J.G. Eales

Application of thyroid and steroid hormones as anabolic agents in fish culture

A basic premise in any intensive culture system is to maximize growth at minimum cost, with an end product that is of high nutritive value and aesthetically acceptable to the consumer. This review is directed towards evaluating the role for thyroid hormones and steroids in fish culture. Particular attention has been given to the following topics: growth, appetite, food conversion, carcass composition, salt water tolerance, species specificity, deleterious responses and economic implications.

Influence of food deprivation on radioiodothyronine and radioiodide kinetics in yearling brook trout, Salvelinus fontinalis (Mitchill), with a consideration of the extent of l-thyroxine conversion to 3,5,3′-triiodo-l-thyronine

Abstract The effects of starvation on the metabolism of l -thyroxine (T 4 ), 3,5,3′-triiodo- l -thyronine (T 3 ) and the deiodination of T 4 to T 3 were studied in 1-year-old brook trout by comparing the metabolism of 125 I-T 4 ( ∗ T 4 ), 125 I-T 3 ( ∗ T 3 ) and 125 I − ( ∗ I) in trout that were fed [0.56% body wt/day; dry wt of food (g)/wet wt of fish (g)] or starved for 12 days before radioisotope injection into the circulation. Water temperature ranged from 11.4 to 12.5° and the trout were maintained on a 12L:12D photperiod regime. In both fed and starved trout ∗ T 3 was cleared less rapidly than ∗ T 4 from plasma and in contrast to ∗ T 4 did not undergo deiodination. Starvation decreased the plasma clearance of ∗ T 4 , ∗ T 3 and ∗ I, reduced biliary-faecal excretion of ∗ T 4 and ∗ T 3 , depressed the monodeiodination of ∗ T 4 to ∗ T 3 , and lowered the T 4 degradation rate. Within 40 min of feeding previously starved trout, a significant increase in ∗ T 4 deiodination to ∗ T 3 was observed. It is concluded that the inhibition of ∗ T 4 deiodination to ∗ T 3 induced by starvation is rapidly removed following ingestion of food.

The influence of testosterone propionate on thyroid function of immature rainbow trout, Salmo gairdneri Richardson.

Testosterone propionate (TP) was injected intramuscularly (five injections at 4-day intervals) into fed immature rainbow trout at 11–12°; controls were sham injected. TP increased thyroxine (T4) degradation rate, T4 deiodination rate, T4 conversion to 3,5,3′-triiodo-l-thyronine (T3), plasma T3 levels, and thyroid uptake of radioiodide 48–96 hr after radioiodide injection. However, plasma T4 levels responded inconsistently to TP, while thyroid radioiodide uptake measured earlier than 24 hr was suppressed by TP. In conclusion, (i) TP, either directly or indirectly, stimulates many aspects of thyroid function, including conversion of T4 to T3, and (ii) plasma T4 level and thyroidal uptake of radioiodide can be highly misleading indexes of thyroid function.

Saturable 3,5,3′-triiodo-l-thyronine-binding sites in liver nuclei of rainbow trout (Salmo gairdneri Richardson)

Saturable binding of 3,5,3′-triiodo-l-thyronine (T3) was demonstrated in liver nuclei of rainbow trout using an in vivo isotope displacement method; saturable T3 sites were not found in mitochondrial, microsomal, or cytosol fractions. Equilibrium constants ranged from 0.9 to 1.2 × 108 kg liver/mol T3, indicating an affinity comparable to that of mammals. Binding capacities ranged from 0.43 to 0.62 × 10−12 mol T3/g liver. Approximately 50% of the sites were occupied at endogenous T3 levels and a 6- to 11-fold increase in plasma T3 levels was required to achieve saturation. A low food ration followed by starvation did not alter the equilibrium constant but reduced the capacity of the nuclear sites. The sites were intranuclear and represented in a macromolecular fraction extracted with 0.4 N KCl. The macromolecule was identified as a heat-labile protein, probably nonhistone in nature. Further study is necessary to determine the significance of these sites in initiating T3 effects at the cellular level.

Influence of bovine TSH on plasma thyroxine levels and thyroid function in brook trout, Salvelinus fontinalis (mitchill)

Abstract The influence of intramuscularly injected bovine thyroid stimulating hormone (TSH) on the plasma thyroxine (T 4 ) level was studied in 1- and 2-year-old brook trout acclimated at 10–12°. Plasma T 4 was highest betwen 9 and 48 hr after a single TSH injection. The plasma T 4 response to TSH was uninfluenced by nutritional state, anaesthesia, type of injection vehicle, or ACTH and was of similar magnitude at 5, 12, or 20°. The response decreased if daily injections extended beyond 5 days. Large fish were more responsive than small fish of the same age to a TSH dose standardized for body weight. A linear relationship existed between log TSH dose and plasma T 4 of intact trout. The precision index was superior to that for other TSH assays involving fish. Radioiodine indices of thyroid activity (plasma PB 125 I, CR) and bile organic 127 I content were also linearly related to log TSH dose with good precision. Thyroid uptake of 125 I was an ambiguous and less precise assay for TSH. An attempt was made to define a “physiologic” TSH dose to administer to trout.

In vivo determination of thyroxine deiodination rate in rainbow trout, Salmo gairdneri Richardson.

An in vivo method is described for measuring the deiodination rate of l-thyroxine (T4) and 3,5,3′-triiodo-l-thyroxine (T3) in rainbow trout. The method is based on a comparison of plasma radioiodide kinetics following injection of either radioiodide or thyroid hormones phenolically labeled with radioiodine. It relies on a rapid turnover of thyroid hormones and a slow turnover of iodide in trout plasma. In two groups of 1-year-old trout starved for 3 days at 12°, 29.6 and 34.4% of the radioiodine were removed from labeled T4 by deiodination; no deiodination of T3 was detected. Assuming that T4 undergoes phenolic monodeiodination to T3, it was estimated that for the two groups of trout 59 and 69% of circulating T4 were converted to T3 and that rates of T3 production from T4 were 1.5 and 1.9 pmol/hr/100 g body weight.

Use of thyroxine- and triiodothyronine-specific antibodies to study thyroxine kinetics in rainbow trout, Salmo gairdneri

A rapid and discriminating method employing 3,5,3′-triiodo-l-thyronine (T3)- and l-thyroxine (T4)-specific antibodies is described for separating T3 and T4 in body fluids. This technique was used in a compartmental analysis of T4 kinetics in 1-year-old rainbow trout held at 12°C and starved for 3 days prior to 125I-T4 (∗T4) injection into the heart region. The multiexponential curve for ∗T4 loss from plasma with time comprised a “fast” and a “slow” component and was used as the basis for a two-compartment model. The following values, normalized to a body weight of 100 g, were obtained: plasma T4, 0.72 ng/ml; plasma T4 distribution space, 27.0 ml; plasma T4 pool, 19.4 ng; extravascular T4 pool, 103 ng; T4 flow rate from plasma to extravascular pool, 2.93 ng/hr; T4 flow rate from extravascular pool to plasma, 0.78 ng/hr; irreversible T4 loss from the extravascular pool (= T4 degradation rate), 2.15 ng/hr. Both plasma 125I- and plasma 125I-T3 (∗T3) levels increased during the first few hours following injection, suggesting a significant deiodination of ∗T4 to ∗T3.

Creation of chronic physiological elevations of plasma thyroxine in brook trout, Salvelinus fontinalis (mitchill) and other teleosts

Immersion and intraperitoneal injection methods were examined for producing small chronic elevations of plasma thyroxine (T4) in brook trout and other teleosts. Continual immersion of starved yearling brook trout in a solution of 10 μg T4/100 ml water at 12–13°C raised plasma T4 within hours from 1.0 μg/100 ml (or less) to 1–3 μg/100 ml. This increase was sustained during 27 days of treatment. An ambient T4 level of 2 μg/100 ml produced a smaller but distinct increase; 50 μg of T4/100 ml water was considered pharmacological. The influence of ambient T4 on plasma T4 depended on body size and temperature, with possible effects due to feeding state and species. Trout immersed in radioactive T4 (5 μg/100 ml) by 3 hr reached a steady state, with plasma T4 considerably less than ambient T4. Both deiodination and biliary excretion of T4 contributed to the steady state by removing radioactive T4 still entering the fish. A single intraperitoneal T4 injection was found to be impractical owing to rapid T4 turnover. Thus frequent injections would be required which would probably not eliminate surges in plasma T4 due to intraperitoneal injection.

Iodothyronine-glucuronide conjugates in the bile of brook trout, Salvelinus fontinalis (Mitchill) and other freshwater teleosts

Abstract After intraperitoneal injection of l -thyroxine ( ∗ T 4 ) or 3,5,3′-triiodo- l -thyronine ( ∗ T 3 ) labeled with 125 I in 3′ or 5′ positions into starved brook trout held at 12–13°C, approximately 50% of bile radioactivity was identified by thin-layer chromatography as free iodothyronine and approximately 50% as thyroxine-glucuronide conjugates with negligible radioiodide. For ∗ T 4 -injected fish, glucuronide conjugation was almost entirely to ∗ T 4 ; for T 3 -injected fish glucuronide conjugation was mainly to ∗ T 3 , but other derivates may be present. ∗ T 4 and ∗ T 4 -glucuronide in variable proportions were the main radioactive products in bile from 9 other species of freshwater teleosts held under similar conditions and intraperitoneally injected with ∗ T 4 .

Biliary excretion of 3,5,3′-triiodo-l-thyronine-125I by the brook trout, Salvelinus fontinalis (Mitchill) ☆

The percentage of administered radioactivity occurring in several organs and in protein-bound and non-protein-bound serum fractions was measured following intraperitoneal injection of 3,5,3′-triiodo-l-thyronine-125I (T3∗), into brook trout acclimated at 10°C. The sequence of maximum liver uptake of 8.0% within 24 hours, followed by maximum gall bladder uptake including bile of 10.9% at 48 hours and by maximum intestinal uptake including contents of 20.7% at 96 hours suggested a biliary excretion route for T3∗. By descending paper chromatography (butanol:acetic acid:H2O, 4:1:1) most bile radioactivity occurred as two adjacent peaks (Rf 0.2–0.3) while approximately 15% corresponded to free thyronines. Bile 125I-levels were negligible. T3∗ loss from the serum was described by fast (t12 = 6.9 hours) and slow (t12 = 65.4 hours) exponents. T3∗ did not appear to bind as extensively as T4∗ (Eales, 1970) to serum proteins of brook trout, but both deiodination and biliary excretion seemed less pronounced for T3∗ than for T4∗.