Regulation of heart rate following genetic deletion of the β1 adrenergic receptor in larval zebrafish

Aim: Although zebrafish are gaining popularity as biomedical models of cardiovascular disease, our understanding of their cardiac control mechanisms is fragmentary. Our goal was to clarify the controversial role of the beta 1-adrenergic receptor (AR) in the regulation of heart rate in zebrafish. Methods: CRISPR-Cas9 was used to delete the adrb1 gene in zebrafish allowing us to generate a stable adrb1(-/-) line. Larval heart rates were measured during pharmacological protocols and with exposure to hypercapnia. Expression of the five zebrafish adrb genes were measured in larval zebrafish hearts using qPCR. Results: Compared with genetically matched wild-types (adrb1(+/+)), adrb1(-/-) larvae exhibited similar to 20 beats min(-1) lower heart rate, measured from 2 to 21 days post-fertilization (dpf). Nevertheless, adrb1(-/-) larvae exhibited preserved positive chronotropic responses to pharmacological treatment with AR agonists (adrenaline, noradrenaline, isoproterenol), which were blocked by propranolol (general beta-AR antagonist). Regardless of genotype, larvae exhibited similar increases in heart rate in response to hypercapnia (1% CO2) at 5 dpf, but tachycardia was blunted in adrb1(-/-) larvae at 6 dpf. adrb1 gene expression was abolished in the hearts of adrb1(-/-) larvae, confirming successful knockout. While gene expression of adrb2a and adrb3a was unchanged, adrb2b and adrb3b mRNA levels increased in adrb1(-/-) larval hearts. Conclusion: Despite adrb1 contributing to the setting of resting heart rate in larvae, it is not strictly essential for zebrafish, as we generated a viable and breeding adrb1(-/-) line. The chronotropic effects of adrenergic stimulation persist in adrb1(-/-) zebrafish, likely due to the upregulation of other beta-AR subtypes.

Automated summary

This study aimed to clarify the role of the beta 1-adrenergic receptor (AR) in heart rate regulation in zebrafish. By using CRISPR-Cas9 technology, a stable adrb1(-/-) line was generated, showing a lower resting heart rate compared to wild-types. However, adrb1(-/-) zebrafish still exhibited positive chronotropic responses to adrenergic stimulation, likely due to the upregulation of other beta-AR subtypes.