bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of zona binding proteins of rabbit, pig, human, and mouse spermatozoa on nitrocellulose blots
Authors: O'Rand Michael G. , Matthews Jessica E. , Welch Jeffrey E. , Fisher Susan J.
Published: June 2005

Mammalian fertilization is a multi‐step process with different requirements for specificity at each step. In the present report we have examined the binding of spermatozoa to homologous and heterologous zonae pellucidae. The homologous zona binding proteins (ZBP) of ejaculated rabbit, pig and human spermatozoa and epididymal mouse spermatozoa have been identified. The rabbit's ZBPs have relative molecular weights (MW) of 32K, 18K, 16K and 14K; the pig's major ZBP is 16K while human spermatozoa bind human zona protein at 17K and 18K. Mouse sperm ZBPs are 19K, 18K and 16K.
Trimethylamine oxide and the maintenance of volume of dogfish shark rectal gland cells
Authors: Kleinzeller Arnost
Published: June 2005

Determinants of the steady‐state vol of the dogfish shark (Squalus acanthias) rectal gland cells were studied. The cellular levels of trimethylamine oxide (TMAO) in fresh tissue and slices incubated aerobically 60 min in standard (TMAO‐free) elasmobranch saline were close to those in the plasma (71 ± 5 mM S.E.M.); therefore, under these conditions, the cell membrane appears to be impermeable to this solute. However, depolarization of the cells in high‐K+ media produced a rapid loss of TMAO. Thus, TMAO is a major, effectively impermeant solute in the rectal gland cells. The osmolarity of cell solutes in tissue water (fresh and incubated slices) did not differ significantly from values in the plasma or incubation medium, demonstrating the absence of an osmotic pressure gradient across the cell membrane. An analysis of a simple model of cell solutes under steady‐state conditions shows that the presence of an (effectively) impermeant osmolyte decreases the cellular concentration of bulk cations. The analysis is consistent with available observations on the distribution of cell Na+ and K+ in tissues containing high concentrations of (nitrogeneous) osmolytes. One simplifying assumption of the model, i.e., identity (or closeness) of the respective reflection coefficients σ for Na+ and K+ passage through the cell membranes could not be verified. Compared to available data on the steady‐state cellular fluxes of 42K+ in slices of the rectal gland, the uptake of 22Na+ by the tissue was slow (the derived rate constant k′ = 0.017 min−1, i.e., about one tenth of that for K+). Hence, σNa < σK. On the other hand, 36Cl− rapidly exchanged with cell Cl− with the first rate constant (k′ = 0.60 + 0.12 min−1) close to that of 3H2O.
Localization of cytoplasmic determinants responsible for primary mesenchyme formation and gastrulation in the unfertilized egg of the sea urchin Hemicentrotus pulcherrimus
Authors: Maruyama Yoshihiko K. , Nakaseko Yukinobu , Yagi Shintaro
Published: June 2005

The jelly canal of the unfertilized egg of the sea urchin H. pulcherrimus was found to serve as a stable landmark of the egg axis. The eggs were bisected with a micro glass needle into two egg‐halves either equatorially or meridionally in relation to the egg axis defined by the jelly canal. The development of both egg‐halves after fertilization was observed 1 or 2 days later until the control embryos reached the gastrula stage. The embryo derived from the vegetal half formed primary mesenchyme and gastrulated while the animal half remained as the permanent blastula forming neither primary mesenchyme nor archenteron. Both embryos obtained by the meridional bisection formed primary mesenchyme and gastrulated. The presence of the female pronucleus or its adjacent cytoplasm was not a prerequisite for the occurrence of primary mesenchyme formation and gastrulation. These results indicate that unfertilized eggs already localize the cytoplasmic determinant responsible for primary mesenchyme formation and gastrulation. Localization of the cytoplasmic determinant occurs independently of formation of the cortical pigmentation polarity.
The trophotaenial placenta of a viviparous goodeid fish. III: Protein uptake by trophotaeniae, the embryonic component
Authors: Lombardi Julian , Wourms John P.
Published: June 2005

Protein uptake and degradation by trophotaenial cells of the viviparous goodeid fish Ameca splendens were studied colorimetrically and ultrastructurally using horseradish peroxidase (HRP) as a tracer and acid (ACPase) and alkaline (ALPase) phosphatase cytochemistry. Trophotaeniae are ribbon‐like external projections of the embryonic gut that are equivalent to greatly hypertrophied intestinal villi. During gestation within the ovarian lumen, trophotaeniae are directly apposed to the internal ovarian epithelium (IOE) where they establish a placental association between the developing embryo and maternal organism. Trophotaenial absorptive cells possess an ALPase reactive brush border, an endocytotic apparatus, and ACPase reactive standing lysosomes. Ultrastructural studies of protein uptake indicate that cells of the trophotaenial epithelium take up HRP by micropinocytosis and degrade it within lysosomes. Initially (from 1.5–10 min), HRP is taken up in vitro at 22° C at the apical cell surface and passes via endocytotic vesicles into an apical canalicular system. From 1.5 to 10 min exposure, HRP passes passes from the apical canalicular system to a series of small collecting vesicles. After 10 min, HRP is detected within large ACPase reactive supranuclear lysosomes. Three hours after an initial 1 h exposure to HRP, most peroxidase activity within supranuclear lysosomes is no longer detected. Presence of Golgi complexes, residual bodies, and secretory granules in the infranuclear cytoplasm suggest that products of protein uptake and hydrolysis are discharged across basal and lateral cell surfaces and into the trophotaenial circulation. Trophotaeniae of embryos incubated in vitro in HRP‐saline take up HRP at an initial rate of 13.5 ng HRP/mg trophotaenial protein/min. The system becomes saturated after 3 h. Trophotaeniae incubated at 4°C show little or no uptake. In trophotaeniae continuously pulsed with HRP for 1 h, then incubated in HRP‐free saline, levels of absorbed peroxidase declined at a rate of 0.5 ng/mg trophotaenial protein/min. HRP does not appear to enter the embryo via extratrophotaenial routes. These findings are consistent with the putative role of trophotaeniae as the embryonic component of the functional placenta of goodeid fishes. Trophotaenial uptake of maternal nutrients accounts for a massive (15,000%) increase in embryonic dry weight during gestation.
Significance of Con A binding sites in the conjugation of Euplotes vannus
Authors: Lueken W. , Oelgemöller B.
Published: June 2005

Aggregations of Con A binding sites that occur on the surface of preconjugant cells have been studied in order to reveal their possible functions in the conjugation process. By means of a newly adapted horseradish peroxidase method, they can be visualized in distinctly shaped oval fields (OF) as soon as 10–15 min after mixing cells of opposite mating types. Cycloheximide (CHX) prevents establishment of new OFs but does not effect preexisting ones. In the absence of previously developed OFs, cells cannot form pairs. However, even if OFs are fully present, pairing still can be inhibited by CHX, high K+ concentrations (≥ 60 mmol/liter), or the K+‐channel blocker tetraethyl‐ammonium (TEA, 10 mmol/liter). A doublet cell with OFs on both halves normally binds to one singlet. This partner, after a stay of 10 min in an angled agglutination position, quickly rotates into the parallel pairing position. At this moment, the doublet suddenly loses its affinity for complementary cells at its free OF. This behavior explains why many doublets pair only with one singlet. The Con A binding sites themselves seem not to be active in cell pairing; they are regarded as a marker for a region of the cell surface whose contact with a complementary cell subsequently results in the unmasking of unidentified compounds that serve as adhesive agents and/or the hinge for the quick rotation of agglutinated cells.
Development of the masseter muscle and oral behavior in the pig
Authors: Herring Susan W. , Wineski Lawrence E.
Published: June 2005

During mastication the adult pig masseter contracts with a complex pattern involving a wave of electromyographic (EMG) activity moving from the ventro‐rostral corner to the dorso‐caudal corner. The present study was undertaken to ascertain the ontogeny of that contraction pattern. Anatomical measurements were made on masseters from fetal, infant, and juvenile pigs. EMG activity from different parts of the masseter was recorded along with oral movements in infant and juvenile pigs as they suckled, drank, and chewed on food and non‐food objects. The basic arrangement of muscle fibers and tendinous aponeuroses was found to be the same in all ages. The longest and most vertical fibers were found rostrally and ventrally, whereas the shortest and most horizontal fibers were found caudally and dorsally. The length of fasciculi decreased with age, relative to muscle weight. Variance in length among different parts of the muscle increased with age. Fetal masseters were oriented generally more horizontally than the masseters of older animals, except that the dorso‐caudal corner, usually the most horizontal portion, is not developed in fetuses. The contraction patterns within the infant masseter were less complex than those of older animals; only the dorso‐caudal corner was distinct. The further development of intramuscular differences in activity may be associated with the increasing anatomical complexity of the masseter, which augments its functional capabilities.
Hormonal regulation of growth and life span of bullfrog tadpole tail epidermal cells cultured in vitro
Authors: Nishikawa Akio , Yoshizato Katsutoshi
Published: June 2005

Epidermal cells were dissociated from tails of the bullfrog tadpole, Rana catesbeiana, and cultured to investigate their response to steroid and thyroid hormones. Charcoal‐treated serum (CTS) was used in the growth medium when cells were to be grown in the absence of steroid and thyroid hormones. The cells could be maintained for 2 weeks with a small increase in cell number in medium that contained CTS (CTS medium). Addition of cortisol to CTS medium increased both cellular attachment to the culture dishes and the proliferation of the attached cells with an optimum concentration of 5 × 10−7 M. The cells remained viable and attached for at least a week. Cortisol stimulated the rate of protein synthesis 1.8‐fold but did not alter the rate of DNA synthesis. The cells did not proliferate in the medium containing triiodothyronine (T3) and detached themselves from the dish within 5 days, which occurred in a dose‐dependent manner with a maximum effect at 10−8 M. It drastically decreased the rate of DNA synthesis but did not influence the rate of protein synthesis. These responses of cells to cortisol and T3 may reflect growth and death of tail epidermal cells in vivo at metamorphosis.
Studies on histocompatibility in natural populations of freshwater sponges
Authors: Mukai Hideo , Shimoda Hitoshi
Published: June 2005

Histocompatibility in natural populations of individuals was examined for the following four Japanese species of freshwater sponges: Ephydatia fluviatilis, Ephydatia muelleri, Radiospongilla cerebellata, and Eunapius fragilis. Gemmules were obtained from field‐grown specimens. Then, using juvenile sponges hatched from gemmules, fusibility was tested in the laboratory. Every interspecific combination among these four species displayed non‐fusion with no cytotoxic interaction. For all species, the occurrence of allogeneic histoincompatibility was assured, but manifestation of incompatibility was different from species to species. Among 36 specimens of E. fluviatilis collected from three localities, two strains nonfusible with each other were distinguished. Each locality was populated by a single strain. Specimens of E. muelleri were obtained from two localities. All combinations of specimens coming from the same locality were compatibly fusible, but those between specimens coming from different localities displayed conspicuous cytotoxic rejection reaction. This reaction occurred asymmetrically, resulting in the destruction of one partner. In R. cerebellata, two types of reactions, fusion and rejection, were also distinguished. In rejection, cytotoxic reaction occurred bilaterally and symmetrically in and around the contact zone. A total of 40 specimens, collected from a single pond, could be divided into 16 groups by their fusibility. As to E. fragilis, some combinations of specimens were compatibly fusible; the others displayed various degrees of incompatibility. In completely incompatible combinations, the paired sponges simply showed nonfusion with no sign of cytotoxic reaction. In more weakly incompatible combinations, the paired sponges were separated after initial fusion, the degree of which was variable depending on combination. A total of 59 specimens, collected from a single pond, could be classed into 14 groups by their fusibility. In both R. cerebellata and E. fragilis, some intergroup combinations were compatible, giving rise to probable genetic chimeras. Such chimeric sponges persisted with no sign of segregation for more than 1 month in the field.
Ca+ + localization in boar spermatozoa by the pyroantimonate technique and X‐ray microanalysis
Authors: Berruti Giovanna , Franchi Emilia , Camatini Marina
Published: June 2005

Intracellular, loosely bound Ca+ + has been localized electron microscopically in freshly ejaculated boar spermatozoa by in situ precipitation with potassium antimonate. Ca+ + was identified as the cation precipitated by testing the EGTA‐sensitivity of the precipitates and by X‐ray microprobe analysis. The data obtained revealed that the outer acrosomal membrane is the preferential site for Ca+ + precipitation in the sperm head.
Morphogenetic waves in the development of the lateral mesoderm in the Mexican axolotl (Ambystoma mexicanum) and their relationship to pronephric duct migration
Authors: Gillespie Laura L. , Armstrong John B.
Published: June 2005

That portion of the lateral mesoderm capable of supporting migration of the pronephric duct was mapped by transplanting secondary ducts ventral to the primary duct. Ducts transplanted more anterior than the primary duct did not migrate, whereas those transplanted more than 2 somitewidths posterior to the primary duct appeared to be delayed in the onset of migration. These transplants thus define an “active” region that extends approximately from the posterior margin of the last‐formed somite to the position of the primary duct tip, about 2 somite‐widths anterior. The anterior boundary of the active region coincides with a wave of shape change that passes in a craniocaudal direction through the lateral mesoderm. In this wave, the mesoderm changes from a two‐cell to a single‐cell layer and then back to a two‐cell layer. The duct tip is always located over the single‐cell layer. We also examined the distribution of extracellular matrix (ECM) between the mesoderm and epidermis. The posterior limit of the ECM was always posterior to the duct tip but did not extend beyond the last‐formed somite.
Biochemical changes in the composition of the testes during spermatogenesis in the sea starAsterias vulgaris
Authors: Smith Frank F. , Walker Charles W.
Published: June 2005

Examination of major biochemical classes during the spermatogenic cycle provides a definition of the changing chemical microenvironment influencing germinal cells and also suggests temporal relationships among successive spermatogenic events. Testes from sea stars collected throughout the year were homogenized and lyophilized, and aliquots were assayed for DNA, RNA, total protein, free amino acids, total lipids, glycogen, and other carbohydrates; spermatogenic stage was determined by examination of paraffin sections. The resulting data, expressed as 1) mg/g dry mass, 2) mg/mg DNA, and 3) total content, were analyzed by weighted periodic regression and circular‐linear correlation with spermatogenic stage and time of year.

With the exception of lipid/DNA, all components show a significant regression against time. DNA concentration increases during the proliferative and differentiative phases, attaining maximal values late in the differentiative phase; DNA synthesis appears to be essentially complete by the end of March. RNA concentration increases in the late aspermatogenic phase, as testicular cells begin preparations for proliferation, and remains high until late in the differentiative phase. Protein accumulates during proliferation, preceding the accumulation of DNA, but then gradually declines during differentiation. Free amino acids, simple sugars, and glycogen all increase during the aspermatogenic phase, possibly as a result of nutritive input.

The observed patterns of change correlate closely with cytological variations observed within the germinal epithelium. Biochemical data thus complement existing data on the cytology of the germinal epithelium.
Plasma levels and renal handling of endogenous amino acids in snakes: A comparative study
Authors: Benyajati Siribhinya , Dantzler William H.
Published: June 2005

Plasma levels of 22 endogenous amino acids were measured by ion‐exchange chromatography in four species of snakes: Thamnophis sirtalis, T. radix, Aipysurus laevis, and Python molurus. Despite considerable interspecific variation in the amino acid composition, all species showed relatively high plasma concentrations of histidine, a feature apparently unique to reptiles. The renal handling of these amino acids was studied by renal clearance methods. As in other vertebrates, net tubular absorption of filtered amino acids predominated. However, net tubular secretion of taurine, cysteic acid and/or phosphoserine and β‐alanine was observed, with taurine being the predominant amino acid secreted. The percentage reabsorption of the total amino acids filtered by the snake kidneys ranged from 79 to 95%. Evidence for the postrenal absorption of amino acids in these reptiles is presented. In species that normally undergo hibernation (Thamnophis spp.), the ability of the kidney to reabsorb amino acids was depressed by cold acclimation. Cold acclimation significantly decreased plasma levels of all amino acids except taurine, whose concentration increased. The increase in plasma taurine level may have resulted from cellular osmoregulation. Under these conditions, renal excretion of taurine increased concomitantly with the increase in plasma taurine concentration.
31P‐NMR studies of Limulus polyphemus: Spermatozoa at rest and after motility
Authors: Robitaille Patricia A. , Robitaille Pierre‐Marie L. , Brown George G.
Published: June 2005

Phosphorus‐31 nuclear magnetic resonance spectroscopy (31PNMR) was used to monitor changes in the levels of several high‐energy compounds both prior to and following the initiation of motility in spermatozoa of the horseshoe crab, Limulus polyphemus L. These spermatozoa are easily collected and are nonmotile until stimulated with a sperm‐motility‐initiating peptide (SMI) which can be extracted from Limulus eggs. Following the addition of SMI, several changes in metabolite levels within the spermatozoa were obtained, the most significant of which were a decrease in nucleotide triphosphate (NTP) and phosphoarginine levels and an increase in the levels of inorganic phosphate (Pi), NDP, and phosphomonoesters. An increase in the intracellular pH (0.45 pH units) of the sperm was also noted following the completion of SMI‐stimulated motility.
Pronounced influence of Hb‐O2 saturation on red cell pH in tench blood in vivo and in vitro
Authors: Jensen Frank B.
Published: June 2005

At an extracellular pH of 7.9 the red cell pH in tench blood exhibits a nonlinear, strongly inverse relationship with Hb‐O2 saturation, both when investigated in vitro and in vivo (in fish exposed to acid water with and without aluminium). The pHi difference between deoxygenated and oxygenated erythrocytes is large (i.e., 0.35 pH units), which can be accounted for by the Haldane effect and the buffering properties of the blood. The major shift in pHi occurs between 50 and 100% Hb‐O2 saturation, indicating an almost full exploitation of the Haldane effect within the normal physological range of blood O2 saturations.
Cyclic AMP stimulation of Cl− secretion by the opercular epithelium: The apical membrane chloride conductance
Authors: Degnan Kevin J.
Published: June 2005

Chloride secretion (Isc) by the opercular epithelium of the teleost, Fundulus heteroclitus, is stimulated by elevations in intracellular cyclic AMP (cAMP) elicited by β‐adrenergic agonists, such as isoproterenol, and is accompanied by a small but significant increase in the transepithelial conductance (Gt). Cupric ions (Cu2+) have been shown to block the apical membrane CI− channels in this epithelium, leading to a reduction in both the Isc and Gt (Degnan, '85). In the present studies, the effects of Cu2+ on cAMP‐elevated and cAMP‐depleted epithelia were observed to define the actions of cAMP in this stimulatory process. At a concentration of 5 × 10−4 M in the mucosal solution, Cu2+ inhibited the Isc 79.8% and reduced the Gt 39.2%. Isoproterenol produced an attenuated stimulation of the Isc in these tissues compared to untreated controls, but had no effect on the Gt. In tissues bathed bilaterally with Cl− ‐free Ringer, the Isc was virtually abolished and the Gt was reduced 37.0%; neither Cu2+ nor isoproterenol had any effects on the Isc or Gt under this condition. Simultaneous 22Na and 36 Cl unidirectional flux determinations indicated that the only effects of both isoproterenol and Cu2+ were on the active Cl‐ secretory flux. An inhibitor of adenylate cyclase, 2′,5′ dideoxyadenosine (DDA), reduced the Isc and Gt 39.8% and 20.8% respectively. This inhibitor had no additional effects in Cu2+ ‐treated tissues and the action of Cu2+ on the Gt was reduced in DDA‐treated tissues. The results indicate that cAMP stimulation of Cl− secretion involves only the active transcellular Cl− pathway and is accomplished in part by an increase in a Cl− ‐dependent, mucosal Cu2+ ‐inhibitable increase in the Gt, which is most likely localized to the apical membrane of the chloride cell.
Attenuated cardiovascular effects of prostaglandin I2 and prostaglandin F2α in cold acclimated American bullfrogs, Rana catesbeiana
Authors: Herman Ceil A. , Robleto Daniel O. , Mata Paula L. , Lujan Monica D.
Published: June 2005

American bullfrogs, Rana catesbeiana respond to prostaglandins with changes in heart rate and blood pressure. These studies compare responses of warm (22°C) and cold acclimated (5°C) bullfrogs to prostaglandins. Gas chromatographic analysis determined equivalent fatty acid profiles in total lipids of heart and artery tissue from warm and cold acclimated animals. Arachidonic acid was the fatty acid precursor found in greatest abundance in both groups. For cardiovascular experiments, bullfrogs were cannulated by using a T‐cannula implanted in the right sciatic artery. In warm acclimated bullfrogs, preinfusion systemic arterial pressure (SAP) was 14.7 ± 0.5 mm Hg, and heart rate was 33.0 ± 1.7 beats/min. Cold acclimated bullfrogs had SAP values of 8.0 ± 0.8 mm Hg, and heart rate was 6.9 ± 0.3 beats/min. Arachidonic and eicosapentaenoic 3‐acid infusions (2,000 μg/kg body weight [bw]) were hypertensive in cold acclimated and hypotensive in warm acclimated animals. These effects were blocked by indomethacin (4 mg/kg bw). In both warm and cold acclimated bullfrogs, prostaglandin F2α (3–100 μg/kg bw) was hypertensive, while prostaglandin I2 (0.03‐3 μg/kg bw) was hypotensive, with both prostaglandins stimulating a greater absolute response in warm acclimated animals. In addition, both prostaglandins increased heart rate in warm but not in cold acclimated bullfrogs. The results suggest diminished cardiovascular sensitivity to prostaglandins at low environmental temperatures.
Cell surface carbohydrate involvement in controlling the adhesion and morphology of neural crest cells and melanophores of Xenopus laevis
Authors: Milos Nadine C. , Wilson H. Chris
Published: June 2005

Pieces of dorsal neural tube (stages 22–23) or late neural crest tissue (stages 24–26) of Xenopus laevis were cultured. Migratory cells moved out of explants to form an outgrowth of multipolar melanophores on the substratum. Treatment with β‐galactosidase (0.1–0.4 U/ml) to remove cell surface galactose was correlated with detahment of melanophores. In the presence of lower concentrations of this enzyme the shapes of these cells were converted to arborized, spidery morphologies and cell movement was inhibited. Unpigmented cells were affected more slowly. Neuraminidase treatment, to remove cell surface sialic acid and expose more galactose, only affected melanophores. These became increasingly spread on the substratum and cell overlap was observed. These results suggest that the relative amounts of galactose and sialic acid at the cell surface become increasingly important in controlling cell adhesion as X. laevis neural crest cells migrate and differentiate into melanophores.
Osmolarity affects oxidation of sarcosine by isolated hepatocytes and mitochondria from a euryhaline elasmobranch
Authors: Ballantyne J. S. , Moyes C. , Moon T. W.
Published: June 2005

Hepatocytes isolated from the little skate, Raja erinacea, oxidize sarcosine at higher rates in hypoosmotic media and at lower rates in hyperosmotic media. A similar pattern is noted for the oxidation of sarcosine by mitochondria isolated from the liver. These observations are consistent with a role for mitochondria in ridding the cell of osmotically active organic solutes during volume regulation in hypoosmotically stressed cells while allowing their accumulation during hyperosmotic volume regulation. Such a mechanism would be quantitatively more imporant in osmoconforming species but may occur in osmoregulating species as well.
The role of spindle microtubules in the timing of the cell cycle in echinoderm eggs
Authors: Sluder Greenfield , Miller Frederick J. , Spanjian Kristin
Published: June 2005

Spindle microtubules play an important role in the mechanisms that control the timing of cell cycle events in the eggs of the sea urchins L. variegatus and L. pictus. However, recent work which used colchicine to block microtubule assembly in the eggs of two other echinoderms, S. purpuratus and D. excentricus, has raised serious questions about the generality of this role for spindle microtubules. Thus, we have systematically examined the role of spindle microtubules in the timing of the cell cycle in the fertilized eggs of these latter species. We treated eggs of both species with 5–10 μM Colcemid for several minutes starting 30 min after fertilization to completely prevent spindle microtubule assembly for several h. We used Colcemid, instead of colchicine, because it is effective at lower doses and, at these doses, shows no detectable toxic side effects. We compared for control and treated eggs the time course of nuclear envelope breakdown/reformation and DNA synthesis. We found for both species that the eggs continue to cycle without spindle microtubules; mitosis is up to twice the normal duration while interphase remains essentially unaffected. To test for the possible toxic side effects of the 1–2 mM colchicine used earlier on S. purpuratus and D. excentricus, we treated eggs of these two species, and also those of L. variegatus, with 1 mM lumi‐colchicine. This photo‐inactivated form of colchicine, which does not bind to tubulin, substantially prolongs mitosis and, to a lesser extent, interphase. Thus, the results of the earlier work are most easily explained by the combination of specific and nonspecific effects of the 1–2 mM colchicine used. Our present results indicate that the importance of spindle microtubules in the mechanisms that control the timing of the mitosis portion of the cell cycle is a general phenomenon.
Requisites for growth and myelination of urodele sensory neurons in tissue culture
Authors: Olsen Cherie L. , Bunge Richard P.
Published: June 2005

We describe a system for long‐term culture of adult salamander dorsal root ganglia and present initial observations on the regulation of nonneuronal cell proliferation in this system. Observations of the region of axonal outgrowth in these cultures indicated that, on the one hand, fibroblasts proliferate equally well with neurons present or absent. Schwann cells, on the other hand, require not only the presence of neurons but also proximity to or contact with their neurites. These results indicate that neural tissue is not mitogenic for all cell types represented in the limb regeneration blastema. Further, we describe, for the first time, myelination of salamander axons in vitro. The critical factors found to be required for this high degree of differentiation were the use of long culture periods and addition to the culture medium of ascorbic acid, which is shown to promote extracellular matrix deposition. These observations correlate with extensive observations made on mammalian tissues in vitro, indicating a linkage between deposition of extracellular matrix and full expression of other Schwann cell functions. We anticipate this culture system will be useful in studies of limb blastema cell proliferation.
Photothermal effects on the testicular cycle in the musk turtle, Sternotherus odoratus
Authors: Mendonça Mary T. , Licht Paul
Published: June 2005

A series of photothermal treatments was initiated at different times of year to examine the relative contributions of exogenous and endogenous factors affecting the timing of the testicular cycle of the kinosternid turtle, Sternotherus odoratus. Elevated temperature appeared to be the major factor stimulating testicular recrudescence (spermatocytogenesis, spermiogenesis, and, to some extent, androgen secretion). However, depending on the time of year, short or long daylengths, in conjunction with elevated temperatures, enhanced testicular responses (recrudesence or regression). Also, high temperatures are not requisite for recrudescence. Spermatogenic activity and testis growth differ in their temperature sensitivity and may become disscociated, and an endogenous mechanism may have affected peak androgen secretion. Testicular regression exhibited a dependence on an interaction among temperature, photoperiod, and endogenous regulation. Turtles with testes in advanced stages of spermiogenesis underwent accelerated regression when exposed to even slightly reduced temperatures, especially at short daylengths. However, regression was delayed by a combination of high temperature and short daylength in fall. Results are compared to other experimental studies of photothermal effects on turtles and discussed in terms of the described latitudinal variations in the timing of the testicular cycle of S. odoratus.
Fructose‐6‐phosphate is not a substrate for glucose‐6‐phosphate dehydrogenase
Authors: Jeffery Jonathan , Wood Irene
Published: June 2005

D‐Fructose‐6‐phosphate was shown not to be a substrate for glucose‐6‐phosphate dehydrogenases (EC. 1.1.1.49) from human erythrocytes, bovine adrenal, rat liver, three yeasts (brewer's yeast, baker's yeast, and Candida utilis), and Leuconostoc mesenteroides. These findings contrast with those of G.M. Kidder (J. Exp. Zool., 226:385–390, '83).
A new enzyme from Drosophila melanogaster: In vitro conversion of xanthommatin into its dihydroform by means of xanthommatin reductase
Authors: Santoro P. , Parisi G.
Published: June 2005

A new enzyme was partially purified from D. melanogaster. The enzyme catalyzes the reduction of xanthommatin to dihydroxanthommatin in the presence of NADH/H+ as confactor. FAD, FMN, p‐chloromercuribenzoate, 2‐amino‐4‐hydroxypteridine and biopterin inhibited the enzyme activity. The inhibitory effect of some pterines suggests a correlation between the synthesis of dihydroxanthommatin and that of pterines. It has been suggested that the enzyme may participate, in vivo, in the reduction of xanthommatin.
Oogenesis in Fundulus heteroclitus IV. Yolk‐vesicle formation
Authors: Selman Kelly , Wallace Robin A. , Barr Valarie
Published: June 2005

Yolk vesicles first appear within the oocyte of Fundulus heteroclitus in association with elaborate smooth endoplasmic reticulum and Golgi elements. Cytochemistry indicates the yolk vesicles contain an acidic glycoconjugate. [3H]Glucose is used to examine the synthesis and accumulation of yolk vesicles in studies performed both in vivo and in vitro, since it is selectively incorporated into nascent yolk vesicles. The endogenous synthesis of yolk‐vesicle glycoconjugate is documented by autoradiography, gel electrophoresis, and fluorography. A large (> 200 kDa) glycoconjugate has been identified, which is thought to reside in yolk vesicles. Incorporation studies done in vitro substantiate that this putative yolk‐vesicle material is synthesized by the oocyte itself and that synthesis occurs not only during the so‐called yolk‐vesicle stage, but also throughout vitellogenesis and early maturation. The view is thus adopted that various synthetic and physiological events are sequentially initiated and subsequently remain active throughout oocyte growth.
Basolateral localization of Na+ + K+‐ATPase in gill epithelium of two osmoregulating crabs, Callinectes sapidus and Carcinus maenas
Authors: Towle David W. , Kays W. Todd
Published: June 2005

Although Na+ + K+‐dependent ATPase, the enzymatic equivalent of the sodium pump, is thought to be restricted to the basolateral membrane of nearly all ion‐transporting epithelial cells, its absence from basolateral membranes of crustacean epithelia and/or its presence in apical membranes have been suggested by evidence from other laboratories. However, ultracytochemical localization of K+‐dependent, ouabain‐sensitive p‐nitrophenylphosphatase (p‐NPPase), using the method of Mayahara et al. ('80), indicated that Na+ + K+‐ATPase resides predominantly in the basolateral membrane of thick epithelial cells from gill lamellae of two euryhaline crabs, Callinectes sapidus and Carcinus maenas. Crab gill is therefore not unique in its Na+ + K+‐ATPase distribution and may be used as a model for experiments designed to investigate mechanisms of transepithelial Na+ uptake.
Isolation of Drosophila mutants with abnormal proboscis extension reflex
Authors: Kimura Ken‐Ichi , Shimozawa Tateo , Tanimura Teiichi
Published: June 2005

Drosophila mutants with defects in their proboscis extension reflex (PER) were isolated for a genetic dissection of the PER system. Five mutant strains were screened from 35,000 mutagenized flies through a twostep screening: a mass test of sugar intake, followed by an individual PER test. Mutants were classified into three complementation groups: aperA (genetical map position, 1–22.1), aperB (1–0.6) and aperC (1–0.4). The aperA mutants showed incomplete penetrance and expressivity. Some flies did not extend their proboscises at all, while others did so only to either the right or the left side; the PERs of the rest were apparently normal. The aperB mutants showed abnormality in the direction of the PER: they extended their proboscises backward, instead of straight forward. The aperB mutation was temperature‐sensitive and its phenotype was expressed at a culture temperature above 25°C. The aperC mutation caused a PER defect in aged flies. Within 2 days of eclosion they showed normal PER, but 2 or more days after eclosion most of them had become unable to extend their proboscises. Later, about half of the aperC mutants recovered the PER ability. The anatomical site of action of these mutations seems to be in the neuromuscular or skeletomuscular system.
Stage‐specific changes in protein phosphorylation accompanying meiotic maturation of mouse oocytes and fertilization of mouse eggs
Authors: Endo Yoshihiro , Kopf Gregory S. , Schultz Richard M.
Published: June 2005

Characteristic changes in the patterns of protein phosphorylation occur during meiotic maturation of mouse oocytes from the time subsequent to germinal vesicle breakdown, through metaphase II, and following fertilization. These changes occur during both in vitro or in vivo maturation or fertilization. Three major classes of changes in total phosphoprotein synthesis are observed. In the first class, protein phosphorylations increase from the germinal vesicle stage until just after germinal vesicle breakdown and then decrease during progression to metaphase II and after fertilization. The second class is characterized by decreases in protein phosphorylation during maturation with subsequent increases in phosphorylation of these proteins after fertilization. The third class is characterized by protein phosphorylations that remain relatively constant during maturation but increase after fertilization; phosphotyrosine phosphoproteins comprise the major species. The radiolabeled protein and phosphoprotein composition of isolated germinal vesicles was also examined, and a phosphoprotein of Mr29,000 is found exclusively associated with the germinal vesicle. Since we have shown previously that 12‐O‐tetradecanoyl phorbol 13‐acetate inhibits fertilization (Y. Endo, R.M. Schultz, and G.S. Kopf, submitted), we examined the effects of this compound on the phosphoprotein patterns of metaphase II eggs. 12‐O‐Tetradecanoyl phorbol 13‐acetate treatment stimulates the phosphorylation of a specific phosphoprotein of Mr80,000.
Pigment cell pattern formation in amphibian embryos: A reexamination of the dopa technique
Authors: Tucker R. P. , Erickson C. A.
Published: June 2005

Neural crest‐derived melanophores form species‐specific patterns in the dermis of amphibian embryos. Melanophore patterns may be generated by one of two general mechanisms: (1) pigment cell precursors disperse throughout the embryo, with melanophores differentiating in certain regions due to environmental cues, or (2) melanoblasts may localize in different regions as a result of a hierarchy of tissue affinities. Both of these mechanisms have been proposed to be responsible for the dorso‐ventral patterning of melanophores in Xenopus laevis. We have reexamined the distribution of melanoblasts in X. laevis and Taricha torosa using the dopa (3,4‐dihydroxyphenylalanine)‐staining technique. We have found that many of the dopa‐positive cells identified as melanoblasts by some researchers are actually not derived from the neural crest: (1) dopa‐positive cells in T. torosa were identified in the transmission electron microscope to be either leukocytes or erythrocytes, (2) in X. laevis dopa‐positive cells are found between the ectoderm and somites where neural crest cells are not found, and (3) X. laevis embryos surgically depleted of neural crest have dopa‐staining patterns identical to control embryos. Melanoblasts are apparently not found in the ventralmost regions of early T. torosa and X. laevis embryos, providing additional evidence for the role of differential tissue affinities in directing the formation of embryonic pigment cell patterns.
Phenotypic switching to long cilia effected by various proteases: Results With Dendraster excentricus and Stronglyocentrotus purpuratus blastulae
Authors: Riederer‐Henderson Mary Ann
Published: June 2005

Incubation in trypsin effects a phenotypic switch from short to long cilia (> 30 μ) in hatched blastulae of the sea urchin, Arbacia punctulata. To determine how trypsin causes such a switch we tested whether the phenomenon was unique to the species, Arbacia, and to the protease, trypsin. With two other echinoderm species, the sand dollar, Dendraster excentricus, and the sea urchin, Strongylocentrotus purpuratus, trypsin incubation increased the percentage of long cilia. During incubation of D. excentricus in trypsin, the percentage of long cilia increased progressively from the normal 10% long cilia of the apical tuft to 45–50% long cilia covering 1/2–3/4 of the embryo. With S. purpuratus blastulae, however, the percentage of long cilia was lower (32–40%) and the results were more variable. Of the additional proteases tested with D. excentricus, elastase was more effective than trypsin in terms of the percentage of long cilia (58%), the mean length, and the broad distribution of lengths fromed. Thermolysin was about as effective as tryspin but chymotrypsin was much less so. Thus, increases in ciliary length were not unique to a particular echinoderm species or to incubation in trypsin. The magnitude of the change in length distribution, however, was species‐ and enzyme‐dependent. An extracellular or membrane component with differential susceptibility to various proteases may, therefore, be involved in altering ciliary length.
Ovary‐stimulating pheromone in the freshwater shrimp, Paratya compressa
Authors: Takayanagi Hiroshi , Yamamoto Yoshihiro , Takeda Naokuni
Published: June 2005

In the female shrimp, Paratya compressa, reared in the absence of males, ovarian development was usually delayed, whereas it occurred normally in females reared with mature males. Ovarian development also occurred in shrimps reared in water that contained an extract of the testis or the vas deferens. Therefore, it is proposed that mature, male shrimps secrete an ovary‐stimulating pheromone which accelerates ovarian development, the source of which may be male organs such as the testis or the vas deferens.
Eye factors and lens‐forming transformations of outer cornea in Xenopus laevis larvae
Authors: Bosco L. , Filoni S. , Cioni C. , Bernardini S.
Published: June 2005

Outer cornea of lensectomized Xenopus laevis tadpoles at state 50 (according to Nieuwkoop, P.D. and Faber, J., ('56) Normal Table of Xenopus laevis, Daudin, North‐Holland, Amsterdam, pp. 1–243) was removed 3, 7 and 10 days after lensectomy and implanted between the outer and the inner cornea of larvae of the same species at stage 51–52. In these conditions, the implanted outer cornea remained isolated from the retinal factor of the vitreous chamber, although it received the nutritional factors normally reaching the outer cornea. Results show that lens‐forming transformation process of the outer cornea is arrested, and lens‐forming structures undergo regression at speed which increases with increasing precocity of the stage of lens‐forming transformation undergone by the implanted cornea. These data suggest that the process of lens‐forming transformation is not a single‐step process, but a sequence of interactions extending over a long period of time requiring the continuous presence of the retinal factor in the vitreous chamber until complete differentiation of the lens is achieved.
Immunochemical evidence that the single lactate dehydrogenase of lampreys is more similar to LDHB4 than to LDHA4 of hagfish
Authors: Baldwin J. , Lake P. S. , Moon T. W.
Published: June 2005

The tetrameric lactate dehydrogenases (LDH) of vertebrates contain several different subunits that arose by gene duplication. While the A and B subunits occur in all classes of gnathostomes, the enzymes of agnathans appear to represent two stages in the evolution of vertebrate LDH. Lampreys of the family Petromyzontidae have a single enzyme classified as LDHA4, while hagfish possess both A and B subunits which form only the two homopolymers LDHA4 and LDHB4. It is generally assumed that the original vertebrate LDH was an A4 type, that duplication to give the B subunit occurred prior to the divergence of lampreys and hagfish, and that modern lampreys subsequently lost expression of the B gene. Lactate dehydrogenases were purified from representatives of all three lamprey families, and it was confirmed that members of the Mordaciidae and Geotriidae also possess single tetrameric LDH enzymes containing one subunit type. The kinetic properties of the lamprey LDH enzymes were compared with the LDH homopolymers of hagfish, skate, and sardine. These properties did not allow the lamprey enzymes to be unequivocally identified as either LDHA4 or LDHB4. Immunochemical titration using antisera against lamprey and hagfish LDH homopolymers demonstrated that the lamprey LDH enzymes showed greater immunochemical similarity to LDHB4 than to LDHA4 of hagfish. It is concluded that there is little evidence for the claim that the original vertebrate LDH was an A4 rather than B4 type.
Protein uptake and transport by trophotaenial absorptive cells in two species of goodeid embryos
Authors: Schindler Joachim F. , De Vries Uwe
Published: June 2005

Near‐term embryos of two goodeid species (Xenotoca eiseni and Girardinichthys viviparus) have been exposed to horseradish peroxidase (HRP) and/or cationized ferritin (CF) within the ovarian cavity. The trophotaenial absorptive cells (TACs) of X. eiseni have an apical endocytotic complex and, therefore, readily ingest tracer molecules from ambient proteinaceous liquids. Internalized solutes accumulate in large apical storage vacuoles. In double‐tracer experiments, the two proteins are mostly sorted each into separate vacuolar compartments. After exposure for several hours, reaction product of HRP can be localized in a lamellar membranous complex, which underlies the peripheral plasma membranes and communicates with the intercellular space. Some vesicles carry small portions of ferritin molecules to the basal or basolateral cell surface and eventually exteriorize their contents. No activity of an acid phosphatase (AcPase) is testable in elements of the endocytotic system before and after exposure to tracer proteins. In G. viviparus, the TACs are lacking an endocytotic apparatus. Consequently, the rate of solute movements into the cells is relatively low, and the total amounts of ingested proteins remain rather moderate, even after 24 h of continuous exposure. The binding of CF to cell surface components apparently stimulates micropinocoytotic activity. During the endocytotic process, an apical canalicular system is formed, which subsequently develops into multivesicular bodies (MVBs). These were seen to arise more numerously if the tracer medium contains CF. The effect of a sorting mechanism, resulting in differently labeled storage compartments, is not observable in the TACs of G. viviparus. Some of the internalized ferritin is transferred to the basal and basolateral plasma membranes. The vesicular or short tubular transport vehicles seem to contain only ferritin granules, even when the marker medium is composed of CF and HRP. Clusters of CF molecules can be seen fairly regularly in the lamina lucida of the basement membrane and in erythrocytes of the underlying capillary plexus. A scheme is presented that depicts the mechanisms of protein uptake and transport of either HRP or CF by TACs of both G. viviparus and X. eiseni.