Abstract
A sensitive experimental protocol using cloned corals (hereafter “microcolonies”) of the branching scleractinian coral Stylophora pistillata and 45Ca has been developed to enable reproducible measurements of physiological and biochemical mechanisms involved in calcium transport and compartmentalization during coral calcification. Cloned S. pistillata microcolonies were propagated in the laboratory from small fragments of parent colonies collected in 1990 in the Gulf of Aqaba, Jordan. Cloned microcolonies have several intrinsic properties that help to reduce unwanted biological variability: (1) same genotype; (2) similar sizes and shapes; and (3) absence of macroscopic boring organisms. Errors specifically associated with long-standing problems to do with isotopic exchange were further reduced by producing microcolonies with no skeletal surfaces exposed to the radioisotope-labelled incubation medium. The value of the technique resides principally in its superior ability to elucidate transportation pathways and processes and not in its ability to quantitatively estimate calcium deposition by corals in nature. We describe here a rapidly exchangeable calcium pool in which up to 90% of the radioactive label taken up during incubations is located. This pool (72.9±1.4 nmol Ca mg-1 protein) is presumably located within the coelenteric cavity as suggested by the following: (1) it has 4-min half-time saturation kinetics; (2) the accumulation of calcium is linearly correlated with the calcium concentration of sea-water; and (3) its insensitivity to metabolic and ion transport inhibitors indicate that membranes do not isolate this compartment. Washout of this large extracellular pool greatly improved estimates of calcium deposition as evidenced by 10 to 40% reduction in coefficients of variation when compared with previous 45Ca2+ methods described in the literature. Comparisons of calcification measurements simultaneously carried out using the alkalinity anomaly technique and the 45Ca protocol described here show that the correlation coefficient of both techniques is close to 1. Unlike previous reports, our 45Ca2+-derived measurements are slightly lower than those computed from the alkalinity depletion technique.
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References
Allemand D, De Renzis G, Ciapa B, Girard JP, Payan P (1984) Characterization of valine transport in sea urchin eggs. Biochim Biophys Acta 772:337–346
Allemand D, Grillo MC (1992) Biocalcification mechanism in gorgonians: 45Ca uptake and deposition by the Mediterranean red coral Corallium rubrum. J exp Zool 262:237–246
Al-Moghrabi S, Allemand D, Jaubert J (1993) Valine uptake by the scleractinian coral Galaxea fascicularis: characterization and effect of light and nutritional status. J Comp Physiol 163:355–362
Barnes DJ, Chalker BE (1990) Calcification and photosynthesis in reef-building corals and algae. In: Dubinsky Z (ed) Coral reefs, ecosystems of the world. Elsevier, Amsterdam, pp 109–131
Barnes DJ, Crossland CJ (1977) Coral calcification: sources of error in radioisotope techniques. Mar Biol 42:119–129
Barnes DJ, Crossland CJ (1982) Variability in the calcification rate of Acropora acuminata measured with radioisotopes. Coral Reefs 1:53–57
Böhm L (1978) Application of the 45Ca tracer method for determination of calcification rates in calcareous algae: effect of calcium exchange and differential saturation of algal calcium pools. Mar Biol 47:9–14
Borle AB (1969) Kinetic analysis of calcium movements in HeLa cell cultures. I. Calcium influx. J gen Physiol 53:43–56
Borowitzka MA, Larkum AWD (1976) Calcification in the green alga Halimeda. II. The exchange of Ca2+ and the occurrence of age gradients in calcification and photosynthesis. J exp Bot 27: 864–878
Buddemeier RW, Kinzie RA (1976) Coral growth. Oceanogr mar Biol A Rev 14:183–225
Chalker BE (1976) Calcium transport during skeletogenesis in hermatypic corals. Comp Biochem Physiol 54A:455–459
Chalker BE, Taylor DL (1975) Light-enhanced calcification, and the role of oxidative phosphorylation in calcification of the coral Acropora cervicornis. Proc R Soc Lond (Ser B) 190:323–331
Chisholm JRM, Gattuso J-P (1991) Validation of the alkalinity anomaly technique for investigating calcification and photosynthesis in coral reef communities. Limnol Oceanogr 36:1232–1239
Claret-Berthon B, Claret M, Mazet JL (1977) Fluxes and distribution of calcium in rat liver cells: kinetic analysis and identification of pools. J Physiol 272:529–552
Clausen CD (1971) Effects of temperature on the rate of 45Calcium uptake by Pocillopora damicornis. In: Lenhoff HM, Muscatine L, Davis LD (eds) Experimental coelenterate biology. Univ of Hawaii Press, Honolulu, pp 246–259
Clausen CD, Roth AA (1975) Estimation of coral growth-rates from laboratory 45Ca-incorporation rates. Mar Biol 33:85–91
Crossland CJ, Barnes DJ (1977) Calcification in the staghorn coral Acropora acuminata: variations in apparent skeletal incorporation of radioisotopes due to different methods of processing. Mar Biol 43:57–62
Ehrenfeld J, Cousin J-L (1982) Ionic regulation of the unicellular green alga Dunaliella tertiolecta. J Membrane Biol 70:47–57
Gibbons IR (1981) Cilia and flagella of eukaryotes. J Cell Biol 91(3):107–124
Gladfelter EH (1983) Circulation of fluids in the gastrovascular system of the reef coral Acropora cervicornis. Biol Bull mar biol Lab, Woods Hole 165:619–636
Hansson I, Jagner D (1973) Evaluation of the accuracy of Gran plots by means of computer calculations. Application to the potentiometric titration of the total alkalinity and carbonate content of seawater. Analytica chim Acta 65:363–373
Hidaka M, Yamazoto K (1982) Effect of light on budding of isolated polyps of the scleractinian coral Galaxea fascicularis. Galaxea 1:65–75
Ip YK, Krishnaveni P (1991) Incorporation of strontium (90Sr++) into the skeleton of hermatypic coral Galaxea fascicularis. J exp Zool 258:273–276
Jacques TG, Pilson MEQ (1980) Experimental ecology of the temperature scleractinian coral Astrangia danae. I. Partition of respiration, photosynthesis and calcification between host and symbionts. Mar Biol 60:167–178
Kingsley RJ, Watabe N (1984) Calcium uptake in the gorgonian Leptogorgia virgulata. The effects of ATPase inhibitors. Comp Biochem Physiol 79A:487–491
Krishnaveni P, Chou LM, Ip YK (1989) Deposition of calcium (45Ca2+) in the coral, Galaxea fascicularis. Comp Biochem Physiol 94A:509–513
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measuement with the folin phenol reagent. J biol Chem 193:265–275
Milne JL, Coukell MB (1991) A Ca2+ transport system associated with the plasma membrane of Dictyostelium discoideum is activated by different chemoattractant receptors. J Cell Biol 112:103–110
Rinkevich B, Loya Y (1984) Does light enhance calcification in hermatypic corals? Mar Biol 80:1–6
Smith SV, Kinsey DW (1978) Calcification and organic carbon metabolism as indicated by carbon dioxide. In: Stoddart DR, Johannes RE (eds) Coral reefs: research methods. Unesco, Paris, pp 469–484
Tambutté E, Allemand D, Jaubert J (1995) The Stylophora pistillata coral microcolony: a model for studying transport process during coral biomineralization. Proc 7th int Biomin Symp Bull Inst Oceanogr no spécial 14 (in press)
Velimerov B, King J (1979) Calcium uptake and net calcification rates in the octocoral Eunicella papillosa. Mar Biol 50:349–358
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Tambutté, E., Allemand, D., Bourge, I. et al. An improved 45Ca protocol for investigating physiological mechanisms in coral calcification. Marine Biology 122, 453–459 (1995). https://doi.org/10.1007/BF00350879
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DOI: https://doi.org/10.1007/BF00350879