Dynamics of glucocorticoids in the ontogenesis of freshwater crawfish <Emphasis Type="Italic">Astacus leptodactylus Esch</Emphasis>

Dynamics of changes in glucocorticoid (hydrocortisone and corticosterone) levels was studied in the ontogenesis of freshwater crawfish Astacus leptodactylus Esch. It was shown that steroid concentrations increase during the embryogenesis period. Decrease in the glucocorticoid levels during postembryonic development is most probably related to the stabilization of young fish growth and the attainment of hormone levels typical for adult individuals.     

Total protein in hemolymph of crawfish <Emphasis Type="Italic">Pontastacus leptodactylus</Emphasis> as a parameter of the functional state of animals and a biomarker of quality of habitat

Analysis of total protein in hemolymph was performed by Lowry method on sexually mature crawfish Pontastacus leptodactylus. The total protein content in the crawfish hemolymph was studied for one year. The protein concentration varied widely, amounted to from 12 to 95 mg/ml, and depended on season and the molting cycle phase. There are presented histograms for distribution of animals for the protein level during different seasons and their character is analyzed. In summer the amount of protein is maximal prior to molt and decreases by 40% at once after it. There is studied the diapason of total protein concentrations in hemolymph, in which survival of crawfish at unfavorable changes in habitat is maximal. The adaptive possibilities of crawfish with the low protein content are attenuated. The crawfish with the protein concentration in hemolymph lower than the «critical» one were submitted for different time by action of hydroquinone (1 g/l) used as a model toxicant. A brief action did not affect the protein content in hemolymph. At a long toxic action the protein level in hemolymph fell, on average, by 40%, which preceded death of the animals. Possible mechanisms of positive correlation of the protein concentration in the crawfish hemolymph and of their survival at deterioration of quality of the water medium are discussed.     

Total protein in crawfish hemolymph as a parameter of functional state of animals and biomarker of quality of habitat

Analysis of total protein in hemolymph was performed by Lowry method on sexually mature crawfish Pontastacus leptodactylus. The total protein content in the crawfish hemolymph was studied for one year. The protein concentration varied widely, amounted to from 12 to 95 mg/ml, and depended on season and the moulting cycle phase. There are presented histograms for distribution of animals for the protein level for different seasons and their character is analyzed. In summer the amount of protein is maximal prior to moult and decreases by 40 % at once after it. There is studied the diapason of total protein concentrations in hemolymph, in which survival of crawfish at unfavorable changes in habitat is maximal. The adaptive possibilities of crawfish with the low protein content are reduced. The crawfish with the protein concentration in hemolymph lower than the "critical" one were submitted for different time by action of hydroquinone (1 g/l) used as a model toxicant. A brief action did not affect the protein content in hemolymph. At a long toxic action the protein level in hemolymph fell, on average, by 40%, which preceded the death of the animals. Possible mechanisms of positive correlation of the protein concentration in the crawfish hemolymph and of their survival at deterioration of quality of the water medium are discussed.     

A study of the cytoplasmic receptors for glucocorticoids in intestine of pre- and postweanling rats.

Glucocorticoids cause both enzymic and morphologic changes in the rat intestine during the time of weaning. To obtain information regarding the mechanism of these actions, we examined the cytoplasmic fraction of intestines from 18-day-old rats for the presence of specific glucocorticoid-binding proteins which are characteristics of target tissues. Incubation of slices of intestine with [3H]dexamethasone in a physiological medium at 2 degrees showed the presence of a cytoplasmic binding macromolecule with high specificity for steroids having glucocorticoid activity. The binding reaction was saturable (concentration of binding sites equals 0.24 pmol per mg of protein) and of high affinity (dissociation constant equals 9.3 nM). Binding was reversible on addition of nonlabeled dexamethasone (t 1/2 equals 5.2 hours), indicating that the usual assay procedure measured both corticosterone-filled and unoccupied binding sites. Sucrose density gradient centrifugation showed that the receptor-dexamethasone complex from intestinal cytosol sedimented at the same rate as that from liver (8.2 S). The receptor-dexamethasone complex was stable at 2 degrees for at least 24 hours in intestinal slices, but in isolated cytosol fractions there was considerable loss of binding even in the presence of high concentrations of [3H]dexamethasone. Furthermore, mixing experiments showed that the presence of cytosol from intestinal mucosa (but not from the muscle layers) caused a dissociation of dexamethasone from receptors of liver cytosol. This suggested the presence of some interfering factor in isolated mucosal cytosol and meant that quantitative studies had to be confined to intact slices. Although the reasons for the instability of steroid-receptor complexes in the presence of isolated intestinal cytosol are not understood, the instability is believed to be associated with homogenization and, therefore, is believed to have no physiological significance. Finally, the ontogenesis of cytoplasmic glucocorticoid receptors in intestinal slices was examined and the pattern compared with that in liver and lung. Receptor activity was present in intestine from late fetal life through adulthood, but concentrations were significantly higher during the first two postnatal weeks than at all other times. By contrast, receptor activity detected in cytosol prepared from rat lung was high around the time of birth, while that in liver rose steadily during the first postnatal week and remained at high levels. Thus specific receptors for glucocorticoids are present in the rat intestine during periods of both responsiveness and unresponsiveness. This suggests that although corticosteroids exert their effects through the cytoplasmic receptors, this early event in glucocorticoid action may not be a controlling step for changes in responsiveness during development.     

Postnatal physiological development of rats after acute prenatal hypoxia

The aim of study was to investigate the physiological development of the brain and behaviour in rats subjected to prenatal hypoxia on the 13.5th day of embryogenesis. We have found that such rats manifested a delayed physiological development and a change in nervous tissue of the sensorimotor cortex, as well a disturbed formation of motor responses during the first month of postnatal ontogenesis. During maturation these modifications were in part compensated, however we observed a decrease of the rats' ability to learn new forepaw movements. The destruction of the brain tissue and the modification of neurons composition in the sensorimotor cortex correlated with changes of behaviour at different stages of ontogenesis. Thus, changes of the conditions under which an organism develops during embryogenesis, predetermine a disturbance in ontogenesis and the learning ability.     

Expression of Na+/K+-ATPase α-subunit mRNA during embryonic development of the crayfish Astacus leptodactylus

Astacus leptodactylus is a decapod crustacean fully adapted to freshwater where it spends its entire life cycle after hatching under huge osmoconcentration differences between the hemolymph and surrounding freshwater. We investigated the expression of mRNA encoding one ion transport-related protein, Na⁺/K⁺-ATPase α-subunit, and one putative housekeeping gene, β-actin, during crayfish ontogenesis using quantitative real-time PCR. A 216-amino acid part of the open reading frame region of the cDNA coding for the Na⁺/K⁺-ATPase α-subunit was sequenced from total embryo, juvenile and adult gill tissues. The predicted amino acid sequence showed a high percentage similarity to those of other invertebrates (up to 95%) and vertebrates (up to 69%). β-actin expression exhibited modest changes through embryonic development and early post-embryonic stage. The Na⁺/K⁺-ATPase α-subunit gene was expressed in all studied stages from metanauplius to juvenile. Two peaks of expression were observed: one in young embryos at 25% of embryonic development (EI = 100 μm), and one in embryos just before hatching (at EI = 420 μm), continuing in the freshly hatched juveniles. The Na⁺/K⁺-ATPase expression profile during embryonic development is time-correlated with the occurrence of other features, including ontogenesis of excretory antennal glands and differentiation of gill ionocytes linked to hyperosmoregulation processes and therefore involved in freshwater adaptation.     

Effect of small doses of gamma radiation on the activity of marker enzymes in the plasma membranes of chick embryo liver

A study was made of marker enzyme activity in plasma membranes of chick liver during embryogenesis in normal conditions and after exposure to ionizing radiation. It was shown that irradiation with small doses of eggs prior to incubation and of chick embryos during different periods of the prenatal ontogenesis caused essential changes in activity of membrane-bound enzymes of liver plasmolemma. Stimulation with small radiation doses was most effective with preincubation irradiation of eggs and most manifest at later stages of the prenatal ontogenesis. Thus, the marker enzymes were shown to play a significant role in the realization of the stimulatory effect of low-level radiation.     

Combined reaction of the lymphoid organs and the adrenals to stress in mature animals subjected to cold in the early period of postnatal ontogeny

Structural-functional organization of the lymphoid organs and functional state of the adrenals have been studied in animals, subjected to cold in early postnatal period, as well as changes of the parameters mentioned to a short and prolonged cooling in mature rats. For the animals increase in the thymus mass and in reproduction rate of lymphocytes in the thymus, spleen and lymph nodes is specific against the background of high corticosteroid secretion. When the control animals are kept in cold for a long time, after the phase of an acute stress, accompanied with hypercorticism and a pronounced lymphatic effect, during the period of an increased cold stability, the high secretion of glucocorticoid hormones is accompanied with a certain activation of thymus-dependent zones in the peripheral lymphoid organs. In the mature rats, subjected to cold at early ontogenesis both stress-reaction to cooling and rearrangement in the regulatory systems studied does not develop at adaptation to cold.     

Adaptation mechanisms of respiratory blood function in Teleostei

Analysis of osmotic resistance of erythrocytes identified the similarity between marine and freshwater Teleostei and inclination of erythrocytes from freshwater fish to intravascular haemolysis. Structural resistance of hemoglobin was significantly higher in marine species and resistance to dehydration was significantly higher in freshwater fish. The resistance of erythrocytes and hemoglobin in freshwater fish widely varyies depending on species’ inhabitation conditions. Several strategies for stabilization of respiratory function in the blood of Teleostei were formulated as following: (1) a strategy of compensatory type in the fish ontogenesis, (2) formation of different quality resistance to haemolysis of young and mature erythrocytes; (3) variability resistance of hemoglobin to dehydration and (4) transformation of homeostasis of an organism in extreme conditions.     

Characteristics of Basic Parameters of the Hormonal Function of the Adrenal Cortex and Its Modification in Rat Ontogenesis

The stress-reactivity of the pituitary-adrenocortical system (PAS) as assessed by the dynamics of the blood corticosterone level changes was studied in rats administered with cortisol at different periods of their pre- and postnatal ontogenesis. The participation of the activation and deactivation mechanisms in this process was estimated by means of a mathematical modeling, using the basic parameters of hormonal wave. It is established that in the one-month old rat pups born from mothers injected with cortisol from the day 14 to 18 of pregnancy, the basal and stress-evoked PAS activity was not essentially changed, whereas the adult animals demonstrated a faster decrease of the stress-induced corticosterone level. Injection of cortisol at the early neonatal ontogenesis (1–5 day of life) decreased the basal and stress-induced corticosterone levels at the morning hours in one-month old rats, whereas in adult rats it increased the PAS stress-reactivity. Injection of cortisol in the late neonatal ontogenesis, i.e., during the period of formation of the sensory systems (opening of the ears, eyes, maximal motor activity) resulted predominantly in changing the time of completion of the stress-induced hormonal response that became longer than in control animals of the same age. With the aid of mathematical modeling, we have found that at the early neonatal period of development the hormonal exposure mainly increases the rate of PAS activation, whereas injection of glucocorticoids at the late neonatal period changes PAS regulation by a feedback mechanism, thus decreasing the rate of system inactivation and increasing the time of completion of PAS stress-induced reaction. It is concluded that the phenotypic reorganization of PAS stress-reactivity by exogenous corticosteroids depends on the time of their action on development of the excitatory and inhibitory mechanisms during the critical periods of their formation.     

Physiological and molecular endocrine changes in maturing wild sockeye salmon, <Emphasis Type="Italic">Oncorhynchus nerka</Emphasis>, during ocean and river migration

Maturing adult sockeye salmon Oncorhynchus nerka were intercepted while migrating in the ocean and upstream in freshwater over a combined distance of more than 1,300 km to determine physiological and endocrine changes associated with ionoregulation. Sockeye migrating through seawater and freshwater showed consistent declines in gill Na⁺/K⁺-ATPase (NKA) activity, plasma osmolality and plasma chloride concentration. In contrast, plasma sodium concentration became elevated in seawater as fish approached the river mouth and was then restored after sockeye entered the river. Accompanying the movement from seawater to freshwater was a significant increase in mRNA for the NKA α1a subunit in the gill, with little change in the α1b subunit. Potential endocrine signals stimulating the physiological changes during migration were assessed by measuring plasma cortisol and prolactin (Prl) concentrations and quantifying mRNA extracted from the gill for glucocorticoid receptors 1 and 2 (GR1 and GR2), mineralocorticoid receptor (MR), growth hormone 1 receptor (GH1R), and prolactin receptor (PrlR). Plasma cortisol and prolactin concentrations were high in seawater suggesting a preparatory endocrine signal before freshwater entry. Generally, the mRNA expression for GR1, GR2 and MR declined during migration, most notably after fish entered freshwater. In contrast, PrlR mRNA increased throughout migration, particularly as sockeye approached the spawning grounds. A highly significant association existed between gill PrlR mRNA and gill NKA α1a mRNA. GH1R mRNA also increased significantly, but only after sockeye had migrated beyond tidal influence in the river and then again just before the fish reached the spawning grounds. These findings suggest that cortisol and prolactin stimulate ionoregulation in the gill as sockeye salmon adapt to freshwater.     

Modulation of Sphingosine 1-Phosphate and Tyrosine Hydroxylase in the Stress-Induced Anxiety

Stress causes endocrinological changes and leads to induce anxiety. It was determined the anxiety and stress-related endocrinological changes through the observation of the level of glucocorticoid and sphingolipid metabolites in serum after stress. Immobilized stress and electric shock was applied to rats for 7 days. This study investigated the induction of anxiety, changes of TH and pERK expression in cortex and amygdala after stress. Also it was determined the changes of glucocorticoid and anxiety when the rats were given stress after amygdala lesion. The stress-given rats spent a lesser percentage of time significantly in the open arm than the control rats. The elevated level of glucocorticoid after stress was suppressed in amygdala lesion group. The expression of TH in the amygdala was decreased, but the expression of TH was not changed in the cortex after stress. To investigate the changes in sphingolipid metabolites after stress, the levels of sphingosine and the phosphate form of sphingolipid (So-1-P) were analyzed in serum. The level of So-1-P was elevated after stress and anxiety was observed after the So-1-P infusion (100 pmol/10 μl/h, i.c.v., for 7 days). Continuous infusion of So-1-P for 7 days led to the significant decrease of TH expression in the amygdala. In conclusion, the results of this study indicate that the lesion of amygdala suppressed the stress-induced anxiety and elevation of glucocorticoid in serum. It was also observed that expression of TH in amygdala as well as increased levels of glucocorticoid in serum might be responsible biomarker, at least in part, of chronic stress. These results suggest that the elevation of So-1-P might be involved in induction of anxiety during stress by the modulation of dopaminergic system in amygdala.     

The role of serotonin in the immune system development and functioning during ontogenesis

In this study, we investigated the influence of serotonin on the development and functioning of T- and B-cell-mediated immunity during ontogenesis using the pharmacological model of serotonin depletion in rat fetuses. It has been demonstrated that prenatal serotonin deficiency resulted in a decrease in thymus and spleen weights, changes in their cellular composition, and long-lasting disturbances in cell-mediated and humoral immunity in postnatal ontogenesis. The data obtained suggest that serotonin may be considered a morphogenic factor in development of the immune system.     

Changes in the content of glutamic, aspartic and gamma-aminobutyric acids in mitochondrial fractions of limbic system structures of the dog brain during postnatal ontogenesis

Calculation of the amount of mitochondria isolated from 1 g of fresh tissue has shown that the amount of glutaminic (GA), aspartic (AsA) and gamma-aminobutyric (GABA) acids in the initial mitochondrial fraction of the limbic structure increased during the postnatal ontogenesis and reached the "adult" level in dogs aged 1 year. The GA amount in all the studied periods of the ontogenesis exceeded the AsA and GABA levels in the mitochondrial fraction of the limbic structure.     

Synchronization of changes in the level of direct current potential and concentration of lipid peroxidation products in the developing rat brain]

The dynamics of the level of direct current potentials and the content of lipid hydroperoxides, conjugated dienes, Schiff bases of phospholipids and lipofuscin in the brain of rats during ontogenesis was studied. It is shown that changes in neurophysiological and biochemical characteristics have an M-shaped character. The synchronization of age-dependent changes in the content of lipid peroxidation products and the dynamics of direct current potentials was revealed.