The Neuroendocrine Mechanisms Analysis of Fish Breeding Regulation Is the Main Basis of Biotech Reproduction Development

Garlov PE

Published on: 2023-04-21

Abstract

The aim of the study is to improve the bio-technique of fish artificial reproduction, based on constructive working scheme of reproductive neuroendocrine regulation. The new bio-technique methods of the main phases of fish-factory reproduction bioengineering, based on environmental and hormonal factors combination were developed. Comparative tests of the innovative and standard sturgeon and salmonids bio-technique effectiveness were produced. Piscicultural and biological results of production tests are discussed.

Keywords

Fish farming; Fish artificial reproduction; Factory sturgeon and salmonids tech breeding; Fish farming in brackish sea water

Introduction

Abbreviations

  • HHNS-hypothalamic-hypophysealneurosecretory system
  • APD - isolated anterior pituitary drug
  • PPD - isolated posterior pituitary drug
  • MPI - method of pituitary injection, LH-RH, GnRH
  • luliberin, gonadotropin-releasing hormone
  • SGM – stage of gonadal maturity.

Mechanisms of neuroendocrine regulation of spawning are well researched only in relation to the specialized influence of neurohormones on adenohypophysial functions and on endocrine and generative gonadal functions [1, 2]. Methods of fish sexual maturing stimulation by purificated gonadotropins and synthetic analogues of luliberin (LH-RH), for example surfagon and combined them with dopamine antagonists, such as pimozid, were developed on the basis of the analysis of these mechanisms. At the same time, the research of neuroendocrinal complex participation in generalized implementation of all phases of spawning, especially with regard to the important role of neurohormones in organism adaptations to the physiological stress-spawn is also needed to manage reproduction [3]. During spawning fish are particularly sensitive to stress, such as to conditions of detention ("handling"), violating all stages of reproductive process. Simplified constructive schemes on vector-temporary basis are especially important to develop biotechnological solutions for managing the timing and quality of fish breeding as a milestone of the cyclic process, in our view. As a result of the analysis the dynamics of the main neuroendocrine links in this process we have  developed simplified working scheme of neuroendocrine regulation of spawning (Figure 1). It reflects the process of hypothalamic neurohormonal integration of spawning, based on the principle of self-regulation [3, 4]Figure 1: The basic principle of participation and functional role of fish preoptical-hypophyseal neurosecretory system in reproduction for example of sturgeon.

 

Based on this scheme and our fishery breeding experience it has been speculatively suggested the following methods of biotechnology of fish reproduction [5]: 1 - to improve the sexual maturation quality (fishery usage) of breeders we offered removing posterior pituitary lobe, retarding sex maturation, from the whole pituitary gland and to use for pituitary injections only isolated anterior pituitary; 2 – to stimulate breeders sex maturation by using extracts of ventral hypothalamus (the tuber cinereum of brain,"median eminence"), containing gonadotropin-releasing hormone, lûliberin (GnRH, LH-RH); 3 - to inhibit breeders sex maturation and to reserve them in the "critical" salinity - 4-8 ‰ as optimal environment for survival; 4 -– to control the process of fish reproduction (its timing and quality) by the complex of adequate environmental factors, primarily threshold: temperature, brightness on the background effects of critical salinity. The main purpose of the suggested biotechnological developments was to enhance the efficiency of existing and development the new methods of valuable fish reproduction management. They are protected by 5 earlier inventor's certificates and the RF patent.

 

Material And Methods

Material and methods development of bioengineering were collected in industrial conditions - the sturgeon fish-farms at lower Volga and Don rivers, salmon fish-farms in North-West region, including sea cage fish farm in Finnish (Vyborg) Bay. Research was performed on the most valuable trade fish, sturgeons: Russian sturgeon (Acipenser g?ldenst?dti Brandt) and stellate sturgeon, sevruga (Acipenser stellatus Pallas), salmons: Atlantic salmon (Salmo salar L.) of Baltic population. Their number (breeders, juvenile fish) is indicated in tables of comparative performance test results of biote?h methods that were evaluated by piscicultural and biological features, morphometrical i?hthyological and some major morpho-physiological indicators (see results). To rate the overall physiological fish state the quantitative morphometry, indicators of water-salt balance of the body, determined blood levels of hemoglobin, total protein, serum, ovary fluid and urine osmolality were used by commonly accepted techniques [4]*/.

Results And Discussion

Improvement Of Bioengineering Stimulation Sex Maturity Breeders of Industrial Fish. The Isolated Anterior Pituitary Drug – APD.

For the quality improvement of sturgeon breeders’ sex maturation we proposed to delete from a pituitary gland its neurointermediate complex (posterior pituitary), exerting its inhibitive neurohormonal effect on the gonads function and to use for pituitary injections (MPI) only isolated anterior pituitary (Figure 1, 2). In the ventral area of pituitary (in the so-called “mesoadenohypophysis”, or “proximal pars distalis” of hypophysis) here is the highest concentration of gonado- stimulating hormone [6]. Anatomical division pituitary into two lobes (anterior and posterior) separated by the hypophysial cavity (in Chondrostei) and their "functional antagonism" (Figure 1) allowed to offer drug APD and to develop, improve and introduce (from 1987) a method of its preparation in sturgeon farming (Figure 2) [7].

Figure 2a: Scheme of sturgeon pituitary gland (hypophysis, pituitary gland drug - PGD: its average optimal dose injection: 30 mg/?). B. Scheme of pituitary division into anterior pituitary drug (APD: 85 ± 5% of PGD mass) and posterior pituitary drug (PPD: 15 ± 5%), after inventor's certificates USSR ? ? 719571, 1163817).

The benignity of this technology for industrial use was proved by comparing histological preparations of isolated pituitary lobes with reference sample [6]. Industrial efficiency audits of the use effectiveness of APD in the sturgeon farming at lower Volga and Don proved a higher degree of breeding use of female breeders in average of 15% (Table 1A, B). While main average indicators (after APD) of physiological state breeders and their offspring remain in the normal range (Table 1B). For the same purpose, including savings source drug glands, is developed a method of application isolated posterior pituitary drug - PPD [8]. It is proved that this drug causes the same benign maturing males, like pituitary (Table 2). The greatest effect of the drug economy sturgeon pituitary is obtained when using the drug in a dose of 5 mg PPD for 1 male and 25 mg APD for 1 female. It is the ratio of drugs obtained by preparation them from one medium dose of a whole pituitary, adopted in production to 1 female - 30 mg, whereas one parent pair requires an average of 50 mg pituitary drag in sturgeon farming. Therefore, using this method allows to reduce the consumption of pituitary in volume used for maturing males, specifically at 35- 40%. Thus, both these natural integrated drugs – APD, PPD, physiologically adequate own fish pituitary, allow to improve wasteless the efficiency of MPI, in average by 15%. It should be reminded, that stimulation of maturing fish breeders, was also suggested for the first time by application the hypothalamic releasing factors (now: GnRH, LH-RH particularly), which are widely used for this purpose in the world piscicultural practice [5].

Table 1a: Results of the maturity stimulation of sturgeon female breeders of the Volga-Caspian population by the isolated anterior pituitary drugs (APD) and the pituitary gland drugs (PGD).

Table 1b: Results of the maturity stimulation of sturgeon female breeders of the Azov-Don population by the APD and PGD (basic physiological and piscicultural-biological indicators).

Table 2: The results of comparative tests of various drugs (anterior pituitary drug - APD, posterior pituitary drug – PPD, pituitary gland drug – PGD) on stellate sturgeon and carp breeders.

Drugs:

PPD (experience)

APD (control)

PGD (control)

 

 

Test-object:

Stellate sturgeon ?? (mg/specimen

)

 

 

Carp

?? (mg/kg

)

Stellate sturgeon ?? (mg/specimen

)

 

 

Stellate

sturgeo n ??

Carp

?? (mg/kg

)

 

Stellate

sturgeo n ??

 

 

Carp

??

 

Dose

 

5

 

10

 

20

1.5

(3.6-4)

 

5

 

10

 

25

1.5(3.6-

4)

 

20

1.5(3.6

-4)

Injected breeders (number)

 

5

 

10

 

5

 

9

 

4

 

4

 

18

 

9

 

19

 

9

Sex matured (numbers)

4

8

5

9

4

3

12

9

19

9

The duration of maturation (in hours)

 

19-30

 

17-24

17-

24

 

45627

 

21-24

 

21-24

 

18-24

 

45627

 

16-24

 

45627

Average sperm movement activity (s)

 

177.5

 

193

 

-

 

42

 

-

 

-

 

-

 

42

 

175

 

42

 

Ejaculate volume (ML)

 

35-150

 

20-90

 

-

3.8-

30.5

 

15-60

 

40-100

 

-

 

-

 

40-100

3.8-

30.5

The quality of artificial insemination of eggs (% fertilization.)

 

 

81

 

 

83.1

 

81.

7

 

 

73.6

 

 

81

 

 

81

 

 

70

 

 

68.7

 

 

82

 

 

68.7

The average consumption of the original drug to 1 parent pair (mg)

 

 

25

 

 

30

 

 

40

 

 

45081

 

 

35

 

 

40

 

 

45

 

 

7.5

 

 

50

 

 

45144

 

Improvement Of Bioengineering Inhibit Sex Maturity and Reserving Breeders

Method to Reserve Fish Breeders in Critical Salinity Environment

Method of long industrial reservation fish breeders in the environment of critical salinity 4-8‰ was developed to inhibit sex maturation, prevent the onset of sexual products resorption, improve breeding quality and the degree of breeding use fish breeders [9]. Critical salinity, as the threshold for the maturation of gametes of marine and freshwater organisms, defines the limit of their physiological stability, as well as a number of important thresholds, borders and gradients of the organism's relationship with the external environment [10]. To determine the degree of breeder’s survival in this environment, preliminary experiments were conducted on the most research object – vobla (Caspian roach, Rutilus rutilus caspicus (Jak). After a 55-day content in concrete pools without feeding at 17.4-23.8oC and concentration 02 - 6.3-7.1 mg/l in more than 350 breeders only in this environment (5‰) a mass inhibited sex maturity (without signs of oocyte resorption) was discovered, while striving to maintain a favourable physiological state and a high degree of survivability (Figure 3).

Figure 3: Breeders roach survival in fresh water and salt solutions: 1. -5 ‰, 2. -12‰, 3. -3 ‰ and 4. -in control (fresh water).

In control and in salinity 3‰ all-out oocyte resorption and loss of fish was stated. The following experiments were conducted on 56 stellate sturgeon females - the most sensitive farmed species to stress - “handling”. Female in size 75-165 cm and weighing 8.5-22 kg in pre-spawning state (IV stage of gonadal maturity) were contained up to 30 days in farm concrete basins at spawning temperatures (15-17, 22-24oC) and above (26-27oC) and oxygen concentration 5.2-7.5 mg/l. While we allowed the possibility of changing the sea water on the "artificial" industrial solution of table salt (NaCl), assuming that imbalances in the ionic composition of the external environment will not have toxic effects on the body in such a low salinity (up to 8‰). The results were evaluated by piscicultural and biological indicators (Table 3).

By production verification method it was finally proved the possibility of industrial reserving sturgeon breeders in salt solutions when spawning temperatures during production and necessary deadlines (Table 4). Thus, it was discovered that the critical salinity 5-7‰ is optimum for reserve breeders and, obviously, for containing fish Brood Stocks. It is possible to discover the potency of reproduction, development, growth and survival of fish by analysing leading mechanisms of the influence of critical salinity on body. Therefore, studying the most important physiological-biochemical characteristics of experimental breeders was started (Table 5).

Table 3: Preliminary results of stellate sturgeon females’ reservation in different salty habitats. 

  Control in fresh water Experience in critical salinity 5-7‰
In sea water In common salt
solution (Na?l)
Version experience 1 2 1 2 2
    25-     25- 17.4-  
Water temperature (?0?) 22-24 26.8 22-24 15-17 26.8 23.8 o C 02 − 6.3-7.1
Durations (days) 7, 16 10, 7 16 14 21 14, 30
14
The number of females 2, 3 8, 3 2X 4 10 4 4, 4
(individuals)
The number of females who have 0 2 0 4 8 2 4, 3
"normal" physiological state
The number of females with no 0 0 0 4 8 2 4, 4
signs of ovarian atresia
The number of fish-farm 0 0 0 2 (2) 3 2 (2) 3, 2 (1, 1)
productive females (from which -1
received a benign offspring)              

Table 4: Results of the industrial reservation stellate sturgeon females in saline solution 5-7‰.

Table 5: The major physiological indicators of vobla and stellate sturgeon breeders in different salinity.

In critical salinity losses in hemoglobin and serum protein are minimal, with maximum retention of salts in the blood and in body cavity fluids, apparently due to the optimization of water-salt balance of the body. Histophysiological analysis of hypothalamo- hypophysial neurosecretory system (HHNS) in experimental females in this environment showed activation of nonapeptide neurohormone release into the bloodstream at the 15-th day of reservation with subsequent stabilization level of its’ functioning (Figure 4)

Figure 4: The degree of functional activity of preoptical- postneurohypophyseal neurosecretory system (PNS) females stellate sturgeon during 1 month in salt solution 5-7 ‰ (experience) and in river water (control).

This phase reaction of HHNS, responsible for day-to-day water- mineral metabolism (diuretic and antidiuretic effects), interrenal tissue and thyroid glands functions (carbohydrate and total metabolism) indicates moderately stress effect of critical salinity on body (?ustress). The minimum level of reduce serum and ovarial fluid osmolarity specifies the importance of moderate amounts of nonapeptide neurohormones, excreting into the bloodstream and stimulating the glands function (Table 5). Biostimulative effect of critical salinity can be explained by the energy-efficient optimal osmotic gradient between the internal and the external environments (mechanisms of water-salt balance, close to isoosmotic osmoregulation type), on the one hand. On the other hand, it can be explained by the fluctuated balance between release of neurohormones into the bloodstream and their synthesis, providing chronic moderate activation of glands-targets and optimum water-salt homeostasis of the organism. Long inhibit ovulation and resorption in critical salinity is also associated with increased content of nonapeptide neurohormones in blood which have the antigonadotropic resistant effect [1, 4]. Indeed, a comparison of the piscicultural and biological indicators with those same morpho-physiological in the experience and control also indicates to their clear relationship with fish quality females (Figure 4; see Table 3-5).

Artificial Reproduction of Fish Populations

Method of Reproduction of Fish Populations

The main objective of farm reproduction of fish stocks is the preservation of fishing fish populations. This is possible only by preservation of biodiversity their biological races (groups, ecological forms, etc) inside population structure, settled all spatio- temporal ecological niches biocenosis in micro evolution, as the most important element of the status of biological species progress [6]. The basis for the development of biotech reproduction intraspecific groups is to use a combination of hormonal and environmental impacts in a single complex adequate natural. Based on results of complex ecological and histo-physiological studies of the functional role HHNS in fish reproduction “Method of reproduction of fish populations" is aimed to preserve their strength by factory breeding of all elements of the population structure in their natural proportions [11]. Essence of the method consists in synchronization of the timing producing heterogeneous offspring of different biological races in a single spawning farm-season by means of controlling the timing of reproduction breeders with different seasonality and different calendar dates of spawning. This principle is implemented impact multi-cultural biotechnical physiologically adequate complex of environmental factors signal and phylogenetic significance for fish. Ecological principle of management is to reserve breeders in specy-specific pre-spawning threshold conditions (light and temperature) on the background of content in critical salinity, universal for different fish species (Figure 5).

Figure 5: Principle of fish breeding, reservation and growing acceleration control by the triad of main habitat conditions: signal (??, L) and phylogenetic (‰) values, based on the primary mechanism of fish migrations (author’s certificate USSR ? 682197 “Method of reproduction of fish population”).

For example, we offer to reserve spring-spawning breeders at temperatures of 1-20? below the lower threshold for the spawning of this species and race with darkening and autumn-spawning - at 1-20? above the upper threshold spawn-temperature (T) and lightness (L) ranging from 5 to 60 lux, but exceeding the 75-80% of the average daily spawning period. All of these conditions are threshold for fish body reactions within the specy physiological norm. Their specific values established on the basis of our own breeding experiences and literature data [2, 4, 6]. After reservation during necessary time breeders are to carry into the spawning environmental complex and to stimulate their sex maturity (Figure 5, Table 1-4). It is clear that these breeding and reservation conditions, generalized for different species with any season of spawning and based on the principles of manage fish responses in moderate climatic zone are very rough and can be adjusted experimentally.

Development of modern method of reproduction valuable fishing species populations, including the initial and final stages of farm bioengineering was performed on the Atlantic salmon because of particular relevance. Artificial reproduction factory populations of salmon in the Northwest, along with the overall aim, significantly differs from that in the Southern and Eastern regions and fundamentally - from the sturgeon factories, isolated from spawning grounds. The vast majority of salmon hatcheries is located on the spawning grounds, there withdrawing mature breeders in damage to natural reproduction. Thus, fishing dependence of farms combined with fishing of valuable and protected species on the spawning grounds and at the spawning period is probably the main reason for the progressive reduction of population numbers, until the annihilation in many rivers. It is important that during river spawning migration all salmon species are undergoing “nuptial changes”, losing food product quality, which are only at sea in the feeding areas and fisheries. Thus, breeders for artificial reproduction are removed not only from natural, spawning, but also from industrial use [6]. Therefore, the possibility to obtain offspring from the silver breeders in the sea feeding areas and effective cultivation here large viable juveniles in optimal habitat (Figure 5) may be crucial to restore salmon populations [6, 12]. Because of the decrease in the number of running upstream breeders of Atlantic salmon, especially in the Northern regions, the creation and maintenance Brood Stocks are often the only way to ensure the hatcheries spawn. Therefore, with a view to developing complete system biotechnology of intensive factory reproduction the Atlantic salmon biodiverse breeders for the first time were captured and reserved in sea cages in the coastal zone of the Finnish (Vyborg) Bay. Here in brackish water salinity 2.5-4‰ their Brood Stock was reserved before full maturing breeders at temperatures 3-70? (Table 6).

Table 6: Basic piscicultural-biological characteristics of Atlantic salmon breeders reserved in sea cages.

As a result, from naturally ripened breeders during the season of spawning (October-November) was firstly received mature eggs and offspring. Fertilized roe was transported in the incubation shop of spawning-growing farm, where it was incubated in fresh water until the larvaes and extend them to an external ("active") food. After full conversion to active food early fries were taken back in sea cages for intensive growing from juveniles to stage large viable smolts. We believe it is necessary because salmon farms usually produce juveniles one year aged weighing up to 26g and often - even non-viable fingerlings due to thermal anomalies, moreover in the absence of developed biotech its release in habitat. Calculation of efficiency of salmon reproduction prove the need for release a larger smolts-over 40g. and in large enough quantities-more than 150 thousand [12]. Atlantic salmon juveniles were grown in cages in the specified salinity up to 3-summer age in more than 3 thousand smolts and were feeded trout fish feeds "Biomar» at the expense of feed 1.3-1.4 kg. As a result young salmon bonitet their main piscicultural-biological growth characteristics were stated (Table 7).

Table 7: The average values of morphometric biological indicators of year-olds and 3 years-olds salmon fry by all parties, grown in brackish water cages.

 

Based on these primary data, the following comparative indicators of growth of salmon youngs grown in cages in brackish water, in factory pools in river water and according to industrial standards can be provided (Figure 6)

Figure 6a: Comparative indicators of body weight (gm) juvenile salmon (0+ - fingerlings, 1 - yearlings, 1+ - 2-summer age), grown in cages in Finish Bay (left column), at Nevsky fish-farm and according to the accepted bio-norms (bright right columns). B. Comparative dynamics of growth of young salmon in cages in brackish water (2.5-4 ‰), in river water at Nevsky fish-farm (NF) and according to norms (N).

Mass comparison of juveniles, grown in brackish water with factory and normative data shows multiple increased growth rates under equal conditions (temperature, feeding, etc.), especially significant since 1 year old, in 5-7 times [12]. It is important that only in brackish water fingerlings smoltify process has a massive simultaneous character and here growing almost excludes the appearance of "river" dwarf males, which reduces the main factory waste products. We tried to develop a method for improving the efficiency of biotech cultivation factory young salmon through additional cultivation in brackish seawater close to critical "salinity since the start of the smoltification (as the signal "marker" state) and to release it in natural feeding areas [12, 13]. Therefore, the application of the modern biotechnology, integrated maintenance and operation of Brood Stocks and intensive rearing of juveniles using adaptations of sea-river system (river – the depletion, mainly, sea - the accumulation of material-energy resources) can unite the interests of all forms of reproduction, fishing (on the feeding areas) and even commercial cultivation in coastal fish farms. As a result, we propose to discuss scheme integrated factory salmon reproduction combining farm industrial and coastal sea-cage cultivation capabilities (Figure 7). It is proposed to incorporate the activities and products of sea-cage cultivation into the production cycles of the model farm salmon as its’ external reproduction plot. This “marine” centralization of starting and final factory tech cycles is possible due to the fact that fish homing is not retained genetically and imprinting, apparently formed already in the first summer of factory growing larvae and early juveniles (fingerlings) since the transition to active feeding [12-14]. For industrial use proposed bioengineering directly in fish factories, year-round fish breeding and protecting products from contamination has developed a “Water supply system of fish farms” by the underground hydro-air conditioning fully managed environment cultivation based on energy saving principles of environmental engineering [15]. All developed methods we consider as biotech reproduction management system. It is based on the practical use of population-specific phylogenetic adaptations to marine life period, implement the hidden potency of fish species and ensuring maximum productivity of the population [6, 12, 16]. Its unity, general direction and the ultimate goal is based on a view of the central place of the HHNS (and the whole neuroendocrine complex) in controlling the material-energy balance at organism and population levels of biological organization [4, 17, 18].

Figure 7: The proposed scheme of combined breeding fish farming, including the additional sea cages (planting plot) to grow repaired Brood Stocks and major salmon smolts to release to pasture in brackish seawater.

References

  1. Pierantoni R, Cobellis G, Meccariello R, Fasano Evolutionari aspects of cellular communication in the vertebrate hypothalamo-hypophysio-gonadal axis Internat. Rev Cytol. 2002; 218: 69-141.
  2. Zohar Y, Munoz-Cueto JA, Elizur A, Kah Neuroendocrinology of reproduction in teleost fish. Gen Comp Endocrinol. 2010; 165: 438-55.
  3. Garlov PE. On possible mechanisms for the participation of the neurosecretory system of sturgeons in the implementation of - Materials for joint scientific session Central Scientific-Research Institute of sturgeon fisheries (CNIORH) and Azov Scientific-Research Institute of fisheries (AZNIIRH). Astrakhan: “Minrybprom”. 1971; 22-24.
  4. Garlov PE. Plasticity of nonapeptidergic neurosecretory cells in fish hypothalamus and neurohypophysis. Internat Rev Cytol. 2005; 245: 123-170.
  5. Garlov PE. On regulatory effects of hypothalamic-hypophysial neurosecretory system of sturgeons on endocrine glands and perspectives of using this phenomenon in fish farming//Abstracts of the reporting session of the Central Scientific-Research Institute of sturgeon fisheries (CNIORH). 1972; 39-40.
  6. Garlov PE, Kuznetsov YK, Fedorov Artificial reproduction of fish. Management of reproduction. Tutorial (SPbGAU, "GOSNIORH"). St.-Petersburg (SPb.): "Deer". 2014; 256.
  7. Garlov PE, Polenov AL. ?ethod of manufacturing a hormonal drug to stimulate fish breeders’ 1976.
  8. Garlov PE, Polenov AL, Altufyev YV, Popov OP, Burenin OK. Method of stimulation male fish 1983.
  9. Garlov PE, Polenov AL, Altufyev YV, Derevyagina Method of reservation fish breeders. 1977.
  10. Khlebovich Critical salinity of biological processes. Leningrad: “Science”. 1974; 235.
  11. Garlov Method of reproduction of fish population. 1977.
  12. Garlov PE, Yanbukhtin DA, Titarenko KA. The experience of containing sturgeons and salmonids breeders in brackish The News of the St.-Petersburg State Agrarian University (SPbGAU). 2015; 40: 122-130.
  13. Garlov PE, Bugrimov BS, Rybalova NB, Turetckiy VI, Torganov Method of reproduction of fish populations. Application for invention. The positive outcome of the formal examination. 2014. Garlov PE, Bugrimov BS, Rybalova NB, Turetckiy VI, Torganov. SV. Method of reproduction of stellate sturgeon and Baltic salmon populations. Patent for invention No. 2582347. 2016.
  14. Hasler AD, Scholz AT. Olfactory imprinting and homing in Investigations into the mechanism of the imprinting process. 1983; 134.
  15. Garlov PE. Water supply system of fish 2010.
  16. Suvorov Use hidden opportunities for fish growth. Informational collection Advisory Bureau VNIIORH. 1940; 7- 9.
  17. Van Winkle W, Rose KA, Shuter Effects of climatic temperature change on growth, survival, and reproduction of rainbow trout: predictions from a simulation model. Can Journ Fish and Aquatic Sci. 1997; 54: 2526-2542.
  18. Balment RJ, Lu W, Weybourne E, Warne Arginine vasotocin a key hormone in fish physiology and behaviour: a review with insights from mammalian models. Gen Comp Endocrinol. 2006; 147: 9-16.