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1.
The binding characteristics of 17,20,21-trihydroxy-4-pregnen-3-one (20-S) to plasma membranes prepared from the testes and sperm of spotted seatrout (Cynoscion nebulosus) were investigated using a filtration method to retain the bound 20-S. A single class of high affinity (Kd = 17.9 nM), low capacity (Bmax = 0.072 nM g-1 testes) binding sites was identified by saturation and Scatchard analyses on testicular membranes of spermiating spotted seatrout. A corresponding receptor (Kd = 22.17 nM, Bmax = 0.00261 nM ml-1 milt) was also detected in spermatozoan membrane preparations. The rates of 20-S association and dissociation were rapid, both had Thalfs of less than 1 min. Competition studies indicated that the receptor was highly specific for 20-S. 17,20-dihydroxy-4-pregnen-3-one, which had the highest affinity of the other steroids tested, had a relative binding affinity (RBA) of 14.3%. Progesterone, 11-deoxycortisol and testosterone competed with an order of magnitude less affinity (RBA's of 7.4, 1.8 and 1.1%, respectively). Estradiol displayed low affinity for the receptor (RBA = 0.4%) and cortisol did not cause any displacement at 1000-fold excess concentration. Specific 20-S receptor binding was detected in plasma membranes from testes of both spermiating and non-spermiating seatrout and on spermatozoa. Prolonged incubation of testicular fragments from a spermiating fish with gonadotropin (15 IU ml-1 human chorionic gonadotropin) or forskolin (10 µM) caused a 2–3 fold increase in membrane receptor binding. Previous studies have shown that gonadotropin-induced upregulation of the 20-S plasma membrane receptor in seatrout ovaries is required for the oocytes to become responsive to 20-S and undergo final maturation. The existence of a 20-S membrane receptor on sperm and its upregulation in the testes by gonadotropin raises the possibility that final maturation of spermatozoa in male seatrout may be regulated by a similar mechanism.  相似文献   

2.
Decreased plasma total cholesterol concentrations are a useful indicator in fish of reduced resistance to bacterial infection. Taking advantage of the hydrophobic properties of silicone rubber, we developed an optical enzyme sensor system for measuring plasma cholesterol concentrations in fish. The system was constructed using an immobilized enzyme membrane, optical oxygen probe, flow system, and personal computer. The enzyme membrane was prepared from cholesterol esterase, cholesterol oxidase, and silicone rubber. The calibration curves were linear in the range of 55–85 mg dl−1 for yellowtail and 20–90 mg dl−1 for rainbow trout. Assays could be completed within 3 min, and the sensor response was stabilized by the hydrophobic properties of silicone rubber. Storage of immobilized enzyme membranes at 5°C permitted stable measurements for 58 days. The sensor system was applied to determine plasma total cholesterol concentrations in fish. Good correlations were obtained between results obtained using the sensor and results obtained using conventional methods (correlation coefficients: yellowtail 0.9751, rainbow trout 0.9947). Our method required much less time than conventional cholesterol assays and can be used economically for continuous determination of plasma total cholesterol in fish.  相似文献   

3.
The fatty acid content, fatty acid molecular species composition, and fluidity of phospholipid vesicles and plasma membranes of livers of fish feeding on phytoplankton (silver carp, Hypophthalmichtys molitrix), zooplankton, (big head carp, Hypophtalmitrix nobilis), higher plants, (grass carp, Ctenopharyngodon idella), and omnivorous fish, (common carp, Cyprinus carpio), adapted to reduced temperatures, was determined. Levels of long chain polyunsaturated fatty acids (PUFAs) were highest in silver and big head carp and lowest in common carp. Differences in long chain PUFAs were poorly reflected in fluidity, determined either by fluorescence anizotropy of 1,6-diphenyl 1,3,5-hexatriene or rotational correlation time of 16-doxyl- stearic acid, of phospholipid vesicles. Phospholipid vesicles of C. caripo and C. idella proved to be slightly more rigid than of H. molitrix and H. nobilis below ca. 15 °C. These differences were not seen with isolated plasma membranes assessed with 1,6-diphenyl 1,3,5-hexatriene. Using electron spin resonance spectroscopy plasma membranes from H. molitrix proved to be more fluid than those from C. carpio below ca. 15 °C. Phosphatidylethanolamines were abundant in 1-monounsaturated, 2-polyunsaturated (18:1/22:6, 18:1/20:4) species with H. molitrix and H. nobilis more rich in these species than C. carpio and C. idella. It is suggested that differences in fluidities of phospholipid vesicles below 15 °C is due to differences in levels of the above molecular species. In native membranes these differences may be masked by proteins and other membrane components. It is also suggested that it is not the fatty acid composition of the ingested food but the specific response of the fish to temperature that is the most important factor controlling the physicochemical properties of membranes during thermal adaptation.  相似文献   

4.
The aim of this work was to verify if, compared to mammals, the lower molecular weight of GH-R previously reported in salmonid is real or due to the experimental process. For this purpose, we compared the apparent molecular weight of GH-R, obtained by SDS-PAGE after cross-linking with 50 I-rtGH, obtained from rainbow trout crude liver membrane preparation, incubated in different buffers with those obtained after purification with affinity chromatography.Using crude liver membrane preparation, two specific bands of 50 I-rtGH-protein complex were observed: the major one corresponds to a MW of 70 kDa and the minor one to 45 kDa. However, the pattern of electrophoresis varied according to the different incubation buffers tested. Digestion of the cross-linked complex with -galactosidase and phospholipase did not significantly modify the position of bands, whilst N-glycosidase F induced a large smear including 4 more pronounced bands (50, 65, 97 and > 130 kDa), the heavier band corresponding to the most intensive signal.GH receptors were purified using solubilisation and affinity chromatography. The yield of the liver GH-R from crude liver membrane preparation by the solubilization technique was optimized (48%) using Triton 1% for 1 h (12°C ). Specific binding sites in the solubilized membrane proteins were saturable when incubated with increasing 50 I-rtGH concentrations, and revealed a high affinity constant (Ka=0.7×109 M–1). After affinity chromatography, specific binding activity was increased 64,000 fold. However, the purity of the preparation was partial and the purification yield was very low (about 0.3%). This enriched fraction, analysed by SDS-PAGE after cross-linking, showed a very intense band (about 63 kDa) which disappeared with an excess of cold rtGH.These results suggest that the lower molecular weight observed in salmonid (41 kDa), compared to mamals, is not due to the experimental process. The significance of GH-R size difference between salmonids and mammals is discussed.  相似文献   

5.
The presence of 17,20-dihydroxy-4-pregnen-3-one (17,20-DHP) oocyte receptor activity has been demonstrated in brook troutSalvelinus fontinalis. Scatchard analyses of the cytosol fraction during various terminal stages of oocyte maturation gave a high equilibrium association constant (Ka) value of 1.394±0.669 108M–1 (n=7) and low maximum binding capacities (Nmax). The association kinetics of the receptor was second order k+1=2.292×106M–1 sec–1. The dissociation rate constant ka was 1.502×10–2 sec–1 for the first order dissociation reaction. The Ka=1.526×108M–1, when it was determined from k+1/k–1 a value close to that found from the Scatchard analysis. Competition studies showed the following binding affinities testosterone > 17-HP > 17,20-DHP > Promegestone > progesterone > estradiol > pregnenolone; cortisol showed no competitive inhibition. Cytosolic extracts when pre-equilibrated with various labelled steroids and eluted from a Sephacryl S-300 column gave multiple specific binding peaks. On sucrose density gradient centrifugation specific binding was observed at 3.05 S in cytosol containing 0.15M sodium chloride buffer. The receptor lost binding activity when incubated with various proteases, but DNase and RNase had no effect. Blood plasma without heparin at (110) dilution also bound [3H]17,20-DHP, Ka was 8.04×107 M–1.The nuclear pellet extract (750×g) gave very little specific binding activity even at high radiolabelled steroid concentrations and a linear Scatchard plot was not obtained. Nevertheless the nuclear extract, after dextran-charcoal treatment, pre-equilibrated with [3H]17,20-DHP, bound specifically to DNA cellulose, and cytosol from the same oocytes also bound to DNA cellulose under similar conditions. Although specific binding to DNA cellulose was obtained the salt concentrations at which the steroid-receptor complex elution took place was not reproducible in both nuclear extracts and cytosol samples. Also binding activity was extremely small compared to the total cytosolic binding. The nuclear extract when pre-equilibrated with high concentrations (20 nM) of the labelled steroid and then chromatographed on Sephacryl S-300 column gave a specific binding peak which was similar to that of the cytosolic preparation.The receptor levels in cytosol decreased progressively during final maturation (Stages 1–7). There is preliminary evidence for the presence of 17,20-DHP receptor activity in cytosol of landlocked Atlantic salmonSalmo salar ouananiche, and rainbow troutSalmo gairdneri. The zona radiata fraction from late stages oocyes 5, 6, and 7 in brook and rainbow trout oocytes were isolated by ultracentrifugation; from this fraction a protein was characterized which covalently bound [3H]R5020 after photoaffinity labelling. The same protein also bound [3H]17,20-DHP after solubilization in Brig 35 buffer. The SDS gel electrophoresis subunit composition of the above protein was similar to the cytosol counterpart binding [3H]17,20-DHP, although the molecular weights were different. The blood sample [3H]R5020 binding component subunit composition was different from that of the membrane extracted protein. These results demonstrate the presence of 17,20-DHP receptor activity in the cytosol and zona radiata membranes of the oocytes during final maturation.A. Maneckjee is presently NSERC postgraduate scholar at MSRL and Ph.D. candidate at Department of Biochemistry, Memorial University of Newfoundland.  相似文献   

6.
A recombinant carp growth hormone (rcGH) was used to develop for a GH radioreceptor binding assay in the goldfish (Carassius auratus). Specific binding of125I-rcGH to goldfish liver membranes was a pH, time, temperature, and membrane protein dependent process. Scatchard and LIGAND analysis indicated a single class of high affinity and low capacity binding site, with an association constant (Ka) of 1.9×1010 M–1 and a maximum binding capacity (Bmax) of 9 fmol mg–1 protein. Liver tissue displayed the highest125I-rcGH binding of all the tissues examined. Displacement of125I-rcGH with various unlabeled teleost and mammalian GHs and prolactins revealed that the goldfish hepatic binding site was highly specific for teleost GH. Intraperitoneal administration of 0.1, 1.0, and 10 g rcGH g–1 body weight to hypophysectomized goldfish resulted in a 27, 52, and 68% decrease in total binding sites, respectively. Injection of a high dose of rat prolactin (rPRL) (5 g rPRL g–1 body weight) also resulted in a 32% decrease in total binding sites. These results suggest that endogenous GH may have a role in the regulation of its own receptors in the goldfish.  相似文献   

7.
PKC activity was detected in spleen extracts from the turbot, Scophthalmus maximus, a teleost flatfish that is farmed commercially in several countries, in assays with the substrate EGF- R651–658 as phosphate acceptor. The activity was purified about 700-fold by a three-step chromatographic procedure (DEAE-cellulose, phenyl-Sepharose and threonine-Sepharose). Maximal activity was obtained in the presence of the typical PKC cofactors Ca2+ (0.1 mM) PtdS (20 g ml–1) and either DAG (2 g ml–1) or PMA (2 g ml–1). Activity was dose-dependently inhibited by H7 and by the PKC-specific inhibitors PKC19–36 and N-myristoylated PKC19–31. The rate of phosphorylation was highest with the PKC-specific substrate MARCKS161–175. In immunoblotting, MC5 (a mouse monoclonal antibody raised against bovine PKC) recognized bands of 80 and 100 kDa. Immunoblotting with antibodies raised against mouse PKC isozymes (, , , , , , and ) indicated the presence of all these isozymes in turbot spleen.  相似文献   

8.
Gill Na+-K+ ATPase and carbonic anhydrase activities were measured, on a fortnightly basis, from February to July, in 0+ age Atlantic salmon (Salmo solar), hatched and reared in a freshwater experimental station in Covas, northern Portugal. Plasma osmolarity and ionic composition were also measured. Gill Na+-K+ ATPase activity increased slowly until April (15–19 moles Pi mg prot–1 h–1). From April to late May there was a great increase in activity (19–32 moles Pi mg prot–1 h–1) followed by a sharp decline in June (15 moles Pi mg prot–1 h–1). In contrast, carbonic anhydrase activity decreased significantly from early April to early June (170-70 moles p-nitrophenol mg prot–1 h–1) and increased in late June, suggesting the existence of a compensatory mechanism for the changes in Na+-K+ ATPase activity. Plasma osmolarity and sodium concentration showed lower levels during the period of high ATPase activity. On the other hand, plasma calcium concentrations showed an increase during the same period (3.47–5.98 mm1–1 of plasma). A transitory decrease in osmolarity and plasma sodium and chlorine concentrations occurred in March, prior to the surge in Na+-K+ ATPase activity, suggesting that the physiological changes, characteristic of parr-smolt transformation can be a consequence of this loss of freshwater osmoregulatory capacity.  相似文献   

9.
Soft water acclimated (Ca2+ 0.02 mM; Na+ 0.03 mM; K+ 0.01 mM; pH 7.0), cannulated brown trout (Salmo trutta) were exposed to various pH and aluminium (Al) regimes (pH 7.0, pH 5.0, pH 5.0 plus Al: 50, 25, and 12.5 g l–1) for up to 5 days in order to determine (i) the sublethal concentration of Al at pH 5.0 for this species (ii) their ionoregulatory and respiratory status. No mortality or physiological disturbances were evident at pH 7.0 or pH 5.0. All trout died within 48 h at pH 5.0 in the presence of Al at 50 g l–1 and 67% died over the 5 day period at pH 5.0 in the presence of Al at 25 g l–1. Fish at these lethal Al concentrations showed significant decreases in arterial blood oxygen content (CaO2) but no changes in plasma osmolarity or the concentrations of plasma Na+, K+ and Cl–. Physiological disturbance was more marked at the 50 g l–1 Al concentration. The surviving fish at 25 g l–1 showed few signs of physiological recovery while continually exposed to this regime. No fish died during the exposure to water of pH 5.0 containing 12.5 g l–1 Al, but physiological disturbance was still apparent. These sublethally-stressed trout showed a transient decline in the plasma concentrations of Na+ and Cl–1. Although CaO2 decreased, recovery was evident. The data suggest that in the brown trout, environmental Al concentration is as important as pH and calcium concentration in determining the physiological status of the fish.  相似文献   

10.
Total lipids, lipid classes and their associated fatty acids were measured in developing eggs, yolksac larvae and starving larvae (from fertilized egg to day 9 after hatch) of the common dentex Dentex dentex (L., 1758). The larvae of common dentex during lecitotrophia and subsequent starvation consumed 1.6 g of total lipid per larvae per day. The overall decrease was mainly due to utilization of the major neutral lipids, TAG and SE (0.5 and 0.6 g larvae–1 day–1, respectively) which was 3.4-fold greater than that of the the major phosphoglycerides (primarily PC by 0.2 g larvae–1 day–1). There was net synthesis/conservation of PE during the first half of the study period before it decreased rapidly (0.2 g larvae–1 day–1) during the second half. PUFAs were principally catabolized (468.6 ng larvae–1 day–1), primarily 22:6(n-3), 20:5(n-3) and 20:4(n-6) (221.8, 58.5 and 12.1 ng larvae–1 day–1, respectively). Saturated and monounsaturated fatty acids were also utilized (227.2 and 256.7 ng larvae–1 day–1, respectively), principally 16:0 and 18:1(n-9) that were both consumed at 149.8 and 156.7 ng larvae–1 day–1, respectively. The rank order of utilization of fatty acids (ng larvae–1 day–1) by D. dentex larvae from total lipids, PC and TAG coincided with the order of abundance of the different fatty acids in the respective lipid fractions. However, in PE, the most abundant fatty acid, DHA, was relatively conserved and 16:0, the second most abundant fatty acid, was catabolized to the greatest extent. D. dentex showed a pattern of lipid metabolism during early development similar to that of marine larval fish from temperate waters whose eggs contain high levels of total lipids, including an oil globule, and which preferentially utilize neutral lipids as the primary energy source.  相似文献   

11.
Vitellogenin (Vtg) was purified from the plasma of 17-estradiol (E2)-injected male greenback flounder,Rhombosolea tapirina. The molecular weight of the native Vtg was estimated by gel filtration as 540 kD. SDS-PAGE and Western blotting analyses indicated that this protein consisted of three bands with molecular weights of 155, 104, 79 kD, respectively. A polyclonal antibody against the highest molecular weight band of putative Vtg was generated in sheep and an indirect antibody-capture competitive enzyme-linked immunosorbent assay (ELISA) was developed. The assay was validatedfor plasma Vtg measurement in greenback flounder. Serial dilutions of plasma from vitellogenic females parallelled the standard Vtg curve, whereas no cross-reaction was observed with the plasma of males in the ELISA. The Vtg ELISA was used to assess the induction of Vtg by E2 in vivo in males. The induction of Vtg in greenback flounder showed a time- and dose-dependent response as in other species. In E2-treated fish, detectable levels of Vtg were first found at 48 h, and reached a peak at 96 h post-injection. Plasma levels of Vtg increased as the E2 dose increased with a threshold of 0.1 mg kg–1.  相似文献   

12.
The present study investigated the relationships between egg viability and ovarian fluid composition, egg physiology and egg metabolism in lake trout, Salmo trutta lacustris, to obtain biomarkers for egg quality determination. The ovarian fluid pH, protein levels and activities of aspartate aminotransferase and -d-glucuronidase were significantly correlated with egg viability expressed as the number of eyed stage embryos. Regression models demonstrated that an ovarian fluid pH between 8.44 and 8.57, protein levels below 235.56 mg 100 ml–1ovarian fluid, aspartate aminotransferase activity below 31.65 m min–1 l–1ovarian fluid and -d-glucuronidase activity below 8.62 m min–1 l–1 ovarian fluid characterized egg batches with high viability (80%).The increase in the egg wet weight during water hardening was also significantly correlated with the number of eyed stage embryos, and egg batches with high egg viability (80%) increased in wet weight by 13% during water hardening.From the investigated metabolic parameters the number of eyed stage embryos was significantly correlated with activities of NADP-dependent isocitrate dehydrogenase (egg viability 80% at 2.07 nM min–1 mg–1 protein) and NAD-dependent malate dehydrogenase (egg viability 80% at 47.25 nM min–1 mg–1 protein), with the respiration rate (egg viability 80% at 8.71 nM min–1 mg–1 protein), with the ratio of NADH to NAD levels (egg viability 80% 0.872), with the levels of free, non-esterified fatty acids (egg viability 80% 72.34 g mg–1 protein), and the ratio of non esterified to esterified fatty acids (egg viability 80% at 0.749). Also, subjective and visual control methods were described to distinguish between batches with viable and non viable eggs.  相似文献   

13.
The aim of this work was to determine the effects of supplemental dietary sodium chloride on salt water acclimation of tilapia Oreochromis niloticus. Fish were fed a basal diet supplemented with NaCl (8%) during three weeks in fresh water (FW) and then transferred to salt water (SW) at 15 and 20. Changes in plasma osmolality, chloride ion concentration (Cl–), plasma level of cortisol and gill Na+, K+-ATPase activity were measured at 6, 12, 24, 48, 72 and 168 h after transfer to 15SW, while the higher strength SW group (20) was only monitored up to 24 h. Morphological changes in the gill mitochondria-rich (MR) cells were examined in relation to environmental salinity. The changes associated with dietary NaCl were sporadic and of small magnitude. The plasma osmolality and Cl– increased immediately after transfer up to 12–24 h, but fish fed dietary salt (S) showed lower values than the control group (C). The S group showed higher plasma levels of cortisol than the control, which maintained its initial levels during the experiment. Gill Na+, K+-ATPase activity of the S group began to increase in the first hours after transfer, reaching maximum at 12 h and returned to basal level at 24 h, while the control group maintained basal levels. The differences between gill Na+, K+-ATPase activity of S and C fish were significant (p < 0.05) at 12 h. Transmission electron microscopy (TEM) revealed that MR cells in SW show more mitochondria and a more developed tubular system arising from the basolateral membrane. The MR cells of both groups frequently formed a multicellular complex in SW, consisting of a main MR and one or more accessory cells. Such complexes are rarely observed in FW. Some MR cells of fish fed supplemented dietary salt displayed convex apical membrane in FW.  相似文献   

14.
The accumulation of immunoreactive corticosteroids in the bile of rainbow trout during stress was monitored by radioimmunoassay and GUMS. Although plasma cortisol levels were elevated by confinement for 1 hour, biliary levels of free and conjugated steroids in the bile were unaffected. However, after 24 hours confinement, in addition to elevated plasma cortisol levels, free and conjugated steroids in the bile were also significantly higher than in control, unstressed fish. The time-course of change in plasma and biliary corticosteroid levels was determined in rainbow trout subject to 96 hours confinement stress. Free steroid levels in the bile of stressed fish were elevated within 2 hours of the onset of stress, while levels of conjugated steroids were significantly elevated within 4 hours of the onset of confinement. Analysis of bile from stressed fish, by GC/MS, established the major conjugated steroids present to be tetrahydrocortisone (230 g ml–1 bile), tetrahydrocortisol (75 g ml–1), cortisone (33.5 g ml–1), cortisol (25 g ml–1) and -cortolone (5 g ml–1). The data are discussed with reference to the role of cortisone and conjugating enzymes in the clearance of cortisol, and further data are presented to suggest that the analysis of biliary steroid content may provide a suitable means of identifying stressed fish under conditions in which an additional sampling stress is unavoidable.  相似文献   

15.
Compared to control fish, gonadotropin releasing hormone-analogue (GnRHa) treatment delivered either by microspheres or cholesterol pellets successfully increased sperm production (cells kg–1) and milt volume (ml kg–1) in mature yellowtail flounder Pleuronectes ferrugineus during the spawning season. Spermatocrit decreased in both treated and control groups between 12 and 29 days post-implantation, indicating a seasonal decrease in sperm concentration, rather than an effect of the GnRHa treatments.Plasma levels of testosterone, 11-ketotestoterone and 17,20dihydroxy-4-pregnen-3-one (1720P) showed no clear pattern either across treatments or over days, however this does not exclude the possiblity that GnRHa had its effect on milt volumes via the stimulation of steroid production since the sampling protocol did not allow for the rapid clearance of steroids from the plasma. GnRHa treatment did not have a negative effect on sperm fertilizing ability, percentage hatch or larval morphology. Sperm motility and seminal plasma pH were increased by GnRHa treatment.  相似文献   

16.
An intracellular ATP-dependent Ca2+ pumping mechanism, distinct from mitochondrial Ca2+ accumulation, was identified within tilapia gill cells. Cell suspensions treated with 0.003% saponin, which selectively permeabilizes the plasma membrane, were used to characterize the Ca2+ sequentering mechanisms as endoplasmic reticulum and mitochondria and to determine the effect of Zn2+ on their Ca2+ storing activity. Of the Ca2+ taken up by the endoplasmic reticulum, 80% was released by IP3 (10 mol l–1). The Ca2+ pump of the endoplasmic reticulum was 2.5 times less sensitive to Zn2+ (IC50=0.05 nmol l–1) than was the mitochondrial uptake mechanism (IC50=0.20 nmol l–1). The results indicate that Ca2+ is stored predominantly within the endoplasmic reticulum at 0.1 mol l–1 and that this storing capacity is seriously attenuated by namomolar concentrations Zn2+.  相似文献   

17.
Distribution and function of hepatic - and -adrenoceptors were examined in rainbow trout (Oncorhynchus mykiss) injected with slow release hydrogenated coconut oil implants alone (sham) or containing cortisol. - and -Adrenoceptors were assayed on purified hepatic membranes 10–14 days post-implantation using 3H-prazosin () and 3H-CGP (). At 10–14 days, plasma cortisol values were significantly elevated to approximately 220 compared with 35.0 ng ml-1 in cortisol implanted vs. sham trout. No significant differences were found between any of the experimental groups for either the affinity (Kd) or maximal number of binding sites (Bmax) for either receptor type. Epinephrine significantly stimulated glucose release from hepatocytes isolated from sham injected trout, but not from cortisol-treated fish. Epinephrine-induced glucose release was blocked by both - and -antagonists. These studies do not support the hypothesis that rainbow trout exposed to chronic cortisol alter properties of hepatic adrenoceptors.  相似文献   

18.
The role of calcium influx through voltage-dependent calcium ion channels in the exocytotic response of single isolated pars intermedia cells from the teleost tilapia (Oreochromis mossambicus) was investigated by means of the whole-cell patch-clamp technique and high resolution electrical measurements. Calcium currents differed from barium currents in several ways: the peak ICa was smaller, the current-voltage relationship for calcium attained its maximum at +10 mV instead of 0 mV, and the inward calcium current inactivated more rapidly. Electron micrographs showed that pars intermedia cells possess dense, encored vesicles with an average diameter of 140 nm. Influx of calcium resulted in an increased cell membrane capacitance (Cm) after the depolarizing period, indicating a fast exocytotic response. Comparison with late recordings revealed the presence of a transient in Cm, presumably attributable to movement of ion channel gates. The average increase in Cm was 13.4 fF, suggesting the fusion of at least 23 vesicles with the plasma membrane during the depolarizing pulse. In contrast to calcium, barium did not support significant exocytosis. We conclude that calcium entry through voltage-dependent calcium channels rapidly leads to the exocytosis of secretory vesicles from tilapia pars intermedia cells.  相似文献   

19.
Immature brook trout (Salvelinus fontinalis) were randomly divided into a pH control, a pH and food control and an acid-stressed group. Fish in the first two groups were held at neutral pH and those in the last group were maintained at pH 4.2 for up to two months. The food supply to the pH and food control group was restricted to simulate the reduction in food intake demonstrated for acid-stressed trout. Plasma insulin levels were significantly decreased from 5–20 ng/ml to 1–2 ng/ml and plasma cortisol levels were significantly increased from 5–10 ng/ml to as high as 70 ng/ml in the acid-stressed brook trout. Concomitantly, a significant decrease of 21–39% in the proportion (volume density) of insulin immunoreactive -cells was observed within the principal pancreatic islets. Somatic growth was stunted and ultrastructural morphometry revealed the suppression of somatotrope secretory activity in the acid-stressed fish. Restriction of food supply induced a smaller but still significant decrease in circulating levels of insulin which was however not accompanied by a reduction in insulin immunoreactive -cells. The rise in plasma cortisol levels was not significant, and the plasma levels of glucose and protein were unaffected. Nevertheless, somatotrope secretory activity was suppressed and somatic growth was stunted. This study demonstrates for the first time the complexity of the endocrine response to acid stress and that some of the response to acid stress can be attributed to the lowering of food intake.  相似文献   

20.
Environmental stressors caused by inadequate aquaculture management strategies suppress the immune response of fish and make them more susceptible to diseases. Therefore, efforts have been made to relieve stress in fish by using various functional feed additives in the diet, including probiotics. The present work evaluates the effects of Lactobacillus rhamnosus (LR) on physiological stress response, blood chemistry and mucus secretion of red sea bream (Pagrus major) under low salinity stress. Fish were fed four diets supplemented with LR at [0 (LR0), 1 × 102 (LR1), 1 × 104 (LR2) and 1 × 106 (LR3) cells g?1] for 56 days. Before stress, blood cortisol, urea nitrogen (BUN) and total bilirubin (T-BIL) showed no significant difference (P > 0.05), whereas plasma glucose and triglyceride (TG) of fish-fed LR2 and LR3 diets were significantly lower (P < 0.05) than those of the other groups. Plasma total cholesterol (T-CHO) of fish-fed LR3 diet was significantly (P < 0.05) lower than that of the other groups. Furthermore, total plasma protein, mucus myeloperoxidase activity and the amount of mucus secretion were significantly enhanced in LR-supplemented groups when compared with the control group (P < 0.05). After the application of the low salinity stress test, plasma cortisol, glucose, T-CHO and TG contents in all groups showed an increased trend significantly (P < 0.01) compared to the fish before the stress challenge. However, plasma total protein and the amount of secreted mucus showed a decreased trend in all groups. On the other hand, BUN, T-BIL and mucus myeloperoxidase activity showed no significant difference after exposure to the low salinity stress (P > 0.05). In addition, the fish that received LR-supplemented diets showed significantly higher tolerance against low salinity stress than the fish-fed LR-free diet (P < 0.05). The physiological status and the detected immune responses, including total plasma protein and mucus myeloperoxidase activity in red sea bream, will provide a more comprehensive outlook of the effects of probiotics to relieve stress in fish.  相似文献   

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