Topography of the organic components in mother-of pearl

1. The topography of the organic components (conchiolin) has been investigated on positive, postshadow-cast, formvar, and carbon replicas of mother-of-pearl from shells of a Cephalopod, of two Gastropods, and of six Pelecypods. All these shells are characterized by a true nacreous inner shell layer. 2. The material included normal shell surfaces, fragments of cleavage obtained by fracture, and surfaces polished tangentially and transversally to the inner surface of the shells. Replicas of these surfaces were prepared before and after etching of graded heaviness, induced by a chelating agent (sequestrene NA 2, titriplex III). Micrographs of the successive steps of the process of corrosion have been recorded. 3. Corrosion unmasked, on the nacreous surfaces, organic membranes or sheets, running as continuous formations in between adjacent mineral lamellae, and separating the individual crystals of aragonite which are aligned in rows and constitute each lamella. 4. The interlamellar sheets of material exhibit a reticulated structure, which is especially visible in preparations orientated tangentially to the lamellae and to the tabular surface of the aragonite crystals. The pattern of this lace-like structure, different in the various species studied, appeared in the same species as closely similar to that reported previously in leaflets of thoroughly decalcified mother-of-pearl, dissociated by ultrasonic waves. The present results support former conclusions with regard to the existence of taxonomic differences between Cephalopods, Gastropods, and Pelecypods in the morphological organization of the organic phase within mother-of-pearl.     

TOPOGRAPHY OF THE ORGANIC COMPONENTS IN MOTHER-OF-PEARL

1. The topography of the organic components (conchiolin) has been investigated on positive, postshadow-cast, formvar, and carbon replicas of mother-of-pearl from shells of a Cephalopod, of two Gastropods, and of six Pelecypods. All these shells are characterized by a true nacreous inner shell layer. 2. The material included normal shell surfaces, fragments of cleavage obtained by fracture, and surfaces polished tangentially and transversally to the inner surface of the shells. Replicas of these surfaces were prepared before and after etching of graded heaviness, induced by a chelating agent (sequestrene NA 2, titriplex III). Micrographs of the successive steps of the process of corrosion have been recorded. 3. Corrosion unmasked, on the nacreous surfaces, organic membranes or sheets, running as continuous formations in between adjacent mineral lamellae, and separating the individual crystals of aragonite which are aligned in rows and constitute each lamella. 4. The interlamellar sheets of material exhibit a reticulated structure, which is especially visible in preparations orientated tangentially to the lamellae and to the tabular surface of the aragonite crystals. The pattern of this lace-like structure, different in the various species studied, appeared in the same species as closely similar to that reported previously in leaflets of thoroughly decalcified mother-of-pearl, dissociated by ultrasonic waves. The present results support former conclusions with regard to the existence of taxonomic differences between Cephalopods, Gastropods, and Pelecypods in the morphological organization of the organic phase within mother-of-pearl.     

Immunocytochemical localization of a calcium-binding phosphoprotein in hemocytes of heterodont bivalves

Calcium-binding phosphoprotein particles are the most abundant extracellular proteins in the hemolymph of heterodont bivalves, and granular hemocytes are the most abundant cells in the same fluid. In this study, the hemocytes of Rangia cuneata were examined ultrastructurally and probed with anti-phosphoprotein IgG to demonstrate that the granulocytes are a probable source of the hemolymph phosphoprotein. The granulocyte cytoplasm is laden with large vesicles containing an amorphous homogenous matrix and variable numbers of electron-dense particles; the latter are ultrastructurally similar to the extracellular phosphoprotein. The vesicle particles and matrix are related forms of the hemolymph phosphoprotein as evidenced by heavy gold labeling when Lowicryl sections were sequentially treated with rabbit-anti-phosphoprotein IgG and colloidal gold-anti-rabbit IgG. The vesicles may be the loci for posttranslational phosphorylation and eventual secretion of the calcium-binding phosphoprotein, or alternatively the vesicles may be digestive structures which degrade internalized phosphoprotein.     

隆线溞[Daphnia(Ctenodaphnia)carinata]卵鞍的超微结构

在不利的环境条件下,枝角类中有一部分种类可以形成卵鞍(ephippium),内含休眠卵。本文应用扫描电镜和透射电镜对隆线溞的卵鞍进行了超微结构的研究。研究表明:卵鞍外面大部分略呈浅的蜂窝状,内面则排布着多数卵石状小突起。卵鞍分为内外两层,两层的超微结构截然不同;各层又可分为三小层。     

A scanning electron microscopic study of the inorganic and organic matrices comprising the mature shell of Amblema, a fresh-water mollusc

The scanning electron microscope has been used to describe the morphology of the mature shell in a fresh-water bivalve. The structure of the organic and inorganic components within the nacre, the myostracum, and the prismatic layer is described. A transitional or intermediate zone, interposed between the prismatic layer and the nacre, was identified. In demineralized samples, the organic component of the nacre was found to consist of parallel matricial sheets interconnected by irregular transverse bridges. The structure of the mineral component of the nacre was found to vary with the method of specimen preparation. With polished-etched samples, brick-like units were seen. When shells were simply broken and fixed in osmium, the layers of nacreous material consisted of fusing rhomboidal crystals of aragonite which demonstrated subconchoidal fractures. On the inner surface of the shell, the rhomboidal crystals showed an apparent spiral growth pattern. The myostracum was characterized by regions of modified nacreous structure consisting of enlarged aragonite crystals with a pyramidal morphology. The peripheral aspect of the muscle scars was characterized by rhomboidal crystals, the latter fusing to form the typical nacreous laminae. The uniqueness of the anterior adductor scar is exemplified by the presence of pores, each pore walled by pyramidal units, for the insertion of adductor fibres. In most regions of the shell, the prismatic layer consisted of one prism unit thickness with a height of approximately 225–250 μm. However, in two specialized regions of the shell, this layer was seen to consist of multiple layers of stacked prisms. The organic matrices of the prismatic layer are arranged in a honeycomb-like arrangement and packed with mineralized spherical subunits.     

Structure of the shell and tertiary membranes of eggs of softshell turtles (Trionyx spiniferus)

Eggs of the turtle Trionyx spiniferus are rigid, calcareous spheres averaging 2.5 cm in diameter. The eggshell is morphologically very similar to avian eggshells. The outer crystalline layer is composed of roughly columnar aggregates, or shell units, of calcium carbonate in the aragonite form. Each shell unit tapers to a somewhat conical tip at its base. Interior to the crystalline layer are two tertiary egg membranes: the outer shell membrane and the inner shell membrane. The outer shell membrane is firmly attached to the inner surface of the shell, and the two membranes are in contact except at the air cell, where the inner shell membrane separates from the outer shell membrane. Both membranes are multi-layered, with the inner shell membrane exhibiting a more fibrous structure than the outer shell membrane. Numerous pores are found in the eggshell, and these generally occur at the intersection of four or more shell units.     

Aminergic and Indoleamine Accumulating Neurons in the Retina of the River Lamprey (Lampetra fluviatilis)

Tissues were processed for fluorescence microscopy of biogenic amines according to the method of Falck and Hillarp. Normal animals, and animals injected with α-methylnoradrenaline or 5,6-dihydroxytryptamine were used. Catecholamine containing neurons (junctional cells) occur in the innermost rows of cell bodies of the inner nuclear layer (INL) and close to the vitreous surface. Catecholamine containing fibers occur in three layers: (1) an outer layer around the innermost perikarya of the INL, which is a condition not found in retinas of gnathostome chordates; (2) a middle layer within the outer third of the inner synaptic layer (ISL), separated from the outer layer by ganglion cell axons; (3) a sparse inner layer within the innermost third of the ISL. A few catecholamine containing fibers were seen to extend from the innermost region of the INL to the outer synaptic layer. The position of the junctional cells in the lamprey corresponds to that in gnathostome chordates, but whereas all catecholamine containing fiber layers in gnathostomes are located sclerally to the optic fiber layer and within the ISL, the middle and the inner fiber layers in the lamprey occur vitreally to the optic fiber layer. Indoleamine accumulating neurons occur in the innermost row of perikarya of the INL and close to the vitreous surface. Those of the INL send fine, varicose branches to the ISL forming a network which is somewhat denser at the inner and outer borders of the ISL than in its middle. The indoleamine accumulating terminals do not ramify within the INL in contrast to the catecholamine containing terminals.     

The Micro/Nanostructure Characteristics and the Mechanical Properties of Hemifusus tuba Conch Shell

<正> The mollusk shell mobilizes calcium from environment for skeletal mineralization.This occurs through synthesizing solidsin solution in the presence of organic molecules of specific interior regions of the conch shell.The ultrastructure and microhardnessof the Hemifusus tuba conch shell living in the Huang/Bo sea area are investigated in the paper.It is shown that thecomposition and microstructure of the mollusk shell vary in different positions.The prodissoconch shell consists only of aragonitewith the crossed-lamellar microstructure.While the spiral shell and the body shell of the Hemifusus tuba conch shell arecomposed of one calcite layer and several aragonite layers.The calcite layer consists of cylindrical grains,but the aragonitelayers are crossed-lamellar ultrastructure at three size scales.The minimum structure size (the third-order lamella) is at about20 nm - 80 nm.The margin of shell aperture is only composed of calcite with cylindrical grains.This natural optimization of theshell microstructure is intimately due to the growth of the Organic matrix.At different positions the microhardness of molluscshell is different due to different crystal structures and crystal arrangements.The growth process of shells allows a constantrenewal of the material,thus enabling their functional adaptation to external environments.     

Ultrastructural morphology of the shell and shell membrane of eggs of common snapping turtles (Chelydra serpentina)

Common snapping turtles (Chelydra serpentina) lay nearly spherical, flexible-shelled eggs having an outer mineral layer composed of calcium carbonate in the aragonite form. The mineral layer is arranged into loosely organized groups of nodular shell units, with numerous spaces (or pores) between adjacent shell units. Shell units are structurally complex, consisting of an inner tip that is morphologically distinct from the main body of the shell unit. Contained within an intact shell unit at the interface of the tip and the main part of the shell unit is the central plaque, an apparent modification of the shell membrane that may serve to nucleate calcification of shell units during shell formation. The tips of shell units are firmly attached to a single, multilayered shell membrane throughout much of incubation. The calcareous layer begins to detach from the shell membrane about half-way through incubation, and changes in shell morphology attending this detachment indicate that snapping turtles may use the shell as a source of calcium during embryogenesis. The arrangement of the mineral layer into groups of shell units, the large number of spaces between shell units, and little or no interlocking of crystallites of adjacent shell units apparently are factors contributing to the ability of these eggs to swell as they absorb water.     

Organization pattern of nacre in Pteriidae (Bivalvia: Mollusca) explained by crystal competition

Bivalve nacre is a brick-wall-patterned biocomposite of aragonite platelets surrounded by organic matter. SEM-electron back scatter diffraction analysis of nacre of the bivalve family Pteriidae reveals that early aragonite crystals grow with their c-axes oriented perpendicular to the growth surface but have their a- and b-axes disoriented. With the accumulation of successive lamellae, crystals progressively orient themselves with their b-axes mutually parallel and towards the growth direction. We propose that progressive orientation is a result of competition between nacre crystals at the growth front of lamellae, which favours selection of crystals whose fastest growth axis (b-axis) is oriented parallel to the direction of propagation of the lamella. A theoretical model has been developed, which simulates competition of rhombic plates at the lamellar growth front as well as epitaxial growth of crystals onto those of the preceding lamella. The model predicts that disordered nacre progressively produces bivalve-like oriented nacre. As growth fronts become diffuse (as is the common case in bivalves) it takes longer for nacre to become organized. Formation of microdomains of nacre platelets with different orientations is also reproduced. In conclusion, not only the organic matrix component, but also the mineral phase plays an active role in organizing the final microstructure.     

Mineralogy of shells from two freshwater snails Belgrandiella fontinalis and B kuesteri

X-ray powder diffraction (XRD) was used to study the mineral composition of shells of snails Belgrandiella fontinalis and Belgrandiella kuesteri collected from three freshwater springs in northeastern Slovenia. The fractions of aragonite, calcite, dolomite and quartz in particular shells were determined. The analysed shells consisted of two or more distinct inorganic layers. The outer shell layer for both species and all sampling localities contained aragonite. The outer layer of B. fontinalis collected at one locality, also contained a small fraction of calcite ( approximately 1 molar%) besides the dominant aragonite. Calcite was identified in the inner layer(s) of both species (2 to 3 molar%), while quartz was found only in B. kuesteri (5-7 molar%). However, both species sampled at one locality showed the presence of dolomite (approx. 20 molar%) in the inner layer(s). The presence of dolomite in the shells of adult gastropods and even molluscs is unusual. A possible formation mechanism and specific ecological factor that could influence the precipitation of dolomite in the shells of different Belgrandiella species is discussed.     

Eggshell formation during prolonged gravidity of the tuatara Sphenodon punctatus

Scanning electron microscopy (SEM) was used to examine the process of shell formation in tuatara. Tuatara carry eggs in the oviducts for ∼ 7–8 mo before nesting, a period of gravidity more than three times as long as in any other oviparous reptile. Our aim was to determine whether shell formation occurred rapidly after ovulation, or whether it occurred gradually throughout gravidity. Eggs were obtained from females in early gravidity (May, ∼ 1 mo after ovulation), midgravidity (August and September, 4–5 mo after ovulation), and late gravidity, immediately prior to nesting (December, 8 mo after ovulation). The shell membrane (fibrous layer) was well formed by May, but calcification of the outer surface had only just begun. Vertical columns of calcium carbonate were embedded in the shell membrane and appeared to erupt through the outer surface between early and midgravidity. Changes in the appearance of the outer calcareous layer were evident as gravidity progressed. In all shells, calcium carbonate was present as calcite. The appearance of the inner boundary (innermost layer of eggshell) was variable; some shells had a smooth and amorphous inner boundary as previously reported for tuatara and other reptiles, whereas other shells had an inner boundary composed of small spherical granules on the inner surface of which small calcareous spicules were scattered. A previously published model of the process of shell formation in tuatara eggshells is refined in light of our observations. We interpret the ability of female tuatara to shell their eggs gradually during winter as further evidence of their unusual physiological tolerance of cold conditions. © 1996 Wiley-Liss, Inc.     

Mollusc larval shell formation: amorphous calcium carbonate is a precursor phase for aragonite

The larval shells of the marine bivalves Mercenaria mercenaria and Crassostrea gigas are investigated by polarized light microscopy, infrared spectroscopy, Raman imaging spectroscopy, and scanning electron microscopy. Both species contain similar shell ultrastructures. We show that larval shells contain amorphous calcium carbonate (ACC), in addition to aragonite. The aragonite is much less crystalline than non-biogenic aragonite. We further show that the initially deposited mineral phase is predominantly ACC that subsequently partially transforms into aragonite. The postset juvenile shell, as well as the adult shell of Mercenaria also contains aragonite that is less crystalline than non-biogenic aragonite. We conclude that ACC fulfills an important function in mollusc larval shell formation. It is conceivable that ACC may also be involved in adult shell formation.     

The Characterisation of Residual Strain in Ensis siliqua Shells

1 Introduction Molluscan species exhibit a range of morphologies associated with their particular function and habitat. The razor clam (Ensis siliqua) is a bivalve mollusc, usually found in fine sands, that burrows up to depths of 20 fathoms (～36 m)[1]. The species is found in several coastal regions of the British Isles, the Norwegian Sea, the Baltic south, the Iberian Peninsula, the Mediterranean and along the Atlantic coast of Morrocco[2]. The protec- tive outer shell, which is often …     

Copillaria hepatica: Fine structure of egg shell

The structure of the Capillaria hepatica egg shell was studied with the electron microscope and correlated with light microscope histochemical observations. The shell is composed of fibrous and nonfibrous components, both of which stain for protein. The fibrous component, the major portion of the shell, consists of submicroscopic fibers. The nonfibrous component is located in the outer region of the shell but is not always visible; when present it has a reticulated appearance in electron micrographs. The fibrous component is divided into outer and inner regions. The outer region is composed of radially arranged pillars which are connected at their outer surface by a beam-like network and are anchored at the base to a compact inner region. The inner region consists of a series of concentrically arranged lamellae above which is located a nonlaminated region where the pillar bases originate. At each polar end of the shell is a single opening plugged with a material which contains acid mucopolysaccharide. The fine structure of the body of the plug is unresolvable with the electron microscope; its outer surface is impregnated with electron dense particles. Externally the shell is covered by a 250 Å thick continuous membrane which is in close opposition to the surrounding host tissue.     

Ultrastructure of lamellae, mineral and matrix components of fish otolith twinned aragonite crystals: implications for estimating age in fish

Atomic force microscopy (AFM) of the crystalline ultrastructure of otoliths fromPagrus major(Sparidae),Macruronus novaezelandiae(Merlucciidae),Oncorhynchus tshawytscha(Salmonidae),Sebastes alutus(Scorpaenidae), andHoplostethus atlanticus(Trachichthyidae) showed regular deposition of lamellae in the size range 13-490 nm. The orientation of lamellae in the {010} plane was the same as lamellae in crystals of mineral aragonite. Lamellae in mineral aragonite were in the size range 15-45 nm. Lamellae observed in the otolith ofM. novaezelandiaeby transmission electron microscopy showed a range of widths (25-225 nm) similar to lamellae observed by AFM. The observed lamella widths were in the size range that has been described for sub-daily and daily microincrements in otoliths. Observed lamellae widths were also in the size range of alpha-recoil trajectories of(222)Rn and provide a potential diffusion sink correction for the(222)Rn losses in radionuclide method of ageing otoliths. Comparison of the orientations of lamellae to templates based on the Bragg unit cell structure of twinned aragonite indicated that the lamellae resulted from polysynthetic twinning on the {010} aragonite crystal face. Additional cyclic twinning occurred on the {110} face of the aragonite crystal and sometimes led to pseudohexagonal crystals, whose sizes were orders of magnitude larger than lamellae. The organic matrix of the otolith was visible by atomic force and transmission electron microscopy at the nanometer level of resolution, but the organic matrix was confined to the {110} twinning plane of symmetry of the otolith crystal.     

Calcineurin Plays an Important Role in the Shell Formation of Pearl Oyster (Pinctada fucata)

Calcineurin (CN) is a multifunctional protein involved in many important physiological processes in mammalians, but the function of CN in mollusks is still largely unknown. In the present study, through the shell regeneration system, the changes of enzymatic activity of CN were determined in the process of shell regeneration in pearl oyster Pinctada fucata. CN was activated immediately and continuously in the shell regeneration process. The speed of shell regeneration was measured and the ultrastructure of inner shell surface was observed by scanning electron microscopy after inhibiting CN by intramuscular injection of immunosuppresant cyclosporine A (CsA). The results showed that the speed of shell regeneration was delayed and the morphology of calcite and aragonite in the inner shell surface became abnormal when CN was inhibited by CsA. Meanwhile, RT-PCR analysis revealed that the expression of P. fucata BMP-2 in mantle tissue decreased with CsA injection. In vitro secretion level of proteoglycans (PGs) in primary cultures of mantle cells was also decreased when mantle cells were exposed to CsA. Taken together, our results, for the first time, show that CN is involved in the shell formation through regulating the expression of Pf-BMP-2 in mantle tissue, which controls the secretion of PGs/GAGs of the mantle epithelial cells.     

Distribution and characterization of mineral-binding phosphoprotein particles in Bivalvia

Representative species of four bivalve subclasses were examined for the presence of mineral-binding phosphoprotein particles in the physiological fluids. The particles were identified in Heterodont bivalves only, and particles from nine different Heterodont species were isolated and characterized. All phosphoprotein particles are internally cross-linked via histidinoalanine residues. In all species over 80% of the amino acid residues in the particles are aspartic acid, phosphoserine (and/or phosphothreonine), and histidine. These amino acids are probably the only residues directly related to mineral ion binding, since all phosphoprotein particles bind mineral irrespective of the minor amino acid content, which is species dependent. In their native state the phosphoprotein particles contain large amounts of calcium, magnesium, and inorganic phosphate ions (up to 45 metal ions and 8 phosphate ions per 100 amino acid residues) and trace amounts of transition elements. Evidence for the presence of calcium phosphate complexes in the native phosphoprotein particles was obtained by observing a concomitant increase in the inorganic phosphate and calcium ion content of the particles with pH in vivo.     

Microtexture of larval shell of oyster,Crassostrea nippona: A FIB-TEM study

The initial formation and subsequent development of larval shells in marine bivalve, Crassostrea nippona were investigated using the FIB-TEM technique. Fourteen hours after fertilization (the trochophore stage), larvae form an incipient shell of 100–150 nm thick with a columnar contrast. Selected-area electron diffraction analysis showed a single-crystal aragonite pattern with the c-axis perpendicular to the shell surface. Plan-view TEM analysis suggested that the shell contains high density of {110} twins, which are the origin of the columnar contrast in the cross-sectional images. 72 h after fertilization (the veliger stage), the shell grows up to 1.2–1.4 μm thick accompanying an additional granular layer between the preexisting layer and embryo to form a distinctive two-layer structure. The granular layer is also composed of aragonite crystals sharing their c-axes perpendicular to the shell surface, but the crystals are arranged with a flexible rotation around the c-axes and not restricted solely to the {110} twin relation. No evidence to suggest the existence of amorphous calcium carbonate (ACC) was found through the observation. The well-regulated crystallographic properties found in the present sample imply initial shell formation probably via a direct deposition of crystalline aragonite.     

Differentiation and Growth of the Brachiopod Mantle

Cell differentiation in the mantle edge of Notosaria, Thecidelhnaand Glottidia, representing respectively, the impunctate andpunctate calcareous articulate and chitinophosphatic inarticulatebrachiopods, is described. Comparison of electron micrographssuggests that outer epithelium which secretes periostracum andmineral shell, is separated from inner epithelium by a bandof "lobate" cells, of variable width, exuding an impersistentmucopolysaccharide film or pellicle. The lobate cells alwaysoccupy the same relative position on the inner surface of theouter mantle lobe; but the outer epithelium is commonly connectedwith the inner surface of the periostracum by papillae and protoplasmicstrands which persist during mineral deposition and ensure thatboth shell and attached mantle remain in situ relative to theoutwardly expanding inner surface of the outer mantle lobe.In the prototypic brachiopod, the lobate cells are likely atfirst to have occupied the hinge of the mantel fold but laterto have been displaced into their present position by the rigidoutward growing edge of the mineral shell.