bluuebunny/crossref_metadata_2025 huggingface dataset:
Subcellular differentiation arrested in Artemia embryos under anoxia: Evidence supporting a regulatory role for intracellular pH
Authors: Hofmann Gretchen E. , Hand Steven C.
Published: June 2005

Previous work has clearly documented that intracellular pH (pHi) mediates metabolic arrest during anaerobic dormancy in Artemia embryos. This study provides evidence that subcellular differentiation during preemergence development (PED) is regulated by transitions in pHi and also reports the first detailed ultrastructural description of PED in these embryos.

Mitochondrial maturation, elaboration of glycogen fields, and alteration in yolk platelet structure are clearly evident during aerobic development (embryo pHi ≥ 7.9). Mitochondrial maturation involves two populations of mitochondria that are spatially segregated in embryonic cells—one in the cytoplasm and the other located inside of yolk platelets. Identification of the latter population is based in part on histochemical staining of DNA. While both mitochondrial populations mature structurally during PED, enzymatic evidence indicates that the intra‐yolk platelet population is not functional.

Subcellular morphogenetic events observed during aerobic development are blocked when embryos are incubated under anoxia (pHi ⩽ 6.8) or aerobic acidosis (a quiescent state induced by artificially elevating CO2 to 60%; pHi = 6.8). Embryonic cells exposed to either of these experimental treatments for 12 h are essentially identical to cells of the undifferentiated, hour 0 embryo. Analogously, cytochrome c oxidase specific activity increases 230% over 12 h of aerobic development, but this ontogenetic increase is arrested by 12 h of anoxia or aerobic acidosis. Returning the acidified embryos to control aerobic conditions causes enzyme proliferation to resume at rates identical to aerobic, control values. A similar arrest of the proliferation of cytochrome c oxidase activity is seen under aerobic acidosis when CO2 is present at only 11%. Finally, alkalinizing the pHi of embryos under anoxia by adding ammonia to the medium results in a 37% increase in cytochrome c oxidase levels relative to those measured under anoxia. These data support pHi‐ dependent regulation of development in preemergence Artemia embryos.
Induction of locomotion in spinal tadpoles by excitatory amino acids and their agonists
Authors: Harrison Philip Hugh
Published: June 2005

Bath application of the excitatory amino acids L‐aspartate and/or L‐glutamate or their agonists N‐methyl‐D,L‐aspartate and/or kainate elicited swimming movements in spinal tadpoles. Swimming cycles induced by the amino acids were in the frequency range of natural movements, and could be evoked after sectioning all dorsal roots in the exposed spinal segments. Locomotion was only elicited by L‐aspartate or L‐glutamate at low concentrations when the bath medium was rapidly circulated over the exposed surface of the spinal cord, and was of much shorter duration than the agonist‐induced movements. These results indicate some differences between the actions of L‐aspartate and L‐glutamate and their agonists on the tadpole spinal cord.
Hemoglobin in the parthenogenetic eggs of Daphnia magna
Authors: Kobayashi Michiyori , Kato Shuichi , Ninomiya Masaki , Fujiki Masahiko , Igarashi Yoshihiko , Kajita Akihiko
Published: June 2005

The hemoglobin (Hb) concentration in a single parthenogenetic egg from Hb‐rich Daphnia magna was about three times that in an egg from a Hb‐poor animal. Hemoglobin‐rich animals first produced Hb‐rich eggs in air‐saturated water. The Hb concentration of the eggs gradually decreased, leading to a decrease in the Hb concentration in hemolymph in adult animals. The P50 values (equilibrium oxygen pressure at half saturation) of solutions of purified Hb were 2.9 and 7.2 torr at 20°C for Hb‐rich and Hb‐poor eggs, respectively. Hemoglobin from the eggs was separated into at least eight Hb components by isoelectric focusing. The pI values (isoelectric point) of the major Hb components of the eggs exceeded those of adult Hb. The relative amounts of Hb components with high pI values in Hb‐rich eggs were higher than those of Hb‐poor eggs. On sodium dodecyl sulfate‐polyacrylamide gel electrophoresis technique (SDS‐PAGE), the Hb from eggs was composed of 37,000 molecular weight subunits.
Homeostatic and developmental control of cell size and shape in an insect epithelium, the epidermis of Manduca sexta
Authors: Bultmann Hermann , Goodman Walter G.
Published: June 2005

In contrast to previously described transformations of insect epidermal cells, morphological changes in the larval epidermis of Manduca sexta involve large changes in cell volume: a threefold increase at the beginning and a twofold decrease at the end of the last feeding phase. The volume changes are determined, in part, or entirely, by changes in cell height (cell elongation and cell shortening). Initial cell elongation occurs in a region‐specific manner, whereas subsequent cell shortening affects all of the epidermis equally. As shown by ligation experiments and hormone treatments in tissue culture, larval changes in cell height, unlike cell elongation and differentiation in prepupae, are not regulated by developmental hormones (juvenile hormones, ecdysteroids). Instead, the maintenance of a normal columnar epithelium in fifth instars depends on continuous growth. Lack of food, especially protein, results in reversible cell shortening at any time during the feeding phase. Intake of water does not mitigate this cellular response; cell shortening is also insensitive to ouabain but inhibited by cold treatments. We propose that during larval growth epidermal cell height is under specific homeostatic control, independent of mechanisms regulating cell width, ploidy levels, or mitotic activity.
Heart induction in wild‐type and cardiac mutant axolotls (Ambystoma mexicanum)
Authors: Smith Steven C. , Armstrong John B.
Published: June 2005

We have re‐examined some of the factors affecting the induction of heart‐forming mesodern in the axolotl. The formation of functional, rhythmically contracting myocardial tissue was used as an assay. We have found that heart‐forming mesoderm is fully induced and capable of completing its developmental repertoire by the end of neurulation. As has been previously reported, pharyngeal endoderm appears to be the major inductor of heart mesoderm. Unlike previous workers, we have found that the inducing activity appears to be highly localized in the mid‐ventral pharyngeal endoderm. The endoderm retains its inductive properties, and the mesoderm retains at least some capacity to respond, long after the heart‐forming mesoderm is apparently fully induced.

We have also found that RNA extracts from pharyngeal endoderm, which are capable of causing cardiac‐lethal (c/c) mutant axolotl hearts to begin beating, are not capable of inducing early wild‐type heart‐forming mesoderm. Based on these results, we speculate that induction of heart‐forming mesoderm is a two‐step process. The first signal, occurring during neurulation, directs the mesoderm to begin differentiating into cardiomyocytes, and the second, beginning in mid‐ to late neurulation and continuing until just prior to the onset of heartbeat, causes myofibrillogenesis and the initiation of rhythmic contractions. The latter signal, which is lacking in c/c mutant embryos, appears to be necessary to override an inhibition present in the embryonic milieu.
Control of key carbohydrate‐metabolizing enzymes by insulin and glucagon in freshly isolated hepatocytes of the Marine teleost Hemitripterus americanus
Authors: Foster G. D. , Moon T. W.
Published: June 2005

The effects of scombroid insulin (10−7 M) and bovine glucagon (10−7 M) on key regulatory enzymes of glycolysis and gluconeogenesis are examined in freshly isolated hepatocytes of the sea raven, Hemitripterus americanus, following a 45 and 180 min incubation. Glucagon increased the Vopt of PEPCK, PK, GPT, and LDH, but decreased the substrate activity ratios of both PK and PFK‐1 (the latter only at inhibitory ATP concentrations) at 180 min. Glucagon also decreased Fru‐2,6‐P2 levels at both 45 and 180 min. Insulin decreased the Vopt for LDH at 180 min and decreased the F6P activity ratio for PFK‐1 at both 0.5 mM and 1.5 mM ATP at 45 and 180 min, with larger decreases occurring at the later time period (to 13% of the control value). Insulin also decreased Fru‐2,6‐P2 concentrations at 180 min and increased alanine uptake. PEPCK and PK were not affected by insulin. These data indicate that glucagon alters the characteristics of a variety of enzymes in the glycolytic/gluconeogenic sequence, while the major impact of insulin is at the level of PFK‐1 and alanine uptake.
Factors influencing the affinity and capacity of T3‐binding sites in hepatic nuclei of rainbow trout, Oncorhynchus mykiss
Authors: Bres O. , Cyr D. G. , Eales J. G.
Published: June 2005

The effects of several variables were tested on the in vitro binding affinity (Ka) and maximum binding capacity (MBC) of saturable 3,5,3′‐triiodo‐L‐thyronine (T3)‐binding sites (putative thyroid hormone receptors) extracted from hepatic nuclei of immature rainbow trout held on a 12L:12D photocycle at 12°C. Neither Ka nor MBC varied during the daily photocycle, nor were they influenced by the growth rate, food ration, or a 13‐fold chronic elevation in plasma T3. However, MBC was inversely correlated with fish size and was depressed significantly by two injections of 17β‐estradiol (E2) or starvation for 3 or 12 days. The Ka was uninfluenced in either case. Since both E2 and starvation also depress T3 levels in the blood, the decrease in MBC could be secondary to this change. The drug ipodate, which depresses plasma T3, also lowered the MBC. However, MBC remained low in ipodate‐treated trout given exogenous T3, thus suggesting that the effect of ipodate was not due to T3 deprivation. It remains to be shown, therefore, that plasma T3 depression itself can modify MBC. It is concluded that increasing body size and catabolic states, both of which are associated with a decrease in specific growth rate, tend to decrease the abundance of hepatic T3 receptors.
Purification and subcellular localization of Zn‐dependent acid p‐nitrophenylphosphatase in frog liver and comparison with other vertebrates
Authors: Panara F. , Massetti N. , Angiolillo A. , Fagotti A. , Pascolini R.
Published: June 2005

Zn2+‐dependent acid p‐nitrophenylphosphatase (Zn‐AcPase) from liver of Rana esculenta was purified to homogeneity. The purified enzyme moves as a single electrophoretic band at pH 8.3 in 7.5% acrylamide and was coincident with the enzyme activity. The enzyme has a molecular weight of 102,000 ± 5,000D and is a dimer with two apparently similar polypeptide chains of 48,000 ± 3,000D as determined by sodium dodecylsulfate gel electrophoresis. Zn‐AcPase from frog liver requires Zn2+ ions for catalytic activity; other bivalent cations have little or no effect. The enzyme with a pI of 7.07 does not appear to be a glycoprotein and was associated with the soluble fraction after liver cell fractionation.

The biochemical and molecular properties of frog liver Zn‐AcPase were compared with that of the enzyme partially purified from carp (Cyprinus carpio), pike (Esox lucius), and rat (Rattus norvegicus).
Nutritional state modulates hormone‐mediated hepatic glycogenolysis in chinook salmon (Oncorhynchus tshawytscha)
Authors: Klee Michael , Eilertson Carmen , Sheridan Mark A.
Published: June 2005

Juvenile chinook salmon were used to investigate the interaction of nutritional state and hormones (glucagon, epinephrine) on hepatic glycogenolysis. Experiments were conducted in vitro on liver removed from animals of varying nutritional states: fed 1 week, fasted 1 week, fed 3 weeks, fasted 3 weeks, and fasted 1 week/refed 2 weeks. Basal glycogen breakdown in liver isolated from continuously fed (1 week and 3 weeks) fish proceeded at insignificant levels, whereas glycogen breakdown in liver isolated from fasted animals progressed rapidly during the course of incubation. However, no difference in the pattern of glycogen breakdown between liver isolated from animals fasted for 1 week and those fasted for 3 weeks was observed. The presence of glucose in the incubation medium retarded glycogen breakdown in the livers from fasted fish. Both epinephrine and glucagon stimulated glucose release from liver isolated from fish fed continuously and from fish fasted 1 week; epinephrine appeared more potent in this regard than glucagon. Epinephrine also stimulated glucose release from liver isolated from fish fasted 3 weeks, whereas glucagon was without significant glycogenolytic effect on this tissue. Insulin inhibited epinephrine‐ and glucagon‐stimuluated glucose release. These results suggest that glucagon plays a role in modulating nutrition‐associated adjustments in metabolism during the early stages of fasting and that, with extended fasting, the liver loses its sensitivity to glucagon with regard to glycogenolysis.
Variability of the immune response in freshwater sponges
Authors: Van De Vyver G. , Holvoet S. , Dewint P.
Published: June 2005

The variability of the immune response of four species of freshwater sponges, common in Belgium, Ephydatia fluviatilis, Ephydatia mulleri, Spongilla lacustris, and Eunapius fragilis, has been analyzed at the cellular level.

For each species, strains, which are the collection of all the sponge individuals able to fuse compatibly their aquiferous system, were collected in natural ponds and isolated.

In most cases, the general immune response corresponds to a three‐step process restricted to the zone of contact. This process includes: the fusion of the sponges coming into contact, a local cell accumulation, and the separation of the alien sponges.

The variability of the immune reaction mainly concerns the existence and the duration of the fusion step and the possible secretion of a collagen barrier between the separated alien sponges. In contrast, for all the combinations studied, the accumulated cells differentiate and organize themselves in two facing layers of pinacocytes.

In some allocombinations, involving S. lacustris strain δ, a general reaction was recorded. In these pairings, after the initial fusion, the alien sponges undergo a temporary involution followed by the segregation and reconstitution of two separated functional sponges.

Our exhaustive analysis leads to the conclusion that the alloimmune responses vary according to the species and even within a single species, according to the strains paired. In contrast, all the xenocombinations undergo a similar process consisting in the direct and complete separation of the sponges.

These results demonstrate the possibility for freshwater sponges to diversify their responses to nonself and consequently to discriminate between different nonself.
Water channels and water canals in the gill of the freshwater mussel, Ligumia subrostrata: Ultrastructure and histochemistry
Authors: Kays W. T. , Silverman H. , Dietz T. H.
Published: June 2005

The apical surface of all epithelial cells of the gills of Ligumia subrostrata either are ciliated or contain microvilli or show both structures. The ciliated cells on the gill filaments direct water through the ostia at the base of each gill filament. All the water passing through the gills and out the exhalent siphon must pass through the narrow water canals and into the central water channels. All of the epithelial cells lining the lateral water canals are mitochondria‐rich. In addition, the central water channel epithelia contain clusters of mitochondria‐rich cells, but many of the epithelial cells are mitochondria‐poor. With the exception of the lateral ciliated cells, the lamellar epithelium of the surface filaments contains relatively mitochondria‐poor cells. The mitochondria‐rich epithelial cells of the water canals and water channel display the most histochemically intense reaction products for carbonic anhydrase and cytochrome oxidase. The mitochondria‐rich cells of the water canal and water channel epithelia may be the principal cells responsible for salt transport in these mussels.
Differential limb regeneration in diploid and triploid Rana pipiens larvae with reference to spinal motor neuron development
Authors: Pollack Emanuel D. , Maheras‐Rarick Helen M.
Published: June 2005

Developmental manipulations that can alter nerve‐limb relationships can assist in understanding the neural control of limb regeneration. Pressure‐induced triploidy in Rana pipiens tadpoles results in alterations of the quantitative characteristics of the spinal motor neurons that innervate the limbs, whereas the limbs appear unaltered. Unilateral midthigh amputations at larval stages IX, XI, and XIII of diploid and triploid animals resulted in complete regeneration for only stage IX animals regardless of ploidy. Nevertheless, triploid limbs regenerated much faster than did diploids, an event that can be related to the differential dynamics of nerve fiber extension and/or the altered numbers and sizes of triploid spinal motor neurons. Although normal limb development from stage IX to the endpoint at stage XVIII was the same in diploids and triploids, the rate of regeneration in triploids was nearly twice that of diploids. The data of this noninvasive means of altering the quantitative relationship of nerve‐to‐peripheral target suggest a unique means of studying nerve‐dependent limb regeneration in an animal that progressively loses its regenerative capability during development.
Effect of temperature on gonadal sex differentiation in the sea turtle Lepidochelys olivacea: An organ culture study
Authors: Merchant‐Larios H. , Villalpando I.
Published: June 2005

Gonadal sex differentiation is controlled by the temperature of egg incubation in the sea turtle Lepidochelys olivacea. The physiological mechanism involved in this phenomenon is not understood. Here, we studied the differentiation of gonads isolated from embryos at different stages of development which previously had been incubated at either male‐ or female‐producing temperatures (28°C or 32°C respectively) and cultured at the alternative temperature. We found that morphologically bipotential gonads can undergo sexual differentiation in vitro. Under these conditions the sexual differentiation of isolated gonads was not influenced by the temperature during culture. However when whole embryos were further incubated, the gonads remained sensitive to incubation temperatures for at least three more stages of development. These results suggest that an extragonadal physiological mechanism may mediate the effect of temperature on gonadal sex determination.
Seasonal shifts in odor acuity by starlings
Authors: Clark Larry , Smeraski Cynthia A.
Published: June 2005

A passerine bird, the European Starling Sturnus vulgaris, demonstrated a seasonal ability to respond to odor cues. Cardiac conditioned responses were most evident when birds were in breeding condition. Once birds were in nonbreeding condition, responding to odors all but ceased. For birds in breeding condition, threshold sensitivity to the odor cyclohexanone was comparable to levels reported for nonpasserine birds (0.3% vapor saturation or 3.778 × 1014 molecules/ml). In contrast, the threshold level during the postreproductive phase of the annual cycle was 10% vapor saturation (1.256 × 1016 molecules/ml). a level high enough to implicate the trigeminal system as the modality mediating chemosensory perception. The strong cyclic responding pattern suggests links between photoperiodically controlled endocrine production, breeding behavior, and olfactory sensitivity.
Testicular response to melatonin or suprachiasmatic nuclei ablation in the spotted skunk
Authors: Berria Matthew , Desantis Mark , Mead Rodney A.
Published: June 2005

Testes of the Western spotted skunk enlarge and regress seasonally. The pineal hormone, melatonin, may be important in timing this seasonal reproductive activity. Likewise, the suprachiasmatic nuclei (SCN) have been implicated as possible neural regulators of pineal and reproductive events. These experiments were conducted to determine whether ablation of the SCN or constant administration of melatonin alters timing of the seasonal pattern of testicular regression and recrudescence. Male skunks (n = 24) were treated as follows: six received two empty Silastic capsules, six received two melatonin‐filled Silastic capsules, six received sham lesions to the SCN, and six received lesions to the SCN (SCNx). All skunks were exposed to a natural photoperiod and had regressed testes at the onset of the experiment. Four of six males from the SCNx group had an average of 94 ± 11.3% of these nuclei destroyed. Sham SCNx, animals with <40% of the SCN ablated, and males with empty capsules did not have fully enlarged testes until October. SCNx and melatonin‐treated skunks exhibited a hastening of testicular recrudescence with maximal testis size being reached in June. Skunks with lesions to the SCN maintained enlarged testes for 5 months while all other groups exhibited rapid regression after attaining maximal testis size. Testicular regression occurred from July through October in animals receiving continuous melatonin, while controls exhibited recrudescence during this same period. Our data suggest that the SCN, melatonin, and thus the pineal gland, play a role in regulating the seasonal testicular cycle of the spotted skunk.
The “love dart” of the snail Helix aspersa injects A pheromone that decreases courtship duration
Authors: Adamo Shelley A. , Chase Ronald
Published: June 2005

During the courtship of Helix aspersa, the snails push calcareous darts into one another. To examine the importance f this unusual action for the normal expression of their sexual behaviour, one group of Helix aspersa was prevented from exchanging darts by surgically removing their dart sacs. Another group of Helix aspersa had their digitiform glands surgically removed. These glands secrete a mucus that coats the dart. Glandless snails were capable of exchanging darts, but the darts lacked their normal mucus coating. Both dartless and glandless pairs required more courtship time to reach copulation than did sham‐operated controls. Therefore, it appears that it is the mucus from the digitiform gland, not the mechanical action of the dart, which affects courtship duration.

Injections of gland homogenate decreased the courtship duration of sexually receptive, dartless, and glandless snails. Gland homogenate also increased the size of the recipient's genital eversion, and retarded locomotion. The mucus was only effective if it entered the circulatory system of a sexually receptive snail at a specific stage of genital eversion.

The active agent in the digitiform gland mucus fulfills the requirements for the substance to be classified as a pheromone. It enters the circulatory system of the conspecific via inoculation by the dart.
Development of monosex female Oreochromis mossambicus broodstock by integrating gynogenetic technique with endocrine sex reversal
Authors: Pandian T. J. , Varadaraj K.
Published: June 2005

All female Oreochromis mossambicus were produced by judiciously combining endocrine sex reversal with selective breeding as well as gynogenesis. The combination of gynogenesis and endocrine sex reversal doubled the number of sex reversed males (XX‐‘♂♂’) and reduced the time cost from 12 to 6 months, in comparison to those produced by the combination of endorcine sex reversal with selective breeding. Programmes for commercial production and maintenance of XX‐♂ and XX‐♀ broodstock for large‐scale production of monosex female and monosex triploid sterile female populations are presented.
Clinal variation in the specific activity of lactate dehydrogenase‐B
Authors: Crawford Douglas L. , Place Allen R. , Powers Dennis A.
Published: June 2005

LDH‐B4 specific activity was determined from Fundulus heteroclitus collected in the field and from laboratory‐acclimated fish from four latitudinally distinct populations. The differences in the enzyme activity between populations for both field‐caught and laboratory‐acclimated fish are significant, with the northern populations having the highest LDH‐B4 enzyme activity. This clinal variation in enzyme activity is consistent with an evolutionary mechanism that compensates for lower environmental temperatures in colder northern latitudes.
Gastrointestinal transit and lipid assimilation efficiencies in three species of sub‐antarctic seabird
Authors: Jackson Sue , Place Allen R.
Published: June 2005

Using tritium‐labeled glycerol triether ([3H] GTE) as a nonabsorbable lipid‐phase marker and tritium‐labeled polyethylene glycol 4000 ([3H] PEG) as a nonabsorbable aqueous‐phase marker, we examined gastrointestinal transit of homogenized Antarctic krill (Euphausia superba) meals fed to white‐chinned petrels (Procellaria aequinoctialis), sooty albatrosses (Phoebetria fusca), and rockhopper penguins (Eudyptes chrysocome). The aqueous‐phase marker was excreted significantly more rapidly than was the lipid‐phase marker by the two procellariiform species, whereas no differential transit rates for the two markers were observed in the penguins. Aqueous‐phase marker recoveries after 48 hours from the three species were statistically indistinguishable (78.6% ± 3.7%, n = 5; 71.9% ± 11.3%, n = 7; and 77.0% ± 9.4%, n = 4, respectively). Lipid‐phase marker recovery from the penguins after 48 hours was nearly complete (83.8% ± 19.3%, n = 5, and 92.7% ± 14.8%, n = 5, for two dietary lipid supplements, see below), whereas less than 50% of the original dose of lipid marker was recovered from the two procellariiform species. Substantial lipid‐phase marker was recovered as stomach oils from the procellariiforms.

Assimilation efficiencies of [1‐14C] tripalmitin dissolved in wax ester and [1‐14C] cetyl oleate dissolved in triglyceride were compared for the same three seabirds by comparing 3H/14C ratios in the food and feces of birds simultaneously fed one of the above 14C‐labeled lipids, and the non‐metabolizable marker [3H] GTE. The petrel and the albatross showed high assimilation efficiencies (> 80%) of both 14C‐labeled neutral lipids. Rockhopper penguins consistently excreted [3H] GTE faster than did adult sooty albatrosses and were significantly less efficient at assimilating both neutral lipids (62% and 45% respectively). Sooty albatross fledglings excreted lipids significantly more slowly than did adults of this species, but lipid assimilation efficiencies did not differ with age. Gut measurements showed that the intestine of the rockhopper penguin was three and six times as long as those of the sooty albatross and white‐chinned petrel respectively.
Regional variations in the tightness of the ectodermal epithelium in the developing chick embryo: A study using ion‐sensitive microelectrodes
Authors: Carnan Elspeth , Gillespie J. I.
Published: June 2005

In 2‐day‐old avian embryos there is a rosto‐caudal gradient of interstitial pH (Gillespie and McHanwell: Cell Tissue Res., 247:445–451, '87). Neither the developmental significance nor the basic cellular mechanisms underlying this phenomenon has been studied. The present paper provides information about the interstitial potassium and calcium ion concentrations and the movement of these ions across the ectodermal epithelium. The data suggests a possible explanation for the longitudinal pH gradient in the embryo.

The concentrations of potassium and calcium ions in the interstitial spaces were measured with ion‐sensitive and conventional microelectrodes. In embryos bathed in solution containing 1 mM potassium, the potassium concentration in the region of the mesencephalon was 5.1 ± 0.7 mM while in the region of the unsegmented mesoderm it was significantly lower at 3.3 ± 0.4 mM (mean ± S.E., n = 16). If embryos are exposed to extra‐embryonic solutions containing 30 mM potassium, the K+ concentration in the mesencephalon is 13.0 ± 0.8 mM and higher at 15.4 ± 1.2 mM in the unsegmented mesoderm (n = 12). In embryos bathed in solutions containing 0.1 mM calcium, the interstitial calcium was found to be 1.1 ± 0.52 mM in the mesencephalon and 0.42 ± 0.19 mM in the unsegmented mesoderm (n = 3). In comparison, embryos bathed in solution containing 10 mM calcium had 1.9 ± 0.2 mM rostrally compared to 3.71 ± 0.63 mM caudally (n = 10). Thus it is possible to generate intra‐embryonic ion gradients dependent upon the extra‐embryonic ion concentration.

These results show, firstly, that the rate of potassium exchange across the ectodermal epithelium is lower in the rostral regions compared to more caudal regions, suggesting a regional variation in the tightness of the epithelium. Secondly, unlike H+ ions, potassium and calcium ions are passively distributed across the ectodermal epithelium.

The properties of the ectodermal epithelium are discussed in relation to the pH gradient and to their possible relevance to normal embryonic development.
Structural spermatophore proteins and a trypsin‐like enzyme from the accessory reproductive glands of the male grasshopper,Melanoplus sanguinipes
Authors: Cheeseman M. T. , Gillott C. , Ahmed I.
Published: June 2005

Some of the major structural spermatophore proteins and a trypsin‐like enzyme have been identified from the accessory reproductive glands of the male migratory grasshopper,Melanoplus sanguinipes.Polyclonal antibodies raised against the purified major soluble spermatophore protein (SP62) of M, 62,000 were used in Western blots to identify a soluble Mr85,000 putative precursor protein and, by immunohistochemistry and by partial proteolysis with staphylococcal V8 protease followed by Western blotting, an insoluble putative precursor protein from short hyaline gland three (SH3). The protein SP62 was formed in vitro by the incubation of crude homogenates of accessory gland tissues, seemingly by an interaction between SH3 secretion and soluble proteins from the remainder of the gland complex. In addition to the immunological similarity between SH3 secretion and SP62, there was also a strong similarity between the amino acid compositions of SH3 secretion and sodium dodecyl sulfate (SDS)‐extracted spermatophores. A family of white gland proteins (designated A‐group white gland proteins; WGPA) that showed intermale and intergland variations were also structural spermatophore proteins. Reserves of the major Mr22,000 and 21,000 forms declined by about 24 μg at mating. Polyclonal antibodies raised against these Mrforms recognized an additional Mr28,200 form in white glands one to three and a Mr9,600 counterpart in white gland four. WGPA‐immunoreactive proteins were evidently incorporated into the spermatophore as insoluble products. Both WGPA and SP62 antigens were localized chiefly in the outermost layers of the spermatophore but also occurred in the inner core. Crude homogenates of accessory glands hydrolyzed three trypsin substrates and an aminopeptidase substrate. The role that these enzymes might play in spermatophore formation is discussed.
An in vitro culture system for studying vitellogenin uptake into ovarian follicles of the rainbow trout, Salmo gairdneri
Authors: Tyler C. R. , Sumpter J. P. , Bromage N. R.
Published: June 2005

The development of a short‐term culture system to study vitellogenin (VTG) sequestration in vitro into vitellogenic follicles of the rainbow trout, Salmo gairdneri, is described. The outer epithelial cell layer of the trout follicle restricted VTG uptake considerably and its removal was a prerequisite to study VTG uptake in vitro. Vitellogenic follicles divested of the surface epithelium sequestered VTG at rates of up to 170 ng·mm2 follicle surface−1·h−1 (ng·mm−2·h−1). Uptake of VTG over an 8 h period of incubation was linear.

Increasing the titre of VTG in the culture medium induced greater rates of sequestration in a dose‐dependent manner; follicles cultured in medium containing 45 mg VTG·ml−1 sequestered 170 ± 9ng·mm−2·h−1 whereas those follicles cultured in medium containing 2 mg·ml−1 only incorporated 6 ± 0.5 ng·mm−2·h−1.

Uptake of VTG was temperature dependent with higher temperatures inducing higher levels of incorporation. Sequestration of VTG persisted at temperatures below 5°C, showing that the uptake mechanisms are adapted to low temperatures in this species.

Size of follicles also had a considerable bearing on the rate of VTG sequestration; follicles of less than 1 mm diameter (pre‐vitellogenic follicles) did not sequester VTG in amounts that were detectable. All follicles measuring 1 mm or greater in diameter sequestered VTG. As the follicles increased in size between approximately 1 and 3 mm in diameter there was an increase in both the total amount of VTG sequestered and the rate of uptake of VTG relative to surface area. Limited data suggested that in follicles closely approaching ovulation the rate of VTG sequestration was considerably reduced.
Purification and some properties of alkaline phosphatase from the hepatopancreas of the shrimp Penaeus japonicus (Crustacea: Decapoda)
Authors: Chuang Nin‐Nin , Shih Su‐Ling
Published: June 2005

Alkaline phosphatase purified from the hepatopancreas of Penaeus japonicus is stable to heating at 65°C for 5 min. The specific activity of the purified enzyme is 25,000 units/mg of protein. After polyacrylamide gel electrophoresis under denaturing conditions, the purified alkaline phosphatase from shrimp was found to consist of deglycosylated monomers of Mr 40,000 and to retain the attachment sites for both sialic acid and phosphatidylinositol. The alkaline phosphatase from shrimp has an isoelectric point (pI) of 7.6 and becomes more alkaline after the removal of either sialic acid or phosphatidylinositol residues.
Regulation of mouse preimplantation development: Inhibitory effect of genistein, an inhibitor of tyrosine protein phosphorylation, on cleavage of one‐cell embryos
Authors: Besterman Brian , Schultz Richard M.
Published: June 2005

We investigated the effects of genistein, an inhibitor of tyrosine protein phosphorylation, on mouse 1‐cell embryos, since in response to mitogenic stimuli tyrosine protein phosphorylation in somatic cells is implicated in initiation of DNA synthesis. Genistein inhibits cleavage of 1‐cell embryos in a concentration‐dependent and reversible manner; biochanin A, which is a less potent inhibitor of tyrosine protein phosphorylation, is a less potent inhibitor of cell cleavage. Genistein does not inhibit [35S]methionine incorporation, but does inhibit [3H]thymidine incorporation. Consistent with genistein's ability to inhibit cleavage by inhibiting DNA synthesis is that the loss of genistein's ability to inhibit cleavage corresponds with exit of the 1‐cell embryos from S phase. Genistein is likely to inhibit tyrosine protein phosphorylation in situ, since it reduces by 80% the relative amount of [32P]phosphotyrosine present in 1‐cell embryos; genistein does not inhibit either [32P]orthophosphate uptake or incorporation. As anticipated, genistein has little effect on inhibiting changes in the pattern of phosphoprotein synthesis during the first cell cycle, since tyrosine protein phosphorylation constitutes a small percentage of total protein phosphorylation. Alkalai treatment of [32P]radiolabeled phosphoproteins transferred to Immobilon reveals a base‐resistant set of phosphoproteins of Mr = 32,000 that displays cell‐cycle changes in phosphorylation. Although these properties suggest that these phosphoproteins may be related to the p34cdc2 protein kinase, phosphoamino acid analysis of [32P]radiolabeled phosphoproteins reveals that they are not enriched for phosphotyrosine; the inactive form p34cdc2 protein kinase contains a high level of phosphotyrosine. Results of these experiments suggest that tyrosine protein phosphorylation in response to the fertilizing sperm may be involved in initiating DNA synthesis in the 1‐cell embryo, as well as converting a meiotic cell cycle to a mitotic one.
Polarization of blastomeres in the cleaving rabbit embryo
Authors: Ziomek Carol A. , Chatot Clare L. , Manes Cole
Published: June 2005

Cellular polarization is believed to be a crucial event in the differentiative divergence of the two cell lineages leading to the blastocyst in rodent embryos. This study was undertaken to determine if rabbit embryos exhibited cellular polarization prior to blastocyst formation and to define the embryonic stage at which polarization was first apparent. Polarity was assayed by observation of the pattern of binding of FITC‐Con A to dissociated blastomeres from three stages of rabbit embryos. Scanning electron microscopy on the dissociated cells confirmed the fluorescence results. Fifty‐one percent of blastomeres in 38‐ to 66‐cell rabbit embryos exhibited an intense pole of FITC‐Con A binding and a single pole of microvilli. Only 2% of blastomeres at the 17‐ to 34‐cell stage were similarly polarized and none were polarized at the 8‐ to 16‐cell stage. In addition, during attempts to remove the mucin coat and zona pellucida from the rabbit embryos prior to their dissociation, it was found that the protease sensitivity of these coats also changed at the 38‐ to 66‐cell stage. Prior to this time, although the mucin coat disappeared after 30 min in 0.5% pronase, the zona required approximately 1.5–2.5 hr in pronase for even partial removal. At the 38‐ to 66‐cell stage, pronase dissolved the mucin coat within 10 min and the zona pellucida within 20 min. The zona was resistant to 0.1% proteinase K at all stages examined.
Sex‐specific embryonic mortality during concurrent pregnancy and lactation in house mice
Authors: Krackow Sven
Published: June 2005

To evaluate the importance of lactational stress for sex ratio manipulation in postpartum inseminated house mice, 163 sexually inexperienced male and female BALB/c ABom inbred mice were mated and stud males were removed the day following the initial parturition. Randomly chosen dams were allowed to suckle their young for either one or 14 or 21 days. The results showed that two weeks of lactation enhance the incidence of total abortion of the litter conceived postpartum. If lactation continued another week, dams which gave birth to a second litter lost relatively more embryos during uterine development and the percentage of lost embryos was increased by the number of pups suckled. Among these dams, the number of embryos lost in utero correlated positively with viable litter sex ratios (percentage male pups) at second parturition, indicating that lactational stress enhances embryonic mortality and that this additional mortality predominantly affects female embryos.
Anaerobiosis and the regulation of glycolytic enzymes in the sea anemone Metridium senile
Authors: Michaelidis Basile , Storey Kenneth B.
Published: June 2005

The regulation of glycolytic enzymes during anaerobiosis was examined in three tissues, muscular body wall, tentacles, and mesoglia, of the sea anemcme Metridium senile. Anoxia‐induced stable modifications of the activities or kinetic and regulatory properties of glycogen phosphorylase, phosphofructokinase (PFK), and pyruvate kinase (PK), were detected. In the two muscular tissues, anoxia had the greatest effect on the properties of PFK; for PFK in body wall So F6P decreased, Km ATP increased, I50 PEP increased, and Ka values for AMP and inorganic phosphate increased, compared to PFK in aerobic muscle. In addition, levels of the PFK activator fructose‐2,6‐bisphosphate decreased sharply after 21 hours of anoxia LO amounts 19, 48, and 39% of aerobic control values in body wall, tentacles, and mesoglia, respectively. Anoxia exposure had fewer effects on the properties of PK, and primary control of the enzyme in muscular tissues may reside with allosteric effectors, F1, 6P2 activation, and strong L‐alanine inhibition. PK in mesoglia, by contrast, was virtually unaffected by allosteric modifiers but shcwed a 4‐fold decrease in S0.5 PEP in anoxia. The results suggest that changes in glycolytic function during anaerobiosis in sea anemones are mediated by covalent modifications of the regulatory enzymes as well as changes in the levels of F2,6P2 and other metabolite effectors of the enzymes.
Chemical detection of predators by a lizard, the broad‐headed skink (Eumeces laticeps)
Authors: Cooper William E.
Published: June 2005

The scincid lizard Eumeces laticeps is shown experimentally 1) to be capable of detecting integumentary chemical stimuli from two congeneric species of snakes that are sympatric predators of lizards and 2) to discriminate these chemical stimuli from similar ones derived from a sympatric snake that does not eat lizards and from a common insect food species. The experimental technique employed in this study, i.e., presentation of odorants on cotton‐tipped applicators, allows rapid assessment of responses to predator odors and is suitable for a survey of these responses in a wide variety of lizards. The skinks showed significantly higher tongue‐flicking rates in response to odors of snake predators and food odors (cricket) than to odors of a snake that does not eat lizards and an odorless control. Tongue‐flick attack score, a measure of response strength to chemical stimuli from food, was much greater for cricket stimuli than for the predator stimuli. Significantly more lizards bit applicators bearing cricket stimuli than bit applicators bearing any of the other stimuli. After biting applicators bearing predator odors, the lizards quickly released. In contrast, they frequently maintained a grasp after biting applicators bearing cricket odors. A few preliminary speculations are made regarding ecological factors that may affect responsiveness by lizards to predator odors.
Rat renal papillary tissue explants survive and produce epithelial monolayers in culture media made hyperosmotic with sodium chloride and urea
Authors: Woolverton W. S. , Githens S. , O'Dell‐Smith R. , Bartell C. K.
Published: June 2005

The capacity of papillary cells to adapt to elevated osmotic concentrations is unusual among mammalian cells. This capacity was evaluated by using primary tissue culture. Viability and growth of cells in rat renal papillary tissue explants were assessed after culture in media adjusted with urea and sodium chloride to various osmotic concentrations between 300 and 1,500 mOsm/kg water. The survival of cells, including cells resembling those of the collecting ducts and the loop of Henle, was greatest in medium adjusted to 1,000 mOsm with equiosmolar amounts of the two solutes. At 1,500 mOsm only cuboidal tubular epithelium resembling collecting duct epithelial cells survived. In contrast, cells of cortical tissue survived and grew at 300 and 640 mOsm, but not at 1,000 mOsm or above. Epithelial monolayers appeared to proliferate from collecting ducts and spread over the surface of the explants as well as onto the glass surface in the culture dish. Epithelial growth of medullary tissue was most rapid at 300 mOsm and was slower at 700 and 1,000 mOsm. Monolayers did not form at 1,500 mOsm; however, epithelial overgrowth of explants did occur. Hydropenia in the donor animal did not significantly affect the viability or growth of cultured papillary tissue. Explants cultured for 5 days at 300 mOsm followed by a stepwise increase in medium osmolality to 1,100 or 1,500 mOsm and cultured for 3 more days showed low or no survival whereas explants cultured at 700 mOsm survived such increases. Explants cultured for 5 days at 1,500 mOsm survived and grew monolayers when lowered to 300 mOsm. Poor viability and no epithelial proliferation were observed in explants cultured in medium adjusted to 900 mOsm with either urea or sodium chloride alone, suggesting that a mixture of the two solutes in the extracellular space, as found in vivo, may be essential in achieving elevated osmolalities.
Ultrastructural evidence for an association between an oviductal glycoprotein and the zona pellucida of the golden hamster egg
Authors: Abe Hiroyuki , Oikawa Taneaki
Published: June 2005

The ultrastructural localization of an oviductal glycoprotein, designated ZP‐0 in golden hamster oviductal eggs, was investigated by immunolabeling methods using a monoclonal antibody (C11E8). Immunofluorescence staining showed that C11E8 specifically reacted with the zona pellucida of the oviductal egg but not the ovarian egg. In an immunoelectron microscopic study applying the protein‐A gold technique, gold particles were distributed throughout the zona pellucida of the oviductal eggs and were also associated with the perivitelline matrix. Structures within the eggs and cumulus cells did not react with C11E8. Quantitative evaluations of the degree of labeling demonstrated that a large number of gold particles was bound to the zone pellucida, especially in the middle layer. Moreover, in bovine testicular hyaluronidase‐treated eggs the density of labeling decreased only in the outer third of the zona pellucida. These results show that ZP‐0 was associated with the zona pellucida and perivitelline matrix of the golden hamster egg after ovulation and suggest that there are topographical differences in the binding activity of ZP‐0 to the zona pellucida. In addition, the decrease in labeling density of ZP‐0 induced by hyaluronidase appears to be related to changes in the properties of the outer layer of the zona pellucida.
Distribution of hyaluronidase activity in the tissues of the normal and the lens‐regenerating iris in the newt
Authors: Zalik Sara E. , Dimitrov Eva
Published: June 2005

During lens regeneration in the newt the synthesis of hyaluronic acid increases as iris cells undergo dedifferentiation, and decreases during the redifferentiation phase. In contrast, the levels of hyaluronidase activity which are high in the normal iris remain constant during regeneration (Kulyk et al: Exp. Cell Res., 172:180–191, 1987). We determined whether differential changes in hyaluronidase activity occurred within discrete cell populations of the iris during lens regeneration. In the normal iris hyaluronidase activity was dominant in the iris epithelium, with low levels of enzyme activity being present in the iris stroma. Regionally within the iris, highest levels of enzyme activity were found in the ciliary iris, while lower levels were found in the pupillary iris. Results suggest that a circumferential gradient of hyaluronidase activity decreasing from the dorsal to the ventral regions is present within the iris; superimposed within this gradient, a meridional gradient of enzyme activity increasing from the pupillary to the ciliary region exists. The levels of hyaluronidase activity increase in the dorsal pupillary iris during redifferentiation stages. This activity could be involved in the removal of the hyaluronic acid that is produced during dedifferentiation of the regenerating iris.
Studies on the decondensation of human, mouse, and bull sperm nuclei by heparin and other polyanions
Authors: Jager S. , Wijchman J. , Kremer J.
Published: June 2005

We report heparin‐induced decondensation of human, mouse, and bull sperm nuclei. Decondensation did not occur if the spermatozoa were intact but only if the membranes were severely damaged by freezing and thawing or by treatment with a detergent. If a disulphide bond reducing agent (thiol) was absent, decondensation of human sperm nuclei was usually a relatively slow process, with large interindividual variation. Mouse and bull sperm nuclei did not decondense in the absence of a thiol. With a thiol relatively low concentrations of heparin induced a rapid decondensation of the sperm nuclei of all three species. The decondensation activity was not specific for heparin; other polyanions were also active, with heparin being the most effective compound.

It is supposed that heparin and other polyanions induce sperm nuclear decondensation because they deplete protamines from the chromatin. Thus the negatively charged phosphate groups of the DNA are no longer opposed by positively charged protamines. Consequently the mutual repulsion of unopposed phosphate groups causes the DNA molecules to stretch, which results in an increase of the sperm nuclear volume. Since heparin and other polyanions induce decondensation under physiological pH and temperature, polyanions might also be active in the oocyte.