bluuebunny/crossref_metadata_2025 huggingface dataset:
The production of cloned fish in the medaka (Oryzias latipes)
Authors: Naruse K. , Ijiri K. , Shima A. , Egami N.
Published: June 2005

The measurement of cellular DNA content by DNA microfluorometry revealed that medaka embryos that were fertilized with normal sperm and exposed to heat shock (41°C for 3 min) or hydrostatic pressure (700 kg/cm2 for 10 min) at 85–95 min after insemination were tetraploid. Embryos fertilized with normal sperm and exposed to heat shock (41°C for 2 min at 2–3 min after insemination) were triploid. These results suggest that heat shock or hydrostatic pressure at 85–95 min after insemination arrests the first cleavage, while heat shock at 2–3 min after insemination arrests the second meiotic division.

Medaka clones have been produced by the following method: Eggs from orange‐red or variegated variety were activated by UV‐irradiated, genetically impotent sperm of wild‐type fish (UV sperm). The haploid eggs obtained were diploidized by preventing the first cleavage with heat shock or hydrostatic pressure to produce homozygous females. Each of the two homozygous females was mated with vasectomized male in isotonic balanced salt solution to collect unfertilized eggs. The collected eggs were activated with UV sperm and converted from haploid to diploid by arrest of the second meiotic division with heat shock. Hatched fry of each homozygous diploid (all females) were fed with a methyltestosterone‐containing diet (40 μg/gm diet) to produce sex‐reversed males, which were mated with brood females, and thus two cloned lines were obtained.
Fsh‐induced gonadal development in juvenile lizards, Eumeces obsoletus
Authors: Masson Greg R. , Guillette Louis J.
Published: June 2005

Juvenile lizards (Eumeces obsoletus) treated with FSH exhibited germ cell proliferation. However, only the first stages of oogenesis or spermatogenesis were elicited. No difference in spermatogenic stage was observed between treatment groups. Vitellogenesis was not observed in treated females although the primary (largest) follicles exhibited a granulosa composed of a single cell type indicative of a follicle just prior to yolking. In addition to gonadal effects, accessory structures which are steroid dependent exhibited changes. Oviductal weight increased significantly with FSH treatment. Histological examination revealed that both oviductal glands and oviductal epithelium increased in size as did epididymal cell heights in males. These structures are steroid dependent in adults. These data suggest that juvenile gonads respond to gonadotropin although not in a typical adult manner. Sexual maturation is not only dependent on gonadal responsiveness but on the maturation of the hypothalamohypophysial axis and thus, on gonadotropin secretion as well. In addition, a critical amount of lipid storage may be required for vitellogenesis and gonadal function.
Seasonal variations in body growth rates and circulating levels of growth hormone in the goldfish, Carassius auratus
Authors: Marchant Tracy A. , Peter Richard E.
Published: June 2005

Several times throughout the year, changes in serum growth hormone (GH) levels over a 24‐h period were determined in goldfish maintained under photoperiods and temperatures simulating natural (Edmonton) environmental conditions. In the goldfish a reproducible daily rhythm in circulating GH levels was not present at any time of the year. The average serum GH level over the daily sampling period and the instantaneous relative growth rate in goldfish sampled at the various times of the year were also determined. The highest mean daily serum GH levels were found in March and June, whereas the lowest level was found in goldfish sampled in November. Changes in mean daily serum GH levels were closely correlated to seasonal changes in daylength. The highest growth rate was found in goldfish sampled in July, whereas the lowest growth rates were found in February and March. Female goldfish exhibited a faster growth rate than male goldfish at certain times of the year, but sexual differences in growth rate were correlated with sexual differences in serum GH levels only in November when female goldfish had a higher serum GH level than male goldfish.
Sperm duct contractions mediate centrally evoked sperm release in goldfish
Authors: Dulka Joseph G. , Demski Leo S.
Published: June 2005

In order to determine the peripheral mechanisms underlying sperm release (SR) in goldfish, the contractile activity of the sperm ducts (SD) and testes were monitored during SR responses evoked by electrical stimulation of the brain. Electrical stimulation of the brain triggered testicular and SD contractions, and SR, while electrical stimulation of the genital nerve branch to the SD evoked only SD contractions and SR. Centrally activated SD contractions and SR were blocked by sectioning the SD genital nerve, while testicular contractions were unaffected. Testicular contractions do not appear necessary for centrally evoked SR since the response can be elicited from preparations in which the testes were separated from the SD. The results indicate that SR in goldfish is primarily mediated by the SD and not the testes. Testicular contractions may, however, serve to load the SD with milt. The functional significance of the central pathway(s) associated with SD and testicular contractions are discussed.
Development of pronuclei from human spermatozoa injected microsurgically into frog (Xenopus) eggs
Authors: Ohsumi Keita , Katagiri Chiaki , Yanagimachi Ryuzo
Published: June 2005

Human spermatozoa were demembranated with Triton X‐100 (TX) and injected into the mature eggs of Xenopus laevis. The nuclei of these spermatozoa decondensed and developed into pronuclei. Chromosomes did not appear in the eggs until the end of a 5‐hr incubation period. When the demembranated human spermatozoa were further treated with dithiothreitol (DTT) before they were injected into the eggs, the sperm nuclear decondensation and pronuclear development took place considerably faster than in spermatozoa treated with the detergent alone. By the end of the 5‐hr incubation period, decondensed chromatin threads or chromosome‐like structures appeared, but none of the eggs cleaved. When human spermatozoa were injected into fullgrown ovarian oocytes with intact germinal vesicle (GV) or oocytes which had matured without GV, the nuclei of a proportion of TX‐treated and all TX‐DTT‐treated sperm decondensed but showed no sign of developing into pronuclei. Sperm nuclei injected into maturing oocytes formed condensed chromatin fragments as long as the oocytes were not activated, but they transformed into pronuclei when the oocytes were stimulated with electric shock. These results indicate that the cytoplasmic factors responsible for the decondensation of human sperm nuclei are present in egg cytoplasm independent of GV‐materials. We also suggest that the factors controlling development of decondensed sperm nuclei into pronuclei are dependent on GV materials.
Cortical granule exocytosis is coupled with membrane retrieval in the egg of Brachydanio
Authors: Donovan Michael J. , Hart Nathan H.
Published: June 2005

Activation of the teleost (Brachydanio) fish egg includes the exocytosis of cortical granules, the construction of a mosaic surface consisting of the unfertilized egg plasma membrane and the limiting membranes of the cortical granules, and the appearance of coated and smooth vesicles in the cytoplasm (Donovan and Hart, '82). Unfertilized and activated eggs were incubated in selected extracellular tracers to (1) determine experimentally if cortical granule exocytosis was coupled with the endocytosis of membrane during the cortical reaction, and (2) establish the intracellular pathway(s) by which internalized vesicles were processed. Unfertilized eggs incubated in dechlorinated tap water or Fish Ringer's solution containing either horseradish peroxidase (HRP; 10 mg/ml), native ferritin (12.5 mg/ml), or cationized ferritin (12.5 mg/ml) were activated as judged by cortical granule breakdown and elevation of the chorion. Cells treated with HRP and native ferritin exhibited a delay in cortical granule exocytosis when compared with wateractivated eggs lacking the tracer. Each tracer was internalized through the formation of a coated vesicle from a coated pit. Since coated pits appeared to be topographically restricted to the perigranular membrane domain of the mosaic egg surface, their labeling, particularly with cationized ferritin, strongly suggested that the retrieved membrane was of cortical granule origin. Cationized ferritin and concanavalin A (Con A) coupled with either hemocyanin or ferritin labeled the surface of the unactivated egg and both domains of the mosaic egg surface. Transformation of the deep evacuated cortical granule crypt into later profiles of exocytosis was accompanied by increased Con A binding. Within activated egg cortices, HRP reaction product, native ferritin, and cationized ferritin were routinely localized in smooth vesicles, multivesicular bodies, and autophagic vacuoles. Occasionally, each tracer was found in small coated vesicles adjacent to the Golgi and within Golgi cisternae. The intracellular distribution of HRP, native ferritin, and cationized ferritin suggests that internalized membrane is primarily processed by organelles of the lysosomal compartment. A second and less significant pathway is the Golgi complex.
Regional differences in the histochemical localization of carbonic anhydrase in the midgut of tobacco hornworm (Manduca sexta)
Authors: Ridgway Richard L. , Moffett David F.
Published: June 2005

Carbonic anhydrase (CA) activity was assayed by a histochemical method in mounted cryostat sections of larval midgut of Manduca sexta (Insecta: Sphingidae). In anterior and middle midgut most CA activity was associated with apical microvilli of goblet cells; with occasional patchy staining of the columnar cell brush border in anterior midgut. In contrast, in the posterior midgut, the apical microvilli of goblet cells showed no CA activity, but intense staining was associated with the brush border of columnar cells. Staining of mitochondria and nuclei was seen in all midgut regions. In addition, midguts from late fifth instar larvae showed intense staining of spherites (mineral concretions). Acetazolamide (10−5M) completely inhibited staining at all sites except nuclei and spherites. Thiocyanate (100 mM) completely inhibited staining at all sites except anterior and middle goblet cell microvilli, which remained partially reactive. These results suggest that the anteriormiddle and posterior regions play different roles in generating and managing the alkalinization of midgut contents characteristic of lepidopteran midgut. The different sensitivities of CA activity in anterior‐middle and posterior midgut to thiocyanate raises the possibility that more than one form of the enzyme may be involved.
Incorporation of 14C‐bicarbonate into the free amino acid pool during hyperosmotic stress in an intertidal copepod
Authors: Burton Ronald S.
Published: June 2005

In response to hyperosmotic stress, the harpacticoid copepod Tigriopus californicus rapidly accumulates high intracellular concentrations of free alanine and proline. Using 14C‐bicarbonate as a tracer, in vivo experiments were carried out to determine if pyruvate or phosphoenolpyruvate carboxylation function in the synthesis of carbon skeletons for free amino acid (FAA) synthesis. Label was incorporated into four FAA (aspartate, glutamate, proline, and alanine) during hyperosmotic stress. Relative specific activities of the FAA synthesized in vivo and in vitro enzyme activities suggest that malic enzyme (EC 1.1.1.40) carries out the carboxylation reaction. Incorporation of label into proline was extensive for at least 24 hr after transfer of adults acclimated to 50% seawater to 100% seawater; the proline pool showed relatively slow turnover following its accumulation (approximately 40% drop in specific activity in 48 hr) compared to the other FAA (complete loss of label in 24 hr). While bicarbonate fixation occurs in animals acclimated to both 50% and 100% seawater, label is observed in glutamate and aspartate but not proline. The results of these experiments suggest that the metabolic pathway by which bicarbonate label is incorporated into proline is activated by hyperosmotic stress and is effectively shut down following 24–48 hr of proline accumulation.
Lack of adrenergic control of dorsal aortic blood pressure in the resting eel, Anguilla australis
Authors: Hipkins Scott F. , Smith David G. , Evans Barbara K.
Published: June 2005

Dorsal aortic and cardinal venous blood pressures, dorsal aortic blood flow, and heart rate were measured in different combinations in conscious unrestrained eels (Anguilla australis). One group of animals was treated with either bretylium (10 mg.kg−1) or saline vehicle and dorsal aortic blood pressure and heart rate were monitored for the next 6 days. Two other groups were treated with phentolamine (5 mg.kg−1) and either dorsal aortic blood pressure and heart rate, or dorsal aortic blood flow and venous pressure monitored for 6 hr. In vivo treatment with either bretylium of phentolamine produced effective blockade of the α‐adrenoceptor‐mediated vasoconstriction of stimulated saline‐perfused trunk preparations; however, neither drug had no lasting effects on arterial blood pressure. In addition, phentolamine had no lasting effects on venous blood pressures or dorsal aortic blood flow. It is concluded that, in the resting eel, systemic vascular resistance is not normally under adrenergic control.
Biochemical responses to temperature in the contractile protein complex of striped bass Morone saxatilis
Authors: Moerland Timothy S. , Sidell Bruce D.
Published: June 2005

The effects of acute and long‐term changes in temprature upon catalytic and calcium regulatory function of red (slow oxidative) and white (fast glycolytic) muscle from striped bass (Morone saxatilis) were determined. Acclimation to 5°C or 25°C had no significant effect on catalytic function (ATPase activity) or regulatory sensitivity (Ca++‐activation) of myofibrils from either muscle type. Substantial differences between red and white muscle were found in the intrinsic thermal sensitivity of maximally‐activated Mg++‐Ca++ myofibrillar ATPase. Arrhenius plots of myofibrillar ATPase from white muscle show one significant breakpoint at 29°C, with activation energies (Ea) of 2.3 and 23.4 kcal mole−1 at temperatures above and below this transition, respectively. Arrhenius plots of myofibrillar ATPase from red muscle show two transitions occurring at 22 and 9°C, with Ea of 7.6 kcal mole−1 above 22°C and 18.3 kcal mole−1 between 9 and 22°C. Activation energies for myofibrils from red muscle increase substantially to approximately 107.3 kcal mole−1 below the 9°C breakpoint. Differences in the intrinsic thermal sensitivity of red and white muscle catalytic function are apparently due to interaction of actomyosins and calcium regulatory proteins which are specific to each muscle type. The results suggest that capacity for sustained swimming in striped bass, which is powered exclusively by red muscle, will be severly impaired at cold temperature unless compensations occur above the level of contractile proteins.
Multiple hemoglobins of the cutthroat trout, Salmo clarki
Authors: Southard Jonathan N. , Berry Charles R. , Farley Thomas M.
Published: June 2005

Nine hemoglobins were purified from blood of Salmo clarki by ion‐exchange chromatography and preparative isoelectric focusing. The subunit structures of eight of the purified hemoglobins were studied by electrophoresis of globins in the presence of urea. Six are α2 β2 tetramers while two appear to be heterotetramers of the type αα′ β2 and αα′ββ′.

The effects of pH, nucleotides, and temperature on the oxygen equilibria of the purified hemoglobins were studied. Five hemoglobins with isoelectric points from 9.1 to 7.1 and one minor hemoglobin with an isoelectric point of 5.9 appear to have essentially identical oxygen binding properties. All have similar oxygen equilibria which are independent of pH and temperature and not affected by saturating amounts of ATP. Another minor hemoglobin with an isoelectric point below 5.9 has similar oxygen equilibria except for a possible pH dependence. Two hemoglobins, with isoelectric points of 6.5 and 6.4, have oxygen binding properties which are strongly pH and temperature dependent. Addition of ATP or GTP causes a large decrease in the oxygen affinity without affecting the cooperativity of oxygen binding. The effect of GTP is slightly greater than that of ATP. No significant differences were observed in the oxygen equilibria of these two hemoglobins.

The red blood cells of S. clarki were found to contain large amounts of both ATP and GTP, with an ATP:GTP ratio of 3:1. Both nucleotides may be important modulators of hemoglobin oxygen affinity in S. clarki, in contrast to the situation in S. gairdneri, in which red blood cell GTP concentrations are considerably lower.

The presence of six or possibly seven hemoglobins with identical oxygen binding properties in S. clarki suggests that, to a large extent, the physiological role of multiple hemoglobins in this species involves phenomena not directly related to the oxygen binding properties of the hemoglobins.
Influence of calcium‐perturbing agents on schistosomes: Comparison of effects of praziquantel and verapamil on worm tegument
Authors: Senft A. W. , Gibler W. B. , Guterman J. J.
Published: June 2005

A “slow calcium channel” blocking agent, Verapamil (VPL), induces tegumental disruption in schistosomes. The response of worms to VPL was compared to that caused by a widely used antischistosomal drug, Praziquantel (PZQ), whose mode of action has been related to a rapid influx Ca++.

Scanning microscopy of worms exposed to micromolar concentrations of either PZQ or VPL in vitro revealed a striking similarity of tegumental damage. At high dosage (10−5 M × 1 h), both compounds cause extensive surface bleb formation, which can proceed to shredding and denudation of the tegument. Early VPL‐induced changes in male Schistosoma mansoni appear to consist of small intertubercular lesions, as well as head and neck microblebs and bullae. After longer exposure, the bleb formations develop into large lesions, which after fixation for scanning microscopy, appear as flower‐like outgrowths on and around the tubercles. Females show much less surface disruption than males.

Physiologically, PZQ typically causes quick contraction of worm pairs into a tight, agonal coil. While VPL also induces spasm and clonic jerking, worms soon lengthen into a gentle coil. Both drugs kill the parasites in vitro.

The findings suggest that both PZQ and VPL have a surface target, whose disruption can be noted visually as lipoidal oozing and eventual tegumental blebbing. The inference drawn is that a direct affect on worm internal calcium metabolism, or on calcium channel activities, can lead to pathological changes which are grossly indistinguishable. These two drugs have markedly dissimilar structures, and thus probably different mechanisms of action. Yet, they result in similar final pathological events. These studies raise the possibility that various calcium‐perturbing agents might be useful candidates in a search for alternative or synergistic chemotherapeutic agents against trematodes and cestodes.
The determination of the shell field cells during the first hour in the sixth cleavage cycle of eggs of Ilyanassa obsoleta
Authors: Labordus V. , van der Wal U. P.
Published: June 2005

To investigate the possibility of a relation between the moment of deletion of macromeres and the morphogenetic potential of the resulting embryo, the 2D‐, the 3D‐, or the 3(A–D)‐macromeres were deleted at regular intervals during the fifth or the sixth cleavage cycle. Ectoblasts, ‐3(A–D), are capable of extensive cellular differentiation and the formation of several organs. The developmental capacity of the ectoblasts is independent of the stage of deletion.

Deletions of the D‐macromere at successive stages result in a decreasing number of internal shells; a minimum is reached 40–60 min after the fifth cleavage (24‐cell stage). Simultaneously, the number of external shells increases.

Deletions of the 3D‐macromere during the first 60 min of the sixth cleavage cycle (24‐cell stage) lead to highly teratomorphic larvae; rather normal‐shaped larvae, but without heart and intestine, develop following deletions after 60 min. The moment of deletion of the D‐macromere affects morphogenesis.

These results lead to the following hypothesis: each quadrant may induce an internal shell, irrespective of factors derived from the polar lobe; an internal shell is a differentiated non‐evaginated shell gland. During the first hour of the sixth cleavage cycle the 3D‐macromere suppresses shell field formation in the A‐, B‐, and C‐quadrants, and induces evagination of the shell gland in the D‐quadrant resulting in the formation of an external shell. At the same time the morphogenetic potential of the embryo is enhanced.
Stress‐induced suppression of testosterone secretion in male alligators
Authors: Lance Valentine A. , Elsey Ruth M.
Published: June 2005

In order to test the effect of acute stress on gonadal hormone secretion in reptiles, six mature male alligators were captured, and a blood sample was taken within 5 min of capture. Additional blood samples were taken at timed intervals for up to 41 hr, and plasma testosterone and corticosterone were measured by radioimmunoassay. Plasma testosterone declined to 50% of the initial value by 4 hr and dropped to less than 10% of initial by 24 hr. Plasma corticosterone increased during the first 12 hr, declined at 24 hr, and rose again at 40 hr. Blood samples from male alligators collected in North and South Carolina, south Florida, and in south Louisiana in two consecutive breeding seasons were also assayed for testosterone and corticosterone. In these populations there were significant differences in mean plasma testosterone and corticosterone levels. Elevated corticosterone levels were consistently seen in alligators caught in traps and from which a blood sample was taken several hours later. Plasma testosterone, although consistently lower in trapped alligators, did not show a negative correlation with plasma corticosterone. Farm‐reared alligators bled once, released, and bled again at 24 hr also showed a highly significant suppression of testosterone secretion. These results demonstrate that stress has a rapid and dramatic effect on testicular steroid secretion in both farm‐reared and wild alligators.
Mouse embryos develop normally Exo utero
Authors: Muneoka K. , Wanek N. , Bryant S. V.
Published: June 2005

Surgical procedures are described which permit direct experimental access to postimplantation mouse embryos. These procedures arose from our finding that development proceeds normally to term after embryos have been released from the confines of the uterus, while remaining attached to it via the placenta. Embryos continue to develop exo utero, within the abdominal cavity of the mother, and are capable of surviving a variety of different surgical manipulations. This finding opens the way for experimental analyses of mouse development in vivo.
Abnormal regeneration in the planarian Dugesia tigrina as a function of the length: Width ratio of the regenerating fragment
Authors: Mead Robert W. , Krump Martha A.
Published: June 2005

Regeneration was examined in different regions of planaria (Dugesia tigrina) in order to determine the effect that the shape of a section exerts on regenerative success. Length:width ratios were used as an index of tissue proportion, and overall regenerative success was measured by percent of abnormal regenerates and number of abnormalities per abnormal regenerate. Frequency and number of abnormalities increased as section width exceeded section length. Frequency of specific abnormalities varied with changes in length:width ratios and followed a predictable pattern. As ratios decreased below 1.0, abnormalities typically consisted of characteristics regarded as being due to excess head formation. These included the presence of two heads, the “head hump” syndrome, and/or lack of pharynx. This excess head formation was due to the proportional shape of the section rather than to overall segment size, and the frequency of such abnormalities increased in proportion to decreases in length:width ratios. Abnormalities which were more typically seen at ratios over 1.0 consisted of the lack of head, pharynx, and/or polarity.
Dependence of GnRH action on Na+, K+, and Ca2+ in the frog, Rana pipiens, pituitary
Authors: Porter David A. , Licht Paul
Published: June 2005

The roles of K+, Ca2+, and Na+ ions in the mechanism of gonadotropin releasing hormone (GnRH) action on frog (Rana pipiens) hemipituitaries were studied using an in vitro superfusion system. The effects of elevated K+ alone or in combination with Ca2+ ‐depleted medium, tetrodotoxin (TTX), or with 100 ng/ml GnRH were examined. The involvement of K+ was also studied indirectly through the use of tetraethyl ammonium chloride (TEA). The importance of Ca2+ was established by the loss of responsiveness to GnRH in Ca2+ ‐depleted medium, or in the presence of the Ca2+ competitor CoCl2. The absence of a major dependence of GnRH on Na+ was revealed by the continued gonadotropin secretion after addition of 1 μM TTX to medium containing GnRH or 36.3 mM KCl, or by replacement of NaCL with choline chloride. High (10 × normal) KCl (36.3 mM) stimulated gonadotropin‐both LH and FSH‐secretion, but the response was more gradual than for GnRH. The inclusion of TEA (to block K+ efflux) in medium with GnRH accentuated the effect of GnRH, and the effects of elevated (36.3 mM) KCl and 100 ng/ml GnRH (a relatively high dose) were additive. Responses to high K+, like GnRH, were abolished by removal of Ca2+ from the medium. Overall, the roles of K+, Ca2+, and Na+ ions in the mechanism of GnRH action are very similar between mammals and frogs; Ca2+ apparently serves a critical function in the mechanism of GnRH action, while Na+ appears not to be involved. K+ can induce gonadotropin secretion, but it is not clear that it plays a direct role in the mediation of the action of GnRH.
Calcium effects on progesterone accumulation and oocyte maturation in cultured follicles of Rana pipiens
Authors: Francisco Susan Kleis‐San , Schuetz Allen W.
Published: June 2005

Pituitary homogenates (FPH) provoke a cascade of responses in the amphibian ovarian follicle, culminating in progesterone biosynthesis and oocyte maturation (GVBD). Calcium may play an important role as an intracellular second messenger in regulating these physiological responses. Experiments were carried out on cultured, isolated follicles of Rana pipiens to assess the effects of varying extracellular calcium on follicular progesterone accumulation and oocyte maturation. In hormonally unstimulated follicles, an increase in extracellular Ca2+ alone produced a significant increase in progesterone in methanol extracts of follicles after 4 hours of culture, and in some cases also provoked oocyte maturation assessed after 24 hours of culture. In no case did elevated Ca2+ alone stimulate maximal progesterone accumulation as compared with FPH‐stimulated follicles, although the time‐course of accumulation was similar. The calcium ionophore, A‐23187, similarly increased progesterone accumulation in a dose‐dependent manner when introduced in amphibian Ringer's (1.35 mM Ca2+), but inhibited progesterone elevation caused by increasing calcium concentrations in the culture media and FPH stimulation. Depleting free calcium from the culture medium with graded doses of the chelator EGTA decreased FPH‐induced progesterone accumulation and inhibited FPH‐ and progesterone‐induced GVBD. The calcium channel blocker, verapamil, also inhibited FPH‐induced progesterone accumulation and GVDB in a dose‐dependent manner, while having no effect on progester‐one‐induced meiotic resumption. These data strongly implicate intracellular calcium levels regulating progesterone production by ovarian follicle cells and subsequent oocyte maturation.
Impact of symbiotic algae on sea anemone metabolism: Analysis by in vivo 31P nuclear magnetic resonance spectroscopy
Authors: Steen R. Grant
Published: June 2005

High‐field pulsed Fourier‐transform nuclear magnetic resonance spectroscopy (NMR) was used to quantify the adenylate levels of sea anemones (Aiptasia pulchella) with and without symbiotic dinoflagellates (Symbiodinium sp.). Animals were fed to repletion, then starved in darkness for up to six days before collection of in vivo NMR spectra. The host adenylate ration of ATP: (ATP + ADP) declined significantly with increasing perioids of starvation in both symbiotic and aposymbiotic hosts (P<0.05). However, the decline in the animal adenylate ratio was significantly more rapid in animals bearing symbiotic algae (P<0.05). This suggests that symbiotic algae in darkness cause more rapid depletion of host energy reserves, possibly by drawing on host pools of organic substrates. In vivo NMR spectroscopy was also used to evaluate the effect on A. pulchella of photosynthesis by zooxanthellae. Symbiotic anemones were fed to repletion, then starved under high irradiance (300 to 320 μEin m−2 s−1) or low irradiance (70 to 80 μEin m−2 s−1) conditions for up to five days. The host adenylate ratio declined significantly (P<0.01) with starvation under both treatments, but no significant difference was detected between treatments (P<0.35). Blotted wet weight of anemones under high and low irradiance declined by 50% over eight days of starvation, but there was no significant difference in the rate of weight loss by anemones in the two treatments. There results suggest that translocation of photosynthate from symbiotic zooxanthellae does not significantly affect host adenylate ratio or have a sparing effect on host biomass during starvation in this symbiotic sea anemone.
Expression of homeotic genes in Bombyx mori estimated from asymmetry of dorsal closure in mutant/normal mosaics
Authors: Shimada Toru , Ebinuma Hiroyasu , Kobayashi Masahiko
Published: June 2005

Genetic mosaics of homeotic genes, ENk and ESw, in the silkworm, Bombyx mori, were made using a hereditary mosaic gene, mo. Deformed segmentation, due to asymmetric integumental fusion during dorsal closure, was observed in the mosaics. Left and right integuments that contained an ENk or ESw gene on either side did not fuse symmetrically on the dorsal center line. The asymmetric fusion was observed only in places where the dorsal center lines corresponded to the border line between genotypes. In mosaics of ENk and +E, the sides of segments containing ENk genes frequently fused with anterior parts of the wild‐type side, in the region from the metathorax to the fourth adbominal segment, while on the sixth through eighth abdominal segments they fused with the posterior parts of the opposite sides. In mosaics of ENk and +E, asymmetric fusion was observed in a narrower region, from the metathorax to the second abdominal segment. In this region, the sides containing ESw genes frequently fused with posterior parts of the wild‐type side, in contrast to the fusion observed for ENk mosaics. The cause of asymmetric fusion and expression of the homoeotic genes in degmental differentiation are discussed in terms of a quantitive analysis of the relationship between mosaic patterns and asymmetric patterns of fusion.
Factors involved in the testicular development from fetal mouse ovaries following transplantation
Authors: Taketo‐Hosotani Teruko
Published: June 2005

We have previously shown that fetal mouse ovaries develop testicular structures after transplantation into adult male mice. The mechanisms of gonadal sex reversal is poorly understood. In the present study, we examined how a host environment is involved in the induction of testicular development in ovarian grafts. Fetal ovaries on the twelfth day of gestation were microencapsulated with semipermeable membranes, transplanted beneath the kidney capsules of adult male mice, and fixed for histological examinations between the sixteenth and twenty‐second day after transplantation. Fifteen of forty‐seven ovarian grafts were found to be completely enclosed in microcapsules, whereas the microcapsule membranes of other grafts were partly broken or had been lost. These differences of microencapsulation conditions made it possible to study the role of host factors in gonadal sex reversal. All ovarian grafts surrounded by microcapsule membranes developed ovarian structures. In contrast, most ovarian grafts which had lost the microcapsules developed testicular structures in addition to ovarian structures. When ovarian grafts were partially enclosed in microcapsule membranes, testicular structures developed only in the area in contact with the host kidney. These results suggest that direct interaction between the ovarian graft and cells or large macromolecules from the host is involved in the development of testicular structures in ovarian grafts.
Post‐testicular change in the reptile sperm surface with particular reference to the snake, Natrix fasciata
Authors: Esponda P. , Bedford J. M.
Published: June 2005

Sperm surface changes occurring in the reptile Wolffian duct have been explored with particular references to the snake, Natrix fasciata. In the snake Wolffian duct there are several proteins not present in serum, the pattern of which changes in concert with the seasonal testicular cycle. Whereas testicular spermatozoa did not bind antibody to duct secretions, all Wolffian duct spermatozoa did so over both head and tail, according to immunofluorescence patterns. Thus, on entering the Wolffian duct, the entire surface of N. fasciata spermatozoa acquires one of more of the duct's secretory components.

As indicated by immunofluorescence, immunoelectrophoresis, and immunodiffusion, epitopes on at least some molecules that bind to spermatozoa or that remain free in the duct fluid are shared with those in other Natrix species, but not in more distant reptiles (turtle, anole lizard), nor chicken, rat, or rabbit. In regard to glycoproteins, one prominent con A‐reactive band was present in polyacrylamide gel electrophoresis (PAGE) of snake fluid and more were evident in fluid collected from the turtle duct. However, such lectin‐reactive elements did not bind to spermatozoa as judged by an absence of any change in snake, turtle and lizard sperm lectin‐binding patterns in passing from the testis into and through the Wolffian duct.

In all, evidence from these and other species studied begins to suggest that the nature of the post‐testicular sperm surface modification displayed in most vertebrates that fertilize internally may differ in sub‐therian and therian groups, respectively. There appears to be a relative emphasis on glycosyl‐rich surface elements in the latter. The possible significance of these changes for sperm function in the different groups is discussed briefly in terms of sperm survival/storage, as well as capacitation and sperm binding to the zona.
Reexamination of metabolic potential in the toadfish sonic muscle
Authors: Walsh Patrick J. , Bedolla Cindy , Mommsen Thomas P.
Published: June 2005

Activities of eight enzymes were measured in the sonic muscle of the gulf toadfish, Opsanus beta, to determine the metabolic poise of this unique tissue and to evaluate potential sex related differences in metabolism. In contrast to a prior study (Pennypacker et al., '85, J. Exp. Zool., 239:259–264), we observed substantial activities of M4‐lactate dehydrogenase, 333 to 482 units/g wet sonic muscle weight. This observation and the presence of high activities of other enzymes of glycolytic and anaerobic metabolism (pyruvate kinase and creatine phosphokinase) lead us to conclude that this tissue has high anaerobic capacity. Also in contrast to the observations of Pennypacker et al. ('85), we found that the activities of some enzymes indicative of aerobic metabolism are relatively low. For example, the activities of citrate synthase found in sonic muscle (1.5 to 2.7 units/g) are only slightly higher than values obtained for toadfish white skeletal muscle (1.2 units/g). The discrepancies between the results obtained by the two studies appear to be methodological ones. Lastly, significant differences in enzyme activities between males and females were observed for lactate dehydrogenase, malate dehydrogenase, and citrate synthase, and possible explantions for these differences are discussed.
Temperature‐sensitive periods of sex determination in a lizard: Similarities with turtles and crocodilians
Authors: Bull J. J.
Published: June 2005

The sex of leopard geckoes is determined by incubation temperature. This study identified their temperature‐sensitive period of sex determination by incubating embryos at a male‐producing temperature (32°C) and a female‐producing temperature (26°C) for various intervals of development and comparing the sex ratios from the different experiments. Eggs incubated at 26° for 14 days (to stage 32) could still develop as either sex, but eggs incubated 30 days (through stage 34) necessarily became female; incubation at 32° did not guarantee male development until 20 days of incubation (to stage 37). The temperature‐sensitive period of irreversible sex determination therefore lies between stages 32–37 and occupies much of the first half of development. The temperature‐sensitive period in geckoes is similar to those in turtles and alligators, which are only distantly related to geckoes, but the reasons for these similarities are obscure.
Role of carbonic anhydrase in calcification in the gorgonian Leptogorgia virgulata
Authors: Kingsley Roni J. , Watabe Norimitsu
Published: June 2005

The relationship between carbonic anhydrase and calcification in the gorgonian Leptogorgia virgulata was examined by analysis of the enzyme's activity, its cellular locations and its response to inhibition by Diamox. Enzyme activity was 2.7 nmol μg−1 protein s−1 at 4°C. Carbonic anhydrase localization at the light level using immunofluorescent techniques and at the ultrastructural level using Hansson's method showed activity in the scleroblasts and axial epithelium. Specifically, at the electron microscopic level this activity was found to be very high on the membranes of the spicule‐forming vacuole and electron‐dense bodies. Membranes of the microvilli and plasma membranes of the desmocytes showed high activity of this enzyme. The activity on the scleroblast plasma membranes was variable. This enzyme is possibly present in the axis. Regenerated branch tips incubated in seawater containing 10−4 M and 10−6 M Diamox showed an increase in 45Ca uptake into the spicules and axes. The tissue o fbranch tips incubated in 10−6 M Diamox showed a decrease in 45Ca uptake. The observation that carbonic anhydrase is located adjacent to calcifying structures and organelles associated with spicule formation suggests that the enzyme is involved in calcification. The increase in 45Ca uptake from enzyme inhibition may suggest that carbonic anhydrase is involved in the regulation of HCO3.
Effects of changes in Ca2+ concentration of the culture medium on the in vitro development of 11‐day mouse embryos
Authors: Nakashima K. , Fujiki Y.
Published: June 2005

The present study was designed to examine the effects of changes in Ca2+ concentration in a culture medium on the development of 11‐day mouse embryos (plug day = day 0). The embryos were individually cultured for 24 hours by rotating bottles using a culture medium of 25% rat serum and 75% Waymouth's medium. The Ca2+ concentration was changed by adding EGTA or CaCl2 to the Waymouth's medium. The Ca2+ concentrations examined were 0.5, 1.0, 1.5, 2.0, and 3.0 mM in 95% oxygen and 1.0, 2.0, and 3.0 mM in 60% oxygen. Development of embryos was evaluated on the basis of heartbeat, survival, growth, and differentiation. In 95% oxygen, the best survival was obtained at 1.5 mM Ca2+, while growth and differentiation were little affected by changes in Ca2+ concentrations except at 0.5 mM. In 60% oxygen, rhythmical heartbeats were difficult to maintain in many of the embryos at all Ca2+ concentrations examined, although reasonable development was obtained at 3.0 mM Ca2+. The optimal conditions were a culture medium containing 1.5 mM Ca2+ and an oxygen concentration of 95%. The addition of Waymouth's medium to the rat serum is useful in controlling the Ca2+ concentration of the culture medium.
Age‐dependent changes in fluorescent neurons in the brain of Notoplana acticola, a polyclad flatworm
Authors: Hauser Melinda , Koopowitz Harold
Published: June 2005

The formaldehyde‐glutaraldehyde‐sucrose (FGS) method for in situ localization of catecholamines has been applied to the nervous system of the marine polyclad flatworm Notoplana acticola. This histochemical fluorescence technique revealed the presence of a small population of fluorescent cells within the brain. The number and positions of these neurons were constant in animals of the same size, but varied with the size of the worm. The brains of small animals (8 mm in length) were found to contain 20 fluorescent cells, whereas the largest animals studied (30 mm in length) were found to have 28 such cells. Various intermediate cell numbers were found in animals between these two sizes. The origin of the newly added fluorescent cells is uncertain. Peripheral fluorescence was found in association with the tentacular ocelli (eyespots) and interneurons within the ventral submuscular nerve plexus. The fluorescent spectrum from these cells measured in situ had a λmax of 526 nm. Treatment with HCl shifts this peak to 530 nm. L‐dopamine fluoresces with a similar peak emission before HCl treatment (525.5 nm) and shifts to the appropriate longer wavelength (530 nm) following acidification. This strongly suggests that the fluorescent substance in the neurons is dopaminergic in nature.
Heat shock protein induction and induced thermal tolerance are independent in adult salamanders
Authors: Easton Douglas P. , Rutledge Paul S. , Spotila James R.
Published: June 2005

Ectothermic vertebrates become thermally tolerant (heat hardened) after exposure to heat shock. Eukaryotic cells show a similar response. Cellular thermal tolerance is correlated with the induction of heat shock proteins (hsps). We have investigated the relationship between heat hardening in salamanders and the induction of hsps in the tissues of these organisms. Although the synthesis of hsps can be induced in these animals by sublethal heat shocks, conditions required for hsp induction and heat hardening often do not coincide. We conclude that induced thermal tolerance in adult salamanders is independent of hsp induction in their tissues.
Some effects of adrenaline on anion transport and nitrite‐induced methaemoglobin formation in the rainbow trout (Salmo gairdneri Richardson)
Authors: Williams E. M. , Eddy F. B.
Published: June 2005

The effects of adrenaline on branchial anion transport and nitrite‐induced methaemoglobinaemia have been investigated in rainbow trout. Nitrite uptake and efflux results suggest that adrenaline effects a net anion efflux principally by stimulation of the unidirectional branchial anion efflux. In oxygenated whole blood nitrite‐induced methaemoglobin was significantly reduced in the presence of adrenaline. The physiological and environmental consequences of nitrite‐induced stress are discussed.
Craniofacial development mutant pi (pinhead) in the Axolotl (Ambystoma mexicanum) which exhibits reduced interocular distance
Authors: Neff Anton W. , Briggs F. , Chung H.‐M.
Published: June 2005

The genetics, gross morphology, histology, and experimental production of a phenocopy of a new mutant gene in the Mexican axolotl (Ambystoma mexicanum) are described. The mutant phenotype, termed pin‐head (pi), is characterized at the hatching stage by craniofacial abnormalities including reduced interocular distance (including cases of cyclopia). Rostral head structures such as the olfactory organs, eyes, forebrain, and oral cavity are primarily affected. More posterior structures beginning with the pharynx, otic vesicles, hindbrain, notochord, and gills appear normal. The mutant gene shows variable penetrance and appears to be a simple recessive with increased penetrance when fertile eggs are reared to the hatching state at 10°C (cold sensitivity). Evidence that this gene may affect prechordal mesoderm and plate development is presented. Phenocopies of the pi mutant were produced by the surgical removal of the prechordal mesoderm and plate from neural plate stage embryos.
Androgenic regulation of epididymal function in the skink, Mabuya carinata (Schn.)
Authors: Shivanandappa T. , Sarkar H. B. Devaraj
Published: June 2005

Androgenic control of epididymal function in reptiles has been studied in the skink, Mabuya carinata, by castration and androgen administration on histological, histochemical, and biochemical parameters. Castration of male skinks during the reproductive phase (Nov.–Dec.) leads to atrophy and regression of the epididymal epithelium with histochemically demonstrable loss of acid and alkaline phosphatase activities in the epithelium. Biochemically, reduction in DNA and protein contents and marked decrease in the activities of acid and alkaline phosphatases were observed. Administration of testosterone propionate (250 μg per animal on alternate days for 24 days) leads to hypertrophy of the epididymis, increased cell height, and histochemically demonstrable induction of acid and alkaline phosphatases in the epithelium. DNA and protein levels were also restored. In the quiescent phase (April–June) when the epididymis is fully regressed, the histological, histochemical and biochemical profile is comparable to that of castrated lizards in the reproductive phase. Administration of testosterone propionate to intact lizards leads to hypertrophy and stimulation of all the parameters, reminiscent of the condition in the reproductive phase. Alkaline phosphatase was found to be highly sensitive to androgenic deprivation and stimulation.
Thyroxine content of eggs and larvae of coho salmon, Oncorhynchus kisutch
Authors: Kobuke Louise , Specker Jennifer L. , Bern Howard A.
Published: June 2005

The whole‐animal content of the thyroid hormone thyroxine (T4) was measured in embryonic and larval coho salmon (Oncorhynchus kisutch) by radioimmunoassay following the validation of an extraction method. The embryonic yolk, at 20 days after fertilization and 10 days before hatching, was found to have 5.3 ± 0.3 ng T4 (mean ± SEM). At hatching, the total T4 content decreased to 4.4 ± 0.3 ng, of which 96% remained in the yolk and 4% was distributed in the larva. T4 content decreased to 1.5 ± 0.1 ng during the 30 days of larval development. The first increases in T4 were detected within 2 weeks following emergence from the substrate of the actively feeding juvenile; the newly emerged salmon responds significantly to bovine TSH. The presence and clearance of T4 from the yolk suggest that T4 may be critical to normal larval morphogenesis, that initially the T4 is of maternal origin, and that the hypothalamo‐adenohypophysial‐thyroid axis begins to function independently at or near the time of emergence.
Lactate dehydrogenase homopolymer of hagfish heart and the single lactate dehydrogenase of lampreys display greater immunochemical similarity to LDHC4 than to LDHB4 of teleost fish
Authors: Baldwin J. , Lake P. S.
Published: June 2005

Antisera produced against the single LDH enzymes from representatives of all three lamprey families and against the LDHA4 and LDHB4 enzymes of hagfish were tested for their ability to cross react with LDHC4 and LDHB4 purified from liver and heart of the Southern bastard red cod (Pseudophycis barbata, Order Gadiformes). The lamprey and hagfish heart LDH enzymes displayed greater immunochemical similarity to cod LDHC4, a result consistent with the view that the C subunit predates the B subunit.