Acute handling stress resulted in significant elevation of plasma cortisol and lactate concentrations within 30 min in both fed and food-deprived rainbow trout (Oncorhynchus mykiss), indicating a typical stress response. Plasma glucose levels rose immediately (30 min) poststress in the food-deprived group, while there was a delayed response (2 h) in the fed group. The low liver glycogen content and total glycogen phosphorylase (GPase) and glycogen synthase (GSase) activities in the food-deprived group indicated an overall depression in glycogen metabolism. Acute handling stress maintained liver glycogen stores for up to 4 h in the food-deprived group, but the combined effects of limited substrate and increased energy demand necessitated mobilization of liver glycogen in the food-deprived group, but not in the fed group. This increased glycogen mobilization in the food-deprived group coincided with a secondary elevation in plasma cortisol concentration 4 h poststress. The results indicated that food-deprived rainbow trout were more sensitive to stress of handling and mobilized glycogen stores to meet the energy demand imposed by the stressor. The elevated plasma cortisol levels noted during acute handling stress could play an important role in energy partitioning, metabolically adapting the fish to handling stress.

The objective of the study was to examine the physiological response to stress in a marine species with a sluggish life-style. The sea raven (Hemitripterus americanus), a marine benthic predator, did not produce elevated catecholamine levels when handled for blood removal, which facilitated repeated blood sampling from the same fish without cannulation. However, this species did release catecholamines in response to an acute stress (1 min of air exposure followed by 1 min of chasing), suggesting a high threshold (degree of external stimulation) for catecholamine release in this species. Plasma cortisol concentration increased significantly only after 1 h and remained elevated 4 h post-stress, showing a delayed response compared with salmonids. Plasma glucose concentration increased significantly at 0.5 h post-stress and remained elevated even at 24 h, while lactate levels dropped between 4 and 24 h post-stress. The delayed cortisol increase may not be due to altered plasma clearance, as no change in the plasma disappearance or tissue uptake of cortisol-derived radioactivity occurred with confinement stress in this species. Also, confinement stress did not alter the plasma disappearance or tissue uptake of radioactivity derived from glucose, indicating a higher production of glucose during stress. Food deprivation significantly increased the plasma disappearance and tissue uptake of both cortisol- and glucose-derived radioactivity in the sea raven. These results indicate that the hormonal response to stress in the sea raven is different from that of salmonids. This altered response may be an adaptation to prevent excess energy mobilization in a species with an inactive life-style and low metabolic activity.

Simultaneous infusion of [6-3H]glucose and [U-14C]lactate was used to calculate the turnover rate of glucose, the irreversible replacement rate of lactate, and the rates of the exchange of carbon atoms between glucose and lactate in free-swimming American eels (Anguilla rostrata) fed or food deprived for 6, 15, and 36 (maturing) mo. The mean turnover rate of glucose in fed animals averaged 1.0 mg X min-1 X 100 g-1, while the lactate irreversible replacement rate was approximately 4.0 micrograms X min-1 X 100 g-1. The conversion of 35% of lactate carbon to glucose implied a substantial Cori cycle activity, but this amounted to less than 1% of total glucose production. Food deprivation for 6 mo altered few kinetic patterns, except for an increased lactate irreversible replacement rate and a minor increase in gluconeogenesis from lactate. After a 15-mo fast, glucose turnover decreased to 0.09 +/- 0.02 mg X min-1 X 100 g-1. Plasma lactate concentrations and production rates continuously increased during the experiment. Maturing eels that had been food deprived for 36 mo maintained glucose and lactate concentrations and kinetics similar to values in animals food deprived for only 6 mo. This study stresses the importance of carbohydrate in the metabolism of this species under fed and food-deprived conditions and further supports the tolerance of Anguillid species to food deprivation.

We report for the first time that -naphthoflavone (BNF) abolishes ACTH stimulation of cortisol production in rainbow trout (Oncorhynchus mykiss). There was significantly higher hepatic cytochrome P450 content and ethoxyresorufin O-de-ethylase and uridine-5 -diphosphoglucuronic acid transferase activities in BNFtreated fish than in sham-treated controls. BNF did not significantly effect either plasma turnover or tissue distribution of [ 3 H]cortisol-derived radioactivity. Hepatic membrane fluidity and hepatocyte capacity for cortisol uptake were not altered by BNF as compared with the sham-treated fish. These results taken together suggest that BNF does not affect cortisol-clearance mechanisms in trout. A 3 min handling disturbance period elicited a plasma cortisol response in the sham-treated fish; however, the response in the BNF-treated fish was muted and significantly lower than in the sham fish. This in vivo response corroborates the lack of interrenal sensitivity to ACTH in vitro in the BNF-treated fish, suggesting that BNF affects the ACTH pathway in trout. Our results suggest the possibility that cytochrome P450-inducing compounds may affect cortisol dynamics by decreasing interrenal responsiveness to ACTH stimulation in fish, thereby impairing the physiological responses that are necessary for the animal to cope with the stressor.

No abstract

We have characterized the specific binding of glucagon in hepatocytes isolated from two teleost species, the American eel (Anguilla rostrata) and the brown bullhead (Ictalurus nebulosus). Specific glucagon binding was 9.3 and 10.7% in bullhead and eel hepatocytes respectively, after a 2-h incubation at 12 degrees C. Curvilinear Scatchard plots suggest the presence of two classes of binding sites with apparent dissociation constants (Kd) of 1.97 nM (high affinity) and 17.3 nM (low affinity) for bullhead and 2.68 and 22.9 nM for eel cells. The number of high-affinity binding sites per cell was significantly higher in the eel (10,413) than in the bullhead (3811). The number of high-affinity insulin-binding sites was approximately two times higher than that for glucagon in bullheads and the opposite in the eel hepatocytes. In competition experiments, insulin did not displace 125I-labelled glucagon binding in the hepatocytes of either species, while glucagon-like peptide-1(7-37) (GLP-1) displaced glucagon but only at high concentrations, suggesting separate glucagon- and GLP-1-binding sites. The rate of dissociation of hepatocyte-bound 125I-labelled glucagon was similar for both species. Preincubation of hepatocytes in 100 nM glucagon decreased the number of high-affinity glucagon-binding sites by approximately 55% in both species, while the Kd values remained unchanged. Glucagon bound to the cell surface is internalized by fish hepatocytes. These properties indicate that the glucagon binding to hepatocytes of these two teleost species is similar to that reported for mammalian hepatocytes.

No abstract

The effects of acclimation temperature and acute temperature changes on the intracellular pH (pHi) of hepatocytes isolated from the American eel, Anguilla rostrata, were studied by the measurement of the distribution ratio of dimethyloxizolidinedione (DMO). Varying the concentration of DMO (10(-7) to 10(-4) M) did not affect estimates of pHi, indicating that DMO acts as an ideal pHi probe in eel hepatocytes. In vitro studies yielded values of liver cell pHi identical to those previously measured in vivo (in vitro pHi = 7.556 +/- 0.010; in vivo pHi = 7.570 +/- 0.049 at 20 degrees C); hepatocyte pHi varied inversely with acclimation temperature (5-20 degrees C) in a manner consistent with alphastat regulation (delta pH/delta T = -0.0182 +/- 0.021). During acute temperature increases (5-20 degrees C) and decreases (20-5 degrees C) hepatocytes regulated pHi to the appropriate (acclimated) value within 30-45 min posttransfer under conditions of constant medium pH (pHe). The effects of medium pH were also studied, and although patterns of pHi regulation differed between 5 and 20 degrees C cells, a pHi difference consistent with alphastat regulation was maintained between 5 and 20 degrees C cells over the pHe range 7.8-8.3.