Brian A. McKeown

Changes in plasma glucose, amino acid nitrogen and growth hormone during smoltification and seawater adaptation in coho salmon, Oncorhynchus kisutch

Abstract Weekly samples of plasma were obtained from coho salmon held under natural temperature and photoperiod conditions during the parr to smolt transformation. The samples were measured for glucose, amino acid nitrogen (AAN) and growth hormone (GH). In laboratory studies, fish from the same population were injected with salmon GH at different stages of the smoltification process. Plasma glucose and AAN were measured in these fish. Coho smolts were also exposed to sea water and after various time intervals, levels of plasma glucose, AAN and GH were determined. During smoltification the weights of the fish, as well as plasma levels of glucose and AAN, varied with peaks at, or near, full moons. Plasma GH also varied but did not appear to be associated with lunar cycles. Changes in plasma GH levels correlated with changes in fish weight as well as with plasma glucose and AAN. Injections of salmon GH caused elevations in plasma glucose but had no effect on plasma AAN. The values of these blood parameters varied depending on the stage the fish were at during the smoltification process. Exposure to sea water caused increases in circulating levels of GH but had little effect on plasma glucose or AAN. These results are discussed with respect to the role of GH during smoltification and subsequent seawater adaptation.

Immunohistochemical identification of pituitary hormone producing cells in the sockeye salmon (Oncorhynchus nerka, Walbaum)

SummaryAntisera to six mammalian pituitary hormones and a synthetic corticotrophin (Synacthen) were prepared and tested for specificity. The gamma-globulin fractions of the antisera were conjugated with fluorescein isothiocyanate and the labelled antibodies were isolated by column chromatography.Fresh-frozen sections of pituitary glands of adult migratory sockeye salmon (Oncorhynchus nerka) were incubated with the antibody solutions and examined with a fluorescence microscope. Anti-ACTH and anti-Synacthen appeared to be bound by theepsilon cells, whereas anti-prolactin reacted with theeta cells. Anti-GH was bound to the acidophil cells of the proximal pars distalis. Anti-LH reacted with some of the basophil cells of the proximal pars distalis. Antibodies to FSH or TSH failed to react.

In vitro responses of rainbow trout (Oncorhynchus mykiss) somatotrophs to carp growth hormone-releasing factor (GRF) and somatostatin

To study the hypothalamic control of growth hormone (GH) release in lower vertebrates, we employed an in vitro technique using a monolayer cell culture system of rainbow trout pituitary glands. Two newly purified carp brain growth hormone-releasing factors, carp GRF(1-45) and carp GRF(1-29), and synthetic somatostatin-14 (SST-14) were applied to the cultured pituitary cells. The results indicate that: (1) The carp GRFs had a dose-related potency in stimulating growth hormone release. The dose of half maximum effect (ED50) for carp GRF(1-45) was 0.107 nM, and an equal potency for carp GRF(1-29) was 0.388 nM. (2) SST-14 inhibited GH release having a dose-dependent potency with an ED50 of 0.186 nM. (3) Osmotic pressure did not influence SST-14 inhibited GH secretion but did affect spontaneous GH release. (4) The response of cultured cells was not affected by length of incubation period with SST-14 or carp GRF but was affected by cell density.

Further characterization of growth hormone from the chum salmon (Oncorhynchus keta)

This report describes the isolation of growth hormone (GH) from the chum salmon (Oncorhynchus keta) pituitary using gel, affinity, and ion exchange chromatography. Chum GH has an estimated molecular weight of 23,500 and an amino acid composition that is consistent with a vertebrate GH. The differentially charged forms of chum GH which are only apparent under alkaline conditions were separated by ion exchange and compared immunologically and biologically; Peak I, which consists of a single band (Rf = 0.35) under alkaline electrophoresis and Peak II which consists of two bands with Rfs of 0.41 and 0.45. Both forms were found to be immunologically identical by immunodiffusion and to have similar growth promoting properties in intact rainbow trout (Salmo gairdneri). Chum GH was also active in the rat tibia test at a daily dosage of 70 micrograms/animal. The results are discussed in relation to previous studies with chum GH and other fish GHs.

The gill calcium transport cycle in rainbow trout is correlated with plasma levels of bioactive, not immunoreactive, stanniocalcin

Stanniocalcin (STC) is an inhibitor of gill Ca2+ transport that is produced by the corpuscles of Stannius, endocrine glands in bony fish. In young rainbow trout (Oncorhynchus mykiss), there are cyclical changes in the rate of gill Ca2+ transport, with alternating phases of accelerated and reduced uptake every 14 days. Previous studies by our laboratory have established that the responsiveness of young trout to the inhibitory effects of exogenous STC is dependent on this cycle. Trout are highly responsive to STC at peaks of Ca2+ uptake and unresponsive at nadirs, which has led us to suggest that the gill Ca2+ transport cycle may be regulated by a reciprocal cycle in the levels of plasma STC. In this report, we have further characterized the gill Ca2+ transport cycle in salmonids and investigated the role of STC in its regulation. Our results showed that the cycle is synchronous and is likely a characteristic feature in all salmonids but that it varies in amplitude between species. Surprisingly, we observed no correlation between circulating levels of radioimmunoassayable STC and the rate of gill Ca2+ transport in trout. To address this apparent contradiction, trout fry were passively immunized with STC antiserum to determine if there were variable amounts of bioactive STC in the circulation, at times when trout were either more or less sensitive to exogenous STC. We observed that during the times when trout were responsive to STC treatment (i.e., cycle peaks), passive immunization had no effect on the rate of gill Ca2+ transport in fish from the same population, indicating that there were low levels of bioactive STC in the circulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Growth hormone and seawater adaptation in coho salmon, Oncorhynchus kisutch

1. Effects of growth hormone (GH) were examined on short-term aspects of seawater adaptation in coho salmon smolts. 2. Injection of somatostatin (SRIF) immediately prior to seawater entry suppressed plasma GH levels, but did not have any significant effects at 6 or 12 hr on hematocrits, plasma glucose or plasma Na+ levels. 3. Plasma GH levels increased 250% within 36 hr after seawater exposure. 4. Plasma glucose levels, in contrast, were significantly lower in the seawater fish after 36 hr post-exposure. 5. Plasma Na+ levels increased to 190 mEq/1 by 24 hr but subsequently returned to freshwater levels while hematocrits showed no significant changes over the 72 hr of exposure. 6. The significance of these results is discussed in terms of successful seawater adaptation in coho salmon.

Plasma growth hormone levels in Salmo gairdneri: Studies on temperature and the exercise intensity/duration relationship

Abstract 1. 1. Experiments were conducted to investigate the response of plasma growth hormone (GH) to seasonal temperature and temperature acclimation, both individually and in combination with sustained exercise. 2. 2. The effect of varying the intensity and duration of sustained exercise on plasma GH levels was also assessed. 3. 3. Seasonal GH levels were lowest at the cold temperature of the winter months and highest at peak summer temperatures. 4. 4. Exercise at 1.5 bl/sec for a 24 hr period at high temperature caused a greater increase in plasma GH levels than the same exercise at low temperature. 5. 5. For exercise bouts of similar total work, plasma GH response was greater in the high intensity, shorter duration bout than for the low intensity, long duration bout. Exhaustive exercise resulted in no change in GH levels post-exercise.

Immunocytochemical localization of hormone-producing cells within the pancreatic islets of the rainbow trout (Salmo gairdneri)

SummaryA histological study of the pancreatic islets in rainbow trout, Salmo gairdneri, was undertaken in which polypeptide hormone-producing cells were localized, using immunocytochemical staining techniques. Four different celltypes were identified in this manner. These were the insulin, somatostatin, pancreatic polypeptide and glucagon/gastric inhibitory polypeptide (GIP) cells. The glucagon/GIP cell was designated thus as antisera to both hormones crossreacted with a common population of cells. A fifth cell-type, commonly referred to as a clear cell, was also identified although its secretory product is as yet undetermined. These functional cell types were compared to the standard tinctorial properties of pancreatic endocrine cells. The relationships of the various cell types with each other was also observed.

Interaction of Carp Growth Hormone-Releasing Factor and Somatostatin on in vitro Release of Growth Hormone in Rainbow Trout (Oncorhynchus mykiss)

Possible antagonism between somatostatin (SS) and carp growth hormone-releasing factor (GRF) on growth hormone (GH) secretion was examined by radioimmunoassay in a dispersed rainbow trout pituitary cell culture system. SS (3 nM) significantly antagonized carp GRF(1-29; 1 nM, 10 nM)-induced GH secretion. The slope of the dose-response curve for carp GRF(1-29) with SS was statistically different from that of carp GRF(1-29) alone (p less than 0.05) suggesting a noncompetitive antagonism of SS to carp GRF. The carp GRF(1-29) was also indicated to be a noncompetitive antagonist to SS (p = 0.056). Carp GRF(1-29; 100 nM) was unable to restore the inhibitory effect of SS on GH release after pre-exposure of SS (30 nM) to the pituitary cells. We conclude that SS antagonizes carp GRF on GH release at the pituitary level in rainbow trout and this antagonism is noncompetitive. SS has a postantagonism to carp GRF which may implicate some important physiological adaptations in teleosts.

The effect of pH and/or calcium-enriched freshwater on gill Ca2+-ATPase activity and osmotic water inflow in rainbow trout (Salmo gairdneri)

Rainbow trout (Salmo gairdneri) were exposed to pH 5.0-5.1, 6.6 and/or calcium-enriched freshwater for 14 days. Hematocrit, gill Ca2+-ATPase enzyme activities, gill osmotic water inflow, plasma calcium and osmolarity were measured. No significant changes in plasma calcium ion levels were found. The typical increase in hematocrit usually associated with exposure of fish to acidified water was not found in the present study and is discussed. Plasma osmolarity decreased in fish exposed to calcium-enriched freshwater (60 mg Ca2+ X 1(-1) ) in comparison to fish exposed to control freshwater conditions (2 mg Ca2+ X 1(1) ), irrespective of the pH level. Gill Ca2+-ATPase enzyme activities were measured for both low affinity (3 mM Ca2+) and high affinity (100 microM) activity. Exposure of rainbow trout to low pH (pH 5.0-5.1) did not affect the specific activity of Ca2+-ATPase enzyme. However, low affinity Ca2+-ATPase activity in fish exposed to calcium-enriched freshwater did show a significant reduction. The increase in gill osmotic water permeability in fish exposed to calcium-enriched freshwater is interpreted as a result of the increase in osmolarity of the ambient media.