Christian Tudorache

Establishing zebrafish as a novel exercise model: swimming economy, swimming-enhanced growth and muscle growth marker gene expression.

Background Zebrafish has been largely accepted as a vertebrate multidisciplinary model but its usefulness as a model for exercise physiology has been hampered by the scarce knowledge on its swimming economy, optimal swimming speeds and cost of transport. Therefore, we have performed individual and group-wise swimming experiments to quantify swimming economy and to demonstrate the exercise effects on growth in adult zebrafish. Methodology/Principal Findings Individual zebrafish (n = 10) were able to swim at a critical swimming speed (Ucrit) of 0.548±0.007 m s−1 or 18.0 standard body lengths (BL) s−1. The optimal swimming speed (Uopt) at which energetic efficiency is highest was 0.396±0.019 m s−1 (13.0 BL s−1) corresponding to 72.26±0.29% of Ucrit. The cost of transport at optimal swimming speed (COTopt) was 25.23±4.03 µmol g−1 m−1. A group-wise experiment was conducted with zebrafish (n = 83) swimming at Uopt for 6 h day−1 for 5 days week−1 for 4 weeks vs. zebrafish (n = 84) that rested during this period. Swimming zebrafish increased their total body length by 5.6% and body weight by 41.1% as compared to resting fish. For the first time, a highly significant exercise-induced growth is demonstrated in adult zebrafish. Expression analysis of a set of muscle growth marker genes revealed clear regulatory roles in relation to swimming-enhanced growth for genes such as growth hormone receptor b (ghrb), insulin-like growth factor 1 receptor a (igf1ra), troponin C (stnnc), slow myosin heavy chain 1 (smyhc1), troponin I2 (tnni2), myosin heavy polypeptide 2 (myhz2) and myostatin (mstnb). Conclusions/Significance From the results of our study we can conclude that zebrafish can be used as an exercise model for enhanced growth, with implications in basic, biomedical and applied sciences, such as aquaculture.

Partition of aerobic and anaerobic swimming costs related to gait transitions in a labriform swimmer

SUMMARY Members of the family Embiotocidae exhibit a distinct gait transition from exclusively pectoral fin oscillation to combined pectoral and caudal fin propulsion with increasing swimming speed. The pectoral–caudal gait transition occurs at a threshold speed termed Up–c. The objective of this study was to partition aerobic and anaerobic swimming costs at speeds below and above the Up–c in the striped surfperch Embiotoca lateralis using swimming respirometry and video analysis to test the hypothesis that the gait transition marks the switch from aerobic to anaerobic power output. Exercise oxygen consumption rate was measured at 1.4, 1.9 and 2.3 L s–1. The presence and magnitude of excess post-exercise oxygen consumption (EPOC) were evaluated after each swimming speed. The data demonstrated that 1.4 L s–1 was below the Up–c, whereas 1.9 and 2.3 L s–1 were above the Up–c. These last two swimming speeds included caudal fin propulsion in a mostly steady and unsteady (burst-assisted) mode, respectively. There was no evidence of EPOC after swimming at 1.4 and 1.9 L s–1, indicating that the pectoral–caudal gait transition was not a threshold for anaerobic metabolism. At 2.3 L s–1, E. lateralis switched to an unsteady burst and flap gait. This swimming speed resulted in EPOC, suggesting that anaerobic metabolism constituted 25% of the total costs. Burst activity correlated positively with the magnitude of the EPOC. Collectively, these data indicate that steady axial propulsion does not lead to EPOC whereas transition to burst-assisted swimming above Up–c is associated with anaerobic metabolism in this labriform swimmer.

Pop Up Satellite Tags Impair Swimming Performance and Energetics of the European Eel ( Anguilla anguilla )

Pop-up satellite archival tags (PSATs) have recently been applied in attempts to follow the oceanic spawning migration of the European eel. PSATs are quite large, and in all likelihood their hydraulic drag constitutes an additional cost during swimming, which remains to be quantified, as does the potential implication for successful migration. Silver eels (LT = 598.6±29 mm SD, N = 9) were subjected to swimming trials in a Steffensen-type swim tunnel at increasing speeds of 0.3–0.9 body lengths s−1, first without and subsequently with, a scaled down PSAT dummy attached. The tag significantly increased oxygen consumption (MO2) during swimming and elevated minimum cost of transport (COTmin) by 26%. Standard (SMR) and active metabolic rate (AMR) as well as metabolic scope remained unaffected, suggesting that the observed effects were caused by increased drag. Optimal swimming speed (U opt) was unchanged, whereas critical swimming speed (U crit) decreased significantly. Swimming with a PSAT altered swimming kinematics as verified by significant changes to tail beat frequency (f), body wave speed (v) and Strouhal number (St). The results demonstrate that energy expenditure, swimming performance and efficiency all are significantly affected in migrating eels with external tags.

Covariation between behaviour and physiology indicators of coping style in zebrafish (Danio rerio)

All vertebrates exhibit physiological responses to a wide variety of stressors. The amplitude and profile of the response depend on the intensity, duration, controllability and predictability of the stressor, but there is also individual variation in the response, termed coping style. A better understanding of the expression of coping styles is of great value for medical applications, animal welfare issues and conservation. Here, we investigated the effect of repeated netting stress on proactive and reactive zebrafish (Danio rerio) as an upcoming model system for stress research. Fish were separated by coping styles according to the order of entering a novel environment. Subsequently, repeated netting stress was applied as stressor, over a period of 21 days. Full-body cortisol levels were determined at 0, 15, 30, 60 and 120 min after the last repeated stress event. Our results show that reactive fish display i) increased basal cortisol concentrations after being repeatedly stressed, ii) higher cortisol secretion over time and iii) slow recovery of cortisol concentration towards basal levels after the last repeated stress event. This study shows for the first time in zebrafish that different coping styles are associated with different cortisol responses during the recovery from stress over time and that coping styles can explain otherwise unaccounted variation in physiological stress responses.

Hormonal and ion regulatory response in three freshwater fish species following waterborne copper exposure.

We evaluated effects of sublethal copper exposure in 3 different freshwater fish: rainbow trout (Oncorhynchus mykiss), common carp (Cyprinus carpio) and gibel carp (Carassius auratus gibelio). In a first experiment we exposed these fishes to an equally toxic Cu dose, a Cu level 10 times lower than their 96 h LC50 value: 20, 65, and 150 microg/L Cu. In a second series we exposed them to the same Cu concentration (50 microg/L). Na+/K+-ATPase activity in gill tissue was disturbed differently in rainbow trout then in common and gibel carp. Rainbow trout showed a thorough disruption of plasma ion levels at the beginning of both exposures, whereas common carp and gibel carp displayed effects only after 3 days. Rainbow trout and common carp thyroid hormones experienced adverse effects in the beginning of the exposure. The involvement of prolactin in handling metal stress was reflected in changes of mRNA prolactin receptor concentrations in gill tissue, with an up regulation of this mRNA in rainbow trout and a down regulation in gibel carp, which was more pronounced in the latter. Overall, rainbow trout appeared more sensitive in the beginning of the exposure, however, when it overcame this first challenge, it handled copper exposure in a better manner then common and gibel carp as they showed more long term impacts of Cu exposure.

Behavioral changes in response to sound exposure and no spatial avoidance of noisy conditions in captive zebrafish.

Auditory sensitivity in fish serves various important functions, but also makes fish susceptible to noise pollution. Human-generated sounds may affect behavioral patterns of fish, both in natural conditions and in captivity. Fish are often kept for consumption in aquaculture, on display in zoos and hobby aquaria, and for medical sciences in research facilities, but little is known about the impact of ambient sounds in fish tanks. In this study, we conducted two indoor exposure experiments with zebrafish (Danio rerio). The first experiment demonstrated that exposure to moderate sound levels (112 dB re 1 μPa) can affect the swimming behavior of fish by changing group cohesion, swimming speed and swimming height. Effects were brief for both continuous and intermittent noise treatments. In the second experiment, fish could influence exposure to higher sound levels by swimming freely between an artificially noisy fish tank (120–140 dB re 1 μPa) and another with ambient noise levels (89 dB re 1 μPa). Despite initial startle responses, and a brief period in which many individuals in the noisy tank dived down to the bottom, there was no spatial avoidance or noise-dependent tank preference at all. The frequent exchange rate of about 60 fish passages per hour between tanks was not affected by continuous or intermittent exposures. In conclusion, small groups of captive zebrafish were able to detect sounds already at relatively low sound levels and adjust their behavior to it. Relatively high sound levels were at least at the on-set disturbing, but did not lead to spatial avoidance. Further research is needed to show whether zebrafish are not able to avoid noisy areas or just not bothered. Quantitatively, these data are not directly applicable to other fish species or other fish tanks, but they do indicate that sound exposure may affect fish behavior in any captive condition.

Conservation physiology of animal migration

Conservation physiology has great potential to help us understand how migratory animals interact with current and future anthropogenic threats. Migration is inherently challenging such that additional stressors derived from altered environments or interaction with human infrastructure or activities could lead to long-term changes to migratory phenotypes.

Pectoral fin beat frequency predicts oxygen consumption during spontaneous activity in a labriform swimming fish (Embiotoca lateralis)

The objective of this study was to identify kinematic variables correlated with oxygen consumption during spontaneous labriform swimming. Kinematic variables (swimming speed, change of speed, turning angle, turning rate, turning radius and pectoral fin beat frequency) and oxygen consumption (MO2) of spontaneous swimming in Embiotoca lateralis were measured in a circular arena using video tracking and respirometry, respectively. The main variable influencing MO2 was pectoral fin beat frequency (r2 = 0.71). No significant relationship was found between swimming speed and pectoral fin beat frequency. Complementary to other methods within biotelemetry such as EMG it is suggested that such correlations of pectoral fin beat frequency may be used to measure the energy requirements of labriform swimming fish such as E. lateralis in the field, but need to be taken with great caution since movement and oxygen consumption patterns are likely to be quite different in field situation compared to a small lab tank. In addition, our methods could be useful to measure metabolic costs of growth and development, or bioassays for possible toxicological effects on fish.

Social interactions, predation behaviour and fast start performance are affected by ammonia exposure in brown trout (Salmo trutta L.)

In fish, fast starts are brief, sudden accelerations during predator-prey encounters. They serve for escape and predation and are therefore ecologically important movements. Fast starts are generated by glycolytic muscle performance and are influenced by many internal and external factors. It is known that ammonia pollution has a major effect on the glycolytic muscle action, thus creating conditions in which fast start performance might be reduced and predation rates altered. Therefore, escape response and predation strikes were investigated in brown trout (Salmo trutta) of 10 and 20 cm body length exposed to an elevated (1 mg l(-1)) ammonia concentration for 24 and 96 h. Various locomotor and behavioural variables were measured. In C-starts, i.e. an escape start where the fish bends into a C-shaped position, ammonia exposure had no effect on response latency. After 96 h of exposure, cumulative distance, maximum swimming speed and turning radius of the prey were all significantly reduced and the escape went in no definite direction. The effect of ammonia exposure was more pronounced in large fish than in small fish. Predation strikes were also affected. Distance, speed and turning radius were significantly lower in exposed fish. Agonistic behaviour of dominant fish was significantly reduced and fish spent more time resting. Predator behaviour was also altered and the number of prey captured was reduced. This study shows that ammonia exposure affects brown trout escape response mainly through a reduction in fast start velocity and through an impairment of directionality. Thus, in addition to a reduced strength of the response, ammonia exposure could also reduce the fishs elusiveness facing a predator. Predation rate and social interactions are disrupted and predator-prey relationships could be altered.

Recent advances in telemetry for estimating the energy metabolism of wild fishes.

Metabolic rate is a critical factor in animal biology and ecology, providing an objective measure that can be used in attributing a cost to different activities and to assessing what animals do against some optimal behaviour. Ideally, metabolic rate would be estimated directly by measuring heat output but, until recently, this has not been easily tractable with fishes so instead metabolic rate is usually estimated using indirect methods. In the laboratory, oxygen consumption rate is the indirect method most frequently used for estimating metabolic rate, but technical requirements preclude the measurement of either heat output or oxygen consumption rate in free-ranging fishes. There are other field methods for estimating metabolic rate that can be used with mammals and birds but, again, these cannot be used with fishes. Here, the use of electronic devices that record body acceleration in three dimensions (accelerometry) is considered. Accelerometry is a comparatively new telemetric method for assessing energy metabolism in animals. Correlations between dynamic body acceleration (DBA) and oxygen consumption rate demonstrate that this will be a useful proxy for estimating activity-specific energy expenditure from fishes in mesocosm or field studies over extended periods where other methods (e.g. oxygen consumption rate) are not feasible. DBA therefore has potential as a valuable tool for attributing cost to different activities. This could help in gaining a full picture of how fishes make energy-based trade-offs between different levels of activity when faced with conflicting or competing demands arising from increased and combined environmental stressors.