Osmoregulatory action of PRL,GH, and cortisol in the gilthead seabream (Sparus aurata L)

The osmoregulatory actions of ovine prolactin (oPRL), ovine growth hormone (oGH), and cortisol were tested in the euryhaline gilthead seabream Sparus aurata. Acclimated to sea water (SW, 40 ppt salinity, 1000 mOsm/kg H(2)O) or brackish water (BW, 5 ppt, salinity, 130 mOsm/kg H(2)O), injected every other day for one week (number of injections, 4) with saline (0.9% NaCl), oPRL (4 microg/g body weight), oGH (4 microg/g body weight) or cortisol (5 microg/g body weight), and transferred from SW to BW or from BW to SW 24h after the last injection. Fish were sampled before and 24h after transfer. Gill Na(+), K(+)-ATPase activity, plasma osmolality, plasma ions (sodium and chloride), plasma glucose, and muscle water moisture were examined. SW-adapted fish showed higher gill Na(+), K(+)-ATPase activity, plasma osmolality, and plasma ions levels than BW-adapted fish. Transfer from SW to BW decreased plasma osmolality and ions levels after 24h, while transfer from BW to SW increased these parameters, whereas gill Na(+),K(+)-ATPase activity was unaffected. oPRL treatment significantly decreased gill Na(+),K(+)-ATPase activity and increased plasma osmolality and ions in SW- and BW-adapted fish. This treatment minimizes loss of osmolality and ions in plasma after transfer to BW and increased these values after transfer to SW. No significant changes were observed in gill Na(+),K(+)-ATPase activity, plasma osmolality, and plasma ions in oGH-treated group with respect to saline group before or after transfer from SW to BW or from BW to SW. Treatment with cortisol induced, in SW-adapted fish, a significant increase of gill Na(+),K(+)-ATPase activity and decrease of plasma osmolality and plasma ions. In BW-adapted fish this treatment induced a significant increases in gill Na(+),K(+)-ATPase activity, plasma osmolality, and plasma ions. After transfer to SW cortisol-treated fish had higher plasma osmolality than the saline group. Our results support the osmoregulatory role of PRL in the adaptation to hypoosmotic environment in the gilthead seabream S. aurata. Further studies will be necessary to elucidate the osmoregulatory role of GH in this species. Cortisol results suggest a "dual osmoregulatory role" of this hormone in S. aurata.     

Changes in gene expression for GH/PRL/SL family hormones in the pituitaries of homing chum salmon during ocean migration through upstream migration

Gene expression for growth hormone (GH)/prolactin (PRL)/somatolactin (SL) family hormones in the pituitaries of homing chum salmon were examined, because gene expression for these hormones during ocean-migrating phases remains unclear. Fish were collected in the winter Gulf of Alaska, the summer Bering Sea and along homing pathway in the Ishikari River-Ishikari Bay water system in Hokkaido, Japan in autumn. The oceanic fish included maturing adults, which had developing gonads and left the Bering Sea for the natal river by the end of summer. The absolute amounts of GH, PRL and SL mRNAs in the pituitaries of the maturing adults in the summer Bering Sea were 5- to 20-fold those in the winter Gulf of Alaska. The amount of GH mRNA in the homing adults at the coastal seawater (SW) areas was smaller than that in the Bering fish, while the amount of PRL mRNA remained at the higher level until fish arrived at the Ishikari River. The gill Na⁺,K⁺-ATPase activity in the coastal SW fish and the plasma Na⁺ levels in the brackish water fish at the estuary were lowered to the levels that were comparable to those in the fresh water (FW) fish. In conclusion, gene expression for GH, PRL and SL was elevated in the pituitaries of chum salmon before initiation of homing behavior from the summer Bering Sea. Gene expression for GH is thereafter lowered coincidently with malfunction of SW adaptability in the breeding season, while gene expression for PRL is maintained high until forthcoming FW adaptation.     

Cortisol promotes and integrates the osmotic competence of the organs in North African catfish (Clarias gariepinus Burchell): Evidence from in vivo and in situ approaches

The short-term in situ and long-term in vivo effects of cortisol were examined in North African catfish (Clarias gariepinus) to identify how this major corticosteroid integrates the osmotic competence of fish organs. In the in situ approach, the hydromineral effects of cortisol perfusion (75-300 ng ml⁻¹) for 20 min were tested and the indices of hydromineral and metabolic regulations were measured in our in vivo experimental fish after three alternate intraperitoneal cortisol injections (40 and 200 ng g⁻¹ body mass) for 5 days. Na⁺, K⁺-ATPase activity, a measure of cellular osmotic competence, responded to in situ and in vivo cortisol treatments. In situ cortisol delivery increased the Na⁺, K⁺-ATPase activity in the gill (P < 0.001) and kidney (P < 0.001) but decreased (P < 0.01) in the liver and showed no effect on intestine. In vivo cortisol treatment, on the contrary, increased Na⁺, K⁺-ATPase activity in the gills (P < 0.01), intestine (P < 0.05) and liver (P < 0.01) but decreased (P < 0.05) in the kidney. As expected, plasma cortisol increased (P < 0.001) with increasing doses of cortisol injections which produced direct effects on the metabolites and the mineral contents including the elevations of glucose (P < 0.05), lactate (P < 0.05) and Mg²⁺ (P < 0.05) and reductions of urea (P < 0.05), Na⁺ (P < 0.05) and K⁺ (P < 0.05) in the plasma. A decline of triiodothyronine (P < 0.01) occurred in the catfish after in vivo cortisol treatment and that implies a direct cortisol action on the homeostatic integration in this fish. Evidence is thus presented that in catfish cortisol regulates the whole body hydromineral and metabolite homeostasis by promoting and integrating the osmotic and metabolic functions of the multiple organ systems including liver.     

Disparate release of prolactin and growth hormone from the tilapia pituitary in response to osmotic stimulation

In most teleost fishes, prolactin (PRL) plays a key role in freshwater (FW) adaptation, whereas growth hormone (GH) is involved in seawater (SW) adaptation in salmonids and certain euryhaline species including the tilapia, Oreochromis mossambicus. Consistent with its osmoregulatory activity, PRL release increases in response to physiologically relevant reductions in extracellular osmolality. When dispersed PRL and GH cells from FW-acclimatized fish were incubated in media of varying osmolalities, PRL release increased significantly in response to a 12% reduction in medium osmolality during 1 and 4h of exposure. By contrast, cells from SW-acclimatized fish responded only to a 24% reduction in osmolality. Growth hormone release on the other hand increased whether medium osmolality was reduced or raised. Cell volume increased together with PRL release during the perifusion of dispersed PRL cells in direct proportion to the reduction in medium osmolality. Growth hormone release increased whether GH cell volume increased or decreased. In in vivo studies, circulating PRL levels increased as early as 1h after the transfer of fish from SW to FW, whereas GH levels remained unchanged during 24h of acclimatization. These results indicate that while PRL and GH cells are osmosensitive, the PRL cells respond to reductions in extracellular osmolality in a manner that is consistent with PRL's physiological role in the tilapia. While the rise in GH release following the reduction in osmolality is of uncertain physiological significance, the rise in GH release with the elevation of medium osmolality may be connected to its role in SW adaptation.     

Differential expression of PTHrP and its receptor in pituitary gland and gills in estradiol-treated gilthead sea bream (Sparus auratus, L.)

In the gilthead sea bream (Sparus auratus) 17β-estradiol (E₂) plays an important role in the synthesis of vitellogenin. During vitellogenesis, vitellogenin as a nutritional precursor protein is loaded with calcium, which requires elevated plasma calcium levels. This is accomplished via E₂-dependent processes. Reports have shown that hypercalcemic effects of E₂ are possibly mediated by another hypercalcemic factor, viz. parathyroid hormone related protein (PTHrP). To further investigate the possibility of PTHrP as a mediator of E₂-induced hypercalcemia, we investigated the local expression levels of the pthrp mRNA and of the gene coding for the PTHrP receptor, PTH1R (pth1r) in two tissues involved in the calcium regulation (gills, pituitary gland) of the sea bream treated with E₂. Compared to control, treatment with E₂ resulted in: significantly increased total calcium and plasma PTHrP levels (P < 0.01), a down-regulation of pthrp mRNA in the pituitary gland (P < 0.01), and up-regulation of expression levels for both pthrp and pth1r in the branchial system (P < 0.05). These findings provide direct evidence for a mediating role of PTHrP in E₂ induced hypercalcemia, and in addition support the idea for the presence of two independent systems, an endocrine pituitary PTHrP system and a peripheral paracrine branchial PTHrP system.     

Ambient salinity modifies the action of triiodothyronine in the air-breathing fish Anabas testudineus Bloch: Effects on mitochondria-rich cell distribution, osmotic and metabolic regulations

The hydromineral and metabolic actions of thyroid hormone on osmotic acclimation in fish is less understood. We, therefore, studied the short-term action of triiodothyronine (T₃), the potent thyroid hormone, on the distribution and the function of gill mitochondria-rich (MR) cells and on the whole body hydromineral and metabolic regulations of air-breathing fish (Anabas testudineus) adapted to either freshwater (FW) or acclimated to seawater (SA; 30 g L⁻¹). As expected, 24 h T₃ injection (100 ng g⁻¹) elevated (P < 0.05) plasma T₃ but classically reduced (P < 0.05) plasma T₄. The higher Na⁺, K⁺-ATPase immunoreactivity and the varied distribution pattern of MR cells in the gills of T₃-treated FW and SA fish, suggest an action of T₃ on gill MR cell migration, though the density of these cells remained unchanged after T₃ treatment. The ouabain-sensitive Na⁺, K⁺-ATPase activity, a measure of hydromineral competence, showed increases (P < 0.05) in the gills of both FW and SA fish after T₃ administration, but inhibited (P < 0.05) in the kidney of the FW fish and not in the SA fish. Exogenous T₃ reduced glucose (P < 0.05) and urea (P < 0.05) in the plasma of FW fish, whereas these metabolites were elevated (P < 0.05) in the SA fish, suggesting a modulatory effect of ambient salinity on the T₃-driven metabolic actions. Our data identify gill MR cell as a target for T₃ action as it promotes the spatial distribution and the osmotic function of these cells in both fresh water and in seawater. The results besides confirming the metabolic and osmotic actions of T₃ in fish support the hypothesis that the differential actions of T₃ may be due to the direct influence of ambient salinity, a major environmental determinant that alters the osmotic and metabolic strategies of fish.     

Effect of extracellular osmolality and ionic levels on pituitary prolactin release in euryhaline silver sea bream (Sparus sarba)

In many euryhaline fish, prolactin (PRL) plays a key role in freshwater adaptation. Consistent with this function, the present study showed a remarkable reduction in pituitary PRL content of silver sea bream abruptly transferred to low salinity (6 ppt). This reduction in pituitary PRL content followed closely the temporal changes in serum osmolality and ion levels. Serum osmolality, Na⁺ and Cl⁻ levels of silver sea bream abruptly transferred to hyposmotic salinity (6 ppt) were markedly reduced 2 h after the transfer. The decline in pituitary PRL content lagged behind the serum changes implying that reduction in pituitary PRL content is a response to the drop in serum ion levels and osmotic pressure. Silver sea bream pituitary cells were dispersed and exposed to a medium with reduced ion levels and osmolality in vitro, and PRL released from pituitary cells was significantly elevated. In hyposmotic exposed anterior pituitary cells, cell volume exhibited a 20% increase when exposed to a medium with a 20% decrease in osmolality. The enlarged pituitary cells did not shrink until the surrounding hyposmotic medium was replaced, a phenomenon suggesting an osmosensing ability of silver sea bream PRL cells for PRL secretion in response to a change in extracellular osmotic pressure. The decrease in pituitary PRL content in vivo and stimulated pituitary PRL release in vitro under reduced osmolality together suggest hyposmotic exposure triggers PRL release from the pituitary.     

Effects of short-term acid and aluminum exposure on the parr-smolt transformation in Atlantic salmon (Salmo salar): disruption of seawater tolerance and endocrine status

Episodic acidification resulting in increased acidity and inorganic aluminum (Al_i) is known to interfere with the parr-smolt transformation of Atlantic salmon (Salmo salar), and has been implicated as a possible cause of population decline. To determine the extent and mechanism(s) by which short-term acid/Al exposure compromises smolt development, Atlantic salmon smolts were exposed to either control (pH 6.7–6.9) or acid/Al (pH 5.4–6.3, 28–64 μg l⁻¹ Al_i) conditions for 2 and 5 days, and impacts on freshwater (FW) ion regulation, seawater (SW) tolerance, plasma hormone levels and stress response were examined. Gill Al concentrations were elevated in all smolts exposed to acid/Al relative to controls confirming exposure to increased Al_i. There was no effect of acid/Al on plasma ion concentrations in FW however, smolts exposed to acid/Al followed by a 24 h SW challenge exhibited greater plasma Cl⁻ levels than controls, indicating reduced SW tolerance. Loss of SW tolerance was accompanied by reductions in gill Na⁺,K⁺-ATPase (NKA) activity and Na⁺,K⁺,2Cl⁻ (NKCC) cotransporter protein abundance. Acid/Al exposure resulted in decreased plasma insulin-like growth factor (IGF-I) and 3,3′,5′-triiodo-l-thyronine (T₃) levels, whereas no effect of treatment was seen on plasma cortisol, growth hormone (GH), or thyroxine (T₄) levels. Acid/Al exposure resulted in increased hematocrit and plasma glucose levels in FW, but both returned to control levels after 24 h in SW. The results indicate that smolt development and SW tolerance are compromised by short-term exposure to acid/Al in the absence of detectable impacts on FW ion regulation. Loss of SW tolerance during short-term acid/Al exposure likely results from reductions in gill NKA and NKCC, possibly mediated by decreases in plasma IGF-I and T₃.     

Mechanism of hCG-induced spermiation in the toad Rhinella arenarum (Amphibia, Anura)

In Rhinella arenarum spermiation occurs as a consequence of LH/FSH increase during the amplexus or by a single dose of hCG, among other gonadotropins. The present study employs an in vitro system to study the mechanism of action of hCG in the spermiation of R. arenarum. Testicular fragments were incubated for 2 h at 28 °C in the presence or absence of 20 IU hCG with or without different PKA/PKC inhibitors and activators as well as ouabain and amiloride as Na⁺/K⁺ ATPase and transcellular Na⁺ transport inhibitors, respectively. Ouabain did not induce spermiation in absence of hCG and inhibited hCG-induced spermiation in a dose-dependent manner, reaching 90% inhibition with the higher concentration. In contrast, amiloride neither affected spermiation nor steroidogenesis. Activation of PKA with 8Br-cAMP induced spermiation in the absence of hCG while its inhibition with H89 blocked hCG action. On the other hand, PKC inhibition with Bi or STP did not affect hCG-induced spermiation although PKC activation significantly decreased hCG-dependent sperm release. These results suggest that PKC inhibits spermiation but also that the inhibition exerted by the kinase could be blocked by hCG. Taken together, these observations could indicate that PKA is involved in the mechanism of the gonadotropin action, mechanism also requiring the activation of a non-pumping Na⁺/K⁺ ATPase pathway.     

Effect of phenylalanine and p-chlorophenylalanine on Na+, K+-ATPase activity in the synaptic plasma membrane from the cerebral cortex of rats

Na⁺, K⁺-ATPase activity was measured in synaptic plasma membrane from cerebral cortex of Wistar rats subjected to experimental phenylketonuria, i.e., chemical hyperphenylalaninemia induced by subcutaneous administration of 5.2 μmol phenylalanine /g body weight (twice a day) plus 0.9 μmol p-chlorophenylalanine /g body weight (once a day). The treatment was performed from the 6^th to the 14^th postpartum day and rats were killed 12 h after the last injection. Synaptic plasma membrane from cerebral cortex was prepared by a discontinuous density sucrose gradient for Na⁺, K⁺-ATPase activity determination. The results showed that the enzyme activity was decreased by 30% in animals subjected to experimental phenylketonuria when compared to control. Thein vitro effects of the drugs on Na⁺, K⁺-ATPase activity were also investigated. Phenylalanine and p-chlorophenylalanine inhibited the enzyme activity and this inhibition was reversed by alanine. In addition, competition between phenylalanine and p-chlorophenylalanine for binding to the enzyme was observed, suggesting a common binding site for these substances. Our results suggest that reduction of Na⁺, K⁺-ATPase activity may be one of the mechanisms related to the brain dysfunction observed in human PKU.     

Expression of pituitary prolactin, growth hormone and somatolactin is modified in response to different stressors (salinity, crowding and food-deprivation) in gilthead sea bream Sparus auratus

Prolactin (PRL), growth hormone (GH) and somatolactin (SL) expression was studied in gilthead sea bream (Sparus auratus) in response to several different stressors (salinity, food deprivation or stocking density). In the first experiment, specimens were acclimated during 100 days at three different environmental salinities: low salinity water (LSW, 6 ppt), brackish water (BW, 12 ppt) and seawater (SW, 38 ppt). Osmoregulatory parameters corresponded to those previously reported for this species under similar osmotic conditions. Pituitary PRL expression increased with decreasing environmental salinity, and was significantly different between SW- and LSW-acclimated fish. Pituitary GH expression was similar between SW- and BW-acclimated fish but decreased in LSW-acclimated specimens. Pituitary SL expression had a “U-shaped” relationship to environmental salinity with the lowest expression in BW-acclimated fish. In a second experiment SW-acclimated specimens were randomly assigned to one of four treatments and maintained for 14 days: (1) fed fish under low density (LD, 4 kg m⁻³); (2) fed fish under high density (HD, 70 kg m⁻³); (3) food deprived fish under LD; and (4) food deprived fish under HD. Plasma glucose and cortisol levels corresponded to those previously reported in S. auratus under similar experimental conditions. Pituitary PRL and SL expression increased in fish maintained under HD and decreased in food deprived fish. In conclusion, an effect of environmental salinity on pituitary PRL and GH expression has been demonstrated. In addition, crowding stress seems to interact with food deprivation in S. auratus and this is reflected by changes in pituitary PRL, GH and SL expression levels.     

The Protective Effect of L-Cysteine and Glutathione on the Adult and Aged Rat Brain (Na<Superscript>+</Superscript>,K<Superscript>+</Superscript>)-ATPase and Mg<Superscript>2+</Superscript>-ATPase Activities in Galactosemia In Vitro

The aim of this study was to evaluate whether the addition of the antioxidants L-cysteine (Cys) or the reduced glutathione (GSH) could reverse the alterations of brain total antioxidant status (TAS) and the modulated activities of the enzymes (Na⁺,K⁺)-ATPase, and Mg²⁺-ATPase in adult or aged rat brain homogenates induced by galactosemia in vitro. Mixture A [mix. A: galactose-1-phosphate (Gal-1-P, 2 mM) plus galactitol (Galtol, 2 mM) plus galactose (Gal, 4 mM) = classical galactosemia] or mixture B [mix. B: Galtol (2 mM) plus Gal (1 mM) = galactokinase deficiency galactosemia] were preincubated in the presence or absence of Cys (0.83 mM) or GSH (0.83 mM) with adult or aged brain homogenates at 37C for 1 h. TAS and the enzyme activities were determined spectrophotometrically. Mix. A or mix. B preincubation with the adult brain resulted in a significant (Na⁺,K⁺)-ATPase inhibition (–30%) and a Mg²⁺-ATPase stimulation (+300% and +33%, respectively), whereas lower modifications of the enzyme activities (p < 0.001) were found in the aged brain. Gal mixtures decreased TAS by 40% (p < 0.001) and by 20% (p < 0.01) in adult and aged samples, respectively. The antioxidants significantly increased TAS resulting in the reversion of (Na⁺,K⁺)-ATPase inhibition and Mg²⁺-ATPase stimulation by mix. B only. The inhibitory effect of Gal and its derivatives on brain (Na⁺,K⁺)-ATPase and their stimulatory effect on Mg²⁺-ATPase are being decreased with age, probably due to the producion of free radicals. Cys and GSH increased TAS resulting in a reversion of the inhibited (Na⁺,K⁺)-ATPase in both models of the in vitro galactosemia and the stimulated Mg²⁺-ATPase in galactokinase deficiency galactosemia only.     

An involvement of vasotocin in oocyte hydration in the catfish Heteropneustes fossilis: A comparison with effects of isotocin and hCG

In the present investigation, in vitro effects of vasotocin (VT) on oocyte (follicular) hydration during germinal vesicle breakdown (GVBD) and ovulation were demonstrated in hCG-primed and non-primed catfish. The data were compared with that of groups incubated with isotocin, and hCG alone or in combination with VT. The priming with hCG resulted in significant increases on percentage GVBD and ovulation, and stimulated follicular hydration, as judged by the increase in diameter, volume, water content, osmolality and Ca²⁺ concentration. However, Na⁺, K⁺ ATPase activity, and concentrations of Na⁺, K⁺ and Mg²⁺ did not alter significantly. The incubations with hCG or VT stimulated all the above parameters. In the non-primed fish, the response of hCG was significantly higher on follicular diameter, volume and osmolality, and that of VT on ovulation. In the primed fish, the VT response was significantly higher on GVBD, ovulation, Na⁺, K⁺ ATPase activity and divalent cation concentrations. The co-incubation with both hCG and VT produced maximal increases in all the parameters with significantly higher effects in the primed fish. The effects of IT on various parameters were relatively low compared to hCG or VT effects. The results indicate that VT may play an important role in oocyte (follicular) hydration, which is consistent with its role in osmoregulation of fish.     

Time course of the GH/IGF axis response to fasting and increased ration in chinook salmon (Oncorhynchus tshawytscha)

Body growth in vertebrates is chiefly regulated by the GH/IGF axis. Pituitary growth hormone (GH) stimulates liver insulin-like growth factor-I (IGF-I) production. During fasting, plasma IGF-I levels decline due to the development of liver GH resistance, while GH levels generally increase. In mammals, decreased insulin during fasting is thought to cause liver GH resistance. However, the sequence of events in the GH/IGF axis response to fasting is not well characterized, especially in non-mammalian vertebrates. We assessed the time course of the GH/IGF axis response to fasting and increased ration in chinook salmon. Fish were placed on Fasting, Increased, or Control rations, and sampled daily for 4 days and at more widely spaced intervals through 29 days. Plasma IGF-I, GH, insulin, and 41 kDa IGF binding protein (putative salmon IGFBP-3), and liver IGF-I gene expression were measured. Control and Increased ration fish did not differ strongly. Plasma IGF-I and 41 kDa IGFBP were significantly lower in Fasted versus Control fish from day 4 onward, and liver IGF-I gene expression was significantly lower from day 6 onward. Liver IGF-I gene expression and plasma IGF-I levels were correlated. Plasma insulin was lower in Fasted fish from day 6 onward. There was a trend toward increased GH in Fasted fish on days 1-2, and GH was significantly increased Fasted fish from day 3 onward. Fasted GH first increased (days 1-3) to a plateau of 10-20 ng/ml (days 4-12) and then increased dramatically (days 15-29), suggesting that the GH response to fasting had three phases. The early increase in GH, followed by the decrease in plasma IGF-I after 4 days, suggests that GH resistance developed within 4 days.     

Changes in Antioxidant Status,Protein Concentration,Acetylcholinesterase, (Na<Superscript>+</Superscript>,K<Superscript>+</Superscript>)-, and Mg<Superscript>2+</Superscript>-ATPase Activities in the Brain of Hyper- and Hypothyroid Adult Rats

It is a common knowledge that metabolic reactions increase in hyperthyroidism and decrease in hypothyroidism. The aim of this work was to investigate how the metabolic reactions could affect the total antioxidant status (TAS), protein concentration (PC) and the activities of acetylcholinesterase (AChE), (Na⁺,K⁺)-ATPase and Mg²⁺-ATPase in the brain of hyper- and hypothyroid adult male rats. Hyperthyroidism was induced in rats by subcutaneous administration of thyroxine (25 g/100 g body weight) once daily for 14 days, while hypothyroidism was induced by oral administration of propylthiouracil (0.05%) for 21 days. TAS, PC, and enzyme activities were evaluated spectrophotometrically in the homogenated brain of each animal. TAS, PC, and Mg²⁺-ATPase activity were found unaffected in hyperthyroidism, while AChE and Na⁺,K⁺-ATPase activities were reduced by 25% (p < 0.01). In contrast, TAS, (Na⁺,K⁺)-ATPase and Mg²⁺-ATPase activities were found to be increased (approx. 23–30%, p < 0.001) in the hypothyroid brain, while AChE activity and PC were shown to be inhibited (approx. 23–30%, p < 0.001). These changes on brain enzyme activities may reflect the different metabolic effects of hyper- and hypothyroidism. Such changes of the enzyme activities may differentially modulate the brain intracellular Mg²⁺, neural excitability, as well as the uptake and release of biogenic amines.     

alpha-MSH acetylation in the pituitary gland of the sea bream (Sparus aurata L.) in response to different backgrounds, confinement and air exposure

MSH is a pituitary hormone derived by post-translational processing from POMC and involved in stress and background adaptation. N-terminal acetylation of MSH to monoacetyl alpha-MSH or diacetyl alpha-MSH increases the bioactivity of the peptide. The aim of this study was to characterize alpha-MSH acetylation in the sea bream (Sparus aurata L.) pituitary gland in response to the stressors air exposure and confinement, as well as in fish adapted for 15 days to a white, gray or black background. Pituitary homogenates were purified by reversed-phase HPLC (RP-HPLC). The alpha-MSH content of fractions was measured by RIA. Immunoreactive RP-HPLC fractions were further analyzed by electrospray mass spectrometry and the peptide sequence determined as SYSMEHFRWGKPV-NH2. In the pituitary gland of sea bream, des-, mono- and diacetyl alpha-MSH were identified. Then plasma alpha-MSH levels were measured in sea bream adapted to different backgrounds. Surprisingly, we found the highest plasma alpha-MSH levels in white-adapted as compared with black-adapted sea bream with intermediate values for gray-adapted fish. This observation is in contrast with results that have been obtained in eel, trout or terrestrial vertebrates. Next, des-, mono- and diacetyl alpha-MSH forms were measured in homogenates of the pituitary gland and in plasma of sea bream exposed to air, to confinement, or to different backgrounds. Monoacetyl alpha-MSH was the predominant form in all control and experimental groups. The lowest content of monoacetyl alpha-MSH relative to des- and diacetyl alpha-MSH was found in white-adapted fish. Levels of des- and diacetyl alpha-MSH forms were similar under all conditions. We observed that monoacetyl alpha-MSH is the most abundant isoform in the pituitary gland after background adaptation, confinement and air exposure, in sea bream. These data indicate that the physiologically most potent isoform of alpha-MSH may vary from species to species.     

Effects of adult-onset streptozotocin-induced diabetes on the rat brain antioxidant status and the activities of acetylcholinesterase, (Na+,K+)- and Mg2+-ATPase: modulation by L-cysteine

Uncontrolled diabetes is known to affect the nervous system. The aim of this study was to investigate the effect of the antioxidant L-cysteine (Cys) on the changes caused by adult-onset streptozotocin (STZ)-induced diabetes on the rat brain total antioxidant status (TAS) and the activities of acetylcholinesterase (AChE), (Na⁺,K⁺)-ATPase and Mg²⁺-ATPase. Thirty-eight male Wistar rats were divided into six groups: C_A (8-week-control), C_B (8-week-control + 1-week-saline-treated), C + Cys (8-week-control + 1-week-Cys-treated), D_A (8-week-diabetic), D_B (8-week-diabetic + 1-week-saline-treated) and D + Cys (8-week-diabetic + 1-week-Cys-treated). All diabetic rats were once treated with an intraperitoneal (i.p.) STZ injection (50 mg/kg body weight) at the beginning of the experiment, while all Cys-treated groups received i.p. injections of Cys 7 mg/kg body weight (daily, for 1-week, during the 9th-week). Whole rat brain parameters were measured spectrophotometrically. In vitro incubation with 0.83 mM of Cys or 10 mM of STZ for 3 h was performed on brain homogenate samples from groups C_B and D_B, in order to study the enzymes’ activities. Diabetic rats exhibited a statistically significant reduction in brain TAS (−28%, D_A vs C_A;−30%, D_B vs C_B) that was reversed after 1-week-Cys-administration into basal levels. Diabetes caused a significant increase in AChE activity (+27%, D_A vs C_A; +15%, D_B vs C_B), that was further enhanced by Cys-administration (+57%, D + Cys vs C_B). The C + Cys group exhibited no significant difference compared to the C_B group in TAS (+2%), but showed a significantly increased AChE activity (+66%, C + Cys vs C_B). Diabetic rats exhibited a significant reduction in the activity of Na⁺,K⁺-ATPase (−36%, D_A vs C_A;−48%, D_B vs C_B) that was not reversed after 1-week Cys administration. However, in vitro incubation with Cys partially reversed the diabetes-induced Na⁺,K⁺-ATPase inhibition. Mg²⁺-ATPase activity was not affected by STZ-induced diabetes, while Cys caused a significant inhibition of the enzyme, both in vivo (−14%, C + Cys vs C_B;−17%, D + Cys vs C_B) and in vitro (−16%, D_B + in vitro Cys vs C_B). In vitro incubation with STZ had no effect on the studied enzymes. The present data revealed a protective role for Cys towards the oxidative effect of diabetes on the adult rat brain. Moreover, an increase in whole brain AChE activity due to diabetes was recorded (not repeatedly established in the literature, since contradictory findings exist), that was further increased by Cys. The inhibition of Na⁺,K⁺-ATPase reflects a possible mechanism through which untreated diabetes could affect neuronal excitability, metabolic energy production and certain systems of neurotransmission. As concerns the use of Cys as a neuroprotective agent against diabetes, our in vitro findings could be indicative of a possible protective role of Cys under different in vivo experimental conditions.     

Curcumin counteracts the aluminium-induced ageing-related alterations in oxidative stress, Na+, K+ ATPase and protein kinase C in adult and old rat brain regions

This study investigated the effect of curcumin on aluminium-induced alterations in ageing-related parameters: lipid peroxidation, superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione-s-transferase (GST), protein kinase C (PKC), Na⁺, K⁺-adenosine triphosphatase (Na⁺, K⁺-ATPase) and acetylcholinesterase (AChE) in the cerebral cortex and hippocampus of the brain of 10- and 24-month-old rats. Measurements taken from aluminium-fed rats were compared with those from rats in which curcumin and aluminium were co-administered. In aluminium-treated rats the levels of lipid peroxidation, PKC and AChE were enhanced while the activities of SOD, GPx, GST and Na⁺, K⁺-ATPase were significantly decreased in both the brain regions of both age-groups. In animals co-administered with curcumin and aluminium, the levels of lipid peroxidation, activities of PKC and AChE were significantly lowered while the activities of SOD, GPx, GST and Na⁺, K⁺-ATPase were significantly enhanced in the two brain regions studied indicating curcumin’s protective effects against aluminium toxicity. Though the magnitudes of curcumin-induced alterations varied in young and old animals, the results of the present study also demonstrated that curcumin exerts a protective effect against aluminium-induced elevation of ageing-related changes by modulating the extent of oxidative stress (by upregulating the activities of antioxidant enzymes) and by regulating the activities of Na⁺, K⁺ ATPase, PKC and AChE. Therefore, it is suggested that curcumin counters aluminium-induced enhancement in ageing-related processes.