Hemoglobin genotype has minimal influence on the physiological response of juvenile atlantic cod (Gadus morhua) to environmental challenges

Hemoglobin (Hb) polymorphism in cod is associated with temperature‐related differences in biogeographical distribution, and several authors have suggested that functional characteristics of the various hemoglobin isoforms (HbIs) directly influence phenotypic traits such as growth rate. However, no study has directly examined whether Hb genotype translates into physiological differences at the whole animal level. Thus, we generated a family of juvenile Atlantic cod consisting of all three main Hb genotypes (HbI‐1/1, HbI‐2/2, and HbI‐1/2) by crossing a single pair of heterozygous parents, and we compared their metabolic and cortisol responses to an acute thermal challenge (10&deg;C to their critical thermal maximum [CTM] or 22&deg;C, respectively) and tolerance of graded hypoxia. There were no differences in routine metabolism (at 10&deg;C), maximum metabolic rate, metabolic scope, CTM (overall mean 22.9&deg; &plusmn; 0.2&deg;C), or resting and poststress plasma cortisol levels among Hb genotypes. Further, although the HbI‐1/1 fish grew more (by 15%&ndash;30% during the first 9 mo) when reared at 10&deg; &plusmn; 1&deg;C and had a slightly enhanced hypoxia tolerance at 10&deg;C (e.g., the critical O2 levels for HbI‐1/1, HbI‐2/2, and HbI‐1/2 cod were 35.56% &plusmn; 1.24%, and 40.20% &plusmn; 1.99% air saturation, respectively), these results are contradictory to expectations based on HbI functional properties. Thus, our findings (1) do not support previous assumptions that growth rate differences among cod Hb genotypes result from a more efficient use of the oxygen supply&mdash;that is, reduced standard metabolic rates and/or increased metabolic capacity&mdash;and (2) suggest that in juvenile cod, there is no selective advantage to having a particular Hb genotype with regards to the capacity to withstand ecologically relevant environmental challenges.<br /

Atlantic cod (Gadus morhua) hemoglobin genes: multiplicity and polymorphism

Background: Hemoglobin (Hb) polymorphism, assessed by protein gel electrophoresis, has been used almost exclusively to characterize the genetic structure of Atlantic cod (Gadus morhua) populations and to establish correlations with phenotypic traits such as Hb oxygen binding capacity, temperature tolerance and growth characteristics. The genetic system used to explain the results of gel electrophoresis entails the presence of one polymorphic locus with two major alleles (HbI-1; HbI-2). However, vertebrates have more than one gene encoding Hbs and recent studies have reported that more than one Hb gene is present in Atlantic cod. These observations prompted us to re-evaluate the number of Hb genes expressed in Atlantic cod, and to perform an in depth search for polymorphisms that might produce relevant phenotypes for breeding programs. Results: Analysis of Expressed Sequence Tags (ESTs) led to the identification of nine distinct Hb transcripts; four corresponding to the α Hb gene family and five to the β Hb gene family. To gain insights about the Hb genes encoding these transcripts, genomic sequence data was generated from heterozygous (HbI-1/2) parents and fifteen progeny; five of each HbI type, i.e., HbI-1/1, HbI-1/2 and HbI-2/2. β Hb genes displayed more polymorphism than α Hb genes. Two major allele types (β1A and β1B) that differ by two linked non-synonymous substitutions (Met55Val and Lys62Ala) were found in the β1 Hb gene, and the distribution of these β1A and β1B alleles among individuals was congruent with that of the HbI-1 and HbI-2 alleles determined by protein gel electrophoresis. RT-PCR and Q-PCR analysis of the nine Hb genes indicates that all genes are expressed in adult fish, but their level of expression varies greatly; higher expression of almost all Hb genes was found in individuals displaying the HbI-2/2 electrophoretic type. Conclusion: This study indicates that more Hb genes are present and expressed in adult Atlantic cod than previously documented. Our finding that nine Hb genes are expressed simultaneously in adult fish suggests that Atlantic cod, similarly to fish such as rainbow trout, carp, and goldfish, might be able to respond to environmental challenges such as chronic hypoxia or long-term changes in temperature by altering the level of expression of these genes. In this context, the role of the non-conservative substitution Lys62Ala found in the β1 Hb gene, which appears to explain the occurrence of the HbI-1 and HbI-2 alleles described by gel electrophoresis, and which was found to be present in other fish such as eel, emerald rockcod, rainbow trout and moray, requires further investigation

Time domains of the hypoxic ventilatory response in ectothermic vertebrates

Over a decade has passed since Powell et al. (Respir Physiol 112:123–134, 1998) described and defined the time domains of the hypoxic ventilatory response (HVR) in adult mammals. These time domains, however, have yet to receive much attention in other vertebrate groups. The initial, acute HVR of fish, amphibians and reptiles serves to minimize the imbalance between oxygen supply and demand. If the hypoxia is sustained, a suite of secondary adjustments occur giving rise to a more long-term balance (acclimatization) that allows the behaviors of normal life. These secondary responses can change over time as a function of the nature of the stimulus (the pattern and intensity of the hypoxic exposure). To add to the complexity of this process, hypoxia can also lead to metabolic suppression (the hypoxic metabolic response) and the magnitude of this is also time dependent. Unlike the original review of Powell et al. (Respir Physiol 112:123–134, 1998) that only considered the HVR in adult animals, we also consider relevant developmental time points where information is available. Finally, in amphibians and reptiles with incompletely divided hearts the magnitude of the ventilatory response will be modulated by hypoxia-induced changes in intra-cardiac shunting that also improve the match between O2 supply and demand, and these too change in a time-dependent fashion. While the current literature on this topic is reviewed here, it is noted that this area has received little attention. We attempt to redefine time domains in a more ‘holistic’ fashion that better accommodates research on ectotherms. If we are to distinguish between the genetic, developmental and environmental influences underlying the various ventilatory responses to hypoxia, however, we must design future experiments with time domains in mind

Effects of predator exposure on baseline and stress‐induced glucocorticoid hormone concentrations in pumpkinseed Lepomis gibbosus

We compared baseline and maximal cortisol concentrations between predator exposure and prey blood samples in pumpkinseed Lepomis gibbosus, captured using a standardised fishing event underneath osprey Pandion haliaetus nests and away from osprey nests. We did not detect differences in cortisol or glucose between sites. These findings suggest that predictable sources of predation risk may not confer stress-related costs in teleosts

Effects of Loma morhua (Microsporidia) infection on the cardiorespiratory performance of Atlantic cod Gadus morhua (L).

The microsporidian Loma morhua infects Atlantic cod (Gadus morhua) in the wild and in culture and results in the formation of xenomas within the gill filaments, heart and spleen. Given the importance of the two former organs to metabolic capacity and thermal tolerance, the cardiorespiratory performance of cod with a naturally acquired infection of Loma was measured during an acute temperature increase (2 °C h−1) from 10 °C to the fish's critical thermal maximum (CTMax). In addition, oxygen consumption and swimming performance were measured during two successive critical swimming speed (Ucrit) tests at 10 °C. While Loma infection had a negative impact on cod cardiac function at warm temperatures, and on metabolic capacity in both the CTMax and Ucrit tests (i.e. a reduction of 30–40%), it appears that the Atlantic cod can largely compensate for these Loma‐induced cardiorespiratory limitations. For example, (i) CTMax (21.0 ± 0.3 °C) and Ucrit (~1.75 BL s−1) were very comparable to those reported in previous studies using uninfected fish from the same founder population; and (ii) our data suggest that tissue oxygen extraction, and potentially the capacity for anaerobic metabolism, is enhanced in fish infected with this microsporidian.publishedVersio

Cardiorespiratory effects and efficacy of morphine sulfate in winter flounder (Pseudopleuronectes americanus)

abstract: Objective - To assess the cardiorespiratory effects of morphine sulfate and evaluate whether morphine blocks cardiac responses to a noxious stimulus in winter flounder. Animals - 42 winter flounder (Pseudopleuronectes americanus) that were acclimated at 10°C. Procedures - Each fish was fitted with a Doppler flow probe around the ventral aorta; cannulae were placed for injection of drug or saline (0.9% NaCl) solution and assessments of respiration. Selected cardiorespiratory variables were measured in morphine-injected (40 mg/kg, IP [n=18] or 17 mg/kg, IV [2]) or saline solution-injected (1.6 mL [22]) fish at various intervals. Heart rate and cardiac output (CO) were also measured in flounder that were injected with saline solution (n=19) or morphine (10) and received a noxious or innocuous stimulus (injection of 5% acetic acid or saline solution SC into a cheek) 50 minutes later. Results - Morphine administration promptly induced marked bradycardia (and a concomitant reduction in CO), followed by prolonged (&gt;48 hours) increases in CO and heart rate. Morphine injection only transiently affected respiratory rate. Application of a noxious stimulus to control flounder resulted in a significant (10%) but transient (&lt;5 minutes&#39; duration) increase in CO, which was completely blocked by prior administration of morphine. Conclusions and Clinical Relevance - Although morphine blocked the response to a noxious stimulus in fish, its cardiovascular effects might preclude its use in many research situations. Investigation of the dose dependency of these cardiovascular effects and their interspecific variation is required to determine the applicability of morphine for use in fish

Cardiac function in Atlantic cod (Gadus morhua) as a function acclimation temperature and during acute thermal challenge

Cardiac function has been hypothesised to be a primary factor influencing swimming performance in teleost fish. Using the in situ perfused heart (Farrell) set-up we examined cardiac function in cod acclimated to 0, 4 and 10°C. Only small differences in maximal cardiac output (Qmax) and the maximal myocardial power (max PO) tests were seen between 4 and 10°C acclimated cod with regards to cardiac output (CO) and maximum power generated, and these were primarily due to significant chrontropic rather than inotropic effects. In both the Qmax and max PO tests, the cardiac output was significantly lower in the 0°C cod compared to the 10°C cod. Again this was due to significant chronotropic effects. Through acute temperature reversal, i.e. 10°C cod at 4°C and the 4°C cod at 10°C we found CO to be significantly larger in 4°C cod at 10°C compared to 10°C cod at 10°C, again due primarily to a significant increase in HR during the Qmax test, although SV also tended to be greater. Again the CO and maximum power generated during the max PO test were significantly elevated, however this was due to alterations in both SV and HR. It appears that heart rate alone is responsible for regulation of CO and power generation in cod when challenged at their acclimation temperatures, while SV remains constant. During the acute temperature change it seems that both the HR and SV are altered to maintain and in fact increase CO

Improved mitochondrial function in salmon (Salmo salar) following high temperature acclimation suggests that there are cracks in the proverbial ‘ceiling’

Mitochondrial function can provide key insights into how fish will respond to climate change, due to its important role in heart performance, energy metabolism and oxidative stress. However, whether warm acclimation can maintain or improve the energetic status of the fish heart when exposed to short-term heat stress is not well understood. We acclimated Atlantic salmon, a highly aerobic eurythermal species, to 12 and 20 °C, then measured cardiac mitochondrial functionality and integrity at 20 °C and at 24, 26 and 28 °C (this species’ critical thermal maximum ± 2 °C). Acclimation to 20 °C vs. 12 °C enhanced many aspects of mitochondrial respiratory capacity and efficiency up to 24 °C, and preserved outer mitochondrial membrane integrity up to 26 °C. Further, reactive oxygen species (ROS) production was dramatically decreased at all temperatures. These data suggest that salmon acclimated to ‘normal’ maximum summer temperatures are capable of surviving all but the most extreme ocean heat waves, and that there is no ‘tradeoff’ in heart mitochondrial function when Atlantic salmon are acclimated to high temperatures (i.e., increased oxidative phosphorylation does not result in heightened ROS production). This study suggests that fish species may show quite different acclimatory responses when exposed to prolonged high temperatures, and thus, susceptibility to climate warming