Glycogenolytic action of glucagon-family peptides and epinephrine on catfish hepatocytes

Glycogenolytic effects of salmon and mammalian glucagons, salmon glucagon-like peptide (GLP) and epinephrine were studied on liver cells isolated from catfish (Ictalurus melas). In spring and summer, salmo-glucagon (3×10^–10 to 3×10^–8 M) was more effective than its mammalian counterpart in the stimulation of glucose release and cAMP synthesis in hepatocytes. GLP was less potent as compared to both glucagons. -amylase activity was not affected by the treatment with either glucagon-family peptides or epinephrine.The comparison of the glycogenolytic effects of salmon glucagon to those of epinephrine reveals a greater potency of the latter hormone in the stimulation of cAMP synthesis, glycogen-phosphorylase activity and glucose release. Glycogen content in the liver cells was equally depleted after treatment with both of the two hormones.     

Metabolic actions of glucagon-family hormones in liver

This review addresses direct and indirect metabolic actions of hormones co-encoded in the preproglucagon gene of fishes. Emphasis is placed on a critical analysis of the effects of glucagon and glucagon-like peptide (GLP) and the current knowledge of the respective modes of action is reviewed. In mammals GLPs exert no direct metabolic actions. In fish liver, GLP and glucagon act on similar targets of intermediary metabolism by enhancing flux through glycogenolysis, lipolysis and gluconeogenesis. Increases in substrate oxidation are not uniform. Hormonal activation of glycogen phosphorylase and triglyceride lipase and inhibition of pyruvate kinase are implicated in these actions. Hormone-dependent hyperglycemia, depletion of hepatic glycogen and increases in free fatty acids are noticeablein vivo. Glucagon also activates hepatic amino acid uptake and ammonia excretion. Glucagon actions are accompanied by large increases in hepatic cAMP and increased phosphorylation of pyruvate kinase. Metabolic effects measured after GLP administration are associated with minor, if any, increases in cAMP and effects on pyruvate kinase are variable. We hypothesize that different hepatic receptors with differing modes of intracellular message transduction are involved in glucagon and GLP actions while targetting identical metabolic routes. Responses of different species of fish cover a wide spectrum, and variation of response with the circannual cycle of experimental animals makes comparisons of results, even within one species, difficult.     

Fish hepatocyte glycogen phosphorylase – a sensitive indicator for hormonal modulation

The absence of a reproducible method for the assay of glycogen phosphorylase (GPase) in isolated fish hepatocytes has made the interpretation of hormone-induced glycogenolysis data difficult. This study presents such an assay and demonstrates its sensitivity to hormonal activation. The enzyme is assayed in the reverse direction using glucose 1-phosphate (G1-P) and glycogen as substrates and uses standard methods for the quantification of the liberated inorganic phosphate. The assay is highly reproducible, sensitive, and provides an excellent means to follow small and rapid changes in enzyme phosphorylation status following the addition of hormones. We show for hepatocytes isolated from rockfish (Sebastes caurinus) and brown bullhead (Ameiurus (Ictalurus) nebulosus) that small concentrations of three model hormones, namely epinephrine (catfish), norepinephrine, and prostaglandin E₂ (rockfish), lead to the rapid, concentration and time-dependent conversion of existing GPase into the active GPase a form. Some of the enzyme seems to be impervious to hormonal activation, as the highest %GPase a never reaches 100%. We provide evidence that changes in enzyme phosphorylation status provide a better short-term insight into hormone-dependent activation than estimates of glucose or some other end products, that usually must accumulate for long periods before detection is possible. Our data also show that GPase in freshly isolated hepatocytes is already in an activated state and cells should be given a period of rest for several hours before hormonal studies involving glycogen breakdown or the cAMP cascade are initiated.     

Cortisol and liver metabolism of immature American eels,Anguilla rostrata (LeSueur)

Plasma and liver composition, liver enzyme activities, and metabolite flux in isolated hepatocytes have been studied in immature American eels,Anguilla rostrata, injected daily IP with saline or cortisol (0.35 mg/kg eel). Plasma cortisol values were significantly increased above saline controls in those eels receiving cortisol at 3h and 6h following the final (tenth) injection. On day 6 and 10 of injection plasma cortisol levels were significantly below saline controls 24h following cortisol injection. Plasma glucose values were significantly depressed in the cortisol-injected eels at both 6 and 24h following the final (tenth) injection. At the 24h sampling time, plasma protein had significantly increased, but there was no change in either plasma amino acid or fatty acid levels. An increased hepatosomatic index was attributed to a major increase in total lipids, as both protein and glycogen contents were decreased. Of the liver enzymes assayed, significant activity changes occurred only for lactate dehydrogenase (decreased), mitochondrial citrate synthase (increased) and phosphoenolpyruvate carboxykinase (increased) 24h following the final (tenth) cortisol injection. Although enzyme activity changes implied increased liver gluconeogenesis, the absolute rate of lactate, alanine, and aspartate incorporation into glucose declined in viable hepatocytes isolated from cortisol-injected eels compared to the saline controls. Relative changes in metabolite flux did support a preferential increase in gluconeogenesis from amino acids. These results are consistent with the increase in amino acid gluconeogenesis as a result of cortisol administration implied in previous studies, but failed to show a definitive cortisol effect on this pathway in the eel liver. It is suggested that other hormones (e.g. thyroxine, catecholamines, glucagon) may interact in a complex way with cortisolin vivo to bring about the biochemical changes observed in this study. The rapid clearance of exogenously injected cortisol noted in this study makes causal relationships between the injected hormone and any observed metabolic effect in the intact animal difficult.     

Effects of cortisol on hepatic carbohydrate metabolism and responsiveness to hormones in the sea raven,Hemitripterus americanus

The sea raven, Hemitripterus americanus, is a sit-and-wait, low metabolic rate, marine teleost. The objective of this study was to determine i) whether cortisol implantation (50 mg. kg^-1) for 7 days altered hepatocyte metabolism, and hepatocyte responsiveness to epinephrine, glucagon and insulin, and ii) whether 8 weeks of food-deprivation modified the above response. Cortisol implantation significantly increased hepatocyte total glucose production and oxidation from alanine compared to the sham group. There was no cortisol effect on glycogen breakdown, suggesting that the activation of other pathways, including gluconeogenesis, are required to account for the increased glucose production. Epinephrine-mediated (10^-5M) glycogen breakdown and insulin-mediated (10^-8M) total glucose production were enhanced in hepatocytes of cortisol implanted sea ravens, but there were no change in any glucagon (10^-7M) effects. The enhanced glycogen breakdown in the absence of similar increases in total glucose production with epinephrine indicates mobilization of carbohydrate reserves for endogenous use by the liver. Food-deprivation for 8 weeks significantly decreased condition factor, plasma cortisol concentration and liver glycogen content in the sea raven, but had no effect on plasma glucose concentration. Hepatocyte total glucose production and flux rates from alanine increased significantly with food-deprivation. Moreover, food-deprivation increased responsiveness of total hepatocyte glucose production to the actions of glucagon and insulin, but not to epinephrine; none of these effects were modified by cortisol implantation. Our results indicate that cortisol in the sea raven exerts both a direct and an indirect or permissive effect on hepatocyte metabolism by modifying hepatocyte responsiveness to epinephrine and insulin stimulation. Cortisol implantation did not modify the effects of glucagon or food-deprivation in this species.     

Are cell redox or lactate dehydrogenase kinetics responsible for the absence of gluconeogenesis from lactate in sea raven,hepatocytes?

Previous studies have reported very low rates of gluconeogenesis from lactate in sea raven (Hemitripterus americanus) hepatocytes compared to other teleosts studied. This study examines whether hepatic cell redox or lactate dehydrogenase (LDH) characteristics may explain this observation. Sea raven hepatic optimal LDH activities (pyruvate reductase direction) were more than 40 times less compared with rainbow trout liver values (40 vs 1914 μmol·min⁻¹·g⁻¹ protein). The Km(lactate) was 9.24 and 0.86 mM for sea raven and trout hepatic LDH, but the Km(pyruvate) was similar between the two species (0.11 and 0.21 mM, respectively). These results suggested that sea raven liver LDH did not favour lactate use and was more indicative of the mammalian M-isozyme. Gel electrophoresis showed a predominant intermediate isozyme, with a small amount of the M-type LDH. Phosphoenolpyruvate carboxykinase (PEPCK) was localized to the mitochondrial compartment, while there was no apparent mitochondrial glutamate-oxaloacetate transaminase (GOT) activity. No in vitro lactate flux to glucose was found in untreated, 10 mM ethanol-treated, or 3 mM NH₄Cl-treated sea raven hepatocytes, although CO₂ production from lactate was decreased by ethanol and increased by NH₄Cl. These results provide evidence that cell redox does not limit gluconeogenesis from lactate, while low activities and the kinetic characteristics of LDH may partially explain the low lactate gluconeogenesis reported in sea raven hepatocytes. To whom correspondence should be addressed at University of Ottawa.     

Effects of epinephrine exposure on contractile performance of compact and spongy myocardium from rainbow trout (<Emphasis Type="Italic">Oncorhynchus mykiss</Emphasis>) during hypoxia

Hypoxia results in elevated circulating epinephrine for many fish species, and this is likely important for maintaining cardiac function. The aims of this study were to assess how hypoxia impacts contractile responses of ventricular compact and spongy myocardium from rainbow trout (Oncorhynchus mykiss) and to assess how and if epinephrine may protect myocardial performance from a depressive effect of hypoxia. Work output and maximum contraction rate of isolated preparations of spongy and compact ventricular myocardium from rainbow trout were measured. Tissues were exposed to the blood PO₂ that they experience in vivo during environmental normoxia and hypoxia and also to low (5 nM) and high (500 nM) levels of epinephrine in 100% air saturation (PO₂ 20.2 kPa) and during hypoxia (PO₂ 2 kPa, 10% air saturation). It was hypothesized that hypoxia would result in a decrease in work output and maximum contraction rate in both tissue types, but that epinephrine exposure would mitigate the effect. Hypoxia resulted in a decline in net work output of both tissue types, but a decline in maximum contraction rate of only compact myocardium. Epinephrine restored the maximum contraction rate of compact myocardium in hypoxia, appeared to slightly enhance work output of only compact myocardium in air saturation but surprisingly not during hypoxia, and restored net work of hypoxic spongy myocardium toward normoxic levels. These results indicate hypoxia has a similar depressive effect on both layers of ventricular myocardium, but that high epinephrine may be important for maintaining inotropy in spongy myocardium and chronotropy in compact myocardium during hypoxia.     

Response of enzyme activities and metabolic intermediate concentrations to epinephrine administration in hepatopancreas and muscle of carp

SUMMARY: Common carp Cyprinus carpio were administered intraperitoneally with 10 μmol epinephrine per 100 g bodyweight, and enzyme activities and metabolic intermediate concentrations were determined in the hepatopancreas and muscle. Glycogen phosphorylase a (GPase a) activity together with cyclic AMP (cAMP) concentration was increased, and glycogen content was decreased in the hepatopancreas and muscle at 2 h after the administration. The epinephrine administration also increased hepatopancreatic glucose-6-phosphatase and fructose-1,6-biphosphatase activities as well as serum glucose, lactate, and free amino acid concentrations. Furthermore, its administration increased phosphofructokinase activity together with lactate, fructose-6-phosphate, adenosine-5'-monophosphate, and adenosine-5'-diphosphate concentrations and decreased citrate concentration in the muscle. Thereafter, almost all parameter concentrations in the hepatopancreas and muscle recovered to the pre-administered levels during 24 h after the administration. These results suggest that epinephrine administration enhanced glycogenolysis and gluconeogenesis in the hepatopancreas, and released glucose into the bloodstream to supply it to the muscle. The blood glucose together with muscle glycogen seems to be metabolized through enhanced glycolysis in the muscle.     

Response of enzyme activities and metabolic intermediate concentrations to a long burst of exercise and following resting in muscle and the hepatopancreas of carp

ABSTRACT: The responses of enzyme activities and metabolic intermediate concentrations to a long burst of exercise and following resting were examined in muscle and the hepatopancreas of carp Cyprinus carpio . A 15 min burst of exercise made the fish so exhausted that they could not swim any more. In muscle, the exercise decreased glycogen content significantly and increased lactate content significantly, resulting in a lowered pH. Furthermore, the burst of exercise decreased phosphofructokinase (PFK) activity significantly, although it changed adenosine-5'-monophosphate, adenosine-5'-diphosphate, adenosine-5'-triphosphate, fructose-6-phosphate and citrate concentrations within ranges that could activate PFK. In the hepatopancreas, the exercise increased glucose-6-phosphatase and fructose-1,6-bis-phosphatase activities, and glucose and lactate concentrations in the serum, and it decreased glycogen content. Even at 3 h resting after the burst of exercise, the fish had not completely restored many parameters. These results suggest that although the fish tried to enhance in vivo muscular glycolysis through the activation of PFK by changes in metabolic intermediate concentrations, glycolysis seemed to decrease markedly through the inhibition of PFK as a result of the lowered pH. However, the hepatopancreas made a contribution to muscular exercise through glucose supplementation by enhanced gluconeogenesis and glycogenolysis, indicating the presence of an interdependence of carbohydrate metabolism between muscle and the hepatopancreas in the fish.     

Insulin stimulates hepatic lipogenesis in rainbow trout,Oncorhynchus mykiss

The effects of the pancreatic hormones, insulin and glucagon, on rates of lipid biosynthesis in liver removed from rainbow trout, Oncorhynchus mykiss, were evaluated in vitro. Livers were removed from animals fasted for 30–36h, cut into ca. 1 mm³ pieces, and incubated in the presence of various concentrations of salmon insulin (sINS), bovine insulin (bINS), or a combination of BINS and bovine/porcine glucagon (GLU). Lipid synthesis was evaluated by total lipid concentration, ³H₂O incorporation into total lipid, and by fatty acid synthetase activity. Both mammalian and sINS tended to increase tissue total lipid concentration in hepatic tissue incubated for 5h. Insulin also stimulated ³H₂O incorporation into total lipid in a dose-dependent manner. Bovine INS (2 × 10^?6 M) stimulated de novo synthesis nearly 6-fold over control rates; sINS (2 × 10^?6 M) stimulated label incorporation more than 7-fold over control rates. Glucagon inhibited INS-stimulated ³H₂O incorporation; whereas, GLU alone had no effect on lipid synthesis in liver pieces incubated 5h. Lipid class analysis indicated that bINS significantly stimulated ³H₂O incorporation into phospholipids, fatty acids, and triacylglycerols. The greatest accumulation of label was in the triacylglycerol fraction, where incorporation was stimulated 17-fold over control levels. Hepatic enzymatic analysis indicated that bINS also significantly stimulated lipogenic enzyme activity 9-fold above control levels. These results indicate that INS is an important regulator of lipid synthesis in the liver of trout.     

In vitro effect of pituitary,interrenal and gonadal hormones on vitellogenin synthesis in primary hepatocyte cultures of Chalcalburnus tarichi

Vitellogenin (Vtg) is an important precursor yolk protein in egg‐laying vertebrates, including fish. The 17β‐oestradiol (E2) plays a crucial role in the Vtg synthesis; moreover, certain hormones can stimulate Vtg synthesis. We investigated the possible role of E2, carp recombinant growth hormone (crGH), insulin (Ins), progesterone (P4) and 11‐deoxycortisol (11‐DOC) hormones in Vtg synthesis on primary juvenile Chalcalburnus tarichi hepatocyte culture. The amount of Vtg in the medium was measured at 2‐day intervals using enzyme‐linked immunosorbent assay (ELISA). The hepatocytes were maintained in culture for more than 2 weeks without the addition of serum components. Vitellogenin localization was visualized with the immunofluorescence method in E2‐supplemented hepatocytes. Among hormones applied to the culture, only E2 had an influence on Vtg synthesis in a time‐dependent manner, while crGH, Ins, P4 and 11‐DOC had no effect. However, in hepatocytes stimulated with E2 in combination with P4, a lower Vtg production was seen compared with Vtg produced when hepatocytes were stimulated with E2 alone. P4 proved to have potentiating effects on co‐treatment with E2‐induced Vtg production. As a result, E2 and P4 are the most important hormones for Vtg synthesis in juvenile C. tarichi hepatocyte culture.     

Carbohydrate metabolism and hepatic zonation in the Atlantic hagfish,Myxine glutinosa liver: effects of hormones

Viable Atlantic hagfish (Myxine glutinosa) hepatocytes were isolated from combined or separated large and small lobes and carbohydrate metabolism was studied. Cells had low levels of glycogen (16–30 μmol·g⁻¹), and low rates of total glucose production (TGP; 0–480 nmol·h⁻¹·g⁻¹ cells). Lactate flux to glucose (5.5 nmol·h⁻¹·g⁻¹) and CO₂ (76 nmol·h⁻¹·g⁻¹) was lower than reported values for teleosts, with a low percentage (30%) of the lactate carbon reaching glucose. Insulin significantly increased total glucose production and gluconeogenesis and decreased 6-phosphofructo 1-kinase (PFK-1) activities and glucose oxidation, while glucagon was without effect on any parameter studied. Forskolin significantly increased TGP. Epinephrine (Epi), norepinephrine (NEpi), isoproterenol (Iso), and phenylephrine (Phe) all decreased CO₂ production from lactate; propanolol blocked the effects of Epi, NEpi, and Iso. The large lobe, accounting for 65% of total liver mass, had a higher glycogen content and higher CO₂ production from lactate compared to the small lobe. Furthermore, enzyme activities in the large lobe were greater than in the small lobe, with the exception of glycogen phosphorylase (GPase) which exhibited smaller %a values in the large lobe. These data indicate the presence of a hormonally-responsive carbohydrate metabolism in hagfish hepatocytes, which is qualitatively and quantitatively different between the two liver lobes.     

Plasma insulin,glucagon, glucagon-like peptide and glucose levels in response to feeding,starvation and life long restricted feed ration in salmonids

Plasma insulin, glucagon, glucagon-like peptide (GLP) and glucose were measured in samples taken from rainbow trout,Oncorhynchus mykiss. (1.5 years of age) before feeding and at selected times up to 9 days after feeding. The feed contained 21.7% carbohydrate (65% digestibe) in the dry matter. The fish responded to feeding with an elevated plasma insulin level (p<0.005) 0.5 h post-feeding, which may account for the unchanged plasma glucose levels. Twentyfour hours after feeding, plasma insulin level had returned to prefeeding levels, while 4–9 days after feeding, a significant reduction compared to pre-feeding levels was observed (p<0.001). During this period plasma glucose levels remained unchanged. The corresponding plasma glucagon or GLP levels showed no significant elevation in response to starvation, the plasma GLP concentration was even significantly reduced on days 4–9 post-feeding (p<0.01–0.001).Atlantic salmon,Salmo salar, (3.5 years of age) fed a calculated satiation ration (RL=100) throughout their lifetime had, in addition to a higher body weight, significantly higher plasma insulin (p<0.005) glucagon (p<0.0001) and GLP levels (p<0.0001) than fish fed half the satiation ration. The plasma glucose levels were, however, not significant different between the groups.     

Insights into Insulin and Glucagon Responses in Fish

The focus of this review is the response of insulin and glucagon to various experimental and physiological conditions in fish. Circulating levels of insulin and glucagon have been analyzed in several fish species, principally salmonids. It is generally accepted that, in fish, the insulin secretion in response to amino acids is stronger than to glucose, although this information has been obtained mainly from carnivorous species. Studies of other species demonstrate that the alimentary pattern affect hormonal secretion and should be taken into account when the effects of secretagogues are analyzed. There are few studies of glucagon secretion, even in salmonids, although it is known that amino acids also stimulate its release. The level of this response and its relationship to insulin secretion depends upon the species of fish. We will discuss the significance of these observations, while also considering other important factors, including the influence of neuropeptides and interactions with other pancreatic and gastro-intestinal hormones. Finally, seasonal, temperature and reproductive stage effects will be discussed, all of which should be taken into account when attempting to understand the role of pancreatic hormones in fish. This short review will not cover the action of these hormones on their target tissues.     

Gelatinized wheat in diets for Atlantic salmon, Salmo salar, L. A full-scale production experiment

Three levels of gelatinized wheat, 70, 140 and 210 g kg^?1, were included in diets for Atlantic salmon, Salmo salar, L., of mean initial weight around 270 g. The experiment was run for 10.5 months and there was an eight- to nine-fold increase in fish weight. No significant differences in weight increase were found between fish. No negative physiological effects, such as hyperglycaemia or excessive hepatic glycogen accumulation were observed. Concentrations of pancreatic hormones were not affected by diet, but positive correlations were found between glucagon-like peptide (GLP) and glucagon, and between insulin and glucagon. Gelatinized wheat in diets for Atlantic salmon did not seem to affect organoleptic traits of the fish flesh in any undesired way.     

Oxidative metabolism in a teleost,Anabas testudineus Bloch: effect of testosterone and estradiol-17β on hepatic enzyme activities

Testosterone (T) administration to maleAnabas testudineus significantly stimulated the activities of cytochrome oxidase, -glycerophosphate dehydrogenase (-GPDH) and Mg²⁺ adenosine triphosphatase (Mg²⁺ ATPase) and inhibited lactate dehydrogenase (LDH), cytosolic and mitochondrial malate dehydrogenases (MDH). The activities of succinate dehydrogenase (SDH), glucose-6-phosphate dehydrogenase (G-6-PDH) and catalase were unaffected by testosterone treatment. Administration of estradiol-17 (E2) in female fish, significantly stimulated cytochrome oxidase activity, inhibited Mg²⁺ ATPase, SDH, catalase and cytosolic and mitochondrial MDH activity, and was without effect on other enzymes studied.The simultaneous injections of actinomycin D or chloramphenicol and T or E2 prevented the hormonal influence on hepatic enzyme activities. The present study demonstrates that inA. testudineus sex steroids influence hepatic oxidative metabolism by a mechanism sensitive to the action of inhibitors of protein synthesis.     

Ammonia and urea excretion in the tidepool sculpin (Oligocottus maculosus): sites of excretion,effects of reduced salinity and mechanisms of urea transport

Tidepool sculpins live in a variable environment where water temperature, salinity, gas tensions, and pH can change considerably with the daily tide cycle. Tidepool sculpins are primarily ammoniotelic, with 8–17% of nitrogen wastes excreted as urea. The majority of net ammonia (J^net _amm; 85%) and urea (J^net _urea; 74%) excretion occurred across the gill, with the remainder excreted across the skin, the kidney, and/or gut. Acute (2h) exposure to 50% seawater significantly increased J^net _urea (2.8-fold), but reduced J^net _amm (3.5-fold). In fish exposed to 50% seawater for 1 week, J^net _urea returned to control values, but J^net _amm remained slightly depressed. Unidirectional urea influx (Jⁱⁿ _urea) and efflux (J^out _urea) were measured using¹⁴C-urea to determine if urea was excreted across the gills by simple diffusion or by a carrier-mediated mechanism. Jⁱⁿ _urea increased in a linear manner with increasing urea water levels (0–11 mmol N l^–1), while J^out _urea was independent of external urea concentrations. As well, J^net _urea and J^{out inurea} were not significantly different from one another, indicating the absence of back transport. Urea analogs and transport inhibitors added to the water did not have any consistent effect on unidirectional urea flux. These results demonstrate that ammonia and urea excretion rates and sites of excretion in tidepool sculpins are very similar to those found in other marine and freshwater teleosts. Urea and ammonia may play a role in osmoregulation as excretion rates and tissue levels were influenced by changes in water salinity. Finally, we found no evidence for a specific urea carrier; branchial urea excretion is likely dependent on simple diffusion.     

Direct actions of serotonin on gonadotropin-II and growth hormone release from goldfish pituitary cells: interactions with gonadotropin-releasing hormone and dopamine and further evaluation of serotonin receptor specificity

In this study, the direct actions of serotonin (5HT) on gonadotropin (GTH)-II and growth hormone (GH) release in the goldfish were tested at the pituitary cell level. 5HT (10 nM - 10 µM) stimulated GTH-II but inhibited GH release from perifused goldfish pituitary cells in a dose-dependent manner. The minimal effective dose of 5HT tested to suppress basal GH secretion (10 nM) was 10-fold lower than that to stimulate GTH-II release (100 nM). The GTH-II releasing effect of 5HT was abolished by repeated 5HT treatment (10 µM) whereas the corresponding inhibition on GH release was unaffected. These results suggest that 5HT receptors on goldfish gonadotrophs and somatotrophs exhibit intrinsic differences in terms of sensitivity to stimulation and resistance to desensitization. Salmon GTH-releasing hormone (sGnRH, 100 nM) stimulated GTH-II and GH release from goldfish pituitary cells. The GTH-II releasing action of sGnRH was unaffected by simultaneous treatment of 5HT (1 µM). However, the corresponding GH response to sGnRH (100 nM) was inhibited. In the goldfish, dopamine is known to stimulate GH release through activation of pituitary D1 receptors. In the present study, the GH-releasing action of dopamine (1 µM) and the D1 agonist SKF38393 (1 µM) was significantly reduced by 5HT (1 µM). To examine the receptor specificity of 5HT action, the effects of 5HT1 and 5HT2 analogs on GTH-II and GH release were tested in goldfish pituitary cells. The 5HT1 agonist 8OH DPAT (0.1 and 1µM) and 5HT2 agonist methyl 5HT (0.1 1µM) mimicked the GTH-II releasing effect of 5HT. The 5HT1 agonist 8OH DPAT (0.1 and 1µM) also stimulated GH release but the 5HT2 agonist methyl 5HT (0.1 and 1µM) was inhibitory to basal GH secretion. In addition, 5HT (1µM) -stimulated GTH-II release was abolished by the 5HT1 antagonist methiothepin (10µM) and 5HT2 antagonist mianserin (10µM). Similarly, the inhibitory action of 5HT (1µM) on basal GH release was blocked by the 5HT2 antagonist mianserin (10µM). The 5HT1 antagonist methiothepin (10µM) was not effective in this regard. These results, taken together, indicate that 5HT exerts its regulatory actions on GTH-II and GH release in the goldfish directly at the pituitary cell level, probably through interactions with other regulators including sGnRH and dopamine. The GTH-II releasing action of 5HT is mediated through 5HT2 and possibly 5HT1 receptors. The inhibition of 5HT on basal GH release is mediated through 5HT2 receptors only. Apparently, 5HT1 receptors are not involved in this inhibitory action. In this study, a paradoxical stimulatory component of 5HT on GH release by activating 5HT1 receptors is also implicated.