stereomicroscope and the data was analyzed by custom-made software. The heart rate was determined by digital motion analysis and power spectral analysis through extraction of frequency characteristics of the cardiac rhythm. The heartbeat regularity, defined as the rhythmicity index, was determined by short-time Fourier Transform analysis.

Results: The heart rate measured by this noninvasive method in zebrafish embryos at 52 hour post-fertilization was similar to that determined by direct visual counting of ventricle beating (p > 0.05). In addition, the method was validated by a known cardiotoxic drug, terfenadine, which affects heartbeat regularity in humans and induces bradycardia and atrioventricular blockage in zebrafish. A significant decrease in heart rate was found by our method in treated embryos (p < 0.01). Moreover, there was a significant increase of the rhythmicity index (p < 0.01), which was supported by an increase in beat-to-beat interval variability (p < 0.01) of treated embryos as shown by Poincare plot.

Conclusion: The data support and validate this rapid, simple, noninvasive method, which includes video image analysis and frequency analysis. This method is capable of measuring the heart rate and heartbeat regularity simultaneously via the analysis of caudal blood flow in zebrafish embryos. With the advantages of rapid sample preparation procedures, automatic image analysis and data analysis, this method can potentially be applied to cardiotoxicity screening assay.

Show MeSH
Related in: MedlinePlus