Description

The Journal of Experimental Zoology,the foremost journal in the field, has a long-standing tradition of reporting the results of original research of an experimental or analytical nature in zoology, including investigations of all levels of biological organization, from the molecular to the organismal. Topical subdivisions are devoted to developmental biology, cellular biology, genetics and comparative genomics, neurobiology, comparative physiology and biochemistry, reproductive biology, and endocrinology. A separate section of the journal, Molecular and Developmental Evolution, is devoted to experimental approaches to evolution and development. Professor Günter Wagner, Yale University, serves as Editor-in-Chief.

Publisher details

John Wiley & Sons

Publications in this journal

• Ultrastructure of the male genital ducts of the clearnose skate Raja eglanteria.

  Authors: Carolyn J P Jones, William C Hamlett

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(12):1018-29.

  Tissues from the male genital ducts of six specimens of the clearnose skate Raja eglanteria, comprising the Leydig gland, upper and lower epididymis, ductus deferens and seminal vesicle, were fixedTissues from the male genital ducts of six specimens of the clearnose skate Raja eglanteria, comprising the Leydig gland, upper and lower epididymis, ductus deferens and seminal vesicle, were fixed and embedded for ultrastructural examination. In the Leydig gland, two types of columnar cells were identified, one bearing microvilli, a basal nucleus and evidence of active secretion with plentiful endoplasmic reticulum and numerous secretory droplets, and the other pyriform with cilia, and swathes of cytofilaments emanating from prominent desmosomes. Occasional crystalloid intramitochondrial inclusions were seen in the first type, with a periodicity of 24 nm. The upper epididymis was composed of cuboidal cells with microvilli and cilia and irregular electron dense granules, some of which were basally situated and extremely large, often within cells resembling intraepithelial leucocytes; such cells were also seen in the stroma underlying the epithelium. The lower epididymis cells also bore microvilli and cilia and were heavily vacuolated with fatty inclusions as well as the granule-laden leucocytes seen previously. In the ductus deferens, cells had masses of long cilia with occasional microvilli; endoplasmic reticulum was well developed, forming complex arrays with sparse secretory droplets and basal mitochondria. In the seminal vesicle there were two cell types, the most common having long cilia and short microvilli and an occasional, paler cell with supranuclear accumulations of small, round mitochondria. These ultrastructural appearances have been related to cell glycosylation and functions including protein secretion, water absorption and waste removal, and illustrate how structure and function vary down the length of the genital tract.

• Nitrogen metabolism and excretion in the aquatic chinese soft-shelled turtle, Pelodiscus sinensis, exposed to a progressive increase in ambient salinity.

  Authors: Serene Min Lin Lee, Wai Peng Wong, Kum Chew Hiong, Ai May Loong, Shit Fun Chew, Yuen Kwong Ip

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(12):995-1009.

  This study aimed to determine effects of 6-day progressive increase in salinity from 1 per thousand to 15 per thousand on nitrogen metabolism and excretion in the soft-shelled turtle, PelodiscusThis study aimed to determine effects of 6-day progressive increase in salinity from 1 per thousand to 15 per thousand on nitrogen metabolism and excretion in the soft-shelled turtle, Pelodiscus sinensis. For turtles exposed to 15 per thousand water on day 6, the plasma osmolality and concentrations of Na+, Cl- and urea increased significantly, which presumably decreased the osmotic loss of water. Simultaneously, there were significant increases in contents of urea, certain free amino acids (FAAs) and water-soluble proteins that were involved in cell volume regulation in various tissues. There was an apparent increase in proteolysis, releasing FAAs as osmolytes. In addition, there might be an increase in catabolism of certain amino acids, producing more ammonia. The excess ammonia was retained as indicated by a significant decrease in the rate of ammonia excretion on day 4 in 15 per thousand water, and a major portion of it was converted to urea. The rate of urea synthesis increased 1.4-fold during the 6-day period, although the capacity of the hepatic ornithine urea cycle remained unchanged. Urea was retained for osmoregulation because there was a significant decrease in urea excretion on day 4. Increased protein degradation and urea synthesis implies greater metabolic demands, and indeed turtles exposed to 15 per thousand water had significantly higher O2 consumption rate than the freshwater (FW) control. When turtles were returned from 15 per thousand water to FW on day 7, there were significant increases in ammonia (probably released through increased amino acid catabolism) and urea excretion, confirming that FAAs and urea were retained for osmoregulatory purposes in brackish water.

• Uterine motility in the reptile Anolis carolinensis: interactive effects of tension, prostaglandins, calcium, and vasotocin.

  Authors: Richard E Jones, Kristin H Lopez, Harriet B Austin, David J Orlicky, Cliff H Summers

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(12):1030-40.

  Uteri of Anolis carolinensis exhibited spontaneous rhythmic contractions in vitro. Addition of arginine vasotocin (AVT) caused an immediate, strong, tonic contraction followed by rhythmicUteri of Anolis carolinensis exhibited spontaneous rhythmic contractions in vitro. Addition of arginine vasotocin (AVT) caused an immediate, strong, tonic contraction followed by rhythmic contractions with the same frequency as spontaneous contractions but of a greater amplitude. At low tension (1.5 g) the AVT-induced tonic contraction was blocked by low dose of indomethacin, suggesting that it is influenced by calcium rather than prostaglandins (PGs). An increase in tension (from 1.5 to 15 g) reduced the duration of the AVT-induced tonic contraction; this stretch-induced decrease was also blocked by indomethacin. Stretch also decreased the duration of the rhythmic contractions, but this stretch effect was not inhibited by indomethacin. The rest interval between rhythmic contractions was decreased by PGF2alpha and PGE2, and indomethacin or stretch blocked these PG effects. Indomethacin, AVT, or stretch alone did not affect PGF2alpha secretion from AVT-treated uteri. Stretch also reduced PGF2alpha secretion from AVT-treated uteri, an effect inhibited by indomethacin.

• Sieving functional morphology of the gill raker feeding apparatus of atlantic menhaden.

  Authors: Kevin D Friedland, Dean W Ahrenholz, Joseph W Smith, Maureen Manning, Julia Ryan

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(12):974-85.

  Menhaden occupy an important position in estuarine food webs, thus the rate processes associated with their feeding are critical to the ecosystem management of fishery and ecological resources.Menhaden occupy an important position in estuarine food webs, thus the rate processes associated with their feeding are critical to the ecosystem management of fishery and ecological resources. Atlantic menhaden feed on a wide range of plankton, the size and food quality of which change ontogenetically. We analyzed the functional morphology of the menhaden feeding apparatus in a size series of menhaden representative of juveniles and the adult migratory stock. The physical dimensions of gill arches and rakers increased isometrically with fish length; however, branchiospinule spacing, the dimension that forms the sieve apertures of the branchial basket, scaled allometrically with fish length. Juvenile menhaden from North Carolina have branchiospinule spacings that averaged 12 microm, with three arch subsections of average spacing < 10 microm. Spacings did not increase with juvenile growth until the first allometric inflection point at approximately 100 mm fork length (FL). Spacing data for juveniles from other locations suggests spacing increases with latitude. Spacings increase with fish length in adults until a second inflection at 200 mm FL, after which spacing averages 37 microm. These data suggest menhaden juveniles filter smaller plankton with higher filtration efficiency than previously considered and that regional recruitment may affect adult distribution through foraging preferences.

• Regulation of immunocompetence by different androgen metabolites in a blenny with alternative reproductive tactics.

  Authors: Albert Frank Huascar Ros, Catarina Ferreira, Ricardo Serrão Santos, Rui Filipe Oliveira

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(12):986-94.

  In Parablennius parvicornis, small reproductive males with relatively low expression of secondary sexual characters (M- morphotype) parasite on the parental investment of the larger nest-holder malesIn Parablennius parvicornis, small reproductive males with relatively low expression of secondary sexual characters (M- morphotype) parasite on the parental investment of the larger nest-holder males which have fully developed secondary sexual characters (M+ morphotype). In comparison with M+ males, M- males have relatively low levels of androgens while having high blood cell percentages of lymphocytes and antigen responsiveness. Here we test the hypothesis that androgens are a causal factor for these differences in immunocompetence between morphotypes. After drawing an initial blood sample, males received a silastic implant containing either oil only (C), or oil with testosterone (T) or 11-ketotestosterone (KT). Males were re-caught 2 weeks later for drawing of the final blood sample. KT but not T induced the development of secondary sexual characters in M- males. M- males treated with KT showed lower swimming activity than the males treated with T or C implants, suggesting that KT also mediates behavioral changes in M- males. As expected, blood cell percentages of lymphocytes, but not of granulocytes, were higher in M- males than in M+ males. Overall, lymphocyte percentages increased in the C group which might have been a response to the surgery/treatment. In concordance with the hypothesis, lymphocyte percentages were suppressed in males treated with T in comparison with controls. However, no significant change was found in KT-treated males. This suggests that androgens modulate central, morphological and immunological traits by partly independent androgen mechanisms in P. parvicornis.

• Caudal autotomy and regeneration in lizards.

  Authors: Amanda R Clause, Elizabeth A Capaldi

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(12):965-73.

  Caudal autotomy, or the voluntary self-amputation of the tail, is an anti-predation strategy in lizards that depends on a complex array of environmental, individual, and species-specificCaudal autotomy, or the voluntary self-amputation of the tail, is an anti-predation strategy in lizards that depends on a complex array of environmental, individual, and species-specific characteristics. These factors affect both when and how often caudal autotomy is employed, as well as its overall rate of success. The potential costs of autotomy must be weighed against the benefits of this strategy. Many species have evolved specialized behavioral and physiological adaptations to minimize or compensate for any negative consequences. One of the most important steps following a successful autotomous escape involves regeneration of the lost limb. In some species, regeneration occurs rapidly; such swift regeneration illustrates the importance of an intact, functional tail in everyday experience. In lizards and other vertebrates, regeneration is a highly ordered process utilizing initial developmental programs as well as regeneration-specific mechanisms to produce the correct types and pattern of cells required to sufficiently restore the structure and function of the sacrificed tail. In this review, we discuss the behavioral and physiological features of self-amputation, with particular reference to the costs and benefits of autotomy and the basic mechanisms of regeneration. In the process, we identify how these behaviors could be used to explore the neural regulation of complex behavioral responses within a functional context.

• The outermost layer of egg-jelly is crucial to successful fertilization in the newt, Cynops pyrrhogaster.

  Authors: Shigekazu Takahashi, Hiromi Nakazawa, Akihiko Watanabe, Kazuo Onitake

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(12):1010-7.

  The significance of egg-jelly layers in internal fertilization was evaluated in the newt, Cynops pyrrhogaster. In this species, six egg-jelly layers, J1, J2, J3, J4, J5 and the outermost J6 layers,The significance of egg-jelly layers in internal fertilization was evaluated in the newt, Cynops pyrrhogaster. In this species, six egg-jelly layers, J1, J2, J3, J4, J5 and the outermost J6 layers, are accumulated on the surface of the fertilizable eggs in pars convoluta of the oviduct. When a large number of sperm (about 6 x 10(5)) were placed on eggs having different numbers of jelly layers, all the eggs were fully fertilized, although many of the eggs developed abnormally. Upon insemination using about 600 sperm, only eggs with the full set of jelly layers were fertilized at a high rate with normal development. Since around 300 (the range of 48-1,192) sperm were observed on and in the egg-jelly in naturally spawned eggs, we conclude that the J6 layer must be present on the outermost surface of the egg-jelly for successful internal fertilization of the newt. Previous studies have suggested that the J6 layer is a prerequisite for the initiation of sperm motility and the acrosome reaction. In the present study, the fertilization rate decreased in eggs with a full set of jelly layers when inseminated using acrosome-reacted and motile sperm. However, the fertilization rate was high when motile sperm with intact acrosome was used. These results suggest that induction of the sperm acrosome reaction in the J6 layer is an important step in the internal fertilization of the newt.

• Patterns of mechanical energy change in tetrapod gait: pendula, springs and work.

  Authors: Andrew A Biewener

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(11):899-911.

  Kinematic and center of mass (CoM) mechanical variables used to define terrestrial gaits are compared for various tetrapod species. Kinematic variables (limb phase, duty factor) provide importantKinematic and center of mass (CoM) mechanical variables used to define terrestrial gaits are compared for various tetrapod species. Kinematic variables (limb phase, duty factor) provide important timing information regarding the neural control and limb coordination of various gaits. Whereas, mechanical variables (potential and kinetic energy relative phase, %Recovery, %Congruity) provide insight into the underlying mechanisms that minimize muscle work and the metabolic cost of locomotion, and also influence neural control strategies. Two basic mechanisms identified by Cavagna et al. (1977. Am J Physiol 233:R243-R261) are used broadly by various bipedal and quadrupedal species. During walking, animals exchange CoM potential energy (PE) with kinetic energy (KE) via an inverted pendulum mechanism to reduce muscle work. During the stance period of running (including trotting, hopping and galloping) gaits, animals convert PE and KE into elastic strain energy in spring elements of the limbs and trunk and regain this energy later during limb support. The bouncing motion of the body on the support limb(s) is well represented by a simple mass-spring system. Limb spring compliance allows the storage and return of elastic energy to reduce muscle work. These two distinct patterns of CoM mechanical energy exchange are fairly well correlated with kinematic distinctions of limb movement patterns associated with gait change. However, in some cases such correlations can be misleading. When running (or trotting) at low speeds many animals lack an aerial period and have limb duty factors that exceed 0.5. Rather than interpreting this as a change of gait, the underlying mechanics of the body's CoM motion indicate no fundamental change in limb movement pattern or CoM dynamics has occurred. Nevertheless, the idealized, distinctive patterns of CoM energy fluctuation predicted by an inverted pendulum for walking and a bouncing mass spring for running are often not clear cut, especially for less cursorial species. When the kinematic and mechanical patterns of a broader diversity of quadrupeds and bipeds are compared, more complex patterns emerge, indicating that some animals may combine walking and running mechanics at intermediate speeds or at very large size. These models also ignore energy costs that are likely associated with the opposing action of limbs that have overlapping support times during walking. A recent model of terrestrial gait (Ruina et al., 2005. J Theor Biol, in press) that treats limb contact with the ground in terms of collisional energy loss indicates that considerable CoM energy can be conserved simply by matching the path of CoM motion perpendicular to limb ground force. This model, coupled with the earlier ones of pendular exchange during walking and mass-spring elastic energy savings during running, provides compelling argument for the view that the legged locomotion of quadrupeds and other terrestrial animals has generally evolved to minimize muscle work during steady level movement.

• Correlation of symmetrical gaits and whole body mechanics: debunking myths in locomotor biodynamics.

  Authors: Audrone R Biknevicius, Stephen M Reilly

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(11):923-34.

  Independent maturation of gait (Hildebrand) and whole body mechanics (Cavagna et al.) traditions in locomotor analyses has led to conflicting terminology. Re-evaluation of these traditions yieldsIndependent maturation of gait (Hildebrand) and whole body mechanics (Cavagna et al.) traditions in locomotor analyses has led to conflicting terminology. Re-evaluation of these traditions yields three primary insights. First, walking and running should be recognized by their fundamentally different mechanics. Because duty factor fails to consistently distinguish these mechanics, its use in discriminating walks from runs should be abandoned in preference to parameters that more accurately reflect the movements of the center of mass (COM; phase difference in external mechanical energy or Froude number). Second, "trot" should be reserved as a descriptor of a particular footfall pattern. This and all gait terms lack explicit information about limb compliance and thus COM movements. Third, symmetrical gait definitions should be broadened to reflect the four primary footfall patterns: the lateral-couplet dominated pattern of the pace, the diagonal-couplet dominated pattern of the trot and the more independent sequencing of footfalls of the two singlefoots. Intermediate gaits (perennially confusing and a mouthful to pronounce) are thereby subsumed by these four discrete gaits. Confusion between gait terminologies would be avoided if limb phase were consistently reported.

• Stability, limb coordination and substrate type: the ecorelevance of gait sequence pattern in primates.

  Authors: Nancy Jeanne Stevens

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(11):953-63.

  The coordination of limb movements during mammalian locomotion has been well documented in the literature. Most mammals use lateral sequence (LS) gaits, in which a forelimb follows an ipsilateralThe coordination of limb movements during mammalian locomotion has been well documented in the literature. Most mammals use lateral sequence (LS) gaits, in which a forelimb follows an ipsilateral hind limb during the stride cycle. Primates, however, tend to utilize diagonal sequence (DS) gaits, whereby a contralateral forelimb follows a given hind limb during the stride cycle. A number of scenarios have been offered to explain why primates favor DS gaits, most of them relating to the use of the arboreal habitat and, in particular, the exploitation of a terminal branch niche. Yet to date, there is surprisingly little evidence to support the advantage of DS gaits for negotiating different aspects of the terminal branch environment. Nonetheless, it is apparent that primates possess unique morphologies and a higher than typically recognized degree of flexibility in gait sequence pattern, both of which likely offer advantages for moving upon discontinuous and unstable terminal branches. This paper reviews potential explanations for the use of DS gaits in primates and considers mechanisms by which gait sequence may be altered during different types of arboreal challenges.

• The tri-segmented limbs of therian mammals: kinematics, dynamics, and self-stabilization--a review.

  Authors: Martin S Fischer, Reinhard Blickhan

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(11):935-52.

  The evolution of therian mammals is to a large degree marked by changes in their motion systems. One of the decisive transitions has been from the sprawled, bi-segmented to the parasagittal,The evolution of therian mammals is to a large degree marked by changes in their motion systems. One of the decisive transitions has been from the sprawled, bi-segmented to the parasagittal, tri-segmented limb. Here, we review aspects of the tri-segmented limb in locomotion which have been elucidated in our research groups in the last 10 years. First, we report the kinematics of the tri-segmented therian limb from mouse to elephant in order to explore general principles of the therian limb configuration and locomotion. Torques will be reported from a previous paper (Witte et al., 2002. J Exp Biol 205:1339-1353) for a better understanding of the anti-gravity work of all limb joints. The stability of a limb in z-configuration will be explained and its advantage with respect to other potential solutions from modeling will be discussed. Finally, we describe how the emerging concept of self-stability can be explained for a tri-segmented leg template and how it affects the design of the musculoskeletal system and the operation of legs during locomotion. While locomotion has been considered as mainly a control problem in various disciplines, we stress the necessity to reduce control as much as possible. Central control can be cheap if the limbs are "intelligent" by means of their design. Gravity-induced movements and self-stability seem to be energy-saving mechanisms.

• What are the relations between mechanics, gait parameters, and energetics in terrestrial locomotion?

  Authors: Donald F Hoyt, Steven J Wickler, Darren J Dutto, Gwenn E Catterfeld, Devin Johnsen

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(11):912-22.

  Are the different energy-conserving mechanics (i.e., pendulum and spring) used in different gaits reflected in differences in energetics and/or stride parameters? The analysis included published dataAre the different energy-conserving mechanics (i.e., pendulum and spring) used in different gaits reflected in differences in energetics and/or stride parameters? The analysis included published data from several species and new data from horses. When changing from pendulum to spring mechanics, there is a change in the slope of metabolic rate (MR) vs. speed in all species, in birds and quadrupeds there is no step increase, and in humans there are conflicting reports. At the trot-gallop transition, where quadrupeds are hypothesized to change from spring mechanics to some combination of spring and pendulum mechanics, there is a change in slope of MR vs. speed in horses but not in other species. Stride frequency (SF) is a logarithmic function of walking speed in all species, a linear function of trotting/running speed, and nearly independent of speed in galloping. In humans and horses there is a discontinuity in SF at the walk-trot (run) transition but not in birds. The slope of time of contact vs. speed does not change with mechanics in most species, but it does in humans. In horses and humans, there is a discontinuity at the walk-trot (run) transition and data for other species do not permit generalization. Duty factor (DF) in humans is greater than 0.5 in walking (pendulum mechanics) and less than 0.5 when running (spring mechanics). However, this is not true in many species that have DF>0.5 at the lowest speeds where they use spring mechanics. When trotting at low speeds, horses use forelimb DF>0.5 and hind limb DF<0.5. Thus, it is confusing to distinguish between walking and running by DF.

• Uptake of dextran-FITC by epithelial cells of the chorioallantoic placentome and the omphalopleure of the placentotrophic lizard, Pseudemoia entrecasteauxii.

  Authors: James R Stewart, Michael B Thompson, Marcus B Attaway, Jacquie F Herbert, Christopher R Murphy

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(10):883-9.

  Placental nutrient provision has evolved in multiple lineages of squamate reptiles and although possible structural specializations for placentotrophy have been described in a variety of species,Placental nutrient provision has evolved in multiple lineages of squamate reptiles and although possible structural specializations for placentotrophy have been described in a variety of species, neither the pathways nor the mechanisms of placental transfer are known. Lizards of the Australian genus Pseudemoia are placentotrophic and have elaborate placental structures that are thought to enhance nutrient transfer. The chorioallantoic placenta, which occupies the embryonic hemisphere of the egg, is regionally diversified into a large area with low epithelial height and a smaller placentome with cuboidal or columnar epithelia. Both regions are underlain by an extensive vascular bed. The abembryonic hemisphere of the egg is covered by an omphaloplacenta, which is similar to the placentome in having cuboidal or columnar epithelia but with a different embryonic vascular supply. We tested the hypothesis that embryonic epithelial cells of the placentome and the omphaloplacenta of Pseudemoia entrecasteauxii are each capable of endocytosis. Embryos (stages 33-39) with intact extraembryonic membranes were surgically removed from the uterus and incubated in a solution containing fluorescein isothiocyanate-dextran (77,000 MW). The fluorescent label was detected in the cytoplasm of scattered populations of epithelial cells in both placental regions of all embryonic stages. We conclude that both the placentome and the omphaloplacenta of P. entrecasteauxii are sites of histotrophic nutrient transport. However, there are histological and cytological differences in the embryonic epithelia of these two placental regions. The histological differences reflect differences in the evolutionary precursors of each tissue. The cytological differences likely portray different functional specializations.

• L-proline transport by purified cell types of lobster hepatopancreas.

  Authors: L Fiandra, P K Mandal, B Giordana, G A Ahearn

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(10):851-61.

  The hepatopancreas of the American lobster, Homarus americanus, has four epithelial cell types that are anatomically distinguishable and can be separated for in vitro investigation of theirThe hepatopancreas of the American lobster, Homarus americanus, has four epithelial cell types that are anatomically distinguishable and can be separated for in vitro investigation of their individual biological roles in the intact organ using centrifugal elutriation. Previous studies employing this separation method have produced hepatopancreatic cell suspensions that have been used to examine the nature of copper transport, 2 Na+/1 H+ exchange, and D-glucose absorption by each cell type in isolation from the other cells comprising the tubular epithelium. The present investigation used this method to study amino acid transport by E-, F-, R-, and B-cells of the lobster hepatopancreas in order to characterize the absorption processes for protein digestion products by this organ and to identify which cell type was most likely the responsible agent for net transcellular transfer of these organic molecules from lumen to blood. Results indicated that heptopancreatic E- and F-cell types were the only cells exhibiting Na+-dependent 3H-L-proline transport. Further examination of 3H-L-proline influx by F-cell suspensions indicated that this cell type possessed plasma membrane Na+-dependent IMINO-like and B0-like transport mechanisms and Na+-independent L-like transport mechanisms. Using selective inhibitors of these separate transport systems (e.g., L-pipecolate, L-alanine, and L-leucine), the IMINO-like transporter appeared to predominate in L-proline influx into F-cells, while lesser amounts of amino acid transport took place by the B0-like and L-like systems. The results of this study suggest that the hepatopancreatic F-cell is the epithelial cell type responsible for the bulk of amino acid absorption by this organ and that the IMINO-like transporter is responsible for most of the L-proline transfer through this agent. It is further suggested that as digestion and absorption proceeds in the hepatopancreas and concentrations of luminal amino acids and sodium fall, Na+-dependent transport systems, like the IMINO-like and B0-like, increase their binding affinities for their substrates to maximize nutrient transfer across the epithelium.

• Galanin and its binding sites in the brain of eels subjected to different osmolar conditions.

  Authors: Maria Angela Masini, Paola Prato, Bianca Maria Uva

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(10):862-71.

  A galanin (GAL)-like peptidergic system was investigated in the brain of Anguilla anguilla subjected to hyperosmolar and hypoosmolar conditions, by using antisera raised against porcine 1-29 GAL. AA galanin (GAL)-like peptidergic system was investigated in the brain of Anguilla anguilla subjected to hyperosmolar and hypoosmolar conditions, by using antisera raised against porcine 1-29 GAL. A group of immunoreactive perikaria was identified in the periventricular hypothalamus, in the ventral thalamus, in the pretectal areas and in the optic tectum. Immunoreactive perikaria were present in the nucleus lateralis of the torus semicircularis in seawater (SW) adapted eels, and were absent in the freshwater (FW) adapted eels. Galaninergic fibres were observed in many areas of the brain. The immunoreactive perikaria and fibres, when localised in the same areas, were more strongly labelled and numerous in the SW adapted eels in comparison to the FW adapted animals. GAL-specific binding sites, investigated by autoradiography using iodinated porcine GAL, occurred in several regions of the brain, in particular in the dorsal telencephalon, in the dorsal thalamus and in the torus semicircularis, where a high density of binding sites was observed in the SW adapted eels. Conversely, a higher density of binding sites was observed in the caudal substantia reticularis and in the corpus cerebelli in the FW adapted eels. We conclude that different distributions and intensity in immunoreactive elements occur in response to the two environmental conditions.

• Interspecific hybridization between Oryzias latipes and Oryzias curvinotus causes XY sex reversal.

  Authors: Ai Shinomiya, Mariko Kato, Miki Yaezawa, Mitsuru Sakaizumi, Satoshi Hamaguchi

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(10):890-6.

  The teleost fish, Oryzias curvinotus, is a closely related species to the medaka, Oryzias latipes, and both species have the DMY gene, which is required for male development in O. latipes. ItThe teleost fish, Oryzias curvinotus, is a closely related species to the medaka, Oryzias latipes, and both species have the DMY gene, which is required for male development in O. latipes. It suggests that the molecular function of the DMY gene and the following molecular events of sex differentiation are conserved between these two species. In the present study, we obtained interspecific hybrids between O. curvinotus and O. latipes and demonstrated sex-reversed XY females in the hybrids. The incidence of sex-reversed females in F1 XY hybrids between O. curvinotus females and O. latipes males, and hybrids between O. latipes females and O. curvinotus males were 21% and 100%, respectively. These results indicate that DMY does not always determine maleness in hybrid fish even though it is able to specify normal male development on its native genetic background and suggest that there are some differences between DMY(latipes) and DMY(curvinotus) alleles. Appearance of XY females in F1 hybrids also suggests that an autosomal or X-liked gene(s) from the maternal species interferes in the function of the paternal DMY gene in the male-determining process of the hybrid fish. These hybrid fish would supply a new experimental approach for investigating the genetic and molecular pathway of testis determination and differentiation.

• Low temperature requirement for embryonic development of Itasenpara bitterling Acheilognathus longipinnis.

  Authors: Kazuhiko Uehara, Koichi Kawabata, Hiromi Ohta

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(10):823-9.

  The Itasenpara bitterling has an embryonic period up to 7 months, when the embryo experiences large seasonal temperature changes. We examined the temperature requisites for normal development duringThe Itasenpara bitterling has an embryonic period up to 7 months, when the embryo experiences large seasonal temperature changes. We examined the temperature requisites for normal development during the embryonic stage. Fertilized eggs reared under any of the constant temperatures ranging from 5 degrees C to 30 degrees C did not achieve complete embryogenesis, and none reached the swim-up stage. The optimum temperature for normal embryonic development was found to be stage-dependent: 10-30 degrees C for fertilization, 15-25 degrees C for hatching, 5 degrees C for the requisite low temperature, 10-15 degrees C for eye pigmentation, and 20-30 degrees C for swim-up. These temperatures correlated well with the embryo's natural environmental conditions. Embryos raised at these temperatures sequentially grew normally, with 70% of the fertilized eggs achieving complete embryogenesis and, for the first time, developed to the swim-up stage. These results indicate that the low temperature, as required by the bitterling embryo, is an essential factor and correlates well with the embryo's natural ambient temperatures. Since the populations of Itasenpara bitterlings have been declining in Japan, this study is the first to provide additional information for successful artificial breeding of this endangered species.

• Oviductal protease and trypsin treatment enhance sperm-envelope interaction in Bufo arenarum coelomic eggs.

  Authors: Ricardo J Llanos, Daniel Barrera, Jorge N Valz-Gianinet, Dora C Miceli

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(10):872-82.

  We describe the morphological and biochemical changes in Bufo arenarum coelomic egg envelopes (CE) following passage through the oviduct. In this species, the transformation of the CE into theWe describe the morphological and biochemical changes in Bufo arenarum coelomic egg envelopes (CE) following passage through the oviduct. In this species, the transformation of the CE into the vitelline envelope (VE) leads to the acquisition of fertilizability and involves the cleavage of a glycoprotein component. Electrophoretic patterns indicate that a pars recta oviductal protease selectively hydrolyzes in vitro the 84 and the 55 kDa glycoproteins of the CE. During the CE to VE transformation, the relative concentrations of gp48, 42 and 39 kDa also change. In in vitro tests, sperm binding to envelope glycoprotein occurs when they are exposed to VE but not when treated with CE, and VE labeled glycoproteins bind to the head and mid piece of the sperm. The gp39 VE component has 100% identity with internal domains of the sequence deduced from ovarian cDNA for the homologous zona pellucida glycoprotein type C (ZPC) protein precursor in B. arenarum. The effects of trypsin as a substitute for oviductal protease were also examined. Trypsin selectively attacks the 84 and the 55 kDa glycoproteins without hydrolyzing other components and renders coelomic eggs fertilizable in a jelly water preparation. Therefore, trypsin can mimic in vitro the biological action of the oviductal protease. However, it does not wholly mimic the biological action of the oviduct which, in B. arenarum at least, exceeds a mere proteolytic effect. This fact was verified by the lower fertility rates and the abnormal embryo development found when trypsin-treated coelomic eggs were fertilized in vitro.

• Maternal exposure to estradiol and endocrine disrupting compounds alters the sensitivity of sea urchin embryos and the expression of an orphan steroid receptor.

  Authors: Troy A Roepke, Ernest S Chang, Gary N Cherr

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(10):830-41.

  Endocrine disrupting compounds (EDCs) are known to affect reproduction and development in marine invertebrates. In previous work, we have shown that developing sea urchin embryos were sensitive toEndocrine disrupting compounds (EDCs) are known to affect reproduction and development in marine invertebrates. In previous work, we have shown that developing sea urchin embryos were sensitive to estradiol and estrogenic EDCs at environmentally relevant concentrations in a tamoxifen-sensitive manner (Roepke et al. 2005. Aquat Toxicol 71:155-173). In this study, we report the effects of maternal exposure to EDCs on embryo sensitivity and regulation of an orphan steroid receptor in sea urchin eggs. Maternal exposures were conducted by injecting female Strongylocentrotus purpuratus sea urchins initiating oogenesis with two concentrations of estradiol, octylphenol, tributyltin and o, p-DDD for 8 weeks with an induced spawning before and after the injection cycle. Developing embryos were less sensitive to estradiol following maternal exposure to estradiol, octylphenol and DDD. The steroidogenesis inhibitor, spironolactone, and the aromatase inhibitor, formestane, affected normal sea urchin development with EC50 values of 18 and 2 microM, respectively. Binding of estradiol was demonstrated in homogenates supernatants of sea urchin embryos by filtration centrifugation and column chromatography, but saturation was not reached until 4-6 hr and was highly variable. Analysis of eggs from pre- and post-injection spawns using real-time Q-PCR for the mRNA of an orphan steroid receptor, SpSHR2, shows that receptor mRNA increased in eggs with estradiol, octylphenol and tributyltin but decreased with DDD. RIA showed that estradiol may be present during gastrulation. In summary, maternal exposure to estradiol and EDCs alters embryo sensitivity and regulates the expression of an orphan steroid receptor in the egg.

• Quantitative changes in branchial carbonic anhydrase activity and expression in the euryhaline green crab, Carcinus maenas, in response to low salinity exposure.

  Authors: Raymond P Henry, Kimberly L Thomason, David W Towle

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(10):842-50.

  Hemolymph osmolality, and changes in gill carbonic anhydrase (CA) activity, relative mRNA expression, and CA protein concentration were measured in the green crab Carcinus maenas acclimated to 32 pptHemolymph osmolality, and changes in gill carbonic anhydrase (CA) activity, relative mRNA expression, and CA protein concentration were measured in the green crab Carcinus maenas acclimated to 32 ppt salinity and transferred to 10 ppt. Hemolymph osomolality stabilized at new, acclimated values, by 24 hr after transfer. There was a large increase in CA mRNA concentrations, as measured by quantitative PCR, in the posterior gills by 24 hr post-transfer that remained elevated through 4 days. By 7 days, however, CA mRNA levels began to decline. CA activity, on the other hand, did not begin to increase until 48 hr after transfer to 10 ppt, but it continued to increase through 7 days. CA protein concentration increased by 5-fold in posterior gills in crabs acclimated to 10 ppt. CA activity, mRNA expression, and CA protein concentrations did not change in anterior gills. These results indicate that low salinity-stimulated CA induction is under transcriptional regulation, and that the increase in CA activity is a result of the increase in gene expression and synthesis of new enzyme. Changes in mRNA appear to be transient, but once synthesized, the CA protein appears to persist in the gill for an extended time. In a separate set of experiments, green crabs acclimated to 32 ppt were transferred directly to salinities of 25, 20, 15, and 10 ppt. CA activity and mRNA concentrations increased with decreasing salinity, peaking at 15 ppt but decreasing between 15 and 10 ppt. The decrease may represent a breakdown in the mechanism of transport-related protein induction near the lower salinity limit of this species.

• Mode of action and functional significance of avian gonadotropin-inhibitory hormone (GnIH): a review.

  Authors: Kazuyoshi Tsutsui, Takayoshi Ubuka, Hong Yin, Tomohiro Osugi, Kazuyoshi Ukena, George E Bentley, Nick Ciccone, Kazuhiko Inoue, Vishwajit S Chowdhury, Peter J Sharp, John C Wingfield

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(9):801-6.

  Neuropeptide control of gonadotropin secretion at the level of the anterior pituitary gland is primarily through the stimulatory action of the hypothalamic decapeptide, gonadotropin-releasing hormoneNeuropeptide control of gonadotropin secretion at the level of the anterior pituitary gland is primarily through the stimulatory action of the hypothalamic decapeptide, gonadotropin-releasing hormone (GnRH). However, a hypothalamic neuropeptide acting at the level of the pituitary to negatively regulate gonadotropin secretion has, until recently, remained unknown in any vertebrate. In 2000, we discovered a novel hypothalamic neuropeptide inhibiting gonadotropin release at the level of the pituitary in quail and termed it gonadotropin-inhibitory hormone (GnIH). A gonadotropin-inhibitory system is an intriguing concept and provides us with an unprecedented opportunity to study the regulation of avian reproduction from an entirely novel standpoint. To elucidate the mode of action of GnIH, we further identified the receptor for GnIH and characterized its expression and binding activity in quail. The identified GnIH receptor possessed seven transmembrane domains and specifically bound to GnIH in a concentration-dependent manner. The expression of GnIH receptor was found in the pituitary and several brain regions including the hypothalamus. These results suggest that GnIH acts directly on the pituitary via GnIH receptor to inhibit gonadotropin release. GnIH may also act on the hypothalamus to inhibit GnRH release. To understand the functional significance of GnIH in avian reproduction, we also investigated the mechanism that regulates GnIH expression. Interestingly, melatonin induced dose-dependently GnIH expression and melatonin receptor (Mel(1c)) was expressed in GnIH neurons. Thus melatonin appears to act directly on GnIH neurons via its receptor to induce GnIH expression. Based on these studies, GnIH is likely an important neuropeptide for the regulation of avian reproduction.

• Comparative genomics in vertebrate evolution and development.

  Authors: Chris T Amemiya, Marta Gomez-Chiarri

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(9):672-82.

  The vast quantities of publicly available DNA sequencing data and genome resources are enabling biologists to investigate age-old problems in biology that were not addressable previously. In thisThe vast quantities of publicly available DNA sequencing data and genome resources are enabling biologists to investigate age-old problems in biology that were not addressable previously. In this review, we discuss how comparative genomics is practiced and how the data can be used to make biological inferences with respect to vertebrate evolution and development. Examples are taken from the well-known HOX clusters, which are always a high-priority target for genomic analyses due to their inferred role in the evolution of metazoans. In addition, we briefly discuss the application of genomic approaches to problems in comparative endocrinology.

• Xenobiotic detoxification in the nematode Caenorhabditis elegans.

  Authors: Tim H Lindblom, Allyn K Dodd

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(9):720-30.

  The nematode Caenorhabditis elegans is an important model organism for the study of such diverse aspects of animal physiology and behavior as embryonic development, chemoreception, and the geneticThe nematode Caenorhabditis elegans is an important model organism for the study of such diverse aspects of animal physiology and behavior as embryonic development, chemoreception, and the genetic control of lifespan. Yet, even though the entire genome sequence of this organism was deposited into public databases several years ago, little is known about xenobiotic metabolism in C. elegans. In part, the paucity of detoxification information may be due to the plush life enjoyed by nematodes raised in the laboratory. In the wild, however, these animals experience a much greater array of chemical assaults. Living in the interstitial water of the soil, populations of C. elegans exhibit a boom and bust lifestyle characterized by prodigious predation of soil microbes punctuated by periods of dispersal as a non-developing alternative larval stage. During the booming periods of population expansion, these animals almost indiscriminately consume everything in their environment including any number of compounds from other animals, microorganisms, plants, and xenobiotics. Several recent studies have identified many genes encoding sensors and enzymes these nematodes may use in their xeno-coping strategies. Here, we will discuss these recent advances, as well as the efforts by our lab and others to utilize the genomic resources of the C. elegans system to elucidate this nematode's molecular defenses against toxins.

• Identification of melanin-concentrating hormone receptor and its impact on drug discovery.

  Authors: Yumiko Saito, Kei Maruyama

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(9):761-8.

  The neuropeptide melanin-concentrating hormone (MCH) was originally isolated from the pituitary of salmon, in which it causes skin paling. MCH is also found abundantly in mammalian neurons, and hasThe neuropeptide melanin-concentrating hormone (MCH) was originally isolated from the pituitary of salmon, in which it causes skin paling. MCH is also found abundantly in mammalian neurons, and has been detected in the lateral hypothalamus and zona incerta, brain regions that are at the center of feeding behavior. Acute central administration of MCH leads to a rapid and significant increase in food intake, while MCH expression changes in states of altered energy balance, such as fasting and obesity. Furthermore, MCH knockout mice tend toward hypophagia and leanness. In 1999, we and four other groups identified an orphan G-protein-coupled receptor (GPCR) as a specific receptor for MCH (MCH-1 receptor). Although a second MCH receptor (MCH-2 receptor) was isolated in humans, it was found to be non-functional or encode a non-functional pseudogene in non-human species, including rodents. The discovery of these MCH receptors permitted the launch of a broad array of drug screening efforts and three MCH-1 receptor antagonists were identified to reduce food intake and body weight. Interestingly, some antagonists unexpectedly produced evidence that blockade of these receptors has antidepressant and anxiolytic activities. The expressions of the MCH receptors, which have been implicated in regulating emotion, stress and motivation, make MCH an excellent candidate for integrating the various homeostatic stimuli necessary for maintaining the proper conditions of energy metabolism and other physiological functions. Finally, the speed at which MCH receptor studies have been undertaken exemplifies the impact that this deorphanized GPCR will have on setting the stage for more detailed physiological studies.

• Interactions of gonadotropin-releasing hormone (GnRH) and gonadotropin-inhibitory hormone (GnIH) in birds and mammals.

  Authors: George E Bentley, Lance J Kriegsfeld, Tomohiro Osugi, Kazuyoshi Ukena, Sara O'Brien, Nicole Perfito, Ignacio T Moore, Kazuyoshi Tsutsui, John C Wingfield

  Journal of experimental zoology. Part A, Comparative experimental biology. 305(9):807-14.

  Gonadotropin-releasing hormone (GnRH) regulates secretion of both of the gonadotropins, luteinizing hormone (LH) and follicle-stimulating hormone. Thus, it is a key hormone for vertebrateGonadotropin-releasing hormone (GnRH) regulates secretion of both of the gonadotropins, luteinizing hormone (LH) and follicle-stimulating hormone. Thus, it is a key hormone for vertebrate reproduction. GnRH was considered to be unusual among hypothalamic neuropeptides in that it appeared to have no direct antagonist, although some neurochemicals and peripheral hormones (opiates, GABA, gonadal steroids, inhibin) can modulate gonadotropin release to a degree. Five years ago, a vertebrate hypothalamic neuropeptide that inhibited pituitary gonadotropin release in a dose-dependent manner was discovered in quail by Tsutsui et al. (2000. Biochem Biophys Res Commun 275:661-667). We now know that this inhibitory peptide, named gonadotropin-inhibitory hormone, or GnIH, is a regulator of gonadotropin release in vitro and in vivo. Its discovery has opened the door to an entirely new line of research within the realm of reproductive biology. In our collaborative studies, we have begun to elucidate the manner in which GnIH interacts with GnRH to time release of gonadotropins and thus time reproductive activity in birds and mammals. This paper reviews the distribution of GnIH in songbirds relative to GnRHs, and our findings on its modes of action in vitro and in vivo, based on laboratory and field studies. These data are simultaneously compared with our findings in mammals, highlighting how the use of different model species within different vertebrate classes can be a useful approach to identify the conserved actions of this novel neuropeptide, along with its potential importance to vertebrate reproduction.

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