Bjørn Tirsgård:
The effects of temperature, hypoxia and hypercapnia on the metabolism of Atlantic cod Gadus morhua

The Atlantic cod (Gadus morhua L. 1758) is an important species both as a major top predator in the ecosystem and economical marine resource in the fishing industry of Northern Europe (Andersen, 2012). As an ectothermic water breather the Atlantic cod is continuously exposed to annual changes in the water conditions affecting their performance, growth and reproduction. Variations in the water conditions between years and due to climatic changes, affects the survival rate, population dynamic and bio-energetic distribution of the Atlantic cod (Hansson et al., 1996; Essington et al., 2001). This thesis is a series of studies investigating how two of the most important abiotic environmental parameters, temperature and low oxygen levels (hypoxia) affect different aspects of larvae and juvenile Atlantic cod metabolism. The temperature studies were developed especially with the aim to be implemented into bio-energetic models used to predict survival, growth and reproduction rates and cod stock dynamics.

Furthermore, due to the growing interest of the continuing global increase in atmospheric and oceanic CO₂ levels (ICES, 2010), and the rapid expansion of fish farming in land-based recirculated aquaculture systems exposing fish to chronically elevated CO₂ levels (Timmons et al., 2010) I also investigated the effect of elevated dissolved CO₂ concentration (termed hypercapnia) on the metabolism and digestion in the Atlantic cod.

Migration between water layers might change water temperature several degrees in the Øresund resulting in a change in the metabolism (Aertebjerg et al., 1998). Specially in the fragile early life stages optimizing the metabolism to either a low maintenance cost in cold water or increased metabolism and potential higher growth rate in warm water might be crucial and thus important for bio-energetic models (Werner et al., 1996; Leising and Franks, 1999; Werner et al., 2001; Lough et al., 2005). The temperature effect on the metabolism of Atlantic cod larvae however have received little attention (Laurence, 1978; Finn et al., 2002; Peck and Buckley, 2008), likely due to hatching and experimental difficulties. In paper I, I determined the effect of temperature (5, 7.5 and 10°C) on the normal range of the routine metabolic rate (RMR, μg O₂ h^-1) by determining a lower (RMR_low) and higher (RMR_high) level of RMR in Atlantic cod larvae at age 14-60 days post-hatch. The study showed that RMR_low and RMR_high increased with age, by scaling exponents ranging between 0.58 and 0.87 at the three temperature tested. In periods with low prey availability a low water temperature and body size increase the survival rate whereas high water temperatures enhance growth rate when prey is abundant. A temperature increase from 5 to 10°C resulted in an average RMR_low value Q₁₀ of 1.66, which is lower than observed in other populations (Q₁₀ of 2.4-5.31) (Laurence, 1978; Finn et al., 2002; Peck and Buckley, 2008). This indicates that Øresund larvae are well adapted to acute changes in water temperature.

Temperature affects most biochemical processes and thus the metabolism. The purpose of paper II was to investigate how thermal changes affect the physiology of juvenile Atlantic cod and thus provide data for bio-energetic models estimating predation and growth rates and predicting stock dynamic (Hansson et al., 1996; Essington et al., 2001). By intermittent respirometry I determined the effect of temperature (2, 5, 10, 15 and 20°C) and body mass (~30-460g) on the standard metabolic rate (SMR, mg O₂ h^-1), maximum metabolic rate (MMR, mg O₂ h^-1) and the metabolic scope (MS, mg O₂ h^-1). The study showed that SMR increased with body mass at all five temperatures, with an average scaling exponent of 0.87 (0.82-0.92). Q₁₀ values were 1.8-2.1 at temperatures between 5 and 15°C but higher (2.6-4.3) between 2°C and 5°C and lower (1.6-1.4) between 15 and at 20°C in 200 and 450 g Atlantic cod. The deviation in Q₁₀ at high and low temperatures compared to Q₁₀ measured between 5 and 15°C increased with body mass indicating an increasing temperature effect on SMR with increasing body mass. MMR increased with temperature in small juvenile Atlantic cod (50g) but in larger Atlantic cod MMR independent of temperature between 10, 15 and 20°C. The temperature where MS was highest (T_opt) was determined by multiple functions describing the combined effect of body mass and temperature on SMR and MMR. Topt decreased with body mass, being respectively 14.5, 11.8 and 10.9°C in a 50, 200 and 450 g Atlantic cod. For cod of all sizes MS was reduced by 30-35% at 2°C compared to at T_opt whereas only larger fish experienced a decrease in MS at temperatures above 10°C. In conclusion, independent of cod size low water temperatures resulted in a decrease in MS whereas the reduction of MS at high temperatures was only evident for larger Atlantic cod (200 and 450g), caused by a plateau in MMR at temperatures above 10°C. High temperatures thus seem favourable for smaller (50g) juvenile Atlantic cod, but not larger conspecifics (200 and 450g).

The purpose of paper III was to improve the understanding of how temperature effects the specific dynamic action (SDA) in juvenile Atlantic cod. Intermittent flow respirometry was used to measure the pre and post-feeding oxygen consumption rate of Atlantic cod fed a single meal of herring corresponding to 5% wet body mass at controlled temperatures of 2, 5, 10, 15 and 20°C. The study showed a statistically increase in pre-feeding MO₂ (SMR) and an increase in maximum post-feeding MO₂ (SDA_scope) with increasing temperature whereas no significant difference was observed concerning the time to reach SDAscope. The SDA duration increased significantly from 80 h at 10°C to 130-160 h at 2, 15 and 20°C and reached a maximum duration of 250 h at 5°C. The SDA coefficients were significantly lower at 2 and 10°C (5.4-6.3%) compared to 5, 15 and 20°C (10.4-12.4%). The conclusion is that within the temperature range from 5 to 20°C the optimum temperature for digestion in juvenile Atlantic cod is around 10°C which have the lowest SDA duration, cost and coefficient. Furthermore the decrease in SDA duration and cost, and coefficient found at 2°C suggests that the digestion is inhibited at low winter temperatures (2°C) probably due to a reduction in the apparent digestion coefficient.

Recent observations have suggested that cod make excursions into sever hypoxic water layers likely to search for benthic fauna (Neuenfeldt et al., 2009). The aim of paper IV was therefore to simulate a 40 min excursion into severe hypoxic water below S_crit and determine a possible additional energy cost. The study showed that the increase in the oxygen consumption rate (excess post-hypoxia oxygen consumption, C_EPHO) above the normal resting level (SMR) when re-entering well-oxygenated water was 7 times higher than the decrease in the oxygen consumption rate (oxygen deficit, Do₂) observed during exposure to severe hypoxia. Based on that the additional energetic cost of foraging one hour per day in severe hypoxic water, was estimated to increase the daily energy costs by 15%. A foraging strategy with excursions to severe hypoxic water is therefore only energetically affordable if the food consumption is increased accordingly.

While many fish species can survive in relatively high dissolved CO₂ concentrations (hypercapnia) (Crocker and Cech, 1996; McKenzie et al., 2003), long-term exposure to sub-lethal environmental hypercapnia is known to affect growth, feed conversion efficiency and body condition index (Smart, 1981; Fivelstad et al., 1998; Cecchini et al., 2001; Hosfeld et al., 2008; Moran and Støttrup, 2011). The physiological basis underlying the pathological effects of environmental hypercapnia are poorly understood, in particular for marine fish species, so we investigated whether changes in energy expenditure and the specific dynamic action (SDA) of digestion and assimilation could account for the lower growth of adult Atlantic cod under environmental hypercapnia. Fish adapted to 800 and 9200 μatm CO₂ exhibited no difference in maintenance metabolic rates, which concurs with previous studies on this and other fish species (Melzner et al., 2009). At 9200 μatm CO2 Atlantic cod had a significantly diminished (14%) maximum aerobic capacity. Hypercapnia prolonged the SDA by 23%, but did not increase the total oxygen demand for the digestion and assimilation of a meal. The longer SDA process time may offer an explanation for the observation of lower feed intake, growth and condition factor in long-term hypercapnia studies (Fivelstad et al., 1998; Moran and Støttrup, 2011). Comparison of aerobic scope and cardiac performance during digestion suggested that reduced oxygen delivery capability under hypercapnia could be one mechanism by which CO₂ prolongs SDA, although our results could not definitively demonstrate this effect.