| Abstract |
Temperature has long been recognized as a major environmental factor contributing to species abundance and geographical distribution. During the last decades, ectotherms’ adaptation to global change is becoming of crucial importance to better understand the diversity of adaptive mechanisms in response to temperature increase. As fish constitute a large group of ectotherms, the susceptibility of their embryonic and larval stages to temperature fluctuations can permanently or temporarily alter growth and survival through phenotypic plasticity. The term phenotypic plasticity, characterizes the ability of a single genotype to produce a variety of phenotypes, as a response to the different environmental stimuli experienced during development.
Within the zone of tolerance, temperature affects fish performance at a variety of levels of biological organization. In some fish species, developmental temperature has already been proven as an acute regulating factor in sex determination, muscle anatomy, body shape, meristic characters, developmental progress, thermal acclimation capacity and swimming performance. Despite the significance of water temperature for the cardiac function in developing fish, to our knowledge, this is the first study demonstrating the programming of cardiac shape by the temperature which was experienced by the fish during embryonic and larval stages.
The purpose of this study was to determine the acute and prolonged changes of cardiac anatomy and swimming performance in response to developmental temperature, several months after exposure to different thermal regimes. On a secondary level, we intended to study the expression of five marker genes involved in heart remodeling, as well as the effect of developmental temperature on zebrafish body shape and relative fecundity. We used the zebrafish as an experimental model since it’s characterized by a variety of plastic responses against developmental temperature and has the same ontogenetic pattern of development with most teleost fish.
Zebrafish embryos were subjected to three developmental temperature treatments (TD = 24, 28 or 32 °C) up to metamorphosis and then all maintained under common conditions (28° C) up to adulthood. Cardiac anatomy measurements were assessed via micro-ct imaging, at the end of the different treatments and at the adult stage. Critical swimming speed (Ucrit) was used as a reliable indicator of aerobic swimming performance. Swimming performance assays took place at the juvenile and adult stages, after 20 and 100 days of acclimation at the common conditions (28 °C). After the swimming performance assays, fish body shape measurements took place. Gene expression analysis was assessed at the end of the different treatment period and after 20 days of acclimation at the common conditions (28 °C). The effect of developmental temperature on zebrafish relative fecundity was measured at the adult stage at 180 and 300 days after the end of the different exposure period.
The elevation of TD induced a significant increase of the ventricle roundness in male (22% decrease of VL/VD ratio), but not in female zebrafish. Sex-related differences were evident in the rest of the under-study morphometrical characters of the zebrafish heart. At 32 °C, females exhibited more elongated ventricles than males of the same temperature treatment (22% increase of VL/VD ratio). Bulbus arteriosus length of males raised at 28 °C was longer than of females raised at the same temperature (7% increase of BaL/SL in males). Relative ventricular volume of males at 24 and 28 °C was significant bigger than of females of the same temperature treatments (26% increase in VeV/SL in both temperature treatments). Bulbus arteriosus relative volume was also found bigger in males of 28 and 32 °C as opposed to females of the same temperature treatments (22% increase of BuLV/SL in males raised at 28 °C and 19% increase in males raised at 32 °C). Developmental temperature exhibited significant effects on the juvenile heart shape (elevation of TD induced a 10% reduction in VL/VD ratio). As for the rest of the heart morphometrics, juveniles exhibited significant acclimation responses to developmental temperature. The elevation of TD resulted in shorter bulbus arteriosus length (12% reduction of BaL/SL) and to a significant reduction of the relative ventricular and bulbus arteriosus volume (45% reduction in VeV/SL and BuLV/SL).
Adults reared at different developmental temperatures exhibited significant differences in critical swimming speed velocities. In males, elevation of TD from 24 to 32 °C resulted in reduction of critical swimming speed by 16%. Females raised at 32 °C exhibited significant lower swimming capacity as opposed to females at 24 and 28 °C (25% and 30% reduction respectively). Sex-related responses were evident in all temperature treatments where males achieved higher velocities than females (16% increase of RUcrit 24 °C, 7% increase of RUcrit at °C, and 26% increase at 32 °C). In juveniles the effect of temperature on swimming performance was not significant but the existence of a similar trend with adults cannot be omitted.
Gene expression analysis that was performed at the end of the temperature treatments revealed significant up-regulation at colder temperatures (24 °C) of nppa (16-fold higher expression levels than 28 °C), myh7 (74-fold higher expression levels than 28 °C), mybpc3 (9-fold hogher expression levels than 28 °C) and actinb2 (4-fold higher experession levels than 32 °C). After 20 days of acclimation at the common conditions (28 °C), differences in most of the genes studied were normalized, while there was a significant up-regulation of nfatc1 and mybpc3 genes at 28 °C opposed to 24 °C (increase of gene expression by 3- and 2-fold, respectively).
Developmental temperature had a significant effect on male body shape, inducing significant differences between the 24 °C group and the 28 and 32°C groups. Males raised at 24 °C had more slender bodies as opposed to the rest of the groups. In females, developmental temperature had also a significant effect on body shape, inducing significant differences between the 32 °C group and the 24 and 28°C groups. Females raised at 32 °C had more swollen bellies as compared to rest of the temperature groups. In juveniles, developmental
temperature induced significant differences between the 32 °C group and the 24 and 28 °C groups. Juveniles raised at 32 °C appeared to have more slender bodies.
With regards to thermal plasticity of fecundity, female zebrafish that were raised at lower developmental temperatures (24 °C) exhibited higher relative fecundity than females raised at high developmental temperatures (32 °C). Results were not confirmed statistically at the second experimental replicate.
Our data show that rises in water temperature at critical early life stages can have detrimental effects on cardiac anatomy and physical capacity of adult fish. Given that critical swimming speed has a vital ecological significance for fish populations (i.e. for migration, foraging, feeding) our findings have relevance to fish populations and the concerns of global warming and seasonal temperature perturbations. Besides fish, our results could also have implications to the plastic responses of other ectothermic vertebrates.
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Μελέτη της επίδρασης της θερμοκρασίας ανάπτυξης στην καρδιά, στην κολυμβητική ικανότητα και στη γονιμότητα του zebrafish Danio rerio (Hamilton 1822)
