bluuebunny/crossref_metadata_2025 huggingface dataset:
Exogenous GnRh overrides the endogenous annual reproductive rhythm in green iguanas, Iguana iguana
Authors: Phillips John A. , Frye Fred , Bercovitz Arden , Calle Paul , Millar Robert , Rivier Jean , Lasley Bill L.
Published: June 2005

Female green iguanas, Iguana iguana, were caught in Belize, Central America (17°N), in December, at the onset of seasonal gonadal activity. The animals were immediately transferred to San Diego (32°N). Ovarian follicular development continued, with peak plasma hormone levels measured in January and February; 200 pg/ml for progesterone (P) and 800 pg/ml for total estrogens (Et = estradiol [E2] + estrone [E1]). E2 was the predominant estrogen throughout the cycle. Follicular atrophy was indicated in April with circulating progesterone and estrogen levels decreasing to baseline (refractory phase) levels (P = 20 pg/ml; Et = 50 pg/ml). Approximately midway through the refractory phase of their annual reproductive cycle (late May), either the D‐Arg6 analog of Chicken II or mammalian GnRH was administered via intraperitoneal osmotic pumps for 14 days to nine females. The analog of chicken II induced a fivefold increase in total circulating estrogens within 3–4 days after induced a fivefold increase in total circulating estrogens within 3–4 days after implantation. Both continuous and pulsatile delivery of the chicken II analog produced a similar pattern of steroidogenic response. A radical sham control animal showed no increase in steroidogenesis. Mammalian GnRH produced a pattern of similar duration, although the magnitude of the steroidogenic response was only half that produced by the chicken II analog. Estrogen titers approached baseline levels in all treatment groups two days after treatment ceased. Progesterone levels increased in all treatment groups during the delivery of exogenous GnRH, although the increases were not consistent. Untreated male cagemates housed with treated females exibited increased territoriality, courtship behavior, and mating, which began on day 4 or 5 of the treatment period. The control female was not courted by its male cagemate.
Immunofluorescent localization of the Ca2+‐dependent proteinase and its inhibitor in tissues of Crotalus atrox
Authors: Kleese William C. , Goll Darrel E. , Edmunds Tim , Shannon John D.
Published: June 2005

The native 108,000 dalton Ca2+‐dependent proteinase (CDP) and its 115,000 dalton protein inhibitor (CDPI) were purified from bovine skeletal muscle using native polyacrylamide gel electrophoresis and were used to elicit antibody production in rabbits and BALB/c mice. Polyclonal antibodies were purified as IgG fractions by column chromatography; monoclonal antibodyies were produced by the hybridoma technique. Indirect immunofluorescence localizations of CDP and CDPI in tissues of Crotalus atrox show both proteins to be ubiquitous. Both occur in the cytoplasm and are absent from the cell membrane and the nucleus; CDPI is also present in the I‐band of skeletal muscle.
Characterization of renal prolactin‐binding sites of two amphibians (Ambystoma tigrinum and Rana catesbeiana) and a reptile( Pseudemys scripta elegans)
Authors: Tarpey John F. , Nicoll Charles S.
Published: June 2005

The specific binding of 125I‐ovine prolactin (oPRL) to renal membrane preparations from Pseudemys scripta elegans, Ambystoma tigrinum, and Rana catesbeiana was characterized. All three membranes showed specific oPRL binding that was dependent upon time, temperature, pH, and membrane concentration. Scatchard analysis of the binding of 125I‐oPRL revealed high‐affinity sites with KA values of 2.1 × 1010, 3.6 × 1010, and 1.25 × 1010 M−1 for kidney membranes of Pseudemys, Ambystoma, and Rana, respectively. In addition, there was a low‐affinity site on the Ambystoma membranes. The binding capacities ranged from 31 to 70 fmol/mg of membrane protein. The hormonal specificity of these membranes was studied by competing increasing amounts of oPRL, human growth hormone (hGH) derived from recombinant DNA techniques, bovine (b) GH, and human placental lactogen (hPL) with 125I‐oPRL. The oPRL standard and hGH were the most potent competitors in all three assays, although hGH was not as potent as the oPRL. Human PL was moderately active in the turtle kidney assay, weakly active in the Ambystoma radioreceptor assay, and inactive in the bullfrog assay. Bovine GH had low potency on the turtle membranes and was inactive in the amphibian assays. The results of these studies indicate that the characteristics of renal PRL receptors of Pseudemys, Ambystoma, and Rana are similar to those of lactogenic receptors throughout the vertebrates. In addition, these data provide evidence for the first time of renal PRL receptors in a reptile.
Seawater‐acclimation and the thyroidal response to thyrotropin in juvenile coho salmon (Oncorhynchus kisutch)
Authors: Specker Jennifer L. , Kobuke Louise
Published: June 2005

Juvenile coho salmon (Oncorhynchus kisutch) were acclimated to seawater beginning at one of two stages of development. The first group was seawater‐acclimated before the 1‐ to 2‐month period of elevated plasma thyroxine (T4) concentration associated with the parr to smolt transformation (smoltification). The second group was seawater‐acclimated midway through the period of elevated T4 levels. Two measures of thyroid activity were used to compare seawater‐acclimated salmon with their controls remaining in fresh water. The baseline concentrations of thyroid hormones were assayed throughout a 9‐ or 10‐week period, and at the end of 9 or 10 weeks the responsiveness to a single injection of 0.2 IU bovine thyrotropin was measured. Transfer to seawater before the smoltification‐associated T4 elevation prevented the rise in thyroid hormone concentrations and reduced the response to bovine thyrotropin. Transfer of salmon during the period of elevated T4 levels did not change thyroid hormone concentrations; however, the response to thyrotropin was greater in seawater‐acclimated fish than in fish still in fresh water. We conclude that the longterm response of the thyroid to seawater acclimation is partly a function of the stage of development at the time of transfer.
Cell size in triploid salmon
Authors: Small Susan A. , Benfey Tillmann J.
Published: June 2005

There is a uniform increase in nuclear and cell size of erythrocytes, leukocytes, brain cells, and retinal cells in triploid coho and Atlantic salmon. In spite of the potentially drastic effects of this on their physiology and behavior, triploid fish appear to function as successfully as normal fish under aquaculture conditions. Triploid fish are easily produced, reared, and identified and should make useful tools for basic and applied research.
Luteal progesterone production and regulation in the viviparous dogfish, Squalus acanthias
Authors: Tsang Paul , Callard Ian P.
Published: June 2005

Although vertebrate viviparity has evolved in parallel on several occasions, the endocrine function of the post‐ovulatory follicle (corpus luteum) is only well understood in eutherian mammals. In this report we present evidence that the corpus luteum is a phylogenetically ancient endocrine organ, which secretes progesterone in response to pituitary hormones in a representative of the oldest extant jawed vertebrate (the dogfish, Squalus acanthias). Corpora lutea from pregnant aplacental viviparous spiny dogfish, Squalus acanthias, were removed, weighed and incubated in Eagle's Basal Medium supplemented with glutamine and urea for between 4 and 8 h at 18°C in 95% oxygen and 5% carbon dioxide. Corpora lutea decreased in weight from early to late pregnancy. Using radioisotopic methods, progesterone was definitively identified as a product of stage C corpora lutea. Basal progesterone synthesis, measured by radiommunoassay, increased from 301 ± 11.8 pg/mg/ corpus luteum/4 h in stage A of pregnancy to 1,881 ± 162.7 at stage B, and decreased markedly as corpora lutea decreased in size (stage C, 179 ± 19.2; stage D, 12 ± 3.2). An aqueous extract equivalent to one twenty‐fifth of the gonadotropic ventral lobe of the pituitary markedly stimulated progesterone in the incubates of stage C corpora lutea but not other stages. Corpus luteum function as an endocrine organ appears to be an ancient mechanism which antedates the appearance of terrestrial vertebrates.
Solute effects on the glycine cleavage system of two osmoconformers (Raja erinacea and Mya arenaria) and an osmoregulator (Pseudopleuronectes americanus)
Authors: Moyes Christopher D. , Moon Thomas W.
Published: June 2005

The effects of several organic solutes on the activity of the mitochondrial enzyme complex known as the glycine cleavage system (GCS) were examined in two osmoconforming species (soft‐shelled clam, Mya arenaria and little skate, Raja erinacea) and an osmoregulator (winter flounder, Pseudopleuronectes americanus). Taurine, beta‐alanine (BALA), and proline, compatible solutes which typically have no effect on enzyme function, significantly affected GCS activity with pronounced interspecific differences in sensitivity evident. Urea had no effect on skate GCS activity but significantly inhibited the enzymes from clam and flounder. The methylamines, trimethylamine oxide (TMAO) and betaine, did not significantly affect GCS from any species examined. Sarcosine significantly inhibited GCS from all three species, although this response may be of a regulatory nature. Inhibition of flounder GCS by urea was overcome by TMAO when these solutes were added in a 2:1 ratio (urea:TMAO). Pronounced inhibition of skate GCS occurred in response to urea + TMAO (2:1), whereas neither solute added alone had an effect on GCS activity. Although significant interspecific differences in solute effects occurred, each osmoconforming species was less sensitive to its own native osmolytes. BALA, however, stimulated skate GCS and may be important in enhancing sarcosine oxidation during hypoosmotic stress. The potential physiological significance and biochemical implications of these solute effects are discussed.
Developmental fate in chimeras derived from highly asynchronous murine blastomeres
Authors: Prather Randall S. , First Neal L.
Published: June 2005

The developmental fate of single blastomeres from eight‐cell murine embryos reaggregated with intact two‐cell embryos was evaluated after culture. Fluorescein isothiocynate was used to follow developmental fate in preblastocyst chimeric embryos. Expression of stage‐specific embryonic antigen 3 was used to assay developmental fate at the blastocyst stage, and glucosephosphate isomerase variants were used to assay at the blastocyst stage and after implantation. The results suggest that the descendents of the 1/8 component stay in a patch area and do not selectively migrate to the inner cell mass (ICM). This is in contrast to many studies that indicate that smaller blastomeres, which are more advanced in development, migrate to the ICM. The differences in experimental designs are discussed. Possible mechanisms for this phenomena are that the eight‐cell blastomere is physically excluded from the ICM by position or polarization, or that it is differentiating ahead of the two‐cell component and becomes trophectoderm.
Gene transfer in swine embryos by injection of cells infected with retrovirus vectors
Authors: Petters Robert M. , Shuman Ruth M. , Johnson Bryan H. , Mettus Richard V.
Published: June 2005

Key components for gene transfer to swine embryos using an avian retrovirus are described. A replication‐defective reticuloendotheliosis (REV) viral vector can infect and be expressed in pig embryo fibroblasts (PEF). Infection with a replication‐competent vector (REV‐A) indicates a presumptive block to viral replication in PEF. Swine embryos obtained at the morula stage can be cultured in vitro to the blastocyst stage, injected with retrovirus helper cells or quail embryo fibroblasts producing REV, and transferred to recipient swine with survival to at least 6 wk of gestation.
Isolation of viable mouse primordial germ cells by antibody‐directed flow sorting
Authors: McCarrey John R. , Hsu Katharine C. , Eddy E. M. , Klevecz Robert R. , Bolen James L.
Published: June 2005

The germ line represents a cell type of unique interest in mammals because it retains complete genotypic totipotency while undergoing significant phenotypic differentiation. Analysis of the mechanism that underlies the maintenance of this totipotency requires the ability to isolate and study all stages of the germ cell lineage. The primordial germ cells (PGC) are the earliest identifiable germ cells in the embryo. It has not previously been possible to isolate PGC in sufficient numbers and purity to facilitate biochemical and/or molecular analysis. We report here that the use of a monoclonal antibody in combination with flow cytometry does permit the isolation of reasonably large and pure yields of viable mouse PGC.
The relationship between plasma proteins and intracellular free amino acids during osmotic regulation in Carcinus maenas
Authors: Zatta Paolo
Published: June 2005

The osmoregulatory behavior of the Adriatic population of the shore crab Carcinus maenas is similar to that of more northern populations. Hyperosmotic regulation of the hemolymph is initiated below about 95–85% seawater (973–871 m Osm/kg) and slight hypo‐osmotic regulation appears to occur above approximately 110% seawater (1,127 m Osm/kg). Extracellular volume is actively regulated, and changes in levels of amino acids imply effective regulation of intracellular volume. Serum protein levels increase immediately after transfer of the crabs from high to low salinity, although the changes are much smaller than those reported by other investigators. Levels of free amino acids in serum decrease after transfer from high to low salinity and labeled leucine is incorporated into hemocyanin at a greater rate after such a transfer than in the absence of a change in salinity. However, the rate of incorporation of leucine is also greater at low than at high ionic strength in vitro, in cell‐free preparations. This result suggests that the experiment does not demonstrate storage of intracellular free amino acids by the extracellular oxygen carrier, but rather reflects “passive” binding to hydrophobic sites on the hemocyanin molecule.
Full‐term development of mouse blastomere nuclei transplanted into enucleated two‐cell embryos
Authors: Tsunoda Y. , Yasui T. , Shioda Y. , Nakamura K. , Uchida T. , Sugie T.
Published: June 2005

The nuclei from four‐ and eight‐cell mouse embryos were transplanted into enucleated two‐cell embryos. It was found that such embryos not only developed to the blastocyst stage in vitro (72% and 35%), but also developed to full term (22% and 8%) after transfer to recipient mice. However, development of embryos which contained nuclei from the inner cell mass was not observed. Since the development of enucleated zygotes which contain advanced nuclei is limited (the present study; McGrath and Solter: Science, 226: 1317–1319, '84; Robl, Gilligan, Critser, and First: Biol Reprod., 34:733–739, '86), it appears that cytoplasmic factors are important for the development of nuclei from advanced cells.
Appearance and developmental kinetics of a unique cell type in Dictyostelium discoideum: Is it the gamete phase of sexual development?
Authors: O'Day Danton H. , McConachie David R. , Rivera Jiji
Published: June 2005

A unique cell type appears during sexual development in Dictyostelium discoideum. These tiny ameboid cells migrate actively making both short‐lived and stable contacts with other cells. When fixed and stained with Hoechst 33258, each of these small amebae is seen to possess a condensed fluorescent nucleus. The cell type demonstrates a transient appearance in mated cultures appearing prior to 8 h and increasing to a peak of about 16% of the cell population by 10 h before decreasing in number. The decrease is correlated with the formation of pre‐zygotic, binucleate cells. In single strain cultures, the small cells are present at 8 h and rise to a plateau by 10–12 h but do not decrease in number, and pre‐zygotic binucleate cells do not form. The events of cell fusion were observed directly using time‐lapse video microscopy. The unique structure, developmental kinetics, and behavior of the cell type suggest that it may be the gametic phase of D. discoideum.
Lingual responces to chemical fractions of urodaeal glandular pheromone to the skink Eumeces laticeps
Authors: Cooper William E. , Grastka William R.
Published: June 2005

Chemical fractions of the female sex pheromone of Eumeces laticeps were extracted from whole urodaeal glands and bioassayed for their ability to induce elevated tongue‐flicking rates in conspecific males. Females were treated with estradiol to maximize pheromone production before removal of the urodaeal glands; males were implanted with testosterone in Silastic capsules to maximize responsiveness to the pheromone. Males emitted signifi‐cantly higher tongue‐flick rates in response to the neutral lipid fraction of urodaeal homogenate than to acidic lipids, basic lipids, or a peanut oil control. No significant differences in ability to induce tongue‐flick rates were observed between the carbohydrate fraction of urodaeal homogenate, the protein fraction, and deionized water. Thus, pheromonal activity appears to reside in the neutral lipid fraction, but in none of the others tested.
Reptilian Skeletal Muscle: Contractile properties of identified, single fast‐twitch and slow fibers from the lizard Dipsosaurus dorsalis
Authors: Gleeson Todd T. , Johnston Ian A.
Published: June 2005

Contractile properties and innervation patterns were determined in identified single fibers from the iliofibularis muscle of the desert iguana, Dipsosaurus dorsalis. Single fibers from both the red and white regions of the iliofibularis muscle were dissected along their length under oil and a portion was mounted on transducers for determination of maximum isometric tension (Po) and unloaded shortening velocity (Vmax) using the slack test method. Fibers were chemically skinned and activated by high Ca++. The remaining portion of the muscle fiber was mounted on a glass slide and histochemically treated to demonstrate myosin ATPase activity. Fibers studied functionally could therefore be classified as fast or slow according to their myosin ATPase activity, and they could also be classified metabolically according to the region of the muscle from which they were dissected. Fast‐twitch glycolytic (FG) fibers from the white region and fast‐twitch oxidative, glycolytic (FOG) and slow fibers from the red region had shortening velocities at 25°C of 7.5, 4.4, and 1.5 l·s−1, respectively. Po did not differ in the three fiber types, averaging 279 kN·m−2. In a second experiment, 10 pm sections were examined every 30 pm through the proximal‐most 7.5 mm of the iliofibularis muscle for motor endplates. Sections were stained to demonstrate regions of acetylcholin‐esterase activity. Fibers with visible endplates were classified in serial sections by histochemical treatment for myosin ATPase and succinic dehydrogenase. All slow fibers examined (n = 22) exhibited multiple endplates, averaging one every 725 pm. All FOG and FG fibers (N = 10) had at most one endplate in the same region. The data suggest that fibers histochemically typed ‘slow’ in Dipsosaurus are a single population of slow‐contracting fibers, all of which are multiply innervated. The available data on these fibers are evaluated and an argument is made that this type of fiber in lizards is best described as ‘slow’ rather than ‘tonic,’ for although they are multiply innervated and capable of graded contractions, they have other characters that are common to fibers capable of a twitch.
Metamorphosis of the American eel, Anguilla rostrata LeSeur: III. Contractile characteristics of skeletal muscle
Authors: Egginton Stuart
Published: June 2005

Previous work has suggested a common migratory strategy among fishes may involve changes in recruitment pattern of skeletal muscle types, allowing fast muscle to function continuously. In this study no evidence was found for changes in adenine nucleotide metabolism, thought to be important in fatigued muscle, with metamorphosis from the nonmigratory yellow to migratory silver eel in either slow or fast muscle tissue. Myofibrillar ATPase activity was found to be lower than reported values for other teleosts, around 0.075 and 0.17 μM inorganic phosphate mg−1 min−1 for slow and fast muscle, respectively. No change was found in the Ca+ +‐kinetics of the enzyme within either muscle type. Likewise, no change in the contractile performance of fast muscle was evident, arguing against changes in activity pattern. In contrast to mammalian endurance exercise training where major changes in aerobic capacity occur in fast muscle, migratory pre‐adaptation in eel appears to be restricted to changes in slow muscle performance. A displacement of the slow muscle force‐velocity curve to the right upon metamorphosis results in 30% increase in the tension developed at maximal power output from 2.4 to 3.2 N cm−2. The difference in migratory potential between yellow and silver eels was shown previously to involve an increased aerobic capacity. The change in contractile characteristics may further improve endurance by permitting a portion of the tissue to periodically replenish endogenous energy stores.
Hormonal control of gluconeogenesis in rainbow trout hepatocytes: Regulatory role of pyruvate kinase
Authors: Petersen T. D. P. , Hochachka P. W. , Suarez R. K.
Published: June 2005

Gluconeogenesis from lactate in hepatocytes from rainbow trout is activated by glucagon and inhibited by insulin in a dose‐dependent fashion. The maximal responses to both hormones occur within their probable physiological concentration ranges. Gluconeogenic activation by glucagon is accompanied by a profound inhibition of pyruvate kinase activity. This is reflected in increases in both the S0.5 and nH for phosphoenolpyruvate and increased sensitivity to inhibition by MgATP compared to the enzyme isolated from cells not treated with hormone (control). Gluconeogenic inhibition by insulin is accompanied by activation of pyruvate kinase activity. This is observed as decreases in both the S0.5 and nH for phosphoenolpyruvate and a decreased sensitivity to inhibition by MgATP compared to enzyme isolated from control cells. These results indicate that insulin and glucagon are involved in the regulation of hepatic gluconeogenesis in trout and that their effects are mediated, at least partly, through the control of pyruvate kinase activity.
Glutamine synthetase in liver and brain tissues of the holocephalan, Hydrolagus colliei
Authors: Ritter Nadine M. , Smith Darwin D. , Campbell James W.
Published: June 2005

Glutamine synthetase was shown to be mitochondrial in liver tissue and cytosolic in brain tissue of the holocephalan Hydrolagus colliei. The liver mitochondrial form has an Mr approximately 1,500 larger than the brain cytosolic form, so the two represent compartmental isozymes. Similar isozymes have been shown to be present in elasmobranchs, and their occurrence in holocephalans suggests the two groups share a common ancestry.
Behavior of “Accessory” somatic and sperm nuclei introduced into physiologically polyspermic newt eggs
Authors: Yamasaki Hisashi , Iwao Yasuhiro
Published: June 2005

In an attempt to understand the mechanism of the accessory sperm nuclei degeneration in the physiologically polyspermic eggs of the newt Cynops pyrrhogaster, nuclei from homologous somatic cells or from sperm of an anuran, Bufo japonicus, were injected into fertilized eggs of Cynops. Homologous blastula nucleus injected into eggs 1–2 hours after fertilization, behaved in synchrony with the zygote nucleus, although all the fertilized Cynops accessory sperm nuclei underwent degeneration. When 1–24 Bufo sperm treated with 0.025% Triton X‐100 were injected into individual Cynops eggs 1–1.5 or 3.5–4 hours after fertilization, none degenerated; they either joined the mitotic plate or the zygote nucleus, or they formed additional mitotic apparatus. However, when the Cynops eggs were inseminated with a higher concentration of sperm from Hynobius nobulosus or H. retardatus, several sperm penetrated each egg, followed by the formation of a zygote nucleus or multipolar spindles. A high frequency of the eggs that were injected with either blastula‐, Bufo sperm‐ or Hynobius sperm‐nuclei underwent an irregular multipolar cleavage. These results suggest that the accessory sperm‐suppressing activity in the cytoplasm of physiologically polyspermic Cynops egg affects only homologous sperm nuclei in a specific way.
Characterization of human sperm plasma membrane: Glycolipids and polypeptides
Authors: Mack S. R. , Zaneveld L. J. D. , Peterson R. N. , Hunt W. , Russell L. D.
Published: June 2005

The plasma membranes from ejaculated human spermatozoa were removed by nitrogen cavitation (600 PSI for 10 min) and isolated by centrifugation followed by a discontinuous sucrose density gradient centrifugation. Glycolipid analysis of the plasma membrane revealed a three‐fold enrichment in gangliosides: GM3 and GD1a/GD1b and neutral glycolipids: globoside and sulfatide as compared to that of whole human sperm. Two dimensional electrophoresis of human sperm plasma membranes revealed about 75 polypeptides. Several of these polypeptides were similar in migration and in display of shape and color to that found in boar sperm plasma membranes.
Development and keratinization of the epidermis in the common lizard, Anolis carolinensis
Authors: Carver Wayne E. , Sawyer Roger H.
Published: June 2005

Epithelial‐mesenchymal interactions play important roles in the development of the vertebrate integument with its diverse appendages. As a result of these interactions, specific morphogenetic events occur which result in the formation of distinct epidermal appendages. Following the early morphogenetic events involving cell proliferation and movement, other developmental events such as stratification, histotypic differentiation, and terminal cytodifferentiation occur in the epidermis. Using the common lizard Anolis carolinensis, we are seeking to obtain a better understanding of the relationship between the various developmental events and the expression of α and β keratins, with the aim of eventually understanding the mechanisms by which tissue‐specific keratinization patterns are established in the integument. As a first step, we have used immunoblot analyses and indirect immunofluorescence procedures with antisera specific for either α or β keratins to determine the temporal and spatial appearance of these keratins at specific developmental stages. We have found that: 1) There are relatively low molecular weight α keratin polypeptides present in the epidermis early in development as morphogenesis is taking place. 2) After morphogenesis occurs and histogenesis is well under way, the α keratins which characterize the adult epidermis appear. 3) Only α keratins are found in the basal cells of all regions of the epidermis. 4) β keratins are found only in the suprabasal layers of well‐developed scales and show region‐specific distribution in overlapping scales.
Tyrosinase activity and the expression of the agouti gene in the mouse
Authors: Movaghar M. , Hunt D. M.
Published: June 2005

Tyrosinase activity at the time of phaeomelanin synthesis in neonatal mice is lower in agouti than in black skin and hair bulb tissue, and this depressed activity is associated with a reduction in the electrophoretically distinct de novo form of the enzyme. Direct chemical measurements of sulphydryl compounds show elevated levels in agouti hair bulb tissue at this stage of development. The addition of exogenous copper to hair bulb extracts raises the activity of tyrosinase in agouti to approximately the black level but has no effect on black itself. These results are discussed in relation to the role of sulphydryl compounds and copper availability in regulating tyrosinase activity and turnover.
Ultrastructural localization of lysosomal enzymes in the egg cortex of Brachydanio
Authors: Hart Nathan H. , Wolenski J. S. , Donovan M. J.
Published: June 2005

The localization of acid phosphatase (E.C. 3.1.3.2), inorganic trimetaphosphatase (E.C. 3.6.1.2), and aryl sulfatase (E.C. 3.1.6.1) in the cortex of unactivated and activated eggs of Brachydanio was examined by ultrastructural cytochemistry. Using a lead capture method, activity for all three acid hydrolases was demonstrated in organelles of the cortex before and after egg activation. Acid phosphatase (AcPase) reaction product was consistently present in primary lysosomes, secondary lysosomes, multivesicular bodies, and yolk bodies. AcPase activity was absent from mitochondria, profiles of the endoplasmic reticulum, coated pits of exocytosed cortical granules, and coated vesicles. Although most cortical granules of the mature, unactivated egg were unreactive for this enzyme, a few showed AcPase reaction product. It is not clear whether the AcPase‐positive granules might be an immature form of cortical granules or a subpopulation of these organelles with lysosomal properties. Most cisternae of the Golgi apparatus did not stain for AcPase; however, reaction product was occasionally localized in a single cisterna as well as several small vesicles at the inner face of the Golgi. The intensity of the reaction product and the pattern of distribution of trimetaphosphatase (TmPase) activity was very similar to that of AcPase. However, TmPase was never observed in cortical granules. Cortices of unactivated and activated eggs showed less overall aryl sulfatase (ArSase) activity when compared with AcPase and TmPase. The presence of ArSase reaction product in lysosomes and multivesicular bodies confirmed the acid hydrolytic nature of these organelles. AcPase and TmPase, and to a lesser extent ArSase, are adequate markers of a cortical lysosomal system in the danio egg. Our results are consistent with the view that membrane, retrieved from the egg surface during the cortical reaction and shuttled into organelles with lysosomal properties (i.e., secondary lysosomes and multivesicular bodies), undergoes degradation. However, our study does not exclude other possibilities for handling internalized membrane, including storage and recycling to the plasma membrane.
Separation of vanadocytes: Determination and characterization of vanadium ion in the separated blood cells of the ascidian, Ascidia ahodori
Authors: Michibata Hitoshi , Hirata Junko , Uesaka Michiko , Numakunai Takaharu , Sakurai Hiromu
Published: June 2005

Ascidian blood cells contain a level of vanadium ion much higher than that in seawater. Blood cells are generally classified into six to nine types based on their morphology. On the basis of a Ficoll gradient for cell fractionation, neutron activation analysis of vanadium ion, and ESR spectra of oxovanadium ion in blood cells, the signet ring cells are the most likely “vanadocytes” in Ascidia ahodori. The vanadyl ion (VO2+), detected in high levels in these cells, binds with some low molecular weight substances through their oxygen donors.
MSORTV imaging of electrotonic conduction in a syncitium: Optical recording of polarization spread in a simple salivary gland
Authors: Senseman D. M. , Horwitz I. S. , Salzberg B. M.
Published: June 2005

The spread of hyperpolarizing current injected into the salivary gland of the snail Helisoma trivolvis was studied with the technique of multiple site optical recording of transmembrane voltage (MSORTV). Using MSORTV it was possible to monitor simultaneously the spread of current passed by an intracellular microelectrode in up to 124 separate gland regions. Our results demonstrate the feasibility of combining conventional intracellular injection techniques with MSORTV to visualize 2‐dimensional current spread in an electrical syncitium.
Comparative studies of Bufo and Xenopus vitelline coat molecular transformations induced by homologous and heterologous oviducal pars recta proteases
Authors: Takamune Kazufumi , Lindsay Leann , Hedrick Jerry L. , Katagiri Chiaki
Published: June 2005

The vitelline coats (VCs) of Bufo japonicus and Xenopus laevis undergo coelomic egg (CEVC)‐ to uterine egg (UEVC)‐type conversions, after passage through the pars recta (PR) portion of oviduct or treatment of eggs with secretory granules from PR (PRGs), as determined by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) analyses of glycoproteins. These conversions include for Bufo the loss of 40K–52K components from the CEVC, concomitant with increasing stainability of a 39K component and the appearance of a 36K component in the UEVC. For Xenopus, a 43K glycoprotein in the CEVC is transformed to a 41K glycoprotein in the UEVC. When coelomic eggs of both Bufo and Xenopus were treated with PRGs from the heterologous species, the macromolecular components of the VCs were processed in exactly the same way as with homologous PRGs. The PRGs from both species showed strikingly similar hydrolytic activities against Boc‐Val‐Leu‐Lys‐MCA and several peptidyl‐Arg‐MCAs. Bufo coelomic eggs treated with Xenopus PRGs were not fertilizable by Bufo sperm, whereas treatment with Bufo PRGs rendered the eggs fertilizable. We conclude that the oviductal enzymes and VC substrates in these species are evolutionarily conserved and that macromolecular alterations of VC glycoproteins as detected by SDS‐PAGE are necessary, but not sufficient, for making coelomic eggs fertilizable.
Formation of cartilage in organ‐cultured leg buds of chick embryo
Authors: Suzuki Hiroaki R. , Ide Hiroyuki
Published: June 2005

When the leg buds from chick embryos at stages 17–23 were cultured as organ cultures in F12 medium, which contained chick serum and an extract of chick embryos, leg cartilage structures developed that included pelvic girdle, femur, tibiofibula, and tarsometatarsus. The proportion of explants with definite distal elements increased when the leg buds from later embryos were used. Supplementation with chick serum was essential. These results suggest that the pattern of formation of limbs in chick embryos can be studied in organ culture.
A Possible contractile role for myoepithelial cells in the trachea of the Australian snake‐necked tortoise, Chelodina longicollis
Authors: Smith R. V. , Hards D. K. , Satchell D. G.
Published: June 2005

The trachea of the Australian snake‐necked tortoise, Chelodina longicollis was studied using histological, immunohistochemical, ultrastructural and organ bath pharmacological techniques. The trachea has complete cartilage rings with dense connective tissue between them. No smooth muscle cells were found. Electrical stimulation of tracheal tube and strip preparations elicited marked contractile responses. Myoepithelial cells containing smooth muscle myosin and bundles of thin filaments were demonstrated between the basement membrane and the mucous and ciliated cells of the epithelium lining the tracheal lumen. These myoepithelial cells could account for the contractions observed and may be involved in the expulsion of mucus.
The effect of nonlimb tissues on forelimb regeneration in the axolotl, Ambystoma mexicanum
Authors: Tank Patrick W.
Published: June 2005

Previous studies with Notophthalmus viridescens have shown that forelimb stumps covered by flank skin do not regenerate when amputated through the graft of nonlimb skin [Tank, P.W. (1984) J. Exp. Zool., 229:143–153]. In an attempt to determine whether or not cells derived from nonlimb skin dedifferentiate and enter the regeneration blastema, four experimental series were prepared using triploid (3N) marked cells grafted into diploid (2N) hosts of the axolotl (Ambystoma mexicanum). In the first series, head skin was used to replace limb skin followed by amputation through the graft 30 days later. Fully 96% (24/25) of the limbs failed to regenerate an externally visible outgrowth and 4.4% of the blastemal mesenchyme cells were observed to be 3N based on the characteristic of three nucleoli per nucleus. In the second series, flank skin replaced limb skin and 38% (8/21) of the limbs failed to regenerate. The remaining limbs regenerated an assortment of hypomorphic, normal, abnormal, or multiple structures. Only 2.4% of the cells in the blastemal mesenchyme were observed to be 3N in the second series. In the third series, head dermis was implanted in place of limb musculature and 20% (3/15) of the limbs failed to regenerate. The other limbs in series three were either abnormal, hypomorphic, or multiple in morphology. Histological analysis revealed that 3.4% of the cells in the blastemal mesenchyme of the head dermis group were 3N. In the fourth series, flank dermis replaced limb musculature and only 7% (1/15) failed to regenerate. Ten limbs (66%) formed hypomorphic regenerates and the remainder were normal, abnormal, or multiple in morphology. Histological analysis revealed that 6.8% of the cells were 3N in the blastemal mesenchyme of limbs in series four. Sham‐operated controls for all four series regenerated normally and histological analysis of contralateral control limb regenerates revealed background 3N marker that averaged from 0.1 to 0.8%. The regenerate epidermis of limbs that were covered by 3N head skin or flank skin contained approximately 25% 3N cells, indicating that the epidermis was of nonlimb origin. The regenerate epidermis of limbs bearing 3N dermis implants did not contain marked cells. These results indicate that non‐limb cells are capable of undergoing dedifferentiation and entering the limb regeneration blastema. An explanation of regenerative failure is provided that is based on interference with short range cellular interactions by nonlimb cells.
A kinematic study of front legs' movements in walking Protura (Insecta)
Authors: Tichy Harald
Published: June 2005

High‐speed cine film of walking Protura shows that a) the forward projected legs 1 move almost parallel to the body; and b) their point of contact with the ground is only in front of the coxal articulation. The average stride length of legs 1 (0.05 mm) measures one‐third that of legs 2 and legs 3 (0.15 mm). These features indicate a relatively small contribution of the front legs in supporting and propelling the animal, which is accomplished by a pulling rather than a pushing action. On a horizontal rough surface of moist plaster, the animal moves by three fashions: a) it holds up the front legs and walks with the middle and rear legs, b) it places occasionally one or both front legs on the ground while walking with the middle and rear legs, and c) it uses all six legs in walking. In the latter case the average promotion duration of legs 1 (0.29 s) is twice that of legs 2 and 3 (0.15 s each), whereas the average remotion duration of legs 1 (0.08 s) is one‐half that of legs 2 and 3 (0.17 s). In addition, the movements of legs 1 are not coordinated with the other two pairs in the typical hexapode stepping pattern. Instead, they are used independently with a tendency to alternating up and down movements. These specializations of legs 1 reflect an exploratory role in walking. On a horizontal slippery surface of transparent paper, the animal walks exclusively with six legs. In all legs, the duration of the remotion period increases as the time elapses in a series of walks. In legs 2 and 3 the remotion duration is longer than that of promotion throughout the experiment, but in legs 1 it changes from shorter to longer. Timing of the front legs' movements does not fit in the hexapode rhythm. Legs 1 are both simultaneously and alternatingly used, thus playing a part in propelling and balancing the body.
Potassium channel blockers unmask electrical excitability of insect follicles
Authors: O'Donnell M. J.
Published: June 2005

Standard techniques for intracellular recording and current injection were used to determine if the developing ovarian follicles of six species of insect were electrically excitable. Follicles were dissected from the panoistic ovarioles of Locusta migratoria and Periplaneta americana, the polytrophic meroistic ovarioles of the fruit fly Drosophila rnelanogaster and the silkworm moths Hyalophora cecropia and H. Columbia, and the telotrophic meroistic ovarioles of the mealworm beetle Tenebrio mlitor. Follicles of all species except Tenebrio were inexcitable in control saline; Tenebrio follicles produced short action potentials which did not outlast the duration of the current pulse. Follicles of all species except Drosophila produced long duration overshooting action potentials when depolarized in salines containing the potassium channel blockers tetraethylammonium (TEA; 10–80 mM), Bazf (10–20 mM) and 4‐aminopyridine (4‐AP; 10 mM). Hyalophora follicles were also excitable in saline containing TEA and Ba2+ but not 4‐AP, and Tenebrio follicles were excitable in saline containing Ba2+ but not TEA and 4‐AP. Excitability in follicles of all species was unaffected by sodium replacement but was reversibly inhibited by the calcium channel blockers La3+ (1 mM), Co2+(10 mM), or verapamil (0.1‐1.0 mM). It is suggested that the action potentials are caused by the movement of calcium or barium through voltage‐dependent channels that are masked in control saline by a large counteracting potassium conductance. Excitability appears to be a property of the oolemma and not the follicle cell membranes. Possible functions of voltage‐dependent ion channels in insect oocytes are discussed.
The physiological basis of slow locomotion in chamaeleons
Authors: Abu‐Ghalyun Younis , Greenwald Lewis , Hetherington Thomas E. , Gaunt Abbot S.
Published: June 2005

The African chamaeleon, Chamaeleo senegalensis, will not move faster than ∼0.1 m/second at 23 °C, whereas the lizard Agama agama, like most lizards its size, runs at speeds more than 10 × as fast. To account for this difference, we measured various physiological parameters of the iliofibularis muscle of both lizards. The maximum speed of tetanic contraction of unloaded Chamaeleo muscle was half as fast as that of Agama muscle (2.5 vs. 5.8 resting lengths per second). Heavily loaded Chamaeleo iliofibularis contracted at nearly ¼ the speed of Agama muscle. Time to peak isometric twitch tension and time to half relaxation were twice as long in Chamaeleo as in Agama (122 vs. 58 msec, and 168 vs. 81 msec). Much more of the Chamaeleo muscle consisted of tonic muscle fibers, and the Chamaeleo muscle, compared to Agama muscle, showed physiological evidence of having a significant amount of tonic fibers (potassium contracture and high tetanus to twitch ratios). Finally, the myofibrillar ATPase activity of the Chamaeleo muscle was ⅓ that of Agama muscle. Thus, these results show that the slow locomotion of old world chamaeleons can, in part, be explained by the physiology, biochemistry, and fiber‐type distribution of their muscles.
Cellular retinoic acid binding protein: Detection and quantitation in regenerating axolotl limbs
Authors: McCormick Anne M. , Shubeita Huda E. , Stocum David L.
Published: June 2005

The concentrations of apo (unoccupied), holo (occupied), and total cellular retinoic acid binding protein (CRABP) were measured at various stages of axolotl limb regeneration. The ratio of apo‐CRABP to holo‐CRABP declined with advancing regenerate stage until the CRABP was all in the holo form. The increase in holo‐CRABP is correlated with a stage‐dependent shift in the effect of exogenous retinoic acid on regenerate pattern, from pattern duplication to inhibition of regeneration. The data suggest, though they do not prove, that these different morphological effects could be due to a shift from a CRABP‐dependent to a CRABP‐independent mechanism of exogenous retinoic acid (RA) action that is related to stage‐specific variations in endogenous RA levels.
Effects of theophylline on expression of the long cilia phenotype in sand dollar blastulae
Authors: Riederer‐Henderson Mary Ann
Published: June 2005

Previously, increases in ciliary length have only been obtained through genetic mutation in Chlamydomonas or by incubation of swimming echinoderm blastulae in trypsin or elastase. We have found that the phenotypic switch from short to long cilia on sand dollar blastulae can also be effected by incubation in theophylline. Cilia detached from control blastulae have a mean length of 21 ± 7 μm with 10% of the cilia being > 30 μm. Upon incubation in 10 mM theophylline additional long cilia appeared after 10 hours and by 24–32 hours 1/2–3/4 of the embryo was covered with long cilia. The percentage of long cilia increased to 65% with a mean length of 40.0 ± 17.6 μm. Incubation in other methylxanthines, such as aminophylline, caffeine, or isobutylmethylxanthine, inhibited development but had no effect on ciliary length distribution. Dibutyryl cAMP, 8‐bromoad‐enosine, and calcium ionophore also had no effect on ciliary length. Cyclic AMP levels were measured and showed only slight differences among controls and embryos incubated in trypsin, caffeine, or theophylline. These data suggest that theophylline may be altering ciliary length control through some mechanism other than elevations in cAMP.
Localization of cortical granule constituents before and after exocytosis in the hamster egg
Authors: Cherr Gary N. , Drobnis Erma Z. , Katz David F.
Published: June 2005

Electrical activation of the hamster egg was used to study cortical granule constituents before and after exocytosis. The activated hamster eggs underwent cortical granule decondensation just prior to and at the time of exocytosis. Some of the cortical granules of aged, unactivated eggs underwent similar changes. FITC‐ and gold‐conjugated Lens Culnaris agglutinin (LCA) bound intensely to the surfaces of activated but not unactivated eggs. This labelling was associated with the microvilli. Permeabilized eggs exhibited discrete cortical labelling before activation, with a subsequent decrease following the cortical reaction. Gold‐conjugated LCA specifically bound to cortical granules when incubated with thin sections. FITC‐soybean trypsin inhibitor (SBTI) bound in discrete foci in the cortex of unactivated eggs. Following activation, cortical labelling by SBTI decreased. Aprotinin and benzamidine hydrochloride inhibited FITC‐SBTI from binding to the egg cortex. Gold‐avidin localization of biotin‐SBTI in the electron microscope demonstrated that condensed cortical granules did not bind SBTI but decondensed or exocytosing granules did. This suggests that a cortical granule protease is exposed just prior to exocytosis. Activated eggs exhibited dramatic decreases in the number of hamster sperm penetrating the cytoplasm, suggesting that a plasma membrane block to polyspermy is temporally related to cortical granule exocytosis.
Properties of standard avian slow muscle grafts following long‐term regeneration
Authors: Hather Bruce M. , Hikida Robert S.
Published: June 2005

The tonic anterior latissimus dorsi (ALD) of adult pigeons was orthotopically homografted and evaluated after 11 months of regeneration for histological, histochemical, electromyographic (EMG), and mechanical properties. The resting EMG activity of the grafts was lower in amplitude than that of the controls, but showed the tonic pattern typical for these tonic muscles. The control and grafted muscles had a histochemically homogeneous population of fibers with moderate myofibrillar adenosine triphosphatase activity. Succinic dehydrogenase activity was moderate for the control muscles, but low for the grafts. The regenerated muscles had fewer and smaller fibers and had much larger intersynaptic distances. Both the regenerated and the contralateral control muscles were slow contracting and maintained tetanic tension for prolonged periods with direct electrical stimulation. The relaxation was slower in the grafted muscle than in the control. The grafts produced 40% of the maximum tension of the control muscles, but the rate of tension development was similar between the two groups. The results indicate that the tonic properties were regenerated, but the innervation pattern was altered and the grafted muscles did not have normal mature fibers even after long‐term regeneration.