Effect of increased calcium concentration in sea water on calcification and photosynthesis in the scleractinian coral Galaxea fascicularis
Analytical Electron Microscopy Laboratory, Department of Zoology, La Trobe University, Bundoora (Melbourne), Victoria 3083, Australia
* Author for correspondence (e-mail: zooam{at}zoo.latrobe.edu.au )
Accepted
![]() |
Summary |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key words: coral, Galaxea fascicularis, calcification, photosynthesis, zooxanthellae, bisphosphonate
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Alternatively it has been suggested that calcification enhances
photosynthesis by providing a source of protons that convert sea water
HCO3- to CO2 and H2O, thereby
supplying some of the CO2 used in photosynthesis
(McConnaughey, 1989;
McConnaughey and Whelan,
1997
). This has been described as the trans-calcification
model. Conceptually, this model is attractive because it explains why an
organism only two cell layers thick produces a massive external skeleton of
calcium carbonate (McConnaughey,
1994
).
Whilst it can be readily demonstrated that the prevention of photosynthesis
results in a marked reduction or cessation of calcification
(Vandermeulen et al., 1972;
Barnes, 1985
), it is only
recently that attempts have been made to determine whether a reduction in
calcification in corals also leads to a reduction in photosynthesis. Yamashiro
(1995
) showed that
bisphosphonate reduced 14C incorporation into the skeleton (i.e.
calcium carbonate deposition) but not into the tissues (i.e. photosynthesis)
of a zooxanthellate coral and concluded that calcification is not necessary
for photosynthesis. Bisphosphonate, however, may prevent the formation of
calcium carbonate crystals but not necessarily the formation of amorphous
calcium carbonate; it is known to prevent calcium phosphate crystallization
but not the formation of amorphous calcium phosphate
(Francis et al., 1969
).
Gattuso et al. (2000
) showed
that artificial sea water with a low calcium concentration lowered
calcification rate but did not reduce the production of photosynthetic oxygen
and concluded that `calcification is not a significant source of
photosynthetic CO2'.
The experimental procedures used so far to investigate the relationship between calcification and photosynthesis have relied upon low or zero concentrations of calcium in artificial sea water. A possible problem with this approach is that low-calcium artificial sea water may have a profoundly deleterious effect on some aspects of coral physiology (A. T. Marshall and P. L. Clode, manuscript submitted for publication). We have reinvestigated the concept of calcification enhancing photosynthesis, therefore, by studying the relationship between calcification and photosynthesis when these processes take place in standard sea water that has an increased calcium concentration.
![]() |
Materials and methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
High-calcium sea water
High-calcium sea water was prepared by adding either 100 mg l-1
or 200 mg l-1 calcium (added as CaCl2.2H2O)
(Swart, 1980) to filtered (0.2
µm Millipore filter) sea water (FSW), which resulted in an increased
calcium concentration of 2.5 mmol l-1 (FSW+100) and 5 mmol
l-1 (FSW+200), respectively. All solutions were made freshly as
required, to pH 8.11 (standard FSW) and pH 8.12 (high-calcium FSW). Standard
artificial sea water (ASW) was prepared according to Benazet-Tambutte et al.
(1996
). The pH of all
incubation media was monitored before and after each incubation and was
observed to be constant to within 0.1 pH units.
Scintillation counting of 45Ca and 14C in
skeleton and tissues
Polyps were incubated in glass jars. The jars were immersed in a large
fibreglass outdoor aquarium through which sea water flowed at a rate
sufficient to maintain a constant temperature. The aquarium was not shaded and
corals were exposed to light levels similar to those they would have naturally
experienced on the reef flat at low water. The incubation sea water was
vigorously stirred and aerated by a diaphragm-operated aquarium pump, except
when NaH14CO3 was present in the sea water, when it was
necessary to prevent exchange with atmospheric CO2. Samples (200
µl) of incubation medium were taken before and after incubation for
determination of specific activities of radioisotopes.
Incubation times and post-incubation processing depended upon the
experiment being done. In all experiments polyps were washed after incubation
for 1.5-2 h in 21 of vigorously aerated, fresh incubation medium to remove
coelenteric 45Ca (Tambutte et
al., 1996). This procedure was carried out at the incubation
temperature and in low light levels [photosynthetic photon flux density (PPFD)
7-45 µmol photons m-2 s-1]. Following the wash, bare
corallite that would have been a site for isotopic exchange was removed and
discarded. Depending on the experiment, polyps were cut into pieces with wire
cutters and tissue was removed by digestion with 5 mol l-1 NaOH at
58 °C for periods of 20 min to 1 h. Where appropriate the digest was
neutralised with HCl and samples taken for scintillation counting and protein
determination. After rinsing in tap water, the skeleton was dried overnight at
60 °C, weighed and dissolved in HCl. Samples of the acid digest were taken
for scintillation counting.
(a) Ten polyps were preincubated in 200 ml aerated FSW and ten polyps in 200 ml FSW+100 for 1 h, under laboratory conditions (PPFD, 8 µmol photons m-2 s-1, 24 °C). Following preincubation, the polyps were transferred to jars containing 200 ml of fresh aerated FSW or FSW+100, prior to the addition of 2 µCi (74 kBq) ml-1 of 45CaCl2 and incubated in the outdoor aquarium (PPFD, 50-1600 µmol photons m-2 s-1, 23 °C) for 4 h. A second experiment was carried out exactly as the first but with three sets of five polyps incubated in each of FSW, ASW and FSW+200 in the outdoor aquarium (PPFD, 1200-1950 µmol photons m-2 s-1, 29 °C) after preincubation in the laboratory (PPFD, 7 µmol photons m-2 s-1, 27 °C).
(b) Five polyps were preincubated in glass jars containing 200 ml aerated FSW and five polyps in 200 ml aerated FSW+100 for 1 h, under laboratory conditions (PPFD, 3 µmol photons m-2 s-1, 23 °C). Following preincubation, the polyps were transferred to jars containing 100 ml of fresh FSW or FSW+100 prior to the addition of 2 µCi (74 kBq) ml-1 of NaH14CO3, and incubated in the outdoor aquarium (PPFD, 1600 µmol photons m-2 s-1, 23 °C) for 2 h. A second experiment was carried out exactly as the first but with five polyps incubated in each of FSW, ASW and FSW+200 in the outdoor aquarium (PPFD, 500-1950 µmol photons m-2 s-1, 29 °C) after preincubation in the laboratory (PPFD, 5.5 µmol photons m-2 s-1, 27.5 °C).
(c) Two series each of five experimental and five control polyps were preincubated in glass jars containing 200 ml of FSW either with or without 0.5 mmol l-1 1-hydroxyethylidene-diphosphonic acid (HEBP) (Fluka Chemika) for 15 min prior to the addition of 1 µCi (37 kBq) ml-1 of 45CaCl2. Following the addition of radioisotope the polyps were then incubated for a further 2.2 h in the outdoor aquarium (PPFD, 1900 µmol photons m-2 s-1, 27.5 °C). One series of polyps was treated after incubation in the standard way, i.e. tissue was removed followed by dissolution in acid. The other series was treated with HCl, after washing, drying and weighing, without prior removal of the tissue. After dissolving the skeleton in acid the remaining tissue was filtered, dried and weighed to facilitate calculation of skeletal mass.
Measurement of photosynthesis and respiration rates
Measurements of oxygen concentration were carried out in small custom-made
Perspex chambers holding 20 ml of FSW (assumed to be free of bacteria), which
was vigorously stirred with a magnetic stirrer. Three polyps, closely matched
for size, were placed in each chamber. Light was provided to each chamber by a
double, fibre optic, light source delivering a PPFD of approximately 1200
µmol photons m-2 s-1. Oxygen concentrations were
measured at intervals of 1 min over a period of 15 min by means of a
Clarke-type electrode (accurate to 0.01 mg l-1) inserted into each
chamber. Readings were made using a YSI Model 58 oxygen meter attached to each
electrode. The temperature of the laboratory, and therefore the chambers, was
maintained at the same temperature as the prevailing inter-reefal sea water
temperature. Following oxygen measurements in the light, the chambers were
shrouded in aluminium foil and oxygen concentrations were recorded as before
to obtain respiratory oxygen consumption. Polyps were then removed from the
chambers and each chamber's complement was incubated separately in 200 ml of
aerated FSW + 100 for 2 h. Following this incubation, the polyps were
reintroduced into the chambers, filled with FSW + 100, and oxygen
concentrations were measured using the same protocol and conditions as
previously. At the termination of the experiment the polyps were frozen and
held at -70 °C until processed by heating at 58 °C in 5 mol
l-1 NaOH for 1 h. The digest was neutralised with a known volume of
hydrochloric acid and samples taken for protein determination.
Rates of gross photosynthesis and respiration were obtained from regression equations and net photosynthesis was calculated from the algebraic sum of gross photosynthesis and respiration.
Oxygen measurements on sets of three polyps were also made with essentially the same equipment except that the recorder output of the YSI oxygen meters was connected to a Macintosh Powerbook via a MacLab interface (AD Instruments). In this way oxygen concentrations were continuously recorded whilst solutions of HEBP were injected (with pressure compensation) into the measuring chambers. The amount of HEBP injected was adjusted to give a final concentration of 0.5 mmol l-1. Rates of change of oxygen concentration were determined directly from the computer traces using the MacLab software.
Protein determination
Protein content in tissue digests was determined without precipitation
using a Protein Assay Kit (Lowry method, procedure no. 5656, Sigma
Diagnostics). A standard curve was constructed from bovine serum albumin
concentrations ranging from 0 to 400 mg ml-1 protein and
absorbances were read using a spectrophotometer (UV-2401PC) at a wavelength of
750 nm. For polyp samples, the neutralised tissue digest was diluted tenfold
to produce a concentration within the range of the standard curve. Following
the addition of Folin and Ciocalteu's reagent, all samples were centrifuged
for 30 min at 1900 g. Absorbances of the supernatant were then
obtained as before, with concentrations (uncorrected for dilution)
automatically calculated from the standard curve.
Statistical analysis
Statistical analyses were carried out using the computer program JMP 3.1.6
(SAS Institute, Cary, North Carolina). Results are expressed as mean ±
standard error of the mean (S.E.M.), N is the sample size. Values of
P<0.05 were taken to indicate significant differences between
means.
![]() |
Results |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
|
Effect of high-calcium sea water on 14C incorporation into
tissues
Incubation of polyps in high-calcium sea water (FSW+100) at 23 °C
resulted in a significant increase in the rate of 14C incorporation
into the tissues compared with polyps incubated in standard sea water (FSW)
(Fig. 3). At 29 °C there
was also a significant increase in 14C incorporation into the
tissues in polyps incubated in FSW+200 compared with polyps incubated in FSW
and ASW (Fig. 3). The rate of
14C incorporation in standard sea water was higher at 29 °C
than at 23 °C.
|
Effect of high-calcium sea water on photosynthetic and respiration
rates
Rates of net photosynthesis (4.2±0.39 µg O2
mg-1 protein min-1) and respiration (-1.9±0.09
µg O2 mg-1 protein min-1) (means ±
S.E.M., N=8) were unaffected (P>0.05) by incubation in
high-calcium sea water (FSW+100) when compared to rates obtained during
incubation in standard sea water (FSW) (4.9±0.4 and -2.1±0.12
µg O2 mg-1 protein min-1 for
photosynthesis and respiration, respectively). It is noticeable that rates in
high-calcium sea water tended to be slightly lower than in standard sea water.
However, pilot experiments, in which polyps were incubated in standard sea
water in place of high-calcium sea water, showed similar variation.
Effect of HEBP on calcium uptake by the skeleton
When the effect of HEBP on calcium incorporation into the skeleton was
measured, using the standard protocol of digesting the tissues prior to
dissolving the skeleton, the rate of incorporation was dramatically reduced
compared to the control (Fig.
4). However, when the processing protocol was changed so that the
skeleton was dissolved prior to digesting the tissues then the reduction in
calcium incorporation was substantially less than with the former protocol
(Fig. 4). The amount of calcium
incorporated was significantly higher when the skeleton was dissolved first
than when the tissues were digested first. We interpret this to indicate that
HEBP does not prevent the formation of amorphous calcium carbonate, which is
lost during the standard processing procedure, but is retained to a much
greater extent by dissolving the skeleton prior to digesting the tissue.
|
Effect of HEBP on photosynthetic and respiration rates
Recordings of oxygen concentration were made from four sets of three polyps
in the light, and a further four sets in the dark, before and after exposure
to 0.5 mmol l-1 HEBP. The gross photosynthetic rate after HEBP
treatment increased by 26±6 % and the respiration rate decreased by
42.5±5.5 %, thus the net photosynthetic rate was reduced by 16.5 %
(data not shown).
![]() |
Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Increasing calcium concentration in sea water by 2.5 mmol l-1
resulted in a marked increase in calcium incorporation into the skeleton of
G. fascicularis. The increase at 23 °C was 61 % and 30 % when
normalised to skeletal mass or to tissue protein content, respectively. The
method of normalising physiological data in corals is a contentious one
(Marshall, 1996). However,
regardless of the method used there was undoubtedly a statistically
significant increase in calcification rate. At 29 °C there was also a
significant increase in calcium incorporation when polyps were incubated in
sea water in which the calcium concentration had been increased by 5 mmol
l-1, regardless of the method of normalisation. The increase,
compared to polyps incubated in standard sea water, was 84 % and 54 % when
normalised to skeletal mass or tissue protein content, respectively.
Calcification rates in standard sea water are very similar at these two
temperatures, which represent the extremes of a Gaussian distribution of
calcification rate versus temperature (A. T. Marshall and P. L.
Clode, manuscript submitted for publication).
In previous studies, as artificial sea water calcium concentration
increased, calcification rate tended to plateau at about 10 mmol
l-1 (Chalker, 1976;
Ip and Krishnaveni, 1991
;
Tambutte et al., 1996
).
Although a continuing increase was recorded
(Chalker, 1976
) in Acropora
cervicornis and in G. fascicularis
(Krishnaveni et al., 1989
),
the increase at a concentration comparable to that in the present
investigation (13.5 mmol l-1) was approximately 5 and 8 %,
respectively, compared with 61 % in the present investigation. At a calcium
concentration of 16 mmol l-1 the increase in calcification rate in
G. fascicularis in the present investigation was even higher, at 84 %
when normalised to skeletal mass and 54 % when normalised to protein content.
The investigations of Chalker
(1976
), Krishnaveni et al.
(1989
), Ip and Krishnaveni
(1991
) and Tambutte et al.
(1996
) were carried out using
artificial sea water under laboratory conditions, with relatively low light
intensities, whereas the present experiments were carried out using natural
sea water and in the open air where light conditions were much higher and
comparable with those on the reef flat. The difference in the experimental
conditions could be presumed to be causal factors in the observed differences
in calcification rate.
The calcium carbonate saturation state of sea water has been suggested to
influence the distribution of zooxanthellate corals by virtue of its effect on
calcium carbonate precipitation and hence the calcification rate of corals
(Buddemeier and Fautin, 1996;
Kleypas et al., 1999
).
Experimental evidence to support this was provided by Gattuso et al.
(1998
), who showed that
increasing the calcium carbonate saturation state, by adding calcium to
artificial sea water, increased the calcification rate in Stylophora
pistillata. The rate increased up to contemporary sea water saturation
levels, but showed no further increase at saturation values above this
threshold. These experiments were again conducted with artificial sea water
and Gattuso et al. (1998
) note
that the calcification rate in S. pistillata measured in standard sea
water was almost twice that measured in artificial sea water with the same
calcium carbonate saturation state. It should be noted that in the present
investigation, exposure of G. fascicularis polyps to the `standard'
artificial sea water formulation of Gattusso et al. (1998) in pilot
experiments had obviously deleterious effects. These effects were not evident
in polyps exposed to artificial sea water formulated according to
Benazet-Tambutte et al.
(1996
).
Inorganic calcification rates can be calculated from the aragonite
precipitation equation of Burton and Walter
(1990):
![]() | (1) |
![]() | (2) |
Consistent with our measurements showing substantial increases in
calcification rates with increases in the calcium concentration of standard
sea water are the measurements obtained by Swart
(1979,
1980
). Swart
(1980
) recorded high growth
rates in several species of corals when exposed to standard sea water with
elevated calcium concentrations and the growth rates peaked at the
concentrations used in the present investigation. Increases in growth rate
ranged from 70 % to 820 %. Swart
(1980
) also found that
response was variable, depending on the locality on the reef flat from which
specimens of a particular species had been collected. This phenomenon may also
have been operating in the present investigation, since the G.
fascicularis polyps used in experiments carried out at 23 °C came
from a locality where this species is prolific, whereas the polyps used at 29
°C were obtained (due to collecting permit restrictions) from a locality
on the reef flat where this species is uncommon. Yamazato
(1966
) also obtained a twofold
increase in calcium uptake in Fungia scutaria following the addition
of 200 mg l-1 of calcium to sea water.
The amount of fixed carbon in coral tissues is generally regarded as a
measure of photosynthetic activity of the zooxanthellae. The effect of
high-calcium sea water was to increase 14C incorporation, and
therefore photosynthesis, by 87 %, after incubation in sea water at 23 °C
with calcium concentration increased by 2.5 mmol l-1, and by 32 %
after incubation in sea water at 29 °C with calcium concentration
increased by 5 mmol l-1. It should be noted that the control rate
of 14C incorporation was markedly higher at 29 °C than at 23
°C. Paradoxically, incubation in high-calcium sea water had no effect on
net photosynthetic oxygen production or on respiratory oxygen consumption. A
full explanation of this is not possible without further investigation. Since
zooxanthellae possess the C-3 carbon-fixation pathway
(Streamer et al., 1993) it may
be conjectured that photorespiration, which is considered to be likely in
zooxanthellae (Black et al.,
1976
; Randall,
1976
), is a possible reason for an apparent lack of increase in
photosynthetic oxygen production. However, an increase in photorespiration
would generally lead to a decline in CO2 fixation
(Tolbert and Osmond,
1976
).
Yamashiro (1995) presented
evidence that appears to be counter to the trans-calcification model.
It was shown that the bisphosphonate (1-hydroxyethylidene-1, 1-biphosphonic
acid, HEBP) reduced calcification, measured by 14C incorporation
into the skeleton, and did not affect photosynthesis, measured by
14C incorporation into the tissues. Whilst HEBP is known to prevent
the growth of calcium phosphate crystals, it does not prevent the formation of
amorphous calcium phosphate (Francis et
al., 1969
). Therefore, it may not prevent the formation of
amorphous calcium carbonate. If this is so then the standard processing
technique used by Yamashiro would not retain the amorphous compound. In this
procedure the tissue is digested in NaOH prior to dissolving the skeleton.
Digestion in NaOH would remove amorphous calcium carbonate and the labeled
carbon would be lost as CO2 during subsequent acid treatment of the
digest. In our experiments, using 45Ca to measure calcium
incorporation, we attempted to prevent losses of amorphous calcium carbonate
by dissolving the skeleton prior to digesting the tissue. The polyps were
first well washed to remove as much 45Ca from the tissue and
coelenteron as possible (Tambutte et al.,
1995
,
1996
). The amount of
45Ca associated with tissue and coelenteric sea water relative to
the skeleton has been shown to be very small
(Marshall and Wright, 1998
).
There was no significant difference, in calcium incorporation into the
skeleton, between polyps in which tissue was removed first and polyps in which
skeleton was removed first when they were incubated in standard sea water.
However, when incubated in sea water containing HEBP, calcium incorporation
was reduced by 91 % in polyps with tissue removed first, compared with control
polyps but the reduction was only 54 % in polyps with skeleton removed first.
The difference between the two treatments was significant. These results
suggest that amorphous calcium carbonate may be formed during treatment with
HEBP and that the loss of amorphous calcium carbonate may have been undetected
in Yamashiro's (1995
)
experiments, thereby invalidating the assumption that a reduction in
calcification has no effect on photosynthetic rate.
The 54 % reduction in calcium incorporation seen after HEBP treatment in
polyps in which the skeleton was dissolved first may be attributable in part
to a direct effect of HEBP on the physiology of G. fascicularis
polyps. That such an effect occurred was revealed by the reduction in
respiration and net photosynthetic rates. It was also observed that polyps
exposed to HEBP then placed in fresh running sea water invariably bleached
within a few hours. Consistent with these observations are reports that
bisphosphonates may inhibit intracellular ATP-dependent enzymes or have
physiological effects on intracellular second messenger pathways
(Rogers et al., 1994;
Russell et al., 1999
). These
effects are distinct from the inhibition of mineralisation due to the binding
of bisphosphonates to mineral surfaces in organisms. Some bisphosphonates also
appear to have a herbicidal action
(Kafarski et al., 2000
).
We conclude that the evidence showing that calcification does not stimulate photosynthesis, based on the use of low-calcium artificial sea water and inhibitors of crystal formation such as bisphosphonates, should be interpreted with caution. Artificial sea water containing low concentrations of calcium caused a marked stress response (A. T. Marshall and P. L. Clode, manuscript submitted for publication) that inevitably must compromise experiments carried out in this medium. Standard sea water in which calcium concentration was increased resulted in higher rates of calcification and photosynthesis. The increase in calcification rate was greater than that previously observed in experiments conducted with artificial sea waters in which calcium concentration was increased above standard sea water concentration. Although it is not possible to say from these data that increased calcification promotes an increase in photosynthesis according to the trans-calcification model, since calcium may act to independently potentiate both processes, it is possible to say that the data are consistent with the model.
![]() |
Acknowledgments |
---|
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Al-Moghrabi, S., Goiran, C., Allemand, D., Speziale, N. and Jaubert, J. (1996). Inorganic carbon uptake for photosynthesis by the symbiotic coraldinoflagellate association. 2. Mechanisms for bicarbonate uptake. J. exp. mar. Biol. Ecol. 199,227 -248.
Allemand, D., Furla, P. and Benazet-Tambutte, S. (1998). Mechanisms of carbon acquisition for endosymbiont photosynthesis in Anthozoa. Can. J. Bot. 76,925 -941.
Barnes, D. J. (1985). The effect of photosynthetic and respiratory inhibitors upon calcification in the staghorn coral, Acropora formosa. In Proceedings of the Fifth International Coral Reef Congress, Tahiti. pp.161 -165.
Barnes, D. J. and Chalker, B. E. (1990). Calcification and photosynthesis in reef-building corals and algae. In Coral Reefs (ed. Z. Dubinsky), pp.109 -131. Amsterdam: Elsevier.
Benazet-Tambutte, S., Allemande, D. and Jaubert, J. (1996). Permeability of the oral epithelial layers in Cnidarians. Mar. Biol. 126,43 -53.
Black, C. C., Burris, J. E. and Everson, R. G. (1976). Influence of oxygen concentration on photosynthesis in marine plants. Aust. J. Plant Physiol. 3, 81-86.
Buddemeier, R. W. and Fautin, D. G. (1996). Saturation state and the evolution and biogeography of symbiotic calcification. Bull. Inst. Oceanogr., Monaco 14, 23-32.
Burton, E. A. and Walter, L. M. (1990). The role of pH in phosphate inhibition of calcite and aragonite precipitation rates in sea water. Geochim. Cosmochim. Acta 54,797 -808.
Chalker, B. E. (1976). Calcium transport during skeletogenesis in hermatypic corals. Comp. Biochem. Physiol. 54A,455 -459.
Francis, M. D., Graham, R., Russell, G. and Fleisch, H. (1969). Diphosphonates inhibit formation of calcium phosphate crystals in vitro and pathological calcification in vivo. Science 165,1264 -1266.[Medline]
Gattuso, J.-P., Allemand, D. and Frankignoulle, M. (1999). Photosynthesis and calcification at cellular, organismal and community levels in coral reefs: a review on interactions and control by carbonate chemistry. Amer. Zool. 39,160 -183.
Gattuso, J.-P., Frankignoulle, M., Bourge, I., Romaine, S. and Buddemeier, R. W. (1998). Effect of calcium carbonate saturation of sea water on coral calcification. Global and Planetary Change 18,37 -46.
Gattuso, J.-P., Reynaud-Vaganay, S., Furla, P., Romaine-Lioud, S., Jaubert, J., Bourge, I. and Frankignoulle, M. (2000). Calcification does not stimulate photosynthesis in the zooxanthellate scleractinian coral Stylophora pistillata. Limnol. Oceanogr. 45,246 -250.
Goreau, T. F. (1959). The physiology of skeleton formation in corals. I. A method for measuring the rate of calcium deposition by corals under different conditions. Biol. Bull. 116,59 -75.
Ip, Y. K. and Krishnaveni, P. (1991). Incorporation of strontium (90Sr2+) into the skeleton of the hermatypic coral Galaxea fascicularis. J. Exp. Zool. 258,273 -276.
Kafarski, P., Lejczak, B. and Forlani, G. (2000). Herbicidally active aminomethylenebisphosphonic acids. Heteroatom Chemistry 11,449 -453.
Kawaguti, S. and Sakumoto, D. (1948). The effect of light on the calcium deposition of corals. Bull. Oceanogr. Inst. Taiwan 4,65 -70.
Kleypas, J. A., Buddemeier, R. W., Archer, D., Gattuso, J.-P.,
Langdon, C. and Opdyke, B. N. (1999). Geochemical
consequences of increased atmospheric carbon dioxide on coral reefs.
Science 284,118
-120.
Krishnaveni, P., Chou, L. M. and Ip, Y. K. (1989). Deposition of calcium (45Ca2+). in the coral Galaxea fascicularis. Comp. Biochem. Physiol. 94A,509 -513.
Marshall, A. T. (1996). Calcification rates in corals. Science 274,117 -118.
Marshall, A. T. and Wright, A. (1998). Coral calcification: autoradiography of a scleractinian coral Galaxea fascicularis after incubation in 45Ca and 14C. Coral Reefs 17,37 -47.
McConnaughey, T. (1989). Biomineralization mechanisms. In Origin, Evolution, and Modern Aspects of Biomineralization in Plants and Animals (ed. R. E. Crick), pp.57 -73. New York: Plenum Press.
McConnaughey, T. A. (1994). Calcification, photosynthesis, and global carbon cycles. In Past and Present Biomineralization Processes. Considerations about the Carbonate Cycle (ed. F. Doumenge, D. Allemand and A. Toulemont), pp.137 -161. Monaco: Monaco Musee Oceanographique.
McConnaughey, T. A. and Whelan, J. F. (1997). Calcification generates protons for nutrient and bicarbonate uptake. Earth Sci. Rev. 42,95 -117.
Millero, F. J. (1996). Chemical Oceanography. Boca Raton, CRC Press.
Randall, D. D. (1976). Phosphoglycollate phosphatase in marine algae: isolation and characterization from Halimeda cylindracea. Aust. J. Plant Physiol. 3, 105-111.
Rogers, M. J., Watts, D. J., Russell, R. G. G., Ji, X. H., Xiong, X. J., Blackburn, G. M., Bayless, A. V. and Ebetino, F. H. (1994). Inhibitory effects of bps on growth of amoebae of the cellular slime mold Dictyostelium discoideum. J. Bone Min. Res. 9,1029 -1039.[Medline]
Russell, R. G., Rogers, M. J., Frith, J. C., Luckman, S. P., Coxon, F. P., Benford, H. L., Croucher, P. I., Shipman, C. and Fleisch, H. A. (1999). The pharmacology of bisphosphonates and new insights into their mechanisms of action. J. Bone Min. Res. 14,53 -65.[Medline]
Streamer, M., McNeil, Y. R. and Yellowlees, D. (1993). Photosynthetic carbon dioxide fixation in zooxanthellae. Mar. Biol. 115,195 -198.
Swart, P. (1980). The effect of sea water chemistry on the growth of some scleractinian corals. In Developmental and Cellular Biology of Coelenterates (ed. P. Tardent and R. Tardent), pp. 203-208. Amsterdam: Elsevier/North Holland Biomedical Press.
Swart, P. K. (1979). The effect of sea water calcium concentrations on the growth and skeletal composition of a scleractinian coral: Acropora squamosa. J. Sed. Petrol. 49,951 -954.
Tambutte, E., Allemand, D., Bourge, I. and Gattuso, J.-P. (1995). An improved 45Ca protocol for investigating physiological mechanisms in coral calcification. Mar. Biol. 122,453 -459.
Tambutte, E., Allemand, D., Mueller, E. and Jaubert, J.
(1996). A compartmental approach to the mechanism of
calcification in hermatypic corals. J. Exp. Biol.
199,1029
-1041.
Tolbert, A. and Osmond, C. B. (1976). The Great Barrier Reef Photorespiration Expedition: introduction. Aust. J. Plant Physiol. 3,1 -8.
Vandermeulen, J. H., Davis, N. D. and Muscatine, L. (1972). The effects of inhibitors of photosynthesis on zooxanthellae in corals and other marine invertebrates. Mar. Biol. 16,185 -191.
Yamashiro, H. (1995). The effects of HEBP, an inhibitor of mineral deposition, upon photosynthesis and calcification in the scleractinian coral, Stylophora pistillata. J. exp. mar. Biol. Ecol. 191,57 -63.
Yamazato, K. (1966). Calcification in a solitary coral, Fungia scutaria Lamarck in relation to environmental factors. PhD Thesis, University of Hawaii, Hawaii.130 pp.