The study by Gräns et al. (Gräns et al., 2014) investigated growth performance and oxygen demand at rest and during recovery from fatiguing exercise in Atlantic halibut (Hippoglossus hippoglossus) under simulated scenarios of ocean warming and acidification. The authors claim that their data, when used to evaluate the aerobic scope for exercise, do not explain temperature-dependent growth. They thus question the general use of the concept of ‘oxygen and capacity limited thermal tolerance’ (OCLTT) in explaining the onset of thermal limitation of fishes under field conditions (Pörtner and Knust, 2007; Pörtner and Farrell, 2008). There are important lessons to learn from this study about how, or how not, to investigate the concept of OCLTT and aerobic scope in thermal limitation.

From a conceptual point of view, the term aerobic scope should not be constrained to use only with exercise. In the context of OCLTT, it makes sense to use the term aerobic scope for all routine performances that draw on aerobic energy such as growth, reproduction and steady-state swimming. The question is whether the chasing protocol imposed on a strictly benthic fish such as halibut provide suitable estimates of performance, of aerobic scope as used by growth and of climate sensitivity.

Methodological issues invariably constrain what experimental data can say. In their paper, Gräns et al. routinely measured growth in active fish. In contrast, aerobic scope was determined in fish exercised to exhaustion and from differences between EPOC (excess post-exercise oxygen consumption) and resting metabolism. An over-riding difficulty with tests using exhaustive exercise is that performance itself is not properly quantified. The physiological state of the animals during steady-state growth clearly differs from that during the non-steady recovery state post-exercise. The latter is characterized by exponentially declining oxygen consumption, very low venous PO2 (Farrell and Clutterham, 2003; Lee et al., 2003), release of catecholamines (Reid et al., 1998), acidosis and shifted metabolite concentrations, pathways and ion equilibria, all of which are non-steady state. Furthermore, the data analysis by Gräns et al. does not build on a clear functional background. Linear regressions are used for non-linear data. At normal water pH and the highest temperature, standard metabolic rate and EPOC indicate a clear decline in metabolic scope (their fig. 1), which is not picked up by the selected polynomial fit. Also, the post hoc statistics do not support a global effect of CO2. For EPOC, statistical significance is reached only at certain temperatures, rather than all test temperatures. Growth was depressed by CO2 only on the cold side of the studied temperature range, matching predictions of synergistic effects at thermal extremes (Pörtner and Farrell, 2008). Thus, the overall CO2 effect could be viewed as small and hardly discernible.

In contrast to the authors' contention, the mechanisms influencing growth have not been investigated. The paper does not provide relevant insight on underlying aerobic scope or the limits important in the field. However, relevant thermal limitation studied in the laboratory should mimic (be ‘calibrated’ by) field observations. The physiological status of experimental organisms should be similar to that of organisms experiencing thermal limitation in their natural environments. For this, the suitability of non-steady state EPOC measurements after chasing in halibut is not clear. In other cases, calibration by field data exists: in benthic eelpout, limited aerobic scope for growth and associated cardiac limitation parallel a reduction in growth performance and a loss in abundance in extreme summer temperatures in the field (Pörtner and Knust, 2007). In Pacific salmon, limited aerobic scope for steady state swimming and associated cardiocirculatory capacity constrain adult spawners during their upstream migration (Eliason et al., 2011). In both cases, the organisms are in routine steady state, as in the field. This indicates how to investigate OCLTT, performance and relevant aerobic scope more successfully, by preferentially capturing the routine situation on relevant time scales. Fatiguing exercise protocols are poorly qualified to elaborate the subtleties of thermal effects, because of low resolution and rapidly shifting physiological states.

Fatigue and recovery involve respiratory (CO2 accumulation) and non-respiratory (metabolic) changes in acid–base status. These phenomena constrain the validity of the chasing protocol to identify and quantify the effects of ocean acidification (OA) caused by elevated ambient CO2 levels. Long-term acclimation to OA increases bicarbonate concentrations and associated buffering in blood and tissues and thereby, reduces the acidosis caused by fatiguing exercise. This may protect the EPOC-derived ‘aerobic scope’ from being depressed by more severe acidosis. The data reported by Gräns et al. do not test such a hypothesis. They cannot easily be compared with or be used as evidence against projected effects of OA on thermal limits of performance and fitness under routine aerobic steady-state conditions (Pörtner, 2012).

Whether the thermal windows of exercise and growth are the same is an important question to keep in mind for future studies. The data reported by Gräns et al. do not allow an answer to this question. Both windows may in fact differ if the physiological backgrounds of the fish body differ, for example, as a result of release of catecholamines under exhaustive conditions. As catecholamines push the organism away from resting and activate functional reserves, this blurs the picture with respect to OCLTT at extreme temperatures. Gräns et al. investigated cardiocirculatory scope and limitations after maximum adrenergic stimulation, in an in situ perfused heart preparation and during a manipulative increase of input pressure. The resulting mobilisation of cardiac functional reserves and pattern of cardiac limitation would fit the condition of enforced fatiguing exercise more than the subtle thermal limitation seen in growth, at first caused by limited cardiocirculatory capacity and associated cost increments, and the resulting shift in energy budget (Pörtner and Knust, 2007). The studies on eelpout and halibut are thus not comparable, to say the least, and the results of the halibut study again, cannot easily be interpreted with respect to their relevance in the field.

Accordingly, we need to clearly find out under which conditions a species becomes thermally limited at the ecosystem level and then the respective situation needs to be simulated in experimental work. In most cases, steady-state aerobic functions such as growth and reproduction and their underlying aerobic scope are more universal indicators of subtle thermal constraints. Steady-state aerobic exercise can also become thermally limiting if used during a constraining life phase (e.g. upstream migration of mature salmon). This should not mislead investigators to impose maximum exercise protocols on all animals for assessing aerobic scope, regardless of whether they exercise in nature and reach steady state or not.

Overall, the paper by Gräns et al. is too much about the conceptual debate rather than about how the data might fit OCLTT or what caused apparent differences. There are other comments to make and emphasize the wider context: a recent metaanalysis provided evidence that across organism domains the highest complexity levels and processes coordinating the largest number of body compartments are the first to be thermally constrained (Storch et al., 2014). Heat limits in animals are in fact lower than in most other organism domains and OCLTT provides an explanation because it suggests coordination of the largest number of body compartments across tissues through O2 supply and demand capacities. OCLTT traces the performance curve of the organism because it primarily focuses on sublethal limits and associated functional constraints, which develop from pejus (mismatch in oxygen supply and demand systems) to critical limits (transition to anaerobic metabolism) and then molecular denaturation, with ecological effects starting beyond pejus limits. Finally, limitations reach lethal values as an end point. These limits, transitions and links need to be further investigated because they are subject to evolutionary adjustments and shifts, for example, during transition to air breathing (Giomi et al., 2014). The evolutionary process and timeline is largely ignored in the current debate but requires consideration. OCLTT is also about linking levels of biological organization, from ecosystem to systemic to molecular, and about oxygen (energy) allocation to specific performances. It thus does not suffice to study whether oxygen affects lethal limits. A reductionist approach limiting the experimental investigation to one level such as the whole organism, regardless of its natural mode of life, physiological background and ecological context, comes with the risk of misleading results and erroneous conclusions. Last, but not least, testing a conceptual framework such as OCLTT requires considering its most recent definition and underlying theory comprehensively. Also, experimental biologists may need to adjust some of their classical concepts and approaches to better link to other subdisciplines, such as ecology and evolutionary biology.

Acknowledgements

The author gratefully acknowledges the constructive criticism and support of this assessment by A. P. Farrell.

References

Eliason
E. J.
,
Clark
T. D.
,
Hague
M. J.
,
Hanson
L. M.
,
Gallagher
Z. S.
,
Jeffries
K. M.
,
Gale
M. K.
,
Patterson
D. A.
,
Hinch
S.
,
Farrell
A. P.
(
2011
).
Differences in thermal tolerance among sockeye salmon populations
.
Science
332
,
109
-
112
.
Farrell
A. P.
,
Clutterham
S. M.
(
2003
).
On-line venous oxygen tensions in rainbow trout during graded exercise at two acclimation temperatures
.
J. Exp. Biol.
206
,
487
-
496
Giomi
F.
,
Fusi
M.
,
Barausse
A.
,
Mostert B
Pörtner H.-O.
,
Cannicci
S.
(
2014
).
Improved heat tolerance in air drives the recurrent evolution of air-breathing
.
Proc. R. Soc. B
218
,
20132927
.
Gräns
A.
,
Jutfelt
F.
,
Sandblom
E.
,
Jönsson
E.
,
Wiklander
K.
,
Seth
H.
,
Olsson
C.
,
Dupont
S.
,
Ortega-Martinez
O.
,
Einarsdottir
I.
,
Björnsson
B. T.
,
Sundell
K.
,
Axelsson
M.
(
2014
).
Aerobic scope fails to explain the detrimental effects on growth resulting from warming and elevated CO2 in Atlantic halibut
.
J. Exp. Biol.
217
,
711
-
717
.
Lee
C. G.
,
Farrell
A. P.
,
Lotto
A.
,
Hinch
S. G.
,
Healey
M. C.
(
2003
).
Excess post-exercise oxygen consumption in adult sockeye (Oncorhynchus nerka) and coho (O. kisutch) salmon following critical speed swimming
.
J. Exp. Biol.
206
,
3253
-
3260
.
Pörtner
H.-O.
(
2012
).
Integrating climate-related stressor effects on marine organisms: unifying principles linking molecule to ecosystem-level changes
.
Mar. Ecol. Progr. Ser.
470
,
273
-
290
.
Pörtner
H.-O.
,
Knust
R.
(
2007
).
Climate change affects marine fishes through the oxygen limitation of thermal tolerance
.
Science
315
,
95
-
97
.
Pörtner
H.-O.
,
Farrell
A. P.
(
2008
).
Physiology and climate change
.
Science
322
,
690
-
692
.
Reid
S. G.
,
Bernier
N. J.
,
Perry
S. F.
(
1998
).
The adrenergic stress response in fish: control of catecholamine storage and release
.
Comp. Biochem. Physiol. C
120
,
1
27
.
Storch
D.
,
Menzel
L.
,
Frickenhaus
S.
,
Pörtner
H.-O.
(
2014
).
Climate sensitivity across the domains of life: Limits to evolutionary adaptation shape species interactions
.
Glob. Chang. Biol.
20
,
3059
-
3067
.