Interactive effects of development and hypoxia on catecholamine synthesis and cardiac function in zebrafish (Danio rerio)

Abstract

The rate-limiting enzyme in the biosynthetic pathway of catecholamines is tyrosine hydroxylase (TH), the activity of which is dependent on molecular oxygen. Zebrafish (Danio rerio) possess two non-allelic TH coding genes, TH1 and TH2. A principal goal of the present study was to determine if the expression of these genes is sensitive to environmental hypoxia. Additionally, we sought to determine if catecholamine content of larvae was changed by environmental hypoxia, and whether the hearts of hypoxic larvae were equally responsive to exogenous catecholamine (norepinephrine) exposure. After 2 days of exposure to hypoxia [5-7 days post-fertilization (dpf); PO(2) = 30 Torr] TH2 mRNA expression was significantly lower and dopamine β hydroxylase (DβH) mRNA was significantly higher in whole larvae. Whole body catecholamine levels were unchanged until after 4 days of hypoxic exposure (5-9 dpf), at which time there was a significant increase in epinephrine and norepinephrine contents. Norepinephrine content was significantly elevated in the hearts of adult fish after 2 and 4 days of hypoxic exposure, and TH1 mRNA expression was increased in the kidney of both groups. After 2 or 4 days of exposure to hypoxia, larvae displayed significantly lower heart rates than normoxic fish. However, application of exogenous norepinephrine caused similar increases in heart rate in both groups. Overall, it is concluded that the mRNA expression of TH1 and TH2 is differentially affected by hypoxia exposure in larvae and adults. Also, catecholamine biosynthesis appears to be activated by 2 dpf and although whole body catecholamine levels increase during hypoxia (possibly promoting downregulation of cardiac β-adrenergic receptors), there is no accompanying decrease in the response of the heart to adrenergic stimulation.

Evidence for a role of heme oxygenase-1 in the control of cardiac function in zebrafish (Danio rerio) larvae exposed to hypoxia

“…The dose of 0.05 mg ml −1 of MS-222 was chosen because it was the lowest dose able to keep the larvae immobilized. Other studies have shown that high doses of MS-222 decrease f H in zebrafish larvae; however, the concentration used in this study is lower than that used in those studies (Steele et al, 2009(Steele et al, , 2011. Furthermore, in our own experiments only a dose of MS-222 above 0.12 mg ml −1 caused a decrease in f H after 10-15 min exposure of 4 dpf larvae.…”

“…Following the hypoxic treatment, larvae were exposed to hypoxic water containing either 40 µmol l −1 CORM or 10 −4 mol l −1 adrenaline for an additional 15 min. The adrenaline concentration used was based on a previous study (Steele et al, 2011) and preliminary trials which demonstrated that it was the lowest concentration to reliably produce a positive cardiac chronotropic effect. The CORM concentration was chosen based on dose−response curves performed with concentrations used previously in mammals (Clark et al, 2003).…”

“…Although bradycardia is the typical response to hypoxia in adult water-breathing fish, the situation is much less clear in developing larvae. Zebrafish (Danio rerio) larvae, in particular, exhibit various cardiac responses depending on the severity of hypoxia and/or the duration of exposure (acute versus chronic) (Barrionuevo and Burggren, 1999;Jacob et al, 2002;Yaqoob and Schwerte, 2010;Bagatto, 2005;Steele et al, 2011). Tachycardia may be the initial response to progressive hypoxia but can develop into bradycardia as the severity or duration of the hypoxia is increased.…”

“…In larval zebrafish, the onset of cardiac control via the parasympathetic and sympathetic nervous systems (or circulating catecholamines) is evident as early as 4 dpf (Steele et al, 2009(Steele et al, , 2011. The development of bradycardia in the face of chronic hypoxia in zebrafish larvae is mediated by cardiac muscarinic receptors (Bagatto, 2005) of the M 2 sub-type (Steele et al, 2009), while the tachycardia response to acute hypoxia is probably controlled by a release of inhibitory vagal tone or stimulation of adrenergic receptors by elevated circulating catecholamine levels or catecholamines released from sympathetic neurons (Steele et al, 2009(Steele et al, , 2011.…”

Neuroendocrine control of ionic balance in zebrafish

“…Catecholamines either can interact with target cells after their secretion as neurotransmitters from nearby nerve endings, or after their release from chromaffin cells for more widespread distribution via the circulatory system. Using high-performance liquid chromatography (HPLC), catecholamines were detectable in whole embryos at 1 h post fertilization (hpf), presumably derived from maternal transfer (Steele et al, 2011). At 2 days post fertilization (dpf), chromaffin cells were found to be dispersed as clusters in the inter-renal organ, which then converged to midline by 3 dpf; the chromaffin cells stay in intimate contact with steroidogenic cells throughout the subsequent development (Liu, 2007;To et al, 2007).…”