Skeletal development inAcropora cervicornis

Scanning electron microscopy, field studies using dyes which become incorporated into the skeleton of living corals as time markers, and petrographic and mineralogic techniques were used to describe the diel pattern of calcium carbonate accretion in the extending axial corallite ofAcropora cervicornis. The axial corallite extends by the formation of randomly oriented fusiform crystals at the distal tip of the branch. Morphological and mineralogical characteristics suggest that these might be calcite crystals. They form a framework upon which needle-like aragonite crystals (initially small tufts) begin to grow. As the needles elongate, groups of them form well defined bundles, fasciculi, which compose the primary skeletal elements. There is a diel pattern in the deposition of the skeleton. At night (1800–0600 hours) the distal spines are pointed and composed primarily of fusiform crystals. During the day (0600–1800 hours) mineral accretion occurs on all surfaces of the skeleton, apparently by epitaxial growth on the aragonite needles of the fasciculi.     

Seasonal variations of pH, pCO2, pO2, HCO3- and Ca2+ in the haemolymph: implications on the calcification physiology in Anodonta cygnea

A study about the relationship between the physical–chemical parameters and the calcium carbonate balance between the haemolymph fluid and mantle calcareous structures was carried out in Anodonta cygnea. An intense peak of HCO₃ ⁻ and a highest pH in December–January months may be understood as a preparation period for creating alkaline conditions. An intense pH decrease from January to February in parallel with the HCO₃ ⁻ reduction seems to indicate the beginning process of carbonate precipitation. On the other hand, the following calcium and HCO₃ ⁻ increases in February–May associated with a continuous and gradual pH fall profile may infer two combined aspects: calcium and HCO₃ ⁻ absorption from external environment and a simultaneous intense calcium carbonate deposition in the haemolymph. So, the pCO₂ peak in this period reflects a subsequent result on equilibrium balance between HCO₃ ⁻ absorption and deposition. The only significant pO₂ increase in the next period, from February to June, is related with an energetic increase to support the metabolic activity favouring the posterior intense pCO₂ peaks. The extended time of CO₂ production in the haemolymph from May to November should induce an increased metabolic acidosis with subsequent intense formation of both HCO₃ ⁻ and Ca²⁺ ions in the same period. This seems to result from CaCO₃ deposits dissolution in the haemolymph, the most direct calcareous source. Additionally, the later increase of metabolic succinic acid during autumn may greatly potentiate this acidosis increasing the dissolution process. Consequently, the pH profile present two simultaneous alkaline peaks in July and October, probably due to a strong HCO₃ ⁻ release from the CaCO₃ dissolution. So, the present seasonal results indicate that in the freshwater bivalve A. cygnea, the low metabolism with higher pH from the early winter is the main cause which may favour a calcareous precipitation, while the high metabolism with lower pH from the early summer may function as an inductor of calcareous dissolution in the haemolymph.     

The skeletal organic matrix from Mediterranean coral Balanophyllia europaea influences calcium carbonate precipitation

Scleractinian coral skeletons are made mainly of calcium carbonate in the form of aragonite. The mineral deposition occurs in a biological confined environment, but it is still a theme of discussion to what extent the calcification occurs under biological or environmental control. Hence, the shape, size and organization of skeletal crystals from the cellular level through the colony architecture, were attributed to factors as diverse as mineral supersaturation levels and organic mediation of crystal growth. The skeleton contains an intra-skeletal organic matrix (OM) of which only the water soluble component was chemically and physically characterized. In this work that OM from the skeleton of the Balanophyllia europaea, a solitary scleractinian coral endemic to the Mediterranean Sea, is studied in vitro with the aim of understanding its role in the mineralization of calcium carbonate. Mineralization of calcium carbonate was conducted by overgrowth experiments on coral skeleton and in calcium chloride solutions containing different ratios of water soluble and/or insoluble OM and of magnesium ions. The precipitates were characterized by diffractometric, spectroscopic and microscopic techniques. The results showed that both soluble and insoluble OM components influence calcium carbonate precipitation and that the effect is enhanced by their co-presence. The role of magnesium ions is also affected by the presence of the OM components. Thus, in vitro, OM influences calcium carbonate crystal morphology, aggregation and polymorphism as a function of its composition and of the content of magnesium ions in the precipitation media. This research, although does not resolve the controversy between environmental or biological control on the deposition of calcium carbonate in corals, sheds a light on the role of OM, which appears mediated by the presence of magnesium ions.     

Multi-scale mineralogical characterization of the hypercalcified sponge Petrobiona massiliana (Calcarea, Calcaronea)

The massive basal skeleton of a few remnant living hypercalcified sponges rediscovered since the 1960s are valuable representatives of ancient calcium carbonate biomineralization mechanisms in basal Metazoa. A multi-scale mineralogical characterization of the easily accessible Mediterranean living hypercalcified sponge belonging to Calcarea, Petrobiona massiliana (Vacelet and Lévi, 1958), was conducted. Oriented observations in light and electron microscopy of mature and growing areas of the Mg-calcite basal skeleton were combined in order to describe all structural levels from the submicronic to the macroscopic scale. The smallest units produced are ca. 50–100 nm grains that are in a mushy amorphous state before their crystallization. Selected area electron diffraction (SAED) further demonstrated that submicronic grains are assembled into crystallographically coherent clusters or fibers, the latter are even laterally associated into single-crystal bundles. A model of crystallization propagation through amorphous submicronic granular units is proposed to explain the formation of coherent micron-scale structural units. Finally, XRD and EELS analyses highlighted, respectively, inter-individual variation of skeletal Mg contents and heterogeneous spatial distribution of Ca ions in skeletal fibers. All mineralogical features presented here cannot be explained by classical inorganic crystallization principles in super-saturated solutions, but rather underlined a highly biologically regulated formation of the basal skeleton. This study extending recent observations on corals, mollusk and echinoderms confirms that occurrence of submicronic granular units and a possible transient amorphous precursor phase in calcium carbonate skeletons is a common biomineralization strategy already selected by basal metazoans.     

Histology and affinity of anaspids,and the early evolution of the vertebrate dermal skeleton

The assembly of the gnathostome bodyplan constitutes a formative episode in vertebrate evolutionary history, an interval in which the mineralized skeleton and its canonical suite of cell and tissue types originated. Fossil jawless fishes, assigned to the gnathostome stem-lineage, provide an unparalleled insight into the origin and evolution of the skeleton, hindered only by uncertainty over the phylogenetic position and evolutionary significance of key clades. Chief among these are the jawless anaspids, whose skeletal composition, a rich source of phylogenetic information, is poorly characterized. Here we survey the histology of representatives spanning anaspid diversity and infer their generalized skeletal architecture. The anaspid dermal skeleton is composed of odontodes comprising spheritic dentine and enameloid, overlying a basal layer of acellular parallel fibre bone containing an extensive shallow canal network. A recoded and revised phylogenetic analysis using equal and implied weights parsimony resolves anaspids as monophyletic, nested among stem-gnathostomes. Our results suggest the anaspid dermal skeleton is a degenerate derivative of a histologically more complex ancestral vertebrate skeleton, rather than reflecting primitive simplicity. Hypotheses that anaspids are ancestral skeletonizing lampreys, or a derived lineage of jawless vertebrates with paired fins, are rejected.     

Biologically controlled mineralization in the hypercalcified sponge Petrobiona massiliana (Calcarea, Calcaronea)

Hypercalcified sponges, endowed with a calcium carbonate basal skeleton in addition to their spicules, form one of the most basal metazoan group engaged in extensive biomineralization. The Mediterranean species Petrobiona massiliana was used to investigate biological controls exerted on the biomineralization of its basal skeleton. Scanning and transmission electron microscopy (SEM, TEM) confirmed that basopinacocytes form a discontinuous layer of flattened cells covering the skeleton and display ultrastructural features attesting intense secretory activity. The production of a highly structured fibrillar organic matrix framework by basopinacocytes toward the growing skeleton was highlighted both by potassium pyroantimonate and ruthenium red protocols, the latter further suggesting the presence of sulfated glycosaminoglycans in the matrix. Furthermore organic material incorporated into the basal skeleton was shown by SEM and TEM at different structural levels while its response to alcian blue and acridine orange staining might suggest a similar acidic and sulfated chemical composition in light microscopy. Potassium pyroantimonate revealed in TEM and energy electron loss spectroscopy (EELS) analysis, heavy linear precipitates 100-300 nm wide containing Ca(2+) and Mg(2+) ions, either along the basal cell membrane of basopinacocytes located toward the decalcified basal skeleton or around decalcified spicules in the mesohyl. Based on the results of the previous mineralogical characterization and the present work, an hypothetical model of biomineralization is proposed for P. massiliana: basopinacocytes would produce an extracellular organic framework that might guide the assemblage of submicronic amorphous Ca- and Mg-bearing grains into higher structural units.     

Hormonal influences on the muscle-bone feedback system: a perspective

Hormones, muscle and bone tissues have co-existed virtually during the whole evolution of vertebrates, and it is obvious that they constitute a complex system able to cope with needs and challenges arising from a variety of physiological and locomotive needs. All body movements are produced by co-ordinated contractions of skeletal muscles, while consequent dynamic muscle work provides the fundamental source of mechanical loading to the skeleton. Mechanical competence of the skeleton is principally maintained by a mechanosensory feedback system that senses the loading-induced deformations within the bones and maintains the skeletal rigidity through structural adaptation. In contrast to the prevalent view suggesting a modulatory effect of hormones on the sensitivity of the mechanosensory system, a new conceptual scheme is proposed. In particular, it is argued that the mechanical and hormonal functions in the skeleton are fundamentally independent but can be seemingly interactive through hormonally-induced modifications in the bone structure, those basically forming a mineral reservoir for maintenance of physiological homeostasis. Whenever needed, utilization of this strategically placed reservoir would not essentially compromise the mechanical competence and locomotive capability of the skeleton. Although plausible, the present view is necessarily speculative and awaits corroborative experimental evidence.     

Stanniocalcin 1 as a pleiotropic factor in mammals

Stanniocalcin (STC)1 is the mammalian homologue of STC which was originally identified as a calcium/phosphate-regulating hormone in bony fishes. STC1 is a homodimeric phosphoglycoprotein with few if any identified unique motifs in its structure with the exception of CAG repeats in the 5'-untranslated region. In contrast to fish STC which is expressed mainly in the corpuscles of Stannius, STC1 is expressed in a wide variety of tissues, but unexpectedly is not detected in the circulation under normal circumstances. Thus, STC1 may play an autocrine/paracrine rather than a classic endocrine role in mammals. Consistent with this, pleiotropic effects of STC1 have been postulated in physiological and measured in pathological situations. There is much current interest in identifying a specific STC1 receptor and putative signaling pathways to which it may be coupled. In this regard, STC1 may regulate intracellular calcium and/or phosphate (Pi) levels. In the skeletal system, for example, Pi uptake in bone-forming osteoblasts via a direct effect of STC1 on expression of the NaPi transporter Pit1 may contribute to bone formation. Here we review current understanding of the role of STC1 and its possible molecular mechanisms in the skeleton and elsewhere.     

Ontogenetic development of an exceptionally preserved Devonian cartilaginous skeleton

Cartilaginous vertebrate skeletons leave few records as fossils, unless mineralized. Here, we report outstanding preservation of early stages of cartilage differentiation, present in the Devonian vertebrate Palaeospondylus gunni. In large specimens of Palaeospondylus, enlarged, hypertrophic cell spaces (lacunae) are dominant in the cartilage matrix, each defined by thin mineralized matrix, where phosphorus and calcium co-occur. This is comparable to living endochondral cartilage, where cell hypertrophy and matrix mineralization mark the end of an ontogenetic process of cell growth and division before bone formation. New information from small individuals of Palaeospondylus demonstrates that the skeleton comprises mostly unmineralized organic matrix with fewer hypertrophic cell spaces, these occurring only in the central regions of each element. Only here has the surrounding matrix begun to mineralize, differing from the larger specimens in that phosphorus is dominant with little associated calcium at these earlier stages. This reflects cellular control of mineralization in living tissues through phosphate accumulation around hypertrophic cells, with later increase in calcium in the cartilaginous matrix. These features are always associated with endochondral bone development, but in the Palaeospondylus skeleton, this bone never develops. This skeletal state is thus far unique among vertebrates, with two alternative explanations: either later stages of endochondral bone development have been lost in Palaeospondylus, or, in a stepwise acquisition of the mineralized skeleton, these late stages have not yet evolved.     

Mammalian hibernation as a model of disuse osteoporosis: the effects of physical inactivity on bone metabolism, structure, and strength

Reduced skeletal loading typically leads to bone loss because bone formation and bone resorption become unbalanced. Hibernation is a natural model of musculoskeletal disuse because hibernating animals greatly reduce weight-bearing activity, and therefore, they would be expected to lose bone. Some evidence suggests that small mammals like ground squirrels, bats, and hamsters do lose bone during hibernation, but the mechanism of bone loss is unclear. In contrast, hibernating bears maintain balanced bone remodeling and preserve bone structure and strength. Differences in the skeletal responses of bears and smaller mammals to hibernation may be due to differences in their hibernation patterns; smaller mammals may excrete calcium liberated from bone during periodic arousals throughout hibernation, leading to progressive bone loss over time, whereas bears may have evolved more sophisticated physiological processes to recycle calcium, prevent hypercalcemia, and maintain bone integrity. Investigating the roles of neural and hormonal control of bear bone metabolism could give valuable insight into translating the mechanisms that prevent disuse-induced bone loss in bears into novel therapies for treating osteoporosis.     

How the osteoclast degrades bone

Osteoclasts are multinucleated monocyte-macrophage derivatives that degrade bone. Their specialized role is central to a process that continuously removes and replaces segments of the skeleton in the higher vertebrates. Osteoclasts allow skeletal mineral to be used to manage extracellular calcium activity, which is an important adaptation for life on land, and solid skeletal structure to be replaced by hollow architecture that has a superior strength-to-weight ratio. Degrading bone also allows periodic repair and remodeling for ordered growth and efficient response to mechanical loads. A fairly comprehensive view of osteoclastic ontogeny and function is emerging from recent studies. Osteoclasts dissolve bone mineral by massive acid secretion and secrete specialized proteinases that degrade the organic matrix, mainly type I collagen, in this acidic milieu. The site of bone dissolution is a high-calcium environment; removal of degradation products by transcytosis of membrane vesicles allows the osteoclast to maintain a normal intracellular calcium. Osteoclastic differentiation is normally balanced with bone formation, although bone formation is the function of unrelated stromal cell-derived osteoblasts. Interactions between osteoclast precursors and bone-forming cells are believed to control osteoclast differentiation under most circumstances, preserving bone architecture over many cycles of bone replacement. BioEssays 20 :837–846, 1998. © 1998 John Wiley & Sons, Inc.     

A comparative view on mechanisms and functions of skeletal remodelling in teleost fish, with special emphasis on osteoclasts and their function

Resorption and remodelling of skeletal tissues is required for development and growth, mechanical adaptation, repair, and mineral homeostasis of the vertebrate skeleton. Here we review for the first time the current knowledge about resorption and remodelling of the skeleton in teleost fish, the largest and most diverse group of extant vertebrates. Teleost species are increasingly used in aquaculture and as models in biomedical skeletal research. Thus, detailed knowledge is required to establish the differences and similarities between mammalian and teleost skeletal remodelling, and between distantly related species such as zebrafish (Danio rerio) and medaka (Oryzias latipes). The cellular mechanisms of differentiation and activation of osteoclasts and the functions of teleost skeletal remodelling are described. Several characteristics, related to skeletal remodelling, distinguish teleosts from mammals. These characteristics include (a) the absence of osteocytes in most species; (b) the absence of haematopoietic bone marrow tissue; (c) the abundance of small mononucleated osteoclasts performing non‐lacunar (smooth) bone resorption, in addition to or instead of multinucleated osteoclasts; and (d) a phosphorus‐ rather than calcium‐driven mineral homeostasis (mainly affecting the postcranial dermal skeleton). Furthermore, (e) skeletal resorption is often absent from particular sites, due to sparse or lacking endochondral ossification. Based on the mode of skeletal remodelling in early ontogeny of all teleosts and in later stages of development of teleosts with acellular bone we suggest a link between acellular bone and the predominance of mononucleated osteoclasts, on the one hand, and cellular bone and multinucleated osteoclasts on the other. The evolutionary origin of skeletal remodelling is discussed and whether mononucleated osteoclasts represent an ancestral type of resorbing cells. Revealing the differentiation and activation of teleost skeletal resorbing cells, in the absence of several factors that trigger mammalian osteoclast differentiation, is a current challenge. Understanding which characters of teleost bone remodelling are derived and which characters are conserved should enhance our understanding of the process in fish and may provide insights into alternative pathways of bone remodelling in mammals.     

Renal tubular acidosis: developments in our understanding of the molecular basis

Renal tubular acidosis is a metabolic acidosis due to impaired acid excretion by the kidney. Hyperchloraemic acidosis with a normal anion gap and normal (or near normal) glomerular filtration rate, and in the absence of diarrhoea, defines this disorder. However, systemic acidosis is not always evident and renal tubular acidosis can present with hypokalaemia, medullary nephrocalcinosis and recurrent calcium phosphate stone disease, as well as growth retardation and rickets in children, or short stature and osteomalacia in adults. Renal dysfunction in renal tubular acidosis is not always confined to acid excretion and can be part of a more generalised renal tubule defect, as in the renal Fanconi syndrome. Isolated renal tubular acidosis is more usually acquired, due to drugs, autoimmune disease, post-obstructive uropathy or any cause of medullary nephrocalcinosis. Less commonly, it is inherited and may be associated with deafness, osteopetrosis or ocular abnormalities. The clinical classification of renal tubular acidosis has been correlated with our current physiological model of how the nephron excretes acid, and this has facilitated genetic studies that have identified mutations in several genes encoding acid and base ion transporters. In vitro functional studies of these mutant proteins in cell expression systems have helped to elucidate the molecular mechanisms underlying renal tubular acidosis, which ultimately may lead to new therapeutic options in what is still treatment only by giving an oral alkali.     

Hypoxia-inducible carbonic anhydrase IX expression is insufficient to alleviate intracellular metabolic acidosis in the muscle of zebrafish, Danio rerio

Recent evidence suggests that carbonic anhydrase (CA) IX in humans is under the regulatory control of hypoxia-inducible factor and is overexpressed in certain cancers. However, little is known of its presence in nonmammalian vertebrates or its physiological function in any vertebrate. The objective of this study was to examine and characterize the presence, distribution, induction by hypoxia, and physiological function of CA IX in the zebrafish. Zebrafish CA IX was highly expressed in the eye, brain, and gastrointestinal tract and showed increased expression in the eye, brain, and muscle in response to hypoxia (water Po(2) = 24 mmHg). The hypothesis that increased CA IX expression during hypoxia would act to attenuate intracellular acidosis was then examined. Muscle intracellular pH (pH(i)) decreased after 4 h of hypoxic exposure (from 7.15 +/- 0.02 to 7.06 +/- 0.01 pH units) and did not recover by 24 h. Manipulation of extracellular CA activity via intraperitoneal injection of either bovine CA or the selective extracellular CA inhibitor F3500 revealed that although increased CA activity could fully restore pH(i), removal of extracellular activity did not result in further acidosis. An exercise-induced acidosis was also attenuated in fish treated with bovine CA; however, the increased extracellular CA expression resulting from hypoxia had no affect. These data suggest that although extracellular CA can potentially minimize the impact of hypoxia on muscle pH(i), the actual level of extracellular CA activity is likely insufficient to achieve this goal, even when enhanced by hypoxia-induced increases in CA IX expression.     

Coralline algal skeletal mineralogy affects grazer impacts

In macroalgal‐dominated systems, herbivory is a major driver in controlling ecosystem structure. However, the role of altered plant–herbivore interactions and effects of changes to trophic control under global change are poorly understood. This is because both macroalgae and grazers themselves may be affected by global change, making changes in plant–herbivore interactions hard to predict. Coralline algae lay down a calcium carbonate skeleton, which serves as protection from grazing and is preserved in archival samples. Here, we compare grazing damage and intensity to coralline algae in situ over 4 decades characterized by changing seawater acidity. While grazing intensity, herbivore abundance and identity remained constant over time, grazing wound width increased together with Mg content of the skeleton and variability in its mineral organization. In one species, decreases in skeletal organization were found concurrent with deeper skeletal damage by grazers over time since the 1980s. Thus, in a future characterized by acidification, we suggest coralline algae may be more prone to grazing damage, mediated by effects of variability between individuals and species.     

Calcium Availability Influences Litter Size and Sex Ratio in White-Footed Mice (Peromyscus leucopus)

The production of offspring typically requires investment of resources derived from both the environment and maternal somatic reserves. As such, the availability of either of these types of resources has the potential to limit the degree to which resources are allocated to reproduction. Theory and empirical studies have argued that mothers modify reproductive performance relative to exogenous resource availability and maternal condition by adjusting size, number or sex of offspring produced. These relationships have classically been defined relative to availability of energy sources; however, in vertebrates, calcium also plays a critical role in offspring production, as a considerable amount of calcium is required to support the development of offspring skeleton(s). We tested whether the availability of calcium influences reproductive output by providing female white-footed mice with a low-calcium or standard diet from reproductive maturity to senescence. We then compared maternal skeletal condition and reproductive output, based on offspring mass, offspring number and litter sex ratio, between dietary treatments. Mothers on the low-calcium diet exhibited diminished skeletal condition at senescence and produced smaller and strongly female-biased litters. We show that skeletal condition and calcium intake can influence sex ratio and reproductive output following general theoretical models of resource partitioning during reproduction.     

New type of organic matrix in cirals formed at the decalcified site: Structure and composition

This is the first demonstration that the organic matrix appears at the decalcified site of the skeleton in a juvenile coral of Fungia fungites (Linnaeus). This matrix was secreted by the calicoblastic layer and is composed of fibrous matters as seen in mesoglea. If formed a thick sheet at the advanced stage. The amino acid composition of this matrix was similar to that in the skeleton rather than mesogleal protein. Morphologic and functional features suggest that the organic sheet must be similar to the sheet which is secreted extracellularly by planula larva at the time of settlement. The sheet in F. fungites may have a role of protecting calicoblastic epithelium from exposure as a result of skeletal dissolution.     

Familial benign hypercalcemia--from clinical description to molecular genetics.

Familial benign hypercalcemia (or familial hypocalciuric hypercalcemia), a syndrome of lifelong hypercalcemia inherited as an autosomal dominant trait, is distinct from the multiple endocrine neoplasia syndromes and other forms of inherited parathyroid disease. Familial benign hypercalcemia results from the inappropriate secretion of parathyroid hormone despite hypercalcemia, enhanced renal tubular reabsorption of calcium (independent of parathyroid hormone), and apparent tissue resistance to adverse effects of hypercalcemia. Heterozygosity for the familial hypercalcemia trait is benign, although homozygosity for the trait may lead to severe neonatal primary hyperparathyroidism. Genetic linkage studies show that most persons affected with familial hypercalcemia have a mutation on the long arm of chromosome 3 (3cen-q21), although one phenotypically indistinguishable family appears to have a mutation on the short arm of chromosome 19 (19p), and another family has neither 3q nor 19p mutations. One group has recently shown mutations in a putative parathyroid cell-surface calcium receptor that are plausible causes for the chromosome 3q variant of the familial hypercalcemia syndrome. Perhaps the other genes for this syndrome encode proteins representing hitherto-unknown regulators of systemic calcium metabolism independent of parathyroid cell calcium sensing or proteins involved in signal transduction from the calcium receptor.