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A large percentage of CO.sub.2 emitted into the atmosphere is absorbed by the oceans, causing chemical changes in surface waters known as ocean acidification (OA). Despite the high interest and increased pace of OA research to understand the effects of OA on marine organisms, many ecologically important organisms remain unstudied. Calcidiscus is a heavily calcified coccolithophore genus that is widespread and genetically and morphologically diverse. It contributes substantially to global calcium carbonate production, organic carbon production, oceanic carbon burial, and ocean-atmosphere CO.sub.2 exchange. Despite the importance of this genus, relatively little work has examined its responses to OA. We examined changes in growth, morphology, and carbon allocation in multiple strains of Calcidiscus leptoporus in response to ocean acidification. We also, for the first time, examined the OA response of Calcidiscusquadriperforatus, a larger and more heavily calcified Calcidiscus congener. All Calcidiscus coccolithophores responded negatively to OA with impaired coccolith morphology and a decreased ratio of particulate inorganic to organic carbon (PIC:POC). However, strains responded variably; C. quadriperforatus showed the most sensitivity, while the most lightly calcified strain of C. leptoporus showed little response to OA. Our findings suggest that calcium carbonate production relative to organic carbon production by Calcidiscus coccolithophores may decrease in future oceans and that Calcidiscus distributions may shift if more resilient strains and species become dominant in assemblages. This study demonstrates that variable responses to OA may be strain or species specific in a way that is closely linked to physiological traits, such as cellular calcite quota. Electronic supplementary material The online version of this article (doi:10.1007/s00227-015-2669-x) contains supplementary material, which is available to authorized users., Author(s): Rachel E. Diner[sup.1] , Ina Benner[sup.1] , Uta Passow[sup.2] , Tomoko Komada[sup.1] , Edward J. Carpenter[sup.1] , Jonathon H. Stillman[sup.1] [sup.3] [sup.4] Author Affiliations: (1) Romberg Tiburon Center, San [...]

The impact of elevated C[O.sub.2] and temperature on photosynthesis and calcification in the symbiont-bearing benthic foraminifer Marginopora vertebralis was studied. Individual specimens of M. vertebralis were collected from Heron Island on the southern Great Barrier Reef (Australia). They were maintained for 5 weeks at different temperatures (28, 32°C) and pC[O.sub.2] (400, 1,000 µatm) levels spanning a range of current and future climate-change scenarios. The photosynthetic capacity of M. vertebralis was measured with [O.sub.2] microsensors and a pulse-amplitudemodulated chlorophyll (Chl) fluorometer, in combination with estimates of Chl a and Chl [c.sub.2] concentrations and calcification rates. After 5 weeks, control specimens remained unaltered for all parameters. Chlorophyll a concentrations significantly decreased in the specimens at 1,000 µatm C[O.sub.2] for both temperatures, while no change in Chl [c.sub.2] concentration was observed. Photoinhibition was observed under elevated C[O.sub.2] and temperature, with a 70-80 % decrease in the maximum quantum yield of PSII. There was no net [O.sub.2] production at elevated temperatures in both C[O.sub.2] treatments as compared to the control temperature, supporting that temperature has more impact on photosynthesis and [O.sub.2] flux than changes in ambient C[O.sub.2]. Photosynthetic pigment loss and a decrease in photochemical efficiency are thus likely to occur with increased temperature. The elevated C[O.sub.2] and high temperature treatment also lead to a reduction in calcification rate (from +0.1 to >-0.1 % [day.sup.-1]). Thus, both calcification and photosynthesis of the major sediment-producing foraminifer M. vertebralis appears highly vulnerable to elevated temperature and ocean acidification scenarios predicted in climate-change models., Introduction Climate change is expected to have significant impacts on near-shore marine ecosystems and their resident organisms (Johnson and Marshall 2007). Human activities, such as burning of fossil fuels, industrialization, [...]

Bryozoans are colonial benthic marine invertebrate calcifiers, important and especially abundant and diverse in southern hemisphere shelf environments. Large heavily calcified colonies can be up to 50 years old, but most longer-lived bryozoans are limited to 10-20 y. Many smaller species are annual. Radial extension in flat encrusting bryozoans is generally on the order of 1-5 mm/y. Erect calcified species generally grow vertically 2-15 mm/y, though articulated species such as Cenaria may reach rates of 40 mm/y. Corresponding calcification rates are generally [10.sup.1]-[10.sup.2] mg/y, but there can be an order of magnitude variation in rate among years in high-latitude bryozoans. Multi-branched bryozoans produce up to 24 g of CaC[O.sub.3]/y. The carbonate produced by bryozoans varies from calcite to aragonite and mixtures of both. Skeletal carbonate mineralogy of bryozoans is complex and appears to be strongly genetically controlled. Global climate change, leading to increasing water temperatures, will generally increase marine bryozoan metabolic rates, and may increase Mg in calcite. On the other hand, decreasing pH (ocean acidification) causes corrosion, changes in mineralogy, and decreased survival. This review of bryozoan growth and calcification allows a general perspective, but also reveals gaps in our knowledge which need to be addressed., Introduction Bryozoans have been described as a minor phylum, and it is true that the combination of being inedible, small, difficult to identify, and currently found mainly in cryptic habitats [...]

Ocean acidification caused by rising atmospheric C[O.sub.2] is predicted to negatively impact growth and calcification rates of coralline algae. Decreases in coralline abundance may have cascading effects on marine ecosystems and on carbon sequestration worldwide. In this study, we measured growth and calcification rates of three common species of articulated coralline algae (Bossiella plu-mosa. Calliarthron tuberculosum, and Corallina vancouv-eriensis) at an intertidal field site in British Columbia. Linear growth rates measured in the field were slow, al-though Bossiella grew significantly faster (0.22 cm [mon.sup.-1]) than Calliarthron and Corallina (0.17 and 0.15 cm [mon.sup.-1], respectively). Growth rates in the field were generally slower than growth rates in the laboratory. Suggesting that data generated in the laboratory may not be representative of natural field conditions. Growth rates did not decrease as fronds approached their maximum observed size, suggest-ing that maximum frond size might be determined not by intrinsic factors but by external factors such as wave-induced drag forces. Using growth data, we estimate that the largest observed Bossiella frond (20 cm2) and Calliarthron frond (40 [cm.sup.2]) were about 4 and 11 years old, respectively, and had deposited approximately 1 and 6 g CaCO3 in that time. Given the great abundance of coralline algae along the coast of British Columbia, deposition rates of Ca[CO.sub.3] are expected to play a significant but poorly characterized role in carbon sequestration., Introduction Coralline algae (Corallinales and Sporolithales, Rhodo-phyta) are important components of marine communities worldwide, cementing coral reefs (Steneck and Adey, 1976) and quickly colonizing bare rock to provide habitat for [...]

The vast majority of ectothermic organisms grow larger when developing at cooler environmental temperatures, a pattern frequently referred to as the temperature-size rule (TSR). Assuming that this reaction norm has adaptive significance, life history theory predicts that converse patterns may evolve if favored by natural selection, namely if the costs associated with complying to the TSR outweigh the benefits. Calcifying ectotherms may comprise such an exception not following the TSR, because calcification is expected to be more costly at lower temperatures thus increasing associated costs. To test this hypothesis, we reared wild-caught juveniles of the intertidal gastropod Monetaria annulus and compared their shell sizes at the end of the juvenile stage between two rearing temperatures. Contrary to our prediction, M. annulus does follow the TSR, suggesting that increased calcification costs at lower temperatures are not high enough to break the TSR. Such plastic responses should be considered when interpreting geographical patterns such as latitudinal size clines, which may be caused at least partly by phenotypic plasticity., Introduction Most ectotherms mature at a larger size at lower developmental temperatures, a pattern of wide taxonomic prevalence which has been coined the temperature-size rule (TSR; Atkinson 1994). The TSR [...]

Using C[O.sub.2] perturbation experiments, we examined the pre- and post-settlement growth responses of a dominant biofouling tubeworm (Hydroides elegans)to a range of pH. In three different experiments, embryos were reared to, or past, metamorphosis in seawater equilibrated to C[O.sub.2] values of about 480 (control), 980, 1,480, and 2,300 µatm resulting in pH values of around 8.1 (control), 7.9, 7.7, and 7.5, respectively. These three decreased pH conditions did not affect either embryo or larval development, but both larval calcification at the time of metamorphosis and early juvenile growth were adversely affected. During the 24-h settlement assay experiment, half of the metamorphosed larvae were unable to calcify tubes at pH 7.9 while almost no tubes were calcified at pH 7.7. Decreased ability to calcify at decreased pH may indicate that these calcifying tubeworms may be one of the highly threatened species in the future ocean., Introduction Carbon dioxide (C[O.sub.2]) produced by anthropogenic activity is absorbed by the ocean, reducing the seawater pH and carbonate chemistry that can adversely affect marine organisms, especially calcifiers, many of [...]

I tested the hypothesis that high pC[O.sub.2] (76.6 Pa and 87.2 Pa vs. 42.9 Pa) has no effect on the metabolism of juvenile massive Porites spp. after 11 days at 28 °C and 545 µmol quanta [m.sup.-2] [s.sup.-1]. The response was assessed as aerobic dark respiration, skeletal weight (i.e., calcification), biomass, and chlorophyll fluorescence. Corals were collected from the shallow (3-4 m) back reef of Moorea, French Polynesia (17°28.614'S, 149°48.917'W), and experiments conducted during April and May 2011. An increase in pC[O.sub.2] to 76.6 Pa had no effect on any dependent variable, but 87.2 Pa pC[O.sub.2] reduced area-normalized (but not biomass-normalized) respiration 36 %, as well as maximum photochemical efficiency ([F.sub.v]/[F.sub.m]) of open RCIIs and effective photochemical efficiency of RCIIs in actinic light (ΔF/[F'.sub.m]); neither biomass, calcification, nor the energy expenditure coincident with calcification (J [g.sup.-1]) was effected. These results do not support the hypothesis that high pC[O.sub.2] reduces coral calcification through increased metabolic costs and, instead, suggest that high pC[O.sub.2] causes metabolic depression and photochemical impairment similar to that associated with bleaching. Evidence of a pC[O.sub.2] threshold between 76.6 and 87.2 Pa for inhibitory effects on respiration and photochemistry deserves further attention as it might signal the presence of unpredictable effects of rising pC[O.sub.2]., Introduction Amidst efforts to evaluate the effects of ocean acidification (OA) on the calcification of scleractinian corals (Erez et al. 2011), much of which has revealed negative implications (Doney et [...]

RhoA/Rho kinases (ROCK) play a critical role in vascular smooth muscle cell (VSMC) actin cytoskeleton organization, differentiation, and function and are implicated in the pathogenesis of cardiovascular disease. We have previously determined that an important step in the regulation of calcification is fetuin-A endocytosis, a process that is dependent on changes in the cytoskeleton, which, in turn, is known to be affected by the RhoA/ ROCK signaling pathway. In the present study, bovine VSMC (BVSMC) were treated with the ROCK inhibitor Y-27632 or transfected with ROCK small interfering (si) RNA to knock down ROCK expression. Both conditions resulted in reduced actin stress fibers and increased Cy5-labeled fetuin-A uptake. Inhibition of ROCK by Y-27632 or siRNA also significantly increased BVSMC alkaline phosphatase (ALP) activity and calcification of BVSMC and rat aorta organ cultures. Cells were then incubated in calcification media in the presence or absence of Y-27632 and matrix vesicles (MV) isolated by collagenase digestion. These MV, isolated from BVSMC incubated with Y-27632, had increased ALP activity and increased ability of MV to subsequently calcify collagen by 66%. In contrast, activation of RhoA, which is upstream of ROCK, by transfecting plasmids encoding the dominant active Rho GTPase mutant (Rho-L63) led to decreased fetuin-A uptake and reduced calcification in BVSMC. These results demonstrate that the RhoA/ROCK signaling pathway is an important negative regulator of vascular calcification. vascular calcification; endocytosis; cytoskeleton; Rho GTPase doi: 10.1152/ajprenal.00730.2009.

Mammalian urine contains a range of macromolecule proteins that play critical roles in renal stone formation, among which Tamm-Horsfall protein (THP) is by far the most abundant. While THP is a potent inhibitor of crystal aggregation in vitro and its ablation in vivo predisposes one of the two existing mouse models to spontaneous intrarenal calcium crystallization, key controversies remain regarding the role of THP in nephrolithiasis. By carrying out a long-range follow-up of more than 250 THP-null mice and their wild-type controls, we demonstrate here that renal calcification is a highly consistent phenotype of the THP-null mice that is age and partially gene dosage dependent, but is gender and genetic background independent. Renal calcification in THP-null mice is progressive, and by 15 mo over 85% of all the THP-null mice develop spontaneous intrarenal crystals. The crystals consist primarily of calcium phosphate in the form of hydroxyapatite, are located more frequently in the interstitial space of the renal papillae than intratubularly, particularly in older animals, and lack accompanying inflammatory cell infiltration. The interstitial deposits of hydroxyapatite observed in THP-null mice bear strong resemblances to the renal crystals found in human kidneys bearing idiopathic calcium oxalate stones. Compared with 24-h urine from the wild-type mice, that of THP-null mice is supersaturated with brushite (calcium phosphate), a stone precursor, and has reduced urinary excretion of citrate, a stone inhibitor. While less frequent than renal calcinosis, renal pelvic and ureteral stones and hydronephrosis occur in the aged THP-null mice. These results provide direct in vivo evidence indicating that normal THP plays an important role in defending the urinary system against calcification and suggest that reduced expression and/or decreased function of THP could contribute to nephrolithiasis. knockout mice; nephrolithiasis; interstitial calcification; supersaturation; urolithiasis doi: 10.1152/ajprenal.00243.2010.

Vascular calcification is a multifaceted process involving gain of calcification inducers and loss of calcification inhibitors. One such inhibitor is inorganic pyrophosphate ([PP.sub.i]), and regulated generation and homeostasis of extracellular [PP.sub.i] is a critical determinant of soft-tissue mineralization. We recently described an autocrine mechanism of extracellular [PP.sub.i] generation in cultured rat aortic vascular smooth muscle cells (VSMC) that involves both ATP release coupled to the ectophosphodiesterase/pyrophosphatase ENPP1 and efflux of intracellular [PP.sub.i] mediated or regulated by the plasma membrane protein ANK. We now report that increased cAMP signaling and elevated extracellular inorganic phosphate ([P.sub.i]) act synergistically to induce calcification of these VSMC that is correlated with progressive reduction in ability to accumulate extracellular [PP.sub.i]. Attenuated [PP.sub.i] accumulation was mediated in part by cAMP-dependent decrease in ANK expression coordinated with cAMP-dependent increase in expression of TNAP, the tissue nonselective alkaline phosphatase that degrades [PP.sub.i]. Stimulation of cAMP signaling did not alter ATP release or ENPP1 expression, and the cAMP-induced changes in ANK and TNAP expression were not sufficient to induce calcification. Elevated extracellular [P.sub.i] alone elicited only minor calcification and no significant changes in ANK, TNAP, or ENPP1. In contrast, combined with a cAMP stimulus, elevated [P.sub.i] induced decreases in the ATP release pathway(s) that supports ENPP1 activity: this resulted in markedly reduced rates of [PP.sub.i] accumulation that facilitated robust calcification. Calcified VSMC were characterized by maintained expression of multiple SMC differentiation marker proteins including smooth muscle (SM) [alpha]-actin, SM22[alpha], and calponin. Notably, addition of exogenous ATP (or [PP.sub.i] per se) rescued cAMP + phosphate-treated VSMC cultures from progression to the calcified state. These observations support a model in which extracellular [PP.sub.i] generation mediated by both ANK- and ATP release-dependent mechanisms serves as a critical regulator of VSMC calcification. ectonucleotide pyrophosphatase/phosphodiesterase-1; ANK; adenosine 3',5'-cyclic monophosphate; ATP release doi:10.1152/ajpcell.00419.2009.

Am J Physiol Heart Circ Physiol 298: H5-H15, 2010. First Published October 23, 2009; doi: 10:1152/ajpheart.00824.2009--Recent epidemiologic studies have revealed the risk factors associated for vascular atherosclerosis, including the male sex, smoking, hypertension, and elevated serum cholesterol, similar to the risk factors associated with the development of AV stenosis. An increasing number of models of experimental hypercholesterolemia demonstrate features of atherosclerosis in the AV, which are similar to the early stages of vascular atherosclerotic lesions. Experimental and clinical studies demonstrate that the hypercholesterolemic AV develops an atherosclerotic lesion which is proliferative and expresses high levels of osteoblast bone markers which mineralize over time to form bone. Calcification, the end-stage process of the disease, is necessary to understand as a prognostic indicator in the modification of this cellular process before it is too late. In summary, these findings suggest that medical therapies may have a potential role in patients in the early stages of this disease process to slow the progression to severe aortic stenosis and to delay the timing of the need for surgery. The translation of these experimental studies to clinical practice will be important to understand the potential for medical therapy for this disease process. valvular heart disease; lipids; pathophysiology atherosclerosis; experimental models

Medial artery calcification, which does not accompany lipid or cholesterol deposit, preferentially occurs in elderly population, but its underlying mechanisms remain unclear. In the present study, we investigated the potential role of senescent vascular smooth muscle cells (VSMCs) in the formation of senescence-associated medial calcification. Replicative senescence was induced by the extended passages (until passages 11-13) in human primary VSMCs, and cells in early passage (passage 6) were used as control young cells. VSMC calcification was markedly enhanced in the senescent cells compared with that in the control young cells. We identified that genes highly expressed in osteoblasts, such as alkaline phosphatase (ALP) and type I collagen, were significantly upregulated in the senescent VSMCs, suggesting their osteoblastic transition during the senescence. Knock-down of either ALP or type I collagen significantly reduced the calcification in the senescent VSMCs. Of note, runt-related transcription factor-2 (RUNX-2), a core transcriptional factor that initiates the osteoblastic differentiation, was also upregulated in the senescent VSMCs. Knockdown of RUNX-2 significantly reduced the ALP expression and calcification in the senescent VSMCs, suggesting that RUNX-2 is involved in the senescence-mediated osteoblastic transition. Furthermore, immunohistochemistry of aorta from the [klotho.sup.-/-] aging mouse model demonstrated in vivo emergence of osteoblast-like cells expressing RUNX-2 exclusively in the calcified media. We also found that statin and Rho-kinase inhibitor effectively reduced the VSMC calcification by inhibiting [P.sub.i]-induced apoptosis and potentially enhancing matrix Gla protein expression in the senescent VSMCs. These findings strongly suggest an important role of senescent VSMCs in the pathophysiology of senescence-associated medial calcification, and the inhibition of osteoblastic transition could be a new therapeutic approach for the prevention of senescence-associated medial calcification. runt-related transcription factor-2; statin doi: 10.1152/ajpheart.00455.2009.

The role of atherosclerotic calcification in plaque rupture remains controversial. In previous analyses using finite element model analysis, circumferential stress was reduced by the inclusion of a calcium deposit in a representative human anatomical configuration. However, a recent report, also using finite element analysis, suggests that microscopic calcium deposits increase plaque stress. We used mathematical models to predict the effects of rigid and liquid inclusions (modeling a calcium deposit and a lipid necrotic core, respectively) in a distensible material (artery wall) on mechanical failure under uniaxial and biaxial loading in a range of configurations. Without inclusions, stress levels were low and uniform. In the analytical model, peak stresses were elevated at the edges of a rigid inclusion. In the finite element model, peak stresses were elevated at the edges of both inclusions, with minimal sensitivity to the wall distensibility and the size and shape of the inclusion. Presence of both a rigid and a soft inclusion enlarged the region of increased wall stress compared with either alone. In some configurations, the rigid inclusion reduced peak stress at the edge of the soft inclusion but simultaneously increased peak stress at the edge of the rigid inclusion and increased the size of the region affected. These findings suggest that the presence of a calcium deposit creates local increases in failure stress, and, depending on relative position to any neighboring lipid pools, it may increase peak stress and the plaque area at risk of mechanical failure. plaque rupture; vulnerable plaque; atherosclerosis; vascular calcification

The purpose of the present study was to test the hypothesis that extraskeletal calcification regresses in uremic rats after reduction in phosphorus intake and treatment with calcimimetics. Extraosseous calcification was induced in five to six nephrectomized rats fed a high-phosphorus (1.2%) diet who received calcitriol (80 ng/kg ip) every other day for a period of 14 days. Next, dietary phosphorus was reduced to 0.6%, and rats were treated with vehicle (n = 20), calcitriol [80 ng/kg ip/48 h (n = 20)], or the calcimimetic AMG 641 [1.5 mg/kg sc/48 h (n = 20)]. Aortic and soft-tissue calcium and phosphorus content was evaluated after 14 and 28 days. At 28 days, reduction of phosphorus intake resulted in a significant decrease in tissue mineral content in vehicle- and AMG 641-treated rats but not in rats receiving calcitriol. Aortic calcium and phosphorus was lower in rats treated with AMG 641 (96.7 [+ or -] 26.4 mg/g) than in rats receiving vehicle (178.3 [+ or -] 38.6 mg/g). An infiltrate of phagocytic cells expressing the calciumsensing receptor was identified in areas surrounding loci of calcification. Additional studies in parathyroidectomized rats demonstrated that AMG 641 increased the urinary excretion of calcium (6.2 [+ or -] 0.6 vs. 3.1 [+ or -] 0.5 mg/day, vehicle) (P < 0.001). In conclusion, experimentally induced extraosseous calcification in uremic rats can be partially resolved by reducing phosphorus intake; the addition of calcimimetics may accelerate the regression process through mechanisms potentially involving a direct stimulatory effect on mineral phagocytic cells plus an increase in urinary calcium excretion. calcitriol; hyperparathyroidism; vascular calcification

Much remains to be discovered about the etiology of heart valve disease and the molecular level mechanisms that drive it. The MAPK/ERK pathway influences calcification in many cell types and has been linked to the expression of a contractile phenotype in valvular interstitial cells (VICs). However, a direct correlation between MAPK/ERK pathway activity and VIC calcification has not been previously described. Thus the role of the MAPK pathway in the calcification of VIC cultures was investigated by measuring ERK activation in both calcifying and noncalcifying VIC environments and then, conversely, analyzing the effects of ERK pathway inhibition on VIC calcification and phenotype. Prolonged elevation of phosphorylated ERK-1/2 was found in calcifying VIC cultures, whereas directly blocking phosphorylation of ERK-1/2 resulted in a dramatic decrease in nodule number, nodule size, and total calcified area. Application of the ERK pathway inhibitor was also associated with a dramatic decrease in apoptosis, which may have contributed to the decreased nodule formation obtained via ERK inhibition. Real-time PCR analysis revealed that calcified samples exhibited significantly elevated expression of several myofibroblastic and osteoblastic markers, while ERK inhibition substantially reduced the expression of these markers, often to levels comparable to the noncalcifying control. These data suggest that the MAPK pathway plays an important role in regulating the phenotype and calcification of VICs, wherein sustained pathway activation is associated with increased VIC calcification. These findings may be used to further elucidate the mechanisms of valvular disease and identify potential treatment targets. heart valve; signaling pathways; extracellular matrix; mineralization

Parathyroid hormone (PTH), which is elevated in patients with chronic renal failure, has been shown to participate in the development of vascular calcification. Previous studies have demonstrated that PTH may promote endothelial expressions of proinflammatory parameters. On the basis of these data, we evaluated whether PTH may have an impact on endothelial osteoprotegerin (OPG), a vascular-protective factor which may control vascular calcification. Endothelial cells were stimulated with [10.sup.-12] to [10.sup.-10] mol/l PTH. PKC and PKA are the main cellular pathways of PTH. Inhibitors and activators of PKC or PKA were used to determine whether these signaling pathways are involved in the control of endothelial OPG. PTH induced a decrease in OPG secretion and mRNA expression. Treatment of PTH-stimulated cells by calphostin C (PKC inhibitor) induced a further decrease in OPG secretion, while Rp-cAMP (PKA inhibitor) had no additional effect. In nonstimulated cells, a PKC activator significantly stimulated OPG secretion, while a PKA activator was associated with a decline. These effects were blunted in the presence of calphostin C and Rp-cAMP, respectively. An increase in OPG secretion induced by a PKC activator indicates that the basal OPG secretion is mediated through PKC. The decrease induced by a PKA activator, which is similar to the decrease observed with PTH, suggests that the action of PTH on OPG secretion and mRNA expression may be due to the PKA pathway. endothelial cells

Several nonsense mutations observed in the FAM83H gene are identified and studied in two families with autosomal-dominant hypocalcified amelogenesis imperfecta (AI). The analysis proves that the FAM83H gene is extremely significant for the proper calcification of the dental-enamel.

Osteoprotegerin (OPG) plays a critical role in regulating postnatal bone mass. Experiments with OPG deficient mice also indicates a corelation between medial calcification of the aorta and renal arteries and the protein. This association between osteoporosis and arterial calcification opens possibilities for studying the impact of OPG regulation on these problems.

For the major coastal ecosystems, the main production, respiration, calcification, carbon burial and exchange with adjacent systems, the atmosphere included, have been reviewed. The coast ecosystems include macrophyte communities, estuaries, coral reefs, mangroves and the rest of the continental shelf. The coastal zone is not great in surface area but plays a large role in biogeochemical cycles. It receives massive inputs of terrestrial organic matter and nutrients and is among the most biologically and geochemically active areas of the biosphere. In coastal areas large amounts of energy and matter are exchanged with the open sea. The ecosystems studied, with the exception of estuaries, are net autotrophic.

Many tropical seaweeds and benthic invertebrates produce both secondary metabolites and calcium carbonate (CaC[O.sub.3]) particles or spicules that serve as possible defenses against consumers. To evaluate the relative defensive value of CaC[O.sub.3], secondary metabolites, and the potential synergistic or additive effects of the two, we made artificial agar-based 'seaweeds' in which we manipulated algal organic content, CaC[O.sub.3], and the secondary metabolites produced by the calcified green seaweeds Rhipocephalus phoenix, Udotea cyathiformis, and Halimeda goreauii, all of which are relatively resistant to herbivores. The effects of these manipulations on herbivore feeding were evaluated using three different types of herbivores, the sea urchin Diadema antillarum, the amphipod Cymadusa filosa, and a mixed-species group of small parrotfishes. Addition of finely powdered CaC[O.sub.3] as 69% of food dry mass had no effect on feeding by parrotfishes, deterred feeding by Cymadusa, and deterred Diadema when food organic content was low but not when it was higher. Although calcification of algal tissues has generally been considered a structural defense that hardens seaweed thalli and makes them more resistant to attack, the decreased feeding on CaC[O.sub.3]-containing foods in our assays occurred without any measurable alteration of food toughness. At natural concentrations, semipurified secondary metabolites from Rhipocephalus or Udotea deterred feeding by all three herbivores. In most assays, feeding was depressed more by the addition of metabolites from Rhipocephalus or Udotea than by the addition of CaC[O.sub.3] even though CaC[O.sub.3] was added at 1.3-2.2 times the natural concentration for these plants. In contrast, the major metabolite from Halimeda goreauii, when tested alone, did not affect feeding by any of the herbivores. In two of our nine assays, the synchronous combination of CaC[O.sub.3] and secondary metabolites acted synergistically and deterred feeding significantly more than the sum of the effects of each tested separately. Mechanisms producing these synergisms are unknown, but it is possible that calcification could also be acting as a chemical defense by altering gut pH in ways that increase the potency of the secondary metabolites. It is common for chemical, structural, morphological, and nutritional deterrents to co-occur in individual prey species. For some plant-herbivore interactions, the combined effects of these characteristics can be much more than the sum of their separate effects.

Effect of addition of free carbon dioxide leading to a decline in pH, and the administering of carbonate and bicarbonate, on calcification, to the articulated coralline alga Corallina pilulifera, is analyzed. Carbondioxide obstructs calcification, while carbonate and bicarbonate promote calcification. Diurnal changes associated with calcification in the alga are also investigated. Photosynthetic and calcification activities increase in the presence of seawater. These processes are characterized by high levels of solubilized inorganic carbon concentrations at a stable pH of 8.2.

Growth form and calcification in marine macrophytes have been suggested to act as structural defenses against grazers. Support for this hypothesis has been largely indirect, consisting of negative correlations of plant and herbivore abundance and the observation that some herbivores prefer less calcified food plants. The hypothesized defensive role of calcification and growth form has generally not been directly tested. Experiments were conducted to test this hypothesis directly for an important group of grazers, the docoglossan limpets, and a large variety of tropical algae with varying degrees of calcification and different thallus forms. This was accomplished by mimicking the mode of feeding of the herbivore using its feeding apparatus (the radula) and directly measuring the forces required to remove tissue. The results were opposite to the predictions. Neither calcification nor the thallus forms that had been predicted to be structural defenses resulted in increased amounts of force to remove tissue. For these particular herbivores, and possibly others, these morphological features are not acting as simple structural defenses. Other explanations for observed correlations of herbivore and plant abundance and the food preferences of herbivores must be considered.

This paper deals with the problem of texture feature extraction in digital mammograms. We use the extracted features to discriminate between texture representing clusters of microcalcifications and texture representing normal tissue. Having a two-class problem, we suggest a texture feature extraction method based on a single filter optimized with respect to the Fisher criterion. The advantage of this criterion is that it uses both the feature mean and the feature variance to achieve good feature separation. Image compression is desirable to facilitate electronic transmission and storage of digitized mammograms. In this paper, we also explore the effects of data compression on the performance of our proposed detection scheme. The mammograms in our test set were compressed at different ratios using the Joint Photographic Experts Group compression method. Results from an experimental study indicate that our scheme is very well suited for detecting clustered microcalcifications in both uncompressed and compressed mammograms. For the uncompressed mammograms, at a rate of 1.5 false positive clusters/image our method reaches a true positive rate of about 95%, which is comparable to the best results achieved so far. The detection performance for images compressed by a factor of about four is very similar to the performance for uncompressed images. Index Terms--Clustered microcalcifications, mammography, optimal filter, texture features.