Intracellular crystal-bearing vesicles in the epidermis of scleractinian corals, Astrangia danae (Agassiz) and Porites porites (Pallas).

Orthorhombic aragonitic crystals, embedded with a granular lipo-protein matrix and surrounded by a trilaminar membrane, are localized in the apical cytoplasm of epidermal cells of Scleractinian corals. Adult specimens of Astrangia danae (Agassiz) and settled planulae of Porites porites (Pallas) contain crystals averaging 0.7 mu by 0.1 mu by 0.3 mu within Golgi-derived vesicles. Short-term labelling with 45Ca reveals distribution of radioactivity amont a basic tissue fraction (92%) an acid tissue fraction (5%) and a skeletal fraction (3%). Identification of the primordial crystal population within membrane-bound visicles provides overwhelming evidence for the intracellular mode of calcification in Scleractinia. Moreover, it permits development of a novel concept of cellular regulation over these dynamic events. The membrane-bound vesicel is a miniature crystal fabrication station and a vehicle responsible for transportation of seed crystals and an organic matrix material to sites of discharge from the cell. The vesicle membrane becomes a probable locus of active transport and enzymatic activity as well as a physical barrier to be penetrated for release of vesicle contents into the extracellular milieu. Contact between the vesicle membrane and the plasmalemma would result in exocytosis and the onset of skeletogenesis. Principles governing crystal growth would prevail from then on. The released crystal becomes a nucleation catalyst and the organic matrix, a supply of ionic calcium for self-limiting crystallization. Crystals are produced by the organism spontaneously and continuously from shortly after larval attachment throughout the life of the polyp. Therefore, these membrane-bound vesicles signal the dynamic process by which initiation, differentiation, growth and limitation of the coral skeleton is regulated.     

Ontogenetic development of skeletal weight in a small delphinid,Delphinus delphis (Cetacea,Odontoceti)

Summary Total body weight and skeletal weight, from clean and dry bones, were measured in 61 common dolphins, Delphinus delphis of known ages (age determinations using both growth lines in the jaw, and growth layer groups in dentine). The relative skeletal weight in adults, expressed as a percentage of total body weight, is notably less than that of any cetaceans previously described. Appearently, this ratio is also smaller than that for any other vertebrate. In particular, skeletons of delphinids tend to be proportionally lighter than those of bony fishes. Classical hypotheses concerning the effect of buoyancy on the mass of supportive tissues can only partially explain such a relative reduction of skeletal weight.     

Directional asymmetry in the limbs,skull and pelvis of the silver fox (V. vulpes)

Directional asymmetry (DA) is a characteristic of most vertebrates, most strikingly exhibited by the placement of various organs (heart, lungs, liver, etc.) but also noted in small differences in the metrics of skeletal structures such as the pelvis of certain fish or sauropsids. We have analyzed DA in the skeleton of the fox (V. vulpes), using ～1,000 radiographs of foxes from populations used in the genetic analysis of behavior and morphology. Careful measurements from this robust data base demonstrate that: 1) DA occurs in the limb bones, the ileum, and ischium and in the mandible; 2) regardless of the direction of the length asymmetry vector of a particular skeletal unit, the vectorial direction of length is always opposite to that of width; 3) with the exception of the humerus and radius, there is no correlation or inverse correlation between vectorial amplitudes or magnitudes of bone asymmetries. 4) Postnatal measurements on foxes demonstrate that the asymmetry increases after birth and continues to change (increasing or decreasing) during postnatal growth. 5) A behavior test for preferential use of a specific forelimb exhibited fluctuating asymmetry but not DA. None of the skeletal asymmetries were significantly correlated with a preferential use of a specific forelimb. We suggest that for the majority of fox skeletal parameters, growth on the right and left side of the fox are differentially biased resulting in fixed differences between the two sides in either the rate of growth or the length of the period during which growth occurs. Random effects around these fixed differences perturb the magnitude of the effects such that the magnitudes of length and width asymmetries are not inversely correlated at the level of individual animals. J. Morphol., 2010. © 2010 Wiley‐Liss, Inc.     

Appendicular skeletal morphology in minute salamanders,genus Thorius (amphibia: Plethodontidae): Growth regulation,adult size determination,and natural variation

Patterns of growth and variation of the appendicular skeleton were examined in Thorius, a speciose genus of minute terrestrial plethodontid salamanders from southern Mexico. Observations were based primarily on ontogenetic series of each of five species that collectively span the range of adult body size in the genus; samples of adults of each of seven additional species provided supplemental estimates of the full range of variation of limb skeletal morphology. Limbs are generally reduced, i.e., pedomorphic, in both overall size and development, and they are characterized by a pattern of extreme variation in the composition of the limb skeleton, especially mesopodial elements, both within and between species. Fifteen different combinations of fused carpal or tarsal elements are variably present in the genus, producing at least 18 different overall carpal or tarsal arrangements, many of which occur in no other plethodontid genus. As many as four carpal or tarsal arrangements were observed in single population samples of each of several; five tarsal arrangements were observed in one population of T. minutissimus. Left-right asymmetry of mesopodial arrangement in a given specimen is also common. In contrast, several unique, nonpedomorphic features of the limb skeleton, including ossification of the typically cartilaginous adult mesopodial elements and ontogenetic increase in the degree of ossification of long bones, are characteristic of all species and distinguish Thorius from most related genera. They form part of a mechanism of determinate skeletal growth that restricts skeletal growth after sexual maturity. Interspecific differences in the timing of the processes of appendicular skeletal maturation relative to body size are well correlated with interspecific differences in mean adult size and size at sexual maturity, suggesting that shifts in the timing of skeletal maturation provide a mechanism of achieving adult size differentiation among species. Processes of skeletal maturation that confer determinate skeletal growth in Thorius are analogous to those typical of most amniotes – both groups exhibit ontogenetic reduction and eventual disappearance of the complex of stratified layers of proliferating and maturing cartilage in long bone epiphyses – but, unlike most amniotes, Thorius lacks secondary ossification centers. Thus, the presence of secondary ossification centers cannot be used as a criterion for establishing determinate skeletal growth in all vertebrates.     

LYSTROSAURUS MURRAYI (THERAPSIDA, DICYNODONTIA): BONE HISTOLOGY, GROWTH AND LIFESTYLE ADAPTATIONS

Abstract:  Examination of the bone microstructure of Lystrosaurus murrayi from India and South Africa reveals a predominance of fibrolamellar bone tissue, which suggests rapid periosteal osteogenesis and an overall fast growth. Four distinct ontogenetic stages have been identified based on tissue type, organization of the primary osteons, incidence of growth rings, secondary reconstruction and endosteal bone deposition. An indeterminate growth strategy is proposed for Lystrosaurus . Inter-elemental histovariability suggests differential growth rate of the skeletal elements within the same individual, and among different individuals. The high cortical thickness of the dorsal ribs, an extensive secondary reconstruction in the cortical region of different skeletal elements that resulted in erosionally enlarged channels from the perimedullary to the midcortical region, and trabecular infilling of the medullary region even in the diaphyseal sections of the limb bones suggest at least a semi-aquatic lifestyle for L. murrayi .     

Relative growth,ontogeny, and sexual dimorphism in gorilla(Gorilla gorilla gorilla and G. g. beringei): Evolutionary and ecological considerations

Gorillas are the largest and among the most sexually dimorphic of all extant primates. While gorillas have been incorporated in broad-level comparisons among large-bodied hominoids or in studies of the African apes, comparisons between gorilla subspecies have been rare. During the past decade, however, behavioral, morphological, and molecular data from a number of studies have indicated that the western lowland (Gorilla gorilla gorilla) and eastern mountain (Gorilla gorilla beringei) subspecies differ to a greater extent than has been previously believed. In this study I compare patterns of relative growth of the postcranial skeleton to evaluate whether differences between subspecies result from the differential extension of common patterns of relative growth. In addition, patterns of ontogeny and sexual dimorphism are also examined. Linear skeletal dimensions and skeletal weight were obtained for ontogenetic series of male and female G.g. gorilla (n = 315) and G.g. beringei (n = 38). Bivariate and multivariate methods of analysis were used to test for differences in patterns of relative growth, ontogeny, and sexual dimorphism between sexes of each subspecies and in same-sex comparisons between subspecies. Results indicate males and females of both subspecies are ontogenetically scaled for postcranial proportions and that females undergo an earlier skeletal growth spurt compared to males. However, results also indicate that the onset of the female growth spurt occurs at different dental stages in lowland and mountain gorillas and that mountain gorillas may be characterized by higher rates of growth. Finally, data demonstrate lowland and mountain gorilla females do not differ significantly in adult body size, but mountain gorilla males are significantly larger than lowland gorilla males, suggesting mountain gorillas are characterized by a higher degree of sexual dimorphism in body size. Thus, although lowland and mountain gorillas do not appear to have evolved novel adaptations of the postcranium which correlate with differences in locomotor behavior, the present investigation establishes subspecies differences in ontogeny and sexual dimorphism which may be linked with ecological variation. Specifically, these findings are evaluated in the context of risk aversion models which predict higher growth rates and increased levels of sexual dimorphism in extreme folivores. Am. J. Primatol. 43:1–31, 1997. © 1997 Wiley-Liss, Inc.     

Skeletal form and growth in Acropora aspera (Dana) from the Pulau Seribu,Indonesia

Linear extension and calcium carbonate accretion were measured in the branching coral Acropora aspera (Dana) from shallow-water sites around Pulau Pari, Pulau Seribu, Indonesia, during both wet and dry monsoon periods. Skeletal density and corallite form were also monitored in specimens collected from sites, variously affected by wave energy resulting from the monsoonal influence. Although the reversing monsoon appeared to exert the greatest effect on skeleton growth (by influencing temperature and possibly number of “sun-hours”) wave energy was also shown to affect skeletal extension, skeletal accretion, and skeletal density. The scale of differences between growth rate measurements was greatest for weight of skeleton accreted between monsoon period (8-fold), followed by between site differences (maximum 3-fold during west monsoon) and finally between station differences (maximum 3-fold during west monsoon at an outer reef flat and reef edge station). Skeletal extension did not appear to be as sensitive to the reversing monsoon influence as skeletal accretion.     

Crystallization, stability, and enzymatic degradation of poly(L-lactide) thin film

Poly(L-lactide) (PLLA) thin film with 100 nm thickness was crystallized at 160 degreesC for 20 min from the melt obtained at 220 degreesC. Hexagonal crystals with three types of growth (derivative growth lamellae, overgrowth multistacked lamellae, and undergrowth multistacked lamellae) were simultaneously observed by atomic force microscopy (AFM). These phenomena are due to the differences of the formative points of secondary crystal nuclei against the basal lamella. Enzymatic degradation of PLLA thin film revealed two types of amorphous regions. These regions were identified as the free amorphous region around the crystals and the restricted amorphous region between the crystal and glass substrate. In situ observation of thermal behavior of lamellar crystals was performed to understand the correlation between the chain folding and stability of the crystal by using temperature-controlled AFM. The morphology of the sectors with [100] growth plane had changed to a comblike morphology despite the fact that the [110] growth plane remained unchanged, suggesting that the stability of the chain folding and the chain-packing state affected the thermal behavior.     

Use of the frontal bone in age determination of Labeo horie (Pisces,Cyprinidae) in Jebel Aulia Reservoir,Sudan

In spite of the exuberance of recent approaches to age determination of fish, many ecologists are convinced that the most valid method is age reading from permanent marks or growth rings laid on skeletal parts. Of the known structures used, only the vertebrae are suitable and readable for a large variety of species and sizes. Examination of different skeletal structures of Labeo horie, a Cyprinid collected from Jebel Aulia reservoir, revealed that the frontal bone carried well-defined growth rings. The validity of these rings was checked against other direct and indirect methods of age determination. Results proved to be statistically valid, and the frontal bone offers an additional device of age determination of L. horie.     

Molecular characterization,expression patterns and subcellular localization of Myotrophin (MTPN) gene in porcine skeletal muscle

Myotrophin (MTPN) is an effective growth factor in promoting skeletal muscle growth in vitro and vivo and has been purified from porcine skeletal muscle. However, in pigs, the information on MTPN gene is very limited. In this study, we cloned cDNA sequences and analyzed the genomic structure of porcine MTPN gene. The deduced amino acid sequence of porcine MTPN contains two the ankyrin repeat domains. RT-PCR analysis revealed that porcine MTPN gene was widely expressed in many tissues, a high expression level was observed in the spleen, liver and uterus, and transient transfection indicated that porcine MTPN proteins was located in cytoplasms within Pig Kidney Epithelial cells (PK15). Quantitative real-time PCR (qRT-PCR) analyses showed that MTPN expression peaked at embryonic 65 day post conception (dpc). During postnatal muscle development, MTPN expression was down-regulated from the 3 day to the 180 day in Yorkshire pigs. This result suggests that the MTPN gene may be important gene for skeletal muscle growth and provides useful information for further studies on its roles in porcine skeletal muscle.     

Youngest early carboniferous (late Visean) shallow-water patch reefs in eastern Australia (Rockhampton Group, Queensland): Combining quantitative micro- and macro-scale data

Summary Although skeletal organisms have received most of the emphasis in studies on Phanerozoic roef history, the roles of non-skeletal (non-enzymatic) carbonates (e.g., synsedimentary cements, automicrite, microbialite, etc.) in reef framework construction are becoming increasingly better understood. One problem in understanding the role of non-enzymatic carbonates in reef construction has been the difficulty in recognizing them in reef facies. Whereas skeletal organisms commonly can be recognized and documented in the field, non-enzymatic carbonates may be recognizable only in thin section. This paper describes the application of a new sampling technique that allows the quantitative comparison of skeletal macrofauna and flora with associated non-enzymatic carbonates and other microfaunal/microfloral constituents. The technique involves the point counting of thin sections made from small diameter cores that are systematically recovered from grids and line transects that cover a reasonable area (m²) of reef facies. Small, shallow-water patch reefs are abundant in scattered oolitic intervals in the Lower Carboniferous strata of eastern Australia. The youngest known Carboniferous reefs in eastern Australia occur in uppermost Visean strata (limestone FC5) near the top of the Rockhampton Group, approximately 50 km west-northwest of Rockhampton, Queensland. The largest sampled reef was 15 m thick and 42 m in diameter, with synsedimentary relief above the sea floor of at least 2 m during the primary growth phase. The reef occurs within bioclasticoolitic grainstones representing a shallow shelf setting and consists of eight common framework microfacies: 1) coral boundstone; 2) bryozoan boundstone; 3) mixed crinoid-bryozoan boundstone; 4) tubular problematica boundstone; 5) sponge-automicrite boundstone; 6) encrusted thrombolite boundstone; 7) mixed automicrite boundstone; and 8) thrombolitic wackestone-packstone. Reef growth was initiated by automicrite-producing biofilms, sponges and a tubular problematic organism. Primary relief building was accomplished by automicrite-dominated frameworks and lithistid sponges, crinoids, and corals. Large cerioidAphrophyllum coral colonies had a heterogeneous distribution through the reef. The framework of the main relief-bearing portion of the reef consists on average of 44.4% automicrite and automicrite-bound detritus, excluding automicrite-bound sponge body fossils, and at most 19.6% skeletal organisms in growth position (minimum of 7.2%). If sponge body fossils are included as automicrite framework, because they are preserved only as a result of automicrite formation, the percentage of automicrite and bound sediment is 54.9%. A smaller sampled reef consisting of the same basic facies had 39.5% automicrite and automicrite-bound sediment in its fremework (50.2% including sponges) and, at most, 33.4% skeletal organisms in growth position (minimum of 22.7%). The greater volume of skeletal framework in the small reef reflects a greater proportion of large corals. Of framebuilding skeletal organisms, automicrite-preserved lithistid and other sponges and cerioid rugose corals provided the greatest volume. However, crinoid holdfasts were the most widespread skeletal framework components. The dominant framework facies are sponge-automicrite boundstone, encrusted thrombolite, boundstone, mixed automicrite boundstone, and coral boundstone. The reefs are similar in overall framework construction and ecological succession to slightly older Visean reefs in eastern Australia and to some of the late Visean reefs of northern England. Surprisingly, framework similarities also exist between the reefs and certain thrombolite-lithistid-coral reefs of the European Jurassic.     

Perlinhibin, a cysteine-, histidine-, and arginine-rich miniprotein from abalone (Haliotis laevigata) nacre, inhibits in vitro calcium carbonate crystallization

We have isolated a 4.785 Da protein from the nacreous layer of the sea snail Haliotis laevigata (greenlip abalone) shell after demineralization with acetic acid. The sequence of 41 amino acids was determined by Edman degradation supported by mass spectrometry. The most abundant amino acids were cysteine (19.5%), histidine (17%), and arginine (14.6%). The positively charged amino acids were almost counterbalanced by negatively charged ones resulting in a calculated isoelectric point of 7.86. Atomic-force microscopy studies of the interaction of the protein with calcite surfaces in supersaturated calcium carbonate solution or calcium chloride solution showed that the protein bound specifically to calcite steps, inhibiting further crystal growth at these sites in carbonate solution and preventing crystal dissolution when carbonate was substituted with chloride. Therefore this protein was named perlinhibin. X-ray diffraction investigation of the crystal after atomic-force microscopy growth experiments showed that the formation of aragonite was induced on the calcite substrate around holes caused by perlinhibin crystal-growth inhibition. The strong interaction of the protein with calcium carbonate was also shown by vapor diffusion crystallization. In the presence of the protein, the crystal surfaces were covered with holes due to protein binding and local inhibition of crystal growth. In addition to perlinhibin, we isolated and sequenced a perlinhibin-related protein, indicating that perlinhibin may be a member of a family of closely related proteins.     

Ubiquitin-specific peptidase 9, X-linked (USP9X) modulates activity of mammalian target of rapamycin (mTOR)

The mammalian target of rapamycin (mTOR) is an atypical serine/threonine kinase that responds to extracellular environment to regulate a number of cellular processes. These include cell growth, proliferation, and differentiation. Although both kinase-dependent and -independent functions of mTOR are known to be critical modulators of muscle cell differentiation and regeneration, the signaling mechanisms regulating mTOR activity during differentiation are still unclear. In this study we identify a novel mTOR interacting protein, the ubiquitin-specific protease USP9X, which acts as a negative regulator of mTOR activity and muscle differentiation. USP9X can co-immunoprecipitate mTOR with both Raptor and Rictor, components of mTOR complexes 1 and 2 (mTORC1 and -2), respectively, suggesting that it is present in both mTOR complexes. Knockdown of USP9X leads to increased mTORC1 activity in response to growth factor stimulation. Interestingly, upon initiation of differentiation of C2C12 mouse skeletal myoblasts, knockdown of USP9X increases mTORC2 activity. This increase in mTORC2 activity is accompanied by accelerated differentiation of myoblasts into myotubes. Taken together, our data describe the identification of the deubiquitinase USP9X as a novel mTORC1 and -2 binding partner that negatively regulates mTOR activity and skeletal muscle differentiation.     

Environmental influences on skeletal banding in eastern Pacific (Panama) corals

The timing of skeletal band formation and concomitant changes in calcification rates and linear skeletal extension were investigated in Pavona corals growing under two distinct thermal regimes along the Pacific coast of Panama: fluctuating, marked by seasonal upwelling (Gulf of Panama) and stable, nonupwelling (Gulf of Chiriqui). The purpose of this study was to test the hypothesis that banding in corals is largely mediated by seasonal variations in temperature (Highsmith 1979). Our results indicate that the timing of band formation is synchronous at these two environmentally distinct locations. The low density (LD) portion of the annual band is accreted over a five month period (January–June) and represents an increase in linear skeletal extension (mm/mo.) as well as a marked increase in calcification rate (g CaCO₃ · cm^-2 · mo^-1) relative to the high density portion which forms over the remaining seven month period (July through December). In contrast to the predictions of the Highsmith model these findings indicate that variations in light levels rather than fluctuation in temperature is a better correlate to changes in skeletal density. Qualitatively, banding patterns were similar at the two sites; however, higher growth rates (particularly with respect to the LD band) for Pavona clavus in the Gulf of Panama indicate that lower water temperatures and higher productivity, or both, may be responsible for quantitative differences in banding between sites. We found that formation of the HD band corresponds to lower light levels and the production of gametes. We propose that banding in corals is a complex phenomenon governed by endogenous processes (e.g. reallocation of energy from growth to reproduction) which may be mediated by exogenous factors (e.g light and productivity).     

Calcium Homeostasis in Myogenic Differentiation Factor 1 (MyoD)-Transformed,Virally-Transduced,Skin-Derived Equine Myotubes

Dysfunctional skeletal muscle calcium homeostasis plays a central role in the pathophysiology of several human and animal skeletal muscle disorders, in particular, genetic disorders associated with ryanodine receptor 1 (RYR1) mutations, such as malignant hyperthermia, central core disease, multiminicore disease and certain centronuclear myopathies. In addition, aberrant skeletal muscle calcium handling is believed to play a pivotal role in the highly prevalent disorder of Thoroughbred racehorses, known as Recurrent Exertional Rhabdomyolysis. Traditionally, such defects were studied in human and equine subjects by examining the contractile responses of biopsied muscle strips exposed to caffeine, a potent RYR1 agonist. However, this test is not widely available and, due to its invasive nature, is potentially less suitable for valuable animals in training or in the human paediatric setting. Furthermore, increasingly, RYR1 gene polymorphisms (of unknown pathogenicity and significance) are being identified through next generation sequencing projects. Consequently, we have investigated a less invasive test that can be used to study calcium homeostasis in cultured, skin-derived fibroblasts that are converted to the muscle lineage by viral transduction with a MyoD (myogenic differentiation 1) transgene. Similar models have been utilised to examine calcium homeostasis in human patient cells, however, to date, there has been no detailed assessment of the cells’ calcium homeostasis, and in particular, the responses to agonists and antagonists of RYR1. Here we describe experiments conducted to assess calcium handling of the cells and examine responses to treatment with dantrolene, a drug commonly used for prophylaxis of recurrent exertional rhabdomyolysis in horses and malignant hyperthermia in humans.     

Chemistry and structure of skeletal growth rings in the black coral Antipathes fiordensis (Cnidaria,Antipatharia)

The skeleton of the black coral Antipathes fiordensis is a laminar structure displaying major and minor growth rings. The skeleton is composed primarily of protein and chitin organized into successive microlayers. Each microlayer is tightly bound to the next by an organic cement that is periodically opaque in section, and occurs as clusters throughout the skeleton. In contrast, the skeletal microlayers themselves are never opaque. The apposition of multiple, opaque cement lines between skeletal layers is primarily responsible for the growth ring pattern.     

Growth requires bone resorption at particular skeletal elements in a teleost fish with acellular bone (Oreochromis niloticus, Teleostei: Cichlidae)

To date, little is known about bone resorption during skeletal development in teleostean fish with acellular bone. We report here about bone resorption with regard to growth in the tilapia Oreochromis niloticus. Nine skeletal elements obtained from growing juveniles were examined using histological and histochemical methods, and transmission electron microscopy (TEM). Tartrate-resistant acid phosphatase (TRAP) served as a marker for bone resorbing cells (osteoclasts), alkaline phosphatase (ALP) was used to identify osteoblasts, and alizarin staining indicated sites of bone formation. TRAP-activity was located at those skeletal elements where growth requires bone resorption, and at sites of cartilage degeneration. No TRAP-activity was found at those skeletal elements where resorption was not required for growth. The examination of the praeopercular shaft leads to a model of bone enlargement, including bone resorption by TRAP-positive cells located at the endosteal bone surface and bone formation by ALP-positive cells located at the periosteal bone surface. TRAP-positive cells were mononucleated and lacked a ruffled border. They appeared either as cell aggregates (resembling the shape of multinucleated giant cells) or as flat cells (resembling bone lining cells). Problems of osteoclast identification in bony fish are discussed.     

Skeletal morphology of the mud star,Ctenodiscus crispatus (Echinodermata: Asteroidea)

The morphology of the following eight major ossicle types is described and illustrated for the goniopectinid asteroid Ctenodiscus crispatus: terminal plates, superomarginal and inferomarginal ossicles, adambulacral and ambulacral ossicles, odontophores, oral intermediate plates, and superambulacral ossicles. Development, variation, and relationships with soft body-parts and with other ossicles are embphasized. Each ossicle type is distinguished by numerous structures related to its function and to articulation with adjoining skeletal elements. Because major structures (such as pustules, alveoli, and articulation surfaces) distinguishing ossicle types develop early during ontogeny, immature ossicles are readily identifiable. However, changes in form and orientation of these structures occur during ossicle growth. Ontogenetic changes are influenced by development of associated skeletal and soft parts. Ambulacral and adambulacral ossicles near the peristome are highly modified but retain the basic characteristics of structure and orientation which define these major types of skeletal elements.     

Cationic distribution in vessels, heart, skeletal muscle, and blood of normotensive (NWR) and spontaneously hypertensive rats (SHR)

Electrolytic analysis carried out with the atomic absorption spectrophotometer permitted to compare the cationic (Ca2+, Mg2+, Zn2+, K+ and Na+) tissue content of spontaneously hypertensive male rats (SHR) and normotensive male Wistar rats (NWR). In all SHR-tissues, Zn2+ is augmented, but mainly in the atria (left atrium), inferior vena cava, left ventricle, and skeletal muscle. The inferior cava vein and the right atrium have a similar, accentuated high bivalent (Ca2+, Mg2+, and Zn2+) cation content; Ca2+ and Mg2+ is present in a minor content in the right ventricle and type II "pale" skeletal muscle while only Mg2+ was also reduced in the left atrium, aorta, and the whole blood. A higher K+ concentration is seen in the right atrium, aorta, and type I "red" skeletal muscle. In the aortic wall and the whole blood a higher Na+- tissue content is found, confirming earlier observations. There is a discret water retention in the tissues from the left ventricle and skeletal muscle, simultaneously with a small depletion in the whole blood. We have concluded, that there is a specific cationic profile in SHR structural and functional different cardiovascular tissues, including skeletal muscle. The cationic tissue distribution is related to underlying genetic and adaptative factors and may be involved into specific drug effects.     

Production, crystallization, and preliminary X-ray analysis of rabbit skeletal muscle troponin complex consisting of troponin C and fragment (1-47) of troponin I.

Troponin is a ternary protein complex consisting of subunits TnC. TnI, and TnT, and plays a key role in calcium regulation of the skeletal and cardiac muscle contraction. In the present study, a partial complex (CI47) was prepared from Escherichia coli-expressed rabbit skeletal muscle TnC and fragment 1-47 of TnI, which is obtained by chemical cleavage of an E. coli-expressed mutant of rabbit skeletal muscle TnI. Within the ternary troponin complex, CI47 is thought to form a core that is resistant to proteolytic digestion, and the interaction within CI47 likely maintains the integrity of the troponin complex. Complex CI47 was crystallized in the presence of sodium citrate. The addition of trehalose improved the diffraction pattern of the crystals substantially. The crystal lattice belongs to the space group P3(1)(2)21, with unit cell dimensions a = b = 48.2 A, c = 162 A. The asymmetric unit presumably contains one CI47 complex. Soaking with p-chloromercuribenzenesulfonate (PCMBS) resulted in loss of isomorphism, but enhanced the quality of the crystals. The crystals diffracted up to 2.3 A resolution, with completeness of 91% and R(merge) = 6.4%. The crystals of PCMBS-derivative should be suitable for X-ray studies using the multiple-wavelength anomalous diffraction technique. This is the first step for elucidating the structure of the full troponin complex.