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Notes: The present study investigated, for the first time, a non-invasive technique for monitoring cortisol levels in fishes that had been implanted with electronic transmitters. This technique involved the measurement of cortisol released into the water by individual fish. Carp Cyprinus carpio and roach Rutilus rutilus that had been tagged with miniature dummy acoustic transmitters responded to the surgical implantation of the tags with an immediate (1 to 4 h) increase in cortisol concentrations in the water. These amounts of cortisol were higher than in ‘handled only’ and ‘handled and anaesthetized’ groups. Estimated cortisol release rates for both species correlated well with the direct measurements in the water. In both species, water cortisol concentration and cortisol release rates returned to pre-stress levels within 12 h of tag-insertion, and remained low for the remainder of the experiment. The calculation of cortisol release rates highlighted a difference of a factor of about four between peak release rates in carp and roach. There was no evident long-term cortisol stress response to the presence of a tag in the body cavity in either species.

Notes: Individual rainbow trout Oncorhynchus mykiss were held in a specially constructed tank that enabled water to be collected separately from the anterior and posterior ends of the fish. Measurement by radioimmunoassay showed that 〉95% of the cortisol and melatonin released into the water originated from the anterior end (dominated by the gills). High performance liquid chromatography confirmed the identity of both hormones.

Notes: The bile of sexually mature female rainbow trout Oncorhynchus mykiss has pheromonal activity which causes a significant increase in concentrations of 17,20β-dihydroxy-4-pregnen-3-one in the plasma of males. Bile from male trout is inactive. The activity in the female bile binds to octadecylsilane and can be eluted with methanol. The synthetic bile acids, taurocholic acid and taurolithocholic acid, are inactive.

Notes: Previous studies have shown that spermiating male rainbow trout respond to the presence of female urine in the water with significant increases in plasma levels of gonadotrophin II, 17,20β-dihydroxy-4-pregnen-3-one (17,20β-P) and testosterone. The present results show that males only need a single brief exposure to female urine in order to respond; levels of 17,20β-P rise significantly within 1 h of exposure, and peak between 3 and 4 h. Also, milt amounts increase significantly following exposure of males to female urine. Levels of 17,20β-P are also related positively to the amount of female urine to which the males are exposed. Furthermore, when live females are placed, out of physical and visual contact, in the same tank as males, levels of 17,20β-P rise in the same way as in males which are exposed to female urine. However, if females are fitted with urinary catheters (which drain the urine outside the tank), males respond more slowly. These results indicate that urine is the main source of the male ‘priming’ pheromone.

Notes: Photoperiods of 19 h light or more induced sexual maturation in male underyearling rainbow trout but did not influence maturation in the females. Early maturation was not a consequence of the increased growth under long photoperiods. The rate of mortality was much higher in the early maturing males than in the immature fish.

Notes: Measurements were made of plasma 11-ketotestosterone and testosterone in control and sex-reversed male rainbow trout, before and during their first spawning season (winter 1978/1979). Plasma concentrations of both androgens (in both groups) were 2–3 ng ml-1 in January 1978 (apart from some precocious spawners), rose slowly to 9–11 ng ml-1 in April and July and then increased rapidly to 100–150 ng ml-1 in November. From this time, testosterone levels declined but those of 11-ketotestosterone continued to rise to a peak of 260 ng ml-1 in February 1979. No significant differences in hormone levels were found between control male and masculinized female fish.

Notes: A procedure previously used for sex steroids was adapted to extract free cortisol and cortisone from water samples taken from rainbow trout Oncorhynchus mykiss tanks. Both corticosteroids could be readily detected by radioimmunoassay (RIA), with cortisol being predominant. All stages of the sampling, extraction and RIA procedure were validated for cortisol. An intermittent problem with poor replication was traced to the use of diethyl ether during the extraction procedure, and was overcome by the use of ethyl acetate. Other modifications were also introduced to speed up the procedure. The concentration and time course of release of both corticosteroids were shown to be related to the degree of stress that the fish had been subjected to. It was confirmed that cortisol concentrations in water and estimated cortisol release rates increased in response to handling stress, and that both were correlated with plasma cortisol concentrations. The potential for using water cortisol concentration and release rates to assess the primary stress response of fishes as a non-invasive alternative to blood sampling is discussed.

Notes: Female-specific markers of reproductive activity [plasma 17β-oestradiol (E2), vitellogenin (VTG) and alkali-labile phosphoprotein phosphorous (ALP)] were measured over 12 months in a captive population of brown trout Salmo trutta. During the early months of the reproductive season (February to May) and using the concentration of plasma E2 or plasma ALP as a marker for females the proportion of fish in which sex was misidentified was high (15–50%). The misidentification rate was considerably lower (1–8%) using plasma VTG. Preliminary evaluation of a commercial immunochromatographic VTG test system as a screen for the presence or absence of VTG in plasma from brown trout provided results that were consistent with those obtained from direct measurement of plasma VTG levels by enzyme-linked immunosorbent assay (ELISA). These preliminary conclusions were verified by sampling upstream-migrating anadromous brown trout, sea trout, and Atlantic salmon Salmo salar trapped over a 6 month period. Plasma E2 levels did not satisfactorily discriminate between male and female sea trout and Atlantic salmon. Plasma VTG levels in both species, however, were bimodally distributed and it was assumed that this divergence corresponded to male (plasma VTG levels 〈10 μg ml−1) and female (plasma VTG levels 〉800 μg ml−1) fishes. Plasma ALP provided a more accurate indication of sex in the wild Atlantic salmon and sea trout than was suggested by the pilot study on captive brown trout. The commercial immunochromatographic VTG test system provided results that were wholly consistent with the data obtained from the trapped fishes by direct measurement of plasma VTG.

Notes: The behavioural responses of reproductive and non-reproductive female round gobies Neogobius melanostomus to water conditioned by reproductive and non-reproductive males and females were tested. The behavioural responses of reproductive female round gobies exposed to odour of reproductive males included increased time spent near the source of the odour, elevated swimming velocities and directed movement to and around the odour source when compared with their responses to control water. These results suggested that pheromones released from reproductive males may induce spawning behaviour in reproductive females. Non-reproductive females exposed to reproductive female odour spent significantly more time near the odour source of reproductive females compared with control water. Non-reproductive females also showed directed movement towards and around the odour source when exposed to reproductive female odour. These results suggested that round gobies use inter-sexual and intra-sexual pheromones and that both sex and reproductive status are important in the detection and release of these pheromones.

Notes: Two experiments were carried out in which male and female tench Tinca tinca were placed in individual containers and tritiated steroids then added to the water. Water samples were collected over the next 6 or 7 h and the fish then sacrificed, bled and the gall bladder removed. Radioactivity was counted in all the samples. Over the course of the exposure period in the first experiment (7 h), radioactivity of 11-ketotestosterone (11-KT) in the water was depleted by 11%, 17,20β-dihydroxypregn-4-en-3-one (17,20ß-P) and 17,20α-dihydroxypregn-4-en-3-one (17,20α-P) by 28%, testosterone (T) by 56% and androstenedione (AD) by 68%. HPLC analysis of water samples at 3 h indicated that none of the steroids was extensively metabolized during the experiment. Females had a faster rate of uptake of AD than males. In the second experiment (6 h), radioactivity of cortisol in the water was depleted by 5%, 11-KT by 7%, 17-hydroxypregnen-4-ene (17-P) by 17%, 17β-oestradiol (E2) by 35%, T by 37% and AD by 44%. In both experiments, the amounts of radioactivity that were recovered from the gall bladder and plasma were positively correlated with the rate of disappearance of radioactivity from the water. The ability of the steroids to bind to sex steroid binding protein (SBP) of tench plasma was tested by incubating plasma with radioactive steroids and then separating bound and free with ice cold dextran-coated charcoal. When plasma at a final dilution of 1 : 60 (v/v) was incubated with 5 nM of each steroid, the percentage of radiolabel bound to SBP was: T 48% AD 44%, E2 30%, 17-P 17%, 11-KT 13·2%, 17,20α-P 10·3%, 17,20β-P 4·5% and cortisol 0%. Saturation analysis established dissociation constants (Kd; mean ± s.e.) of 3·4 ± 0·4, 2·2 ± 0·2, 4·0 ± 0·3. 9·0 ± 2·8 and 51·8 nM and binding capacities (Bmax) of 201 ± 29, 201 ± 33, 165 ± 3, 187 ± 15 and 13·4 nM for T, AD, E2,17-P and 17,20β-P respectively. The ability of steroids to displace tritiated T and AD from SBP was in the rank order AD 〉 T 〉 E2 〉 17,20αP = 17,20β-P = 11-KT = 17-P 〉 cortisol. Thus, the ability of tench plasma to bind certain steroids showed a relatively strong correlation with the ability of the fish to take up these steroids from water. Modelling of data for AD and 17,20β-P helped to show why and how plasma binding had a strong influence on the rate of uptake (and hence release) of the steroids.