Adaptation to environmental fluctuations, through sensing and appropriate physiological responses, is crucial to homeostasis. Neuroepithelial cells (NECs) are putative chemoreceptors resembling mammalian Type I (glomus) cells. They have been shown to respond in vitro to changes in O2, CO2, NH3 and pH. Cytosolic carbonic anhydrase (Ca17a) is thought to be involved in CO2 sensing owing to its presence in NECs. A mutant line of zebrafish (Danio rerio) lacking functional Ca17a was generated using CRISPR/Cas9 technology and used to assess the role of Ca17a in initiating the cardiorespiratory responses to elevated CO2 (hypercapnia). Unfortunately, the homozygous knockout mutants (ca17a-/-) did not survive longer than ~12-14 days post fertilization (dpf), restricting experiments to early developmental stages (4-8 dpf). Changes in ventilation (fV) and cardiac (fH) frequency in response to hypercapnia (1% CO2) in wild type (ca17a+/+), heterozygous (ca17a+/-) and ca17a-/- fish were used to investigate Ca17a-dependent CO2 sensing and downstream signalling. Wild type fish exhibited hyperventilation during hypercapnia as indicated by an increase in fV. In the ca17a-/- fish, the hyperventilatory response was attenuated markedly, but only at 8 dpf. Hypercapnic tachycardia was observed for all genotypes and did not appear to be influenced by the absence of Ca17a. Interestingly, ca17a-/- fish exhibited a significantly reduced resting fH¬. This effect of knockout became more pronounced as the fish aged. Anesthesia did not contribute to the decreased fH in the ca17a-/- fish, nor did changes in cardiac adrenergic or cholinergic tone, which were probed using propranolol (-adrenergic receptor blocker) or atropine (muscarinic receptor blocker). The decrease in resting fH was prevented (“rescued”) when ca17a-/- embryos were injected with ca17a mRNA. Collectively, the results of this thesis support a role for Ca17a in promoting hyperventilation during hypercapnia in larval zebrafish and suggest a previously unrecognized role for Ca17a in determining resting heart rate.
| Identifer | oai:union.ndltd.org:uottawa.ca/oai:ruor.uottawa.ca:10393/42098 |
| Date | 07 May 2021 |
| Creators | Kunert, Emma |
| Contributors | Gilmour, Kathleen, Perry, Steve |
| Publisher | Université d'Ottawa / University of Ottawa |
| Source Sets | Université d’Ottawa |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | application/pdf |
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