JOURNAL OF GEOPHYSICAL RESEARCH, VOL. 110, C09S09, doi:10.

1029/2004JC002564, 2005

Physiological effects on fishes in a high-CO2 world

Atsushi Ishimatsu, Masahiro Hayashi, and Kyoung-Seon Lee
Marine Research Institute, Nagasaki University, Nagasaki, Japan

Takashi Kikkawa
Central Laboratory, Marine Ecology Research Institute, Chiba, Japan

Jun Kita
Research Institute of Innovative Technology for the Earth, Kyoto, Japan
Received 27 June 2004; revised 19 December 2004; accepted 26 January 2005; published 23 August 2005.

[1] Fish are important members of both freshwater and marine ecosystems and constitute
a major protein source in many countries. Thus potential reduction of fish resources by
high-CO2 conditions due to the diffusion of atmospheric CO2 into the surface waters or
direct CO2 injection into the deep sea can be considered as another potential threat to the
future world population. Fish, and other water-breathing animals, are more susceptible to a
rise in environmental CO2 than terrestrial animals because the difference in CO2 partial
pressure (PCO2) of the body fluid of water-breathing animals and ambient medium is
much smaller (only a few torr (1 torr = 0.1333 kPa = 1316 matm)) than in terrestrial
animals (typically 30–40 torr). A survey of the literature revealed that hypercapnia acutely
affects vital physiological functions such as respiration, circulation, and metabolism, and
changes in these functions are likely to reduce growth rate and population size through
reproduction failure and change the distribution pattern due to avoidance of high-CO2
waters or reduced swimming activities. This paper reviews the acute and chronic effects of
CO2 on fish physiology and tries to clarify necessary areas of future research.
Citation: Ishimatsu, A., M. Hayashi, K.-S. Lee, T. Kikkawa, and J. Kita (2005), Physiological effects on fishes in a high-CO2 world,
J. Geophys. Res., 110, C09S09, doi:10.1029/2004JC002564.

1. Introduction recent years CO2 ocean sequestration has increasingly been

[2] Our present knowledge is still too scarce to predict
what will happen to fishes in a high-CO2 world on both
short and long timescales. On the basis of the available
evidence, high CO2 will affect fishes directly through acute,
potentially lethal impacts on vital physiological functions,
and chronic, sublethal impacts on more subtle but still
essential aspects of the life cycle. High CO2 levels in water
will also influence fishes indirectly through impacts on the
aquatic environment such as increasing water temperature,
and on ecosystem structure and function. On the timescale
of 100– 200 years, shallow water fishes will incur greater
effects of CO2 diffused from the atmosphere, whereas deep
sea species may be affected by high CO2 levels, if CO2 is
injected into the deep sea to reduce the rapid increase of
atmospheric CO2 concentration [Ohsumi, 2004]. Over lon-
ger timescales of 500 – 1000 years, the ocean CO2 will
equilibrate with the atmosphere, thereby affecting all marine
biota. Our previous review pointed out that information
about the effects of CO2 on fishes was very limited for
marine species as compared with that for freshwater fishes,
and virtually nothing was known for deep sea species
[Ishimatsu and Kita, 1999]. This is unfortunate because in
Copyright 2005 by the American Geophysical Union.
0148-0227/05/2004JC002564
suggested as a potential mitigation method, and knowledge
about the impacts of CO2 on marine organisms is essential
to discuss the feasibility of this strategy [Seibel and Walsh,
2001, 2003].
[3] Water-breathing animals, including fish, are more
susceptible to a rise in environmental CO2 concentration
than terrestrial animals, because of the lower CO2 partial
pressures (PCO2) of their body fluids [Ultsch and Jackson,
1996]. Ventilation of water-breathing animals is usually
governed by O2 stimuli because of the low O2 concentra-
tions in water, even in equilibrium with air (O2 concentra-
tion in water is 1/30 of that in air at the same PO2
[Dejours, 1988]). This situation necessarily results in hy-
perventilation of fish in terms of CO2 excretion, because
CO2 solubility in water far exceeds O2 solubility. As a
result, the PCO2 of fish body fluids is only a few torr, which
is one order lower than in air-breathing animals (for
example 40 torr in man). Thus elevations of CO2 (hyper-
capnia) in water will easily reverse the normal outward
diffusion of CO2 from the fish body. CO2, as a small,
uncharged gaseous molecule, rapidly crosses cell mem-
branes, and acidifies the body fluids through its rapid
conversion into carbonic acid catalyzed by a ubiquitous
enzyme, carbonic anhydrase. As almost all cellular processes
are highly dependent on pH, CO2 undoubtedly has far-
reaching impacts on living organisms.

C09S09 1 of 8
C09S09 ISHIMATSU ET AL.: CO2 EFFECTS ON FISHES
Figure 1. Comparison of mean mortalities of embryos
(open bars, n = 5) and larvae (solid bars, n = 3) of red
seabream, Pagrus major, exposed to two seawater pH levels
lowered by either CO2 or HCl (condition A, pH 6.2;
condition B, pH 5.9). PCO2 levels of the CO2 group for the
conditions A and B are 37 and 74 torr, respectively.
Seawater in the acid group was vigorously aerated before
experimentation to assure equilibrium with aerial PCO2.
Exposure periods for embryos and larvae were 6 hours and
24 hours, respectively. Asterisks show significant difference
between test groups (condition A, Welch’s t-test; condition
B, Student’s t-test; one asterisk, p < 0.05; four asterisks, p <
0.0001 (reprinted from Kikkawa et al. [2004], with
permission from Elsevier).Vertical lines indicate standard
deviation.
[4] Even though the physiological effects of high CO2
in water can be largely ascribed to decreases in body
fluid pH, increased proton concentrations in the ambient
water per se cannot explain the physiological perturba-
tions caused by elevated levels of CO2. The effects of
water acidification by mineral acids such as HCl and
H2SO4 are less than those caused by high CO2, when
tested at the same water pH. This has been demonstrated
clearly by our recent results that compared mortalities of
eggs and larvae of red seabream, Pagrus major, at two
levels of seawater pH lowered by either CO2 or HCl
[Kikkawa et al., 2004]. Exposure to high CO2 resulted in
far higher mortalities than acid exposure at both pH
levels (Figure 1). This is due to a much higher biomem-
brane permeability of gaseous CO2 compared with that of
H+ [Heisler, 1986; Morris et al., 1989]. Although earlier
studies used mortality data based on acid exposure to
evaluate impacts of CO2 ocean sequestration on marine
animals [e.g., Auerbach et al., 1997], the predicted
impacts based on such studies may be milder than those
caused by hypercapnia.
2. Lethal CO2 Levels of Fish
2.1. Short-Term Exposure
[5] Acutely lethal PCO2 appears to vary largely be-
tween fish species, although a quantitative evaluation of
LC50 (median lethal concentration) has rarely been con-
ducted. Grøttum and Sigholt [1996] reported 96 hour
LC50 of 50 torr and 120 hour LC50 of 47 torr for
2 of 8
C09S09
European seabass, Dicentrarchus labrax, at 15C. Be-
cause there was no increase in blood lactate concentration
in the dead fish, they concluded that the death was not
due to asphyxiation. A 22 hour LC50 value for red
seabream can be estimated to be 42 torr at 25C from
the reported survival-PCO2 relationship [Takeda and
Itazawa, 1983]. When exposed to seawater equilibrated
with a gas mixture containing 5% CO2 (PCO2 37 torr) at
20C, 100% mortality occurred at 8 and 48 hours for
yellowtail, Seriola quinqueradiata, and Japanese flounder,
Paralichthys olivaceus, respectively. The elasmobranch,
starspotted dogfish, Mustelus manazo, was more toler-
ant, because only 20% mortality occurred by 72 hour
exposure to PCO2 of 52 torr at 17C [Hayashi et al.,
2004]. European eel, Anguilla anguilla, is highly toler-
ant to hypercapnia, showing no mortality during expo-
sure to progressively increasing CO2 conditions up to
80 torr at 23C [McKenzie et al., 2002].
[6] Embryos and larvae are often more vulnerable to
environmental stress than adults. For example, fish eggs
and larvae are known to be particularly sensitive to low
water pH [Brown and Sadler, 1989]. Therefore even if
elevated CO2 levels in aquatic environments are tolerable
for adults, fish populations may still be affected through
the decreased survival of eggs and larvae. We recently
studied the lethal effects of elevated CO2 on the early
development of both eggs and larvae of several marine
fish [Kikkawa et al., 2003]. The 24 hour LC50 for the
egg stage ranged widely from 10 torr to 70 torr among
species. A similar pattern of developmental changes in
CO2 susceptibilities was found for two species tested (red
seabream and Japanese whiting, Sillago japonica); the
egg and juvenile stages were more susceptible to CO2
than the stages between them (Figure 2) and adults (e.g.,
24 hour LC50 for Japanese flounder eggs was 21 torr
[Kikkawa et al., 2003], while no adult mortality of the
same species occurred during 72 hour exposure to PCO2
of 22 torr [Hayashi et al., 2004]). Therefore hypercapnic
exposure during the egg and juvenile stages could have
profound impacts on population size of the affected stock.
In contrast, Wagner et al. [2002] reported that CO2
anesthesia of rainbow trout broodstock had no effect on
the survival, hatching and the percentage of abnormal fry
reared under normocapnic conditions.
2.2. Long-Term Exposure
[7] Lethal CO2 effects during long-term exposure are also
poorly known. Fivelstad et al. [1999] reported 5 and 8%
mortality at the end of 62 day exposure to 5 and 9 torr,
respectively for freshwater Atlantic salmon (Salmo salar)
smolts, while 1 and 5% mortalities were found for seawater
postsmolts of the same species at 12 and 20 torr after
43 days [Fivelstad et al., 1998]. Smart et al. [1979] found
little difference in mortality for freshwater rainbow trout,
Oncorhynchus mykiss, reared for 275 days at PCO2 of 4 to
17 torr. No mortality occurred by 10 week exposure of
juvenile spotted wolf fish, Anarhichas minor, to PCO2 of up
to 20 torr [Foss et al., 2003]. Life-long CO2 exposure
experiments have not been conducted nor is there any
information on effects of CO2 exposure over generations.
Assessing long-term effects is of utmost importance because
the fate of a population depends on how the population
C09S09 ISHIMATSU ET AL.: CO2 EFFECTS ON FISHES
Figure 2. Ontogenetic changes of the median lethal PCO2
in red seabream, Pagrus major (solid circles), and Japanese
whiting, Sillago japonica (open circles). Horizontal lines
show range. (a) Exposure of 360 min. (b) Exposure of
24 hours. Abbreviations are as follows: cle, cleavage stage;
emb, embryo stage; pre, preflexion stage; fle, flexion stage;
pos, postflexion stage; juv, juvenile stage (after Kikkawa et
al. [2003], Environmental Toxicology # 2003 (John
Wiley)).
responds through reproduction, early development, and
recruitment of the young.
3. Physiological Effects of CO2 on Fish
3.1. Acid-Base Regulation
[8] Elevation of blood PCO2 by environmental hypercap-
nia results in a transient but significant drop of the arterial pH,
even if ambient PCO2 remained at high levels. This drop of
blood pH is attenuated by chemical buffering by proteins and
other low-molecular-weight buffering substances, and sub-
sequently compensated for within 3 – 24 hours by the transfer
of acid-base relevant ions with the surrounding water,
mainly through the gills [Heisler, 1986]. The transport of
HCO3 and/or H+ across the body surface inevitably is
accompanied by the exchange of counterions to maintain
the electroneutrality of body fluids, but the precise mecha-
nisms of, and the cell types responsible for, the transport
process remain obscure, especially in marine fish.
[9] For freshwater fish, the role of H+-ATPase coupled
with Na+ channels has increasingly been recognized in
recent years as an important mechanism of acid-base
regulation. Usually, H+-ATPase is localized to the pavement
cells of the gills [Claiborne et al., 2002]. On the other hand,
several studies have demonstrated morphological alterations
of mitochondria-rich chloride cells by hypercapnic expo-
sure, suggesting the possible involvement of chloride cells
3 of 8
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in acid-base regulation in freshwater fish [Goss et al., 1992;
Cameron and Iwama, 1987; Kaneko et al., 1999]. In vivo
blood analysis from the vasculature draining filamental
tissues rich in chloride cells also suggested the involvement
of chloride cells in acid-base regulation for rainbow trout
exposed to hypercapnic waters [Iwama et al., 1993].
[10] For marine fish, the current model of acid-base
regulation emphasizes the Na+/H+ exchange brought about
by Na+/H+ ion exchangers [Claiborne et al., 2002]. How-
ever, plasma Cl invariably decreases with a nearly equi-
molar increase in plasma HCO3 under low (1 – 2%) levels
of hypercapnia for marine teleosts, with little or no change
in plasma Na+ concentration [Toews et al., 1983; Larsen
and Jensen, 1997; Hayashi et al., 2004]. Only at higher,
possibly lethal levels of hypercapnia, plasma Na+ increased
significantly [Hayashi et al., 2004]. This attests to the
importance of an acid-base compensation mechanism that
involves Cl ions as the main counterion. Marine fish
actively secrete Na+ and Cl through chloride cells to
counterbalance diffusional entry of ions [Zadunaisky,
1984]. We have recently found that Na+/K+ ATPase activity
in the gills increased significantly by exposure to 1% and
5% CO2 and that the apical opening area of branchial
chloride cells significantly increased by exposure to 5%
CO2 in the Japanese flounder (M. Hayashi et al., unpub-
lished manuscript, 2005). These observations support the
hypothesis that chloride cells are also involved in acid-base
regulation of marine fish during hypercapnic exposure.
[11] Elasmobranchs appear to employ different acid-base
regulatory mechanisms, because responses of plasma sodi-
um and chloride ion concentrations to hypercapnia differ
from those in teleosts. Previous studies on elasmobranchs
reported either no change in plasma Cl during hypercapnia
for skate [Graham et al., 1990] and larger spotted dogfish
[Randall et al., 1976], or a small but significant decrease for
spiny dogfish, Squalus acanthias [Cross et al., 1969]. We
also found significantly smaller decreases in Cl , as com-
pared with rises in plasma HCO3 in starspotted dogfish
[Hayashi et al., 2004].
[12] It should be noted that long-term CO2 effects de-
scribed above most likely became manifest long after
extracellular (and certainly intracellular [Larsen et al.,
1997]) pH had been completely restored to preexposure
levels, which usually needs only several hours to a few days
depending on CO2 levels used, salinity, and acid-base
regulatory capacity of fish species, among others. Thus
longer-lasting physiological responses, such as new steady
state ionic concentrations of body fluids, and/or energetic
costs necessary to maintain such new steady state might be
responsible for the observed long-term adverse effects of
CO2 [Cameron and Iwama, 1988].
3.2. Respiratory System
[13] Although the decisive role of O2 for controlling fish
gill ventilation is unquestionable, there is increasing evi-
dence for the direct role of CO2 for stimulating ventilation
in fish [Gilmour, 2001]. CO2 and/or pH sensitive chemo-
receptors that enhance gill ventilation during hypercapnia
have been proposed to occur in the gills, and there are some
indications of the occurrence of such chemoreceptors also in
the central nervous system of some fish [Milsom, 2002;
Remmers et al., 2001].
C09S09 ISHIMATSU ET AL.: CO2 EFFECTS ON FISHES
[14] Changes in PCO2 do not generally affect the O2
consumption rate of resting fish (carp Cyprinus carpio
[Takeda, 1991]; mummichog Fundulus heteroclitus and
spot fish Leiostomus xanthurus [Cochran and Burnett,
1996]), although a transient increase in O2 consumption
rate was reported upon application of hypercapnia for some
species (larger spotted dogfish Scyliorhinus stellaris
[Randall et al., 1976]; skate Raja ocellata [Graham et al.,
1990]; rainbow trout [Thomas et al., 1983]). Thus even in
the face of reduced oxygen-binding affinity and capacity of
hemoglobin by CO2 (Bohr and Root effects), fish appear to
4 of 8
C09S09
be able to satisfy the oxygen demand of tissues as long as
no exercise is required.
[15] On the other hand, active metabolism, determined as
oxygen consumption rate during swimming, decreased
significantly with increasing water PCO2 in several teleosts
[Basu, 1959; Beamish, 1964]. However, CO2 levels used in
these early studies were extremely high (up to 70 torr in
Basu [1959] and 165 torr in Beamish [1964]). More
recently, McKenzie et al. [2003] reported insignificant
effect on active metabolism and swimming speed when
the European eel, Anguilla anguilla, was acclimated for
6 weeks to water PCO2 of 15– 45 torr, although eels are
exceptional in many physiological aspects (extreme toler-
ance to hypoxia, hypercapnia and low plasma Cl concen-
trations), and therefore the results may not be generalized.
The effect of CO2 on swimming performance and active
metabolism requires additional investigation using moderate
CO2 levels and fish more representative of the dominant
marine species.
3.3. Blood Circulation
[16] In spite of the well established negative inotropic
effect (i.e., reduction of contractility) of hypercapnia on fish
myocardium in vitro (see Farrell and Jones [1992] for
review), in vivo cardiac responses to hypercapnia vary
among fishes, and so do the blood pressure responses (see
Perry and Gilmour [2002] for a review). Obviously, in vivo
cardiovascular responses to hypercapnia vary with the
severity as well as the duration of hypercapnia imposed
on the fish, let alone interspecific variability, and probably
the experimental temperature. We have recently shown that
cardiac output (blood volume pumped per unit time) de-
creased rapidly when yellowtail was exposed to a lethal
level of hypercapnia (seawater equilibrated with a gas
mixture containing 5% CO2 (Figure 3)). Heart rate did not
change during the CO2 exposure, and it was a reduction of
the stroke volume (blood volume pumped per contraction)
that caused a reduction in cardiac output. Cardiac output
began to fall before blood pressure started to rise [Lee et al.,
2003]. The high solubility of CO2 will quickly lower
intracellular pH of the myocardium, reducing contractility
through an antagonism between hydrogen ions and intra-
cellular calcium ions [Gesser and Poupa, 1983]. Therefore
Figure 3. Time-dependent changes in (a) cardiac output
_ (b) heart rate (HR), (c) stroke volume (SV), and
(Q),
(d) arterial blood pressure (BP) in yellowtail, Seriola
quinqueradiata, exposed to two levels of hypercapnia.
Cardiac output and blood pressure were measured for fully
recovered fish that were chronically equipped with a Doppler
flow probe in the ventral aorta (cardiac output) and a cannula
in the dorsal aorta (blood pressure). Heart rate and stroke
volume were obtained from the blood flow signal. Circles
represent data at 1% CO2 (water PCO2 = 7 torr, n = 5), and
triangles represent data at 5% CO2 (water PCO2 = 38 torr, n =
6). Open symbols indicate a significant difference from
the control values (P < 0.05). Downward triangles indicate
that n was decreased due to fish death. No statistical analysis
was applied to these points. Means ± standard error of the
mean (after Lee et al. [2003], Zoological Science # 2003
(Zoological Society of Japan)).
C09S09 ISHIMATSU ET AL.: CO2 EFFECTS ON FISHES
it is conceivable that the reduced myocardial contractility is
responsible for the observed lowering of the stroke volume.
[17] Most previous studies of the effects of hypercapnia
on cardiac function have examined only acute responses
(experimental period commonly shorter than 30 min). No
long-term data are available. It has been shown that con-
tractility of the heart muscle strip of some fish (Platichthys
(=Pleuronectes) flesus and P. platessa) restores to normo-
capnic control levels under sustained hypercapnia, if bicar-
bonate concentration of the bathing medium is elevated
[Gesser and Poupa, 1983]. Bicarbonate concentration
increases during acid-base compensation and therefore it
remains to be investigated if cardiac output may be restored.
[18] Effects of hypercapnia on regional blood flow in fish
are little studied. Söderström and Nilsson [2000] reported
increases in brain blood flow during hypercapnia in rainbow
trout and crucian carp, Carassius carassius. Iwama et al.
[1993] found an unchanged blood flow partitioning between
two vascular circuits in the gills (arterial-arterial and arterio-
venous pathways) when rainbow trout was exposed to
PCO2 of 15 torr.
3.4. CO2 Receptors
[19] Konishi et al. [1969] demonstrated the occurrence of
CO2 receptors in the palatine surface of carp. The receptors
are sensitive also to acids, but the sensitivity was highest for
CO2 under the same pH conditions. Gill ventilatory move-
ment was suppressed by stimulation of these CO2 receptors.
Similar CO2 receptors were demonstrated for Japanese eel,
Anguilla japonica [Yoshii et al., 1980] and rainbow trout
[Yamashita et al., 1989]: Thresholds of these CO2 receptors
are 0.5 torr and 0.6 torr, as calculated from the reported
dissolved CO2 concentrations (2  10 5 mol L 1 and 4 
10 5 mol L 1) for the eel and trout, respectively. In
addition, fish have branchial CO2 receptors that seem to
be involved in cardiac and ventilatory responses to changes
in water CO2 levels [Perry and Gilmour, 2002].
[20] Knowledge about detection limits of CO2 has impli-
cations in understanding the avoidance behavior of fish
subjected to high-CO2 water. Such avoidance may affect the
distributions or migration patterns of fish, and result in
changes in marine ecosystem structure. Jones et al. [1985]
studied preference/avoidance responses to H+ and CO2 of a
freshwater fish, Arctic char Salvelinus alpinus. The fish
avoided CO2 plumes at plume/ambient gradients greater
than 1.5 torr, but was attracted at lower levels. The results
also demonstrated that fish behaviorally discriminate be-
tween H+ and CO2, relying principally on CO2 for orienta-
tion within H+/CO2 gradients. This is in sharp contrast to
the in situ observation at a depth of 625 m off California
that deep sea fish and invertebrates were in fact attracted to
CO2 plumes of presumably very high PCO2 (seawater pH
near the plume was 5.6 [Tamburri et al., 2000]). Fish show
a stress response when exposed to hypercapnic conditions
as evidenced by elevations of blood adrenaline and nor-
adrenaline concentrations [Perry et al., 1986]. Therefore
biological significance of the observations by Tamburri et
al. [2000] remains to be investigated.
3.5. Reproduction and Early Development
[21] There are some reports on the effects of CO2 on
sperm motility and fertilization ability of fish, mainly by
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Figure 4. Comparison of 24 hour LC50 among fish egg
(filled circles) (cle, cleavage; emb, embryo), larva/juvenile
(open circles) (fle, flexion; juv, juvenile; pre, preflexion;
pos, postflexion) [Kikkawa et al., 2003], and invertebrate
juvenile (asterisks) (T. Kikkawa et al., unpublished data,
2004). LC50 values are expressed as CO2 percent of gas
mixture used to equilibrate the experimental tank water
(1%CO2 = 7.4 torr). Species are as follows: Elt, eastern
little tuna, Euthynnus affinis; Jf, Japanese flounder,
Paralichthys olivaceus; Jw, Japanese whiting, Sillago
japonica; Kc, Kisslip cuttlefish, Sepia lycidas; Kp,
Kuruma prawn, Penaeus japonicus; Os, oval squid,
Sepioteuthis lessoniana; Rs, red seabream, Pagrus major.
Japanese whiting was measured with a 16.7 hour LC50.
aquaculture scientists. High CO2 applied to milt inhibited
sperm motility upon subsequent dilution of the milt in a
CO2-concentration-dependent manner [Ingermann et al.,
2002; Bencic et al., 2001] and thus the fertilizing ability
[Bencic et al., 2000]. The addition of acetazolamide (car-
bonic anhydrase inhibitor) to milt solution blocked the CO2
inhibition of sperm motility, suggesting that lowering of
intracellular pH is responsible for the inhibition [Bencic et
al., 2000].
[22] Sperm motility was stopped immediately when CO2
is applied to sperms already activated by dilution [Inaba et
al., 2003]. According to the authors, this type of inhibition
was observed only for flatfish species (Pleuronectida:
Limanda yokohamae, L. hersensteini, Psetta maxima,
Verasper moseri, and V. variegates), among those examined.
Sperm motility can also be arrested by the addition of
NaHCO3 to the diluting medium (buffered seawater), and
can be reactivated by the addition of a carbonic anhydrase
inhibitor (ethoxyzolamide [Inaba et al., 2003]).
[23] Effects of elevated CO2 on fish eggs are less well
known. Figure 4 summarizes the 24 hour LC50 values of
several marine fish and invertebrates in early stages [Kikkawa
et al., 2003; T. Kikkawa, unpublished data, 2004]. It is quite
clear that CO2 sensitivity varies largely among species as well
as developmental stages within a species.
3.6. Growth
[24] Effects of CO2 on fish growth have been investigated
by aquaculture scientists in recent years because of the
C09S09 ISHIMATSU ET AL.: CO2 EFFECTS ON FISHES
Table 1. Effect of High Water CO2 on Fish Growtha
Species % Body Weight
Atlantic salmon (Salmo salar) 63
Spotted wolffish (Anarhichas minor) 64
Rainbow trout* (Oncorhynchus mykiss)
Low-mineral diet 52
Normal mineral diet 76
White sturgeon (Acipenser transmontanus) 62
a
Effect of CO2 is expressed as percentages of final body weight of the experimental fish to control fish at the end of experimental period. The % body
weight values for rainbow trout were estimated from the reported figure. Abbreviations are as follows: SW, seawater; FW, freshwater.
development of intensive aquaculture systems. In these
systems, high fish density, low water exchange and low
bubbling rate of pure oxygen may cause the accumulation
of metabolic end products such as CO2 and ammonia in the
rearing water to an extent that they may affect growth, food
intake and health of the reared fish. Table 1 summarizes the
results of growth reduction by high water CO2. Growth was
suppressed by 24 – 49% in these experiments. Since a
similar level of PCO2 was used in these studies, the
relationship of growth suppression and ambient CO2 levels
cannot be determined. High CO2 appears to reduce foraging
activity, thereby decreasing growth [Crocker and Cech,
1996; Ross et al., 2001; Foss et al., 2003]. There are also
some indications that CO2 suppresses taste nerve responses
to taste stimulants in Japanese eel [Yoshii and Yotsui, 1997].
Thus in nature, CO2 may reduce fish growth through
decreased chances to detect food.
[25] These growth suppressions are likely, at least in part,
to be a result of metabolic suppression and reduced protein
synthesis under elevated CO2 conditions, as recently
reported for Antarctic fish, Pachycara brachycephalum
and Lepidonotothen kempi [Langenbuch and Pörtner,
2003]. The effect was best explained by lowering of
extracellular/intracellular pH.
4. Future Studies
4.1. Effects of High CO2 on Shallow Water Fishes
[26] CO2 emitted into the atmosphere will diffuse into the
surface water of oceans and freshwater environments. Acid-
base regulation of freshwater fishes is in general slower and
less efficient, because of the low concentrations of counter-
ions available for the transfer of acid-base relevant ions
through the epithelium. On the other hand, some freshwater
animals may be adapted and highly tolerant to hypercapnic
conditions because at least in some areas, such as tropical
stagnant waters covered by thick vegetation, water PCO2
can be as high as 40 torr [Ultsch, 1976].
[27] Experiments concerning the biological impacts of
increased CO2 diffusion into ocean surface waters should
reflect predicted CO2 concentrations and temperatures
under different future scenarios [Caldeira and Wickett,
2003; Kheshgi, 2003]. Since predicted future CO2 con-
centrations in the atmosphere are lower than the known
lethal concentrations for fish (for example, the expected
peak value is about 1.4 torr around the year 2300
according to Caldeira and Wickett [2003]), research
emphasis should be placed on long-term sublethal effects,
such as those on growth and reproduction. As discussed
above, nothing is known about what happens to fish
populations if they are exposed to low but sustained
CO2 conditions for long periods of time.
C09S09
PCO2, torr Temperature, C Duration, days Medium Reference
20 15 – 16 43 SW Fivelstad et al. [1998]
20 6 70 SW Foss et al. [2003]
17 9 275 FW Smart et al. [1979]
17 9 275 FW Smart et al. [1979]
20 19 28 FW Crocker and Cech [1996]
[28] Field experiments elevating CO2 levels of a restricted
area are probably the only way to understand consequences
of long-term hypercapnia on aquatic ecosystems. Under-
standing biological responses of field communities to water
acidification was only possible when some streams were
purposefully acidified [Sutcliffe and Hildrew, 1989]. Cer-
tainly, care must be taken to control CO2 concentrations at
desired levels, and the size of study area must be carefully
determined by thoroughly considered experimental plans.
[29] In relation to long-term CO2 impacts, effects of CO2
on endocrine systems that regulate maturation, gonadal
development, and reproductive behavior require investiga-
tion. Mechanisms for the developmental changes in CO2
sensitivity should also be clarified to understand why some
stages are more susceptible than others.
4.2. Effects of High CO2 on Deep Sea Fishes
[30] CO2 ocean sequestration can locally elevate PCO2 to
high levels. Injected CO2 will then be diluted with a vast
volume of ocean water. Thus both acute and chronic
impacts are of major concern in considering the biological
impacts of CO2 ocean sequestration.
[31] Animals living in the deep sea are thought to be more
sensitive to environmental changes than shallow water ones,
because the former have adapted to the stable deep sea
environment. Deep sea organisms are characterized by low
biological activities, long life span, high sensitivity to
environmental perturbations, high biodiversity, and low
density. Therefore once population size is reduced by
environmental perturbations, its recovery, if it should occur,
will need a long period of time [Childress, 1995; Haedrich,
1997; Shirayama, 1998]. Physiological characteristics of
deep sea animals have been discussed in detail by Seibel
and Walsh [2003].
[32] Despite repeated arguments that deep sea animals are
more sensitive to high CO2 than shallow water animals,
experimental verification is still lacking. This is certainly
due to the difficulty in capturing these animals alive, and
conducting experiments either in situ or in the laboratory;
the latter also requires transportation with minimal distur-
bance to the animals. We have recently found that adults of
some deep sea fishes, such as rough snailfish, Careproctus
trachysoma (Liparidae, distribution: 300 – 800 m), and
Tanaka’s eelpout Lycodes tanakai (Zoarcidae, distribution:
>300 m) in the Japan Sea (water temperature 1 –2C below
depths of greater than 200 m), can be captured and kept
alive under atmospheric pressure for extended periods, as
long as the rearing water temperature is maintained below
5C. In fact, the snailfish tolerates surgical procedures
required for chronic implantation of a cannula for blood
sampling. We have recently conducted a preliminary CO2
exposure experiment on the snailfish using a high-pressure
6 of 8
C09S09 ISHIMATSU ET AL.: CO2 EFFECTS ON FISHES
chamber (max. pressure 375,000 torr (=50 MPa)) at 5C,
and successfully recorded electrocardiograms from fish
exposed to seawater equilibrated with 1% CO2 at a hydro-
static pressure of 75,000 torr for several hours, although the
results are premature to be included here (A. Ishimatsu et
al., unpublished manuscript, 2005).
[33] It should be noted that animals would be subjected to
fluctuating PCO2 when CO2 droplets are released from the
lower end of the nozzle used for CO2 ocean sequestration
[Sato and Sato, 2002]. Our preliminary studies indicated
that mortality differed considerably when fish were exposed
to fluctuating levels of environmental CO2, as compared
with data obtained using constant CO2 levels (T. Kikkawa et
al., unpublished data, 2004). For example, a step-wise
increase in ambient PCO2 resulted in lower mortalities as
compared with a sudden elevation to the same final PCO2.
Also, a sudden drop of PCO2 can cause nearly immediate
mortalities of surviving fish. This should also be included in
experimental protocols to assess the effects of CO2 under
realistic conditions of CO2 ocean sequestration.
[34] In situ deep sea CO2 exposure experiments such as
one conducted by Barry et al. [2004] are certainly another
highly valuable and informative approach to the issue if
successfully applied to fish. With this approach, deep sea
fish and other fauna can be exposed to hypercapnia under in
situ conditions of low temperature and high pressure, even
though it requires specialized equipments and ROVs.
[35] Long-term CO2 effects on deep sea fish are more
difficult to assess. However, an experimental system was
developed more than 10 years ago that allowed long-term
experiments under high pressures for aquatic animals
[Sébert et al., 1990]. In this system, fish was confined in
a fish chamber that can be pressurized in a hyperbaric
chamber. Water in the fish chamber was supplied from an
external high-pressure water circulation system, which
allowed a long-term confinement of the fish. Sébert et al.
[1990] reported the oxygen consumption of the eel Anguilla
anguilla over a period of 31 days. Using such systems for
deep sea fish experiments is certainly a possibility for
assessing the effects of chronic CO2 under high pressures,
although measurable physiological parameters in these
systems are limited. Again, in situ deep sea CO2 exposure
[Barry et al., 2004] may be a useful method for evaluating
long-term CO2 effects. Barry et al. [2004] reported changes
in biovolume of meiofauna taxa exposed to CO2 for
4.5 weeks.
[36] Acknowledgment. This study was supported by New Energy
and Industrial Technology Development Organization (NEDO) and
Research Institute of Innovative Technology for the Earth (RITE).
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