Mortality of embryos, developmental disorders and changes in biochemical parameters in marsh frog (Rana ridibunda) tadpoles exposed to the water-soluble fraction of Kazakhstan crude oil and O-Xylene.

The effects of different concentrations of water-soluble fraction of crude oil (WSFO) from the Zhanazhol oil field (Aktobe region, Kazakhstan) and compared to o-xylene, prevalent in this oil, on growth and development of marsh frog (Rana ridibunda) were assessed. In subchronic experiments (7 d), a dose-related increase in mortality and incidence of deformities in embryos were observed. In chronic experiments (60 d; starting from the Gosner stage 26), a dose-dependent decrease in body weight, size and developmental delay by 3-4 stages were also detected. In addition, the content of lipid hyperoxide (LHO) and malondialdehyde (MDA), as well as activities of superoxide dismutase (SOD) and catalase (CAT) enzymes in liver of the tadpoles were determined at the end of chronic experiment. Exposure to 0.5 mg/L or 1.5 mg/L WSFO elevated the content of LHO by 76% and 86%, and MDA by 47% and 58% but decreased SOD activity by 26% and 49%, and CAT by 35% and 46%, respectively. A less pronounced adverse effect was found after chronic exposure to the same concentrations of o-xylene. In tadpole liver exposed to o-xylene levels of LHO was increased by 40% and 51%, MDA by 11% and 29%, while the activity of SOD was lowered by 18% and 41%, and CAT - by 13% and 37% in the 0.5 mg/L and 1.5 mg/L treatment groups, respectively. Data demonstrated the embryotoxic and teratogenic effects attributed to WSFO and o-xylene exposure which may involve oxidative stress mechanisms.

[Ploidy and genetic structure of hybrid populations of water frogs Pelophylax esculentus (L., 1758) complex (Amphibia, Ranidae) of Ukraine].

The present study of green frog hybrid populations of Ukraine, including analysis of allozyme variability and planimetric analysis oferythrocytes size has confirmed that the unique region in this area is the Severski Donets basin The allopolyploid individuals there are met very frequently (5.7% of all investigated frogs). In other areas of Ukraine only two polyploid hybrids have been recorded. Beside that, one frog was defined as triploid Rana ridibundus. According to our investigations, all triploid hybrids from the Severski Donets basin are identified as P. esculentu (=lessonae)--2 ridibundus males.

A skeletochronological study of age, growth and longevity in a population of the frog Rana ridibunda from southern Europe.

Age at sexual maturity and longevity in a population of Rana ridibunda from north-eastern Greece were studied by skeletochronology performed on the phalanges. Analysis of the age structure was based on counting the lines of arrested growth (LAGs). Sexual maturity for both sexes arises during the first year or after the first hibernation. Ages ranged from 1 to 5 years (mean=2.96) among 52 males and from 1 to 5 years (mean=3.73) among 56 females. The mean snout-vent length was 69.03+/-12.6mm in males and 82.38+/-13.27 mm in females. The difference between the sexes in age and size was significant. Growth of individuals was fitted on? The von Bertalanffy model. The growth coefficient (K) was 0.57 in males and 0.54 in females, mainly due to faster male growth between metamorphosis and maturation.

Population genetics of a successful invader: the marsh frog Rana ridibunda in Britain.

We investigated the genetic outcome of successful invasion by an alien species, the marsh frog Rana ridibunda, in Britain. Twelve adults translocated from Hungary into Kent (Romney) in 1935 resulted rapidly in a large localized population. A further successful translocation in 1973 from Romney to Sussex (Lewes), together with other range extensions, provided an opportunity to test bottleneck effects during colonization events. Romney and Lewes frogs had similar genetic diversities to those in Hungary at 14 random amplified polymorphic DNA marker (RAPD) and five microsatellite loci. The introduced populations were, however, differentiated genetically from each other and from a reference population in Hungary. Fitness assessments (larval growth and survival) revealed no differences between the Lewes and Romney populations. Despite starting with few founders, significant bottleneck effects on R. ridibunda in Britain were therefore undetectable, presumably because population expansions were rapid immediately after the translocations.

Glutathione redox system, GSH-Px activity and lipid peroxidation (LPO) levels in tadpoles of R.r.ridibunda and B.viridis.

Total glutathione (t-GSH), reduced glutathione (GSH), glutathione disulphide (GSSG) levels, t-GSH/GSSG ratio, glutathione peroxidase (GSH-Px) activity and lipid peroxidation (LPO) levels were investigated during the development period of a predominantly aquatic amphibian R.r.ridibunda and a predominantly terrestrial amphibian B. viridis. While t-GSH and GSH showed a similar trend, GSSG concentration increased significantly (p<0.05) during the larval stages in R.r.ridibunda larvae. In contrast to R.r.ridibunda larvae, there was no significant (p>0.05) change between 1 and 5 weeks in the t-GSH and GSH concentrations of B. viridis. t-GSH and GSH concentrations of B. viridis larvae became sharply elevated after the fifth week, GSSG levels increased 3.25-fold during the metamorphosis. The t-GSH/GSSG ratio fluctuated and the lowest t-GSH/GSSG ratios were observed at the third week for both species. GSH-Px activities for both species increased significantly (p<0.05) during the growing period. The highest GSH-Px activities in R.r.ridibunda and B.viridis were observed at the eighth week and they were 3.45 +/- 0.17 and 4.1 +/- 0.21 IU mg(-1), respectively. The membrane LPO levels in the R.r.ridibunda and B. viridis tadpoles significantly (p<0.001) decreased from 206 +/- 10.3 to 146 +/- 7.3 and from 198 +/- 9.9 to 23 +/- 1.15 nmol MDA g(-1) w.w., respectively.

[Cardiomyogenesis in Rana ridibunda tadpoles during metamorphosis in normal conditions and after exposure to various concentrations of cadmium ions]. / Kardiomiogenez u ozernoi liagushki v period metamorfoza golovastikov v norme i pri vozdeistvii ionov kadmiia v razlichnykh kontsentratsiiakh.

During metamorphosis period Rana ridibunda ventricular cardiomyocytes possess strong ability to adapt to altering developmental conditions, certain peculiarities of differentiation were observed during this process, cell death and mitotic activity increases. The action of different concentrations of cadmium ions on Rana ridibunda larvae changes interrelation and intensity of basic processes of cardiac muscular histogenesis, leading to differentiation disorders, suppression of proliferative activity and intensification of cardiomyocytes death.

Delayed costs of an induced defense in tadpoles? Morphology, hopping, and development rate at metamorphosis.

Models for the evolution of plasticity predict that individuals having phenotypes induced by exposure to enemies should experience relatively low fitness when enemies are absent. However, costs of induced phenotypes have been difficult to find in both plants and animals, perhaps because costs are expressed at later stages in the life cycle. We searched for delayed costs of an induced defense in larvae of the water frog Rana ridibunda, which exhibits strong phenotypic responses to predators. Tadpoles grew to metamorphosis in outdoor artificial ponds, in either the presence or absence of Aeshna dragonfly larvae confined within cages. We collected metamorphs at forelimb emergence, estimated their development rate until tail resorption was complete, and measured their body and leg shape and hopping performance. Development rate through metamorphosis reflects the duration of a transitional period during which metamorphs are especially vulnerable to predators, and hopping performance may reflect ability to escape predators. Froglets from the dragonfly treatment lost mass through metamorphosis significantly faster than those from predator-free ponds, but they resorbed their tails at about the same rate, despite the fact that their tails were relatively large to begin with. Froglets developing from predator-induced tadpoles had shorter, more muscular legs, and hopped 5% longer distances (difference not significant). Therefore, producing an induced defense against insect predators during the tadpole stage did not exact a cost during or immediately after metamorphosis; if anything, tadpoles with the predator-induced phenotype gave rise to more vigorous froglets. These results focus attention on other costs of the induced phenotype, as well as alternative explanations for plasticity that do not rely on direct fitness trade-offs.

Ontogeny of pituitary adenylate cyclase-activating polypeptide (PACAP) in the frog (Rana ridibunda) tadpole brain: immunohistochemical localization and biochemical characterization.

The anatomic distribution and biochemical characteristics of the neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) were investigated in the central nervous system of the frog, Rana ridibunda, during development. Three to four days after hatching, at stages IV-VII, PACAP-immunoreactive perikarya were detected in the dorsal thalamus within the anterior ventral area, and a few fibers were found in the medial pallium. Positive cell bodies were first observed in the hypothalamus at stages VIII-IX, at the level of the dorsal and ventral infundibular nuclei. In these regions, the number of positive perikarya increased during ontogeny. In tadpoles, during the mid- and late premetamorphosis, a more complex organization of the PACAP-immunoreactive system was found in the thalamus with the appearance, at stages IX-XII, of two additional groups of positive neurons in the ventrolateral area and posterocentral nucleus. At stages XIII-XVIII of larval development and subsequent larval stages, PACAP-immunoreactive fibers were found in the median eminence. In newly metamorphosed animals, several additional groups of positive perikarya appeared in the medial pallium, the preoptic nucleus, the torus semicircularis, the tegmentum of the mesencephalon, and the cerebellum. The immunoreactive peptide contained in the tadpole brain was characterized by high performance liquid chromatography analysis combined with radioimmunoassay quantification. At all stages investigated, the predominant form of PACAP-immunoreactive material coeluted with synthetic frog PACAP38. The occurrence of PACAP soon after hatching indicates that the peptide may exert neurotrophic activities. The existence of immunoreactive elements in several thalamic regions at mid- and late premetamorphic stages suggests that PACAP may act as a neurotransmitter, neuromodulator, or both, during ontogenesis. Finally, the presence of PACAP-immunoreactive perikarya in hypothalamic nuclei and nerve fibers in the median eminence supports the view that PACAP may play a role in the control of pituitary hormone secretion during larval development.

Ontogeny of catecholamine systems in the central nervous system of anuran amphibians: an immunohistochemical study with antibodies against tyrosine hydroxylase and dopamine.

To get more insight into developmental aspects of catecholamine systems in vertebrates, in particular anuran amphibians, these systems were studied immunohistochemically in embryos and larvae of Xenopus laevis and Rana ridibunda. Antisera against tyrosine hydroxylase (TH) and dopamine (DA) revealed that catecholamine systems are already present at early embryonic stages. The first dopamine group to be detected was found ventral to the central canal of the spinal cord of Xenopus, soon followed by DA cell groups in the posterior tubercle, the hypothalamic periventricular organ, the accompanying cell group of the periventricular organ, and the suprachiasmatic nucleus. Although weakly TH-immunoreactive cells were found in the olfactory bulb at about the same embryonic stages, DA immunoreactivity was not detected until premetamorphic stage 49. Dopamine cell groups in the caudal brainstem, midbrain, and pretectum appeared at late premetamorphic and prometamorphic stages, whereas the preoptic group was first observed at the metamorphic climax stage. Rana showed an almost similar timetable of development of catecholamine cell groups, except for the caudal brainstem group which was already present at the end of the embryonic period. When compared with previous studies by means of formaldehyde-induced fluorescence technique, it becomes clear that TH/DA immunohistochemistry enables an earlier detection of catecholamine cell groups and fiber systems in anuran amphibians. The present study also revealed that the DA-immunoreactive cells of the hypothalamic periventricular organ never stained with the TH antiserum during development, thus supporting their putatively DA accumulating nature. Another notable result is the site of origin and rather late appearance of the midbrain dopaminergic cell group. It is suggested that the latter cell group only partly corresponds to the ventral tegmental area and substantia nigra of amniotes.

Different levels of hyperoxia reversibly induce catalase activity in amphibian tadpoles.

Studies about the proposed antioxidant physiological role of the catalase (CAT) enzyme in relation to different environmental oxygen tensions are reported for the first time in amphibian larvae of Discoglossus pictus and Rana ridibunda perezi during their development. The CAT levels of whole tadpoles increased constantly in both species during the larval period, reaching a maximum during the metamorphic climax. All through development, CAT activity levels were always greater in D. pictus than in R. ridibunda perezi. This correlates well with the already reported higher SOD activity and hyperoxia resistance of the D. pictus species when compared to R. ridibunda perezi. Long-term acclimation to different levels of hyperoxia (40, 60, and 100% O2) showed dose-related increases in the CAT activity of D. pictus tadpoles. These increases did not take place when the animals were subjected to acute hyperoxia (24 h). The increase in CAT activity observed after 15 days of acclimation to acute hyperoxia (710 mm Hg: 100% O2) was reversed after 15 additional days of postacclimation to normal air (149 mm Hg O2). When recently metamorphosed frogs were acclimated to acute hyperoxia, significant increases in CAT activity were observed after 15 days, but not after 7 days. The results are interpreted as supporting a protective role for the CAT enzyme in amphibian larvae and froglets against oxygen toxicity.

Direct and indirect calorimetry measurements during the annual growth cycle of juvenile frogs Rana ridibunda P.

A thermodynamic study of the energetic metabolism in juvenile amphibians was conducted over an annual cycle of growth. The total energy flux (delta H = enthalpy variation) was measured with direct microcalorimetry and accounted for the sum of all metabolic pathways. Indirect calorimetry, oxygen consumption measurements (VO2), provides the values for the oxidative metabolism-dependent energy. A comparison between the results of both techniques simultaneously performed, provides for the calculation of the energetic efficiency, which indicates the respective importance of aerobic and anaerobic metabolisms during the annual biological cycle. It is clear that oxidative metabolism does not represent all, nor even a constant percentage of the total energy production of the frog, Rana ridibunda. The ration (Formula: see text) varied between 0.72 and 1.00, according to the season and the feeding or fasting periods. These results are discussed with special reference to the ecophysiology and behaviour of this species.