The Physical Responses to Acute Stress (HUMAN)

Following a threat, the part of the brain called the hypothalamic-pituitary-adrenal (HPA) system releases certain neurotransmitters (chemical messengers) called catecholamines, particularly those known as dopamine, norepinephrine, and epinephrine (also called adrenaline). The HPA systems also trigger the production and release of steroid hormones (glucocorticoids), including cortisol -- the primary stress hormone. Cortisol affects systems throughout the body. Catecholamines also activate an area inside the brain called the amygdala, which apparently triggers an emotional response to a stressful event and also signals the hippocampus -- a nearby area in the brain -- to store the emotionally loaded experience in long-term memory. In primitive times, this combination of responses would have been essential for survival, when long-lasting memories of dangerous stimuli (such as a large animal) would be critical for avoiding such threats in the future. During a stressful event, catecholamines also suppress activity in areas at the front of the brain concerned with short-term memory, concentration, inhibition, and rational thought. This sequence of mental events allows a person to react quickly -- either to fight or to flee -- in emergency situations; however, this also hinders a person's ability to handle complex social or intellectual tasks and behaviors.

The heart rate and blood pressure increase instantaneously in response to stressful situations. Breathing becomes rapid and the lungl take in more oxygen. Blood flow may actually increase 300% to 400%, priming the muscles, lung, and brain for added demands. In addition, the spleen discharges red and white blood cells, allowing the blood to transport more oxygen.

The immediate effect of stress is to dampen parts of the immune system. In addition, certain factors in the immune system -- including important white blood cells -- are redistributed, much like marshaling soldiers to potentially critical areas. In the case of stress, these immune-boosting troops are sent to the body's front lines where injury or infection is most likely, such as the skin, the bone marrow, and the lymph nodes.

During stress, fluids are diverted from nonessential locations, including the mouth, causing dryness and difficulty in talking. In addition, stress can cause spasms of the throat muscles, making it difficult to swallow and fight infection.

Stress commonly results in cool, clammy sweaty skin and in a tightening of the scalp that makes the hair seem to stand on end. The skin is cool because blood flow is diverted away so it can support the heart and muscle tissues. As a result, physical capacity is increased and blood loss is reduced in the event of injury.

Stress shuts down digestive activity, a nonessential body function during short-term periods of physical exertion or crisis.

Two constraints to aquaculture productivity are the very low seawater temperatures  and stress-related problems in fish held in captivity.  Studying an ability of a small fish (e.g the rainbow smelt) that could prove useful in avoiding both of these problems in aquaculture. Smelt are not currently a farmed species but they have the capacity to accumulate very high levels of glycerol, an ability that could be of great use in the species that we do farm such as salmon, halibut and others.

Low seawater temperatures in winter limit coastal aquaculture because they can cause fish such as Atlantic salmon and rainbow trout to freeze in sea cages. The risk of freezing makes sea cage aquaculture impossible and this limits the industry to very narrow regions. Many species do survive naturally in the icy waters by means of biochemical adaptations. One of these adaptations is the production of very high glycerol levels in smelt during winter. The glycerol acts in the same way as ordinary windshield antifreeze, lowering the freezing point of the fish and protecting it from freezing.

Fish can also suffer from stress and this is something we wish to prevent in aquaculture because it reduces survival and growth. In mammals, such as humans, stress can elicit increases in an enzyme that helps to make glycerol. We have found that smelt seem to use the same enzyme to make their glycerol. In simpler life forms, such as yeast, enormous amounts of glycerol are produced in the same way during exposure to excess salt or other sources of stress. Glycerol can help to stabilise proteins and membrane fats from almost any life form if they are perturbed so it makes sense to produce glycerol when there is a problem.

We are working to determine how smelt make their large amounts of glycerol and we intend to find means of generating this in other fish that are grown in aquaculture to protect them from freezing and to help them respond well to stress. We are also studying the effect of glycerol on fish cells in culture to determine the nature of its protective effects and how best to deliver it to the fish.

RELATED STUDIES STRESS IN FISH BY Department of Animal Physiology in Nijmegen, The Netherdland.

1 Development and application of methods for detection of stress in Mediterranean fish species in aquaculture The overall objective of the project is to develop a non-invasive method for detecting general and specific stressor-related stress effects in fish reared in aquaculture. Within the general research framework the project aims at two interrelated objectives: To detect, analyze and quantify stress effects, under both laboratory and fish farm conditions, in two Mediterranean marine fish species, gilthead seabream (Sparus auratus) and European sea bass (Dicentrarchus labrax), which are of major economic importance for European aquaculture, using as parameters changes in the skin and gill epithelia, and the mucus layer covering the skin: (a) changes in tissue morphology, (b) marker enzymatic activities in skin and skin mucus, (c) concentrations of the stress hormone cortisol in blood serum and skin mucus and (d) immunological parameters. The stressors to be applied (low dissolved oxygen levels, high ammonia levels, and handling stress) are relevant to aquaculture. To select subsequently the analytical methods and stress parameters that will prove to be reliable and easy to perform, using a sampling procedure from which the sampled fish can easily recover. The selected methods will be combined as bio-indicators through the establishment of a new stress-index, the "Fish Stress Condition Index" (FISCI), that will enable the characterization of stress condition in cultured fish and will reflect both fish health and water quality. This will provide fish farmers and managers of nature reserves with a protocol to check the stress condition of their fish. The participants will study systematically the effect of low dissolved oxygen levels, high ammonia levels, and handling stress, which are three aquaculture relevant stressors, on structure and ultrastructure of the skin and gills of gilthead sea bream and European sea bass, reared under both summer and winter temperature and light regime conditions. The analysis will be both qualitative and quantitative, in order to identify general tissue responses to the stressors and specific responses to each stressor separately. The combination of histopathological and biochemical methods to be applied on skin and gill biopsies and mucus samples is meant to lead to the development of the FISCI index. The FISCI will be used as an (almost) non-invasive method that will not require killing the fish. This method may enable the detection of fish stress status and will enable in many cases, through the stressor specific responses, the determination of the detrimental factor in the water.   2 The immuno-neuroendocrine interactions in fish strains selected for stress response or immune capacity Improvement of genetic disease resistance will reduce fish mortalities caused by diseases, and thus improve fish welfare. However, because of the complicated interaction between the immune system and the neuroendocrine system, it is expected that the stress response of fish will affect disease resistance, even in fish strains genetically selected for improved fish health. In this project we will investigate how this selection has changed the interaction between the immune system and the brain-pituitary-interrenal (BPI) axis. Interleukin-like factors produced by the immune system, and cortisol, the end product of the BPI-axis, are two of the most important mediating factors involved in this interaction. Interleukin-like factors have been reported to stimulate the BPI-axis. Cortisol, at least at the high levels typical for severe stress in fish, has suppressive actions on the immune system, which is explained as moderating mechanisms to prevent overreaction of the immune system. The central hypothesis of this project is that selection for either stress response or immune competence will disturb the interaction between both systems, with negative effects for fish health and welfare. First, the effects of the standard stressor (crowding) on the activity of the BPI axis (ACTH, MSH, b-END and cortisol production) will be compared for different strains selected for either stress response or immune competence (the same strains as used in the other projects). Second, the concentration/response relationship between IL-1 and this axis, and between cortisol and different immune parameters will be investigated in these strains in order to elucidate the physiological mechanisms involved.   3 Stress resistance in two commercially aquacultured marine fish species: turbot and Atlantic halibut Geographical variation in growth capacity with a general tendency towards lower temperature optima and higher growth capacities at higher latitudes has been described for many marine fish species. A potential implication from these variations for marine fish aquaculture could be the use of Northern strains in Southern environments, resulting in an improved growth rate of farmed fish. The use of strains with different growth capacities to improve growth under aquaculture conditions has been demonstrated for several salmonid species. However, for turbot and Atlantic halibut, two finfish species of highly commercial interest and with a wide geographical range, comparative data of optimum temperature ranges for growth from different strains is very scarce. The purpose of this 3-year EU project, which will run from 1998-2002, is to investigate the effects of different rearing conditions on growth, growth efficiency, immune capacity and the stress resistance in geographically distinct populations of turbot and halibut. The project is a collaboration between the Universities of Bergen (Norway), Cork (Ireland) and the Department of Animal Physiology in Nijmegen. The rearing and sampling of the fish will be done in the laboratories of the University of Bergen, where also parts of the analysis will be done. Part of the analyses, however, will be done in Nijmegen. Measurements on stress parameters will include: physiological parameters in blood: haematocrit, leucocrit, plasma osmolality, plasma ions (Na, K, Ca, Mg, Cl), plasma glucose; biochemical parameters in gill epithelia: NaK-ATPase activity; microscopical analysis of gill and skin epithelia: light microscopy (LM), confocal laser-scanning microscopy (CLSM) and electron microscopy (EM): quantification of apoptosis necrosis and leucocyte infiltration with EM and advanced fluorescent probes for CLSM (TUNEL method; biotinylated mAB to annexine); quantification of mitosis with fluorescent probes for LM and CLSM; microscopic analysis of catecholamine and cortisol producing tissues: proliferation measurements with PCNA-techniques, rate of apoptosis of leucocytes. 4 The interaction between stress and colour pattern in a Mediterranean fish species with high potential for aquaculture The red porgy (Pagrus pagrus) is a new commercial (food)fish species with a high potential for successful culture. A major problem met in aquaculture of this species is the skin darkening observed in culture, compared with wild counterparts. This EU-project aims to try and develop a natural hue for this fish in aquaculture, by determining appropriate nutrition, accommodation and handling procedures. The part of Nijmegen in this project is to determine the impact of aquaculture-related stressors on the colour patterns of fish, by investigating the hormonal regulation with a major emphasis on melanotrope cells in the pituitary (which regulate the darkness of the skin) in wild and cultured fish, as well as the regulatory role of these cells during stress. Aquaculture-related stressors are applied and the stress-response of the fish will be evaluated. Also, the environmental situation (background colour, illumination, fish density) will be investigated to find the most suitable artificial habitat for this species. Experiments will be performed at Marine Institutes in Crete (Greece) and in Gran Canaria (Spain). Related research will be performed in Nijmegen University with tilapia (Oreochromis mossambicus), a species well known for its ability to adapt quickly to different background colour; the melanotrope cells in the pituitary gland playing an important role in colour and stress regulation.

RELATED STUDIES STRESS and BEHAVIOUR IN HUMAN.

Repeated exposure to psychosocial stressors as well as exaggerated reactivity to stress have been implicated as factors in the development of hypertension and heart disease. In addition, chronically elevated levels of stress-related hormones (catecholamines and glucocorticoids) are known to inhibit the activity of D6D, the enzyme needed for metabolism of EFAs.

Several early studies on rats found that dietary omega-6 and omega-3 fatty acids reduced the cardiovascular reaction to stress (Mills et al., 1985, 1986). Hence it is not surprising that both GLA and DHA have been found to reduce blood pressure and heart rate responses to psychosocial stress in humans.

Mills extended his findings to humans in a four-week study on 30 male university students (1989). Three different treatment groups were given either borage oil (1.3g/d), fish oil (1.6 g/d) or olive oil (as placebo). Borage oil significantly reduced stress-induced systolic blood pressure and heart rate after four weeks of supplementation, whereas olive oil and fish oil were without effect. Task performance was also significantly improved in the borage oil group, while un-changed by olive oil and fish oil, in a test that required a high level of attention and was designed to measure the cardiovscular response to psychological stress.

These results were similar to findings in the earlier animal studies and suggest that borage oil supplementation is effective in reducing cardiovascular reactions to stressors of all kinds, of both short and long-term, psychological and physical nature.

As a follow-up to earlier findings that DHA intake prevents aggression from increasing at times of mental stress (Hamazaki et al., 1996) Sawazaki et al. (1999) conducted an excellent double-blind study to test the effect of DHA intake on the level of stress hormones (epinephrine and norepinephrine). Fourteen medical students were studied over a stressful nine-week period when they underwent over 20 final exams. The participants in the DHA group were given 1.5 g DHA/day, while the control group members were given a mix of plant oils, all in capsules taken with meals.

The norepinephrine levels were high in both groups at the beginning of the study, since the students had already been under stress for some time, preparing for the exams. At the end of the test period the DHA group showed significantly reduced (-31%) norepinephrine levels, which is believed to be protective and beneficial for the cardiovascular system. In the control group the norepinephrine levels were still high. Epinephrine and cortisol showed no significant changes in either group. (Elevated norepinephrine levels are associated with chronic stress, while epine-phrine increases in situations of acute “survival” stress).

Similar findings of reduced norepinephrine levels related to EFA intake have been reported by other authors (Singer et al., 1990; Christensen et al., 1994). In Singer’s study on 47 hypertensive individuals, norepinephrine levels were reduced 80% after treatment with omega-3 fatty acids compared to the control groups. Christensen’s study showed that norepinephrine levels of men who died from cardiovascular disease were significantly higher than those of survivors.

Interestingly, the students in Awazaki’s study were under considerable stress even long before the testing began, and the baseline levels of norepinephrine were already high at the start. This means that DHA was able to modulate catecholamine meta-bolism even after the appearance of stress. This is a noteworthy point when applying these results to daily life, as we usually do not try to counteract stress until after it starts.

Insulin resistance

Insulin resistance is a common phenomenon in aging and in simple overweight. It is a primary factor in the so called metabolic syndrome X and is strongly linked to the development of a cluster of common age-related disorders including type 2 diabetes, obesity, hypertension, hyperlipidemia and heart disease. Insulin resistance is found in approximately 25% of apparently healthy humans.

Insulin resistance means that cells are desensitized to insulin signaling that normally leads to glucose uptake. The body tries to compensate for higher levels of circulating glucose by increasing insulin production. When this temporary compensatory mechanism fails, the glucose levels stay elevated, leading to diabetes and other degenerative complications.

Research has now shown a strong connection between the intake of essential fatty acids, in particular GLA and DHA, and improved insulin sensitivity (reduced insulin resistance).

Both human and animal studies show that a dietary intake of EFAs both increases the unsaturated fatty acids in membrane phospholipids and makes the individual more insulin sensitive (Storlien et al., 1986, 1987; Borkman et al., 1993; Vessby et al., 1994; Pan et al., 1995; Storlien et al., 1996).

Until recently, however, scientists did not understand the deeper mechanisms behind the influence of EFAs on insulin resistance. The discovery of a fundamental mechanism for the regulation of fat metabolism in the body has shed light on the effect of EFAs: the nuclear receptors and transcription factors called peroxisome proliferator-activated receptors or PPARs (See side bar on previous page).

Recently developed drugs, called glitazones or thiazolidinediones, that bind to and activate PPAR, increase insulin sensitivity. We now know that GLA and DHA, as well as certain other EFAs work in the same way, binding to and activating PPAR.

Brain development and learning

In the last decade DHA has been discovered to be of major importance for the development and maintenance of brain function, both in young and old individuals. As the major structural and functional EFA of the central nervous system, including the retina of the eye (Connor et al., 1992), it constitutes as much as 30% to 50% of the total fatty acid content of the human brain and is essential for optimal neurological function. Part of the reason for this unique function is the role of DHA in the synthesis of phospholipids in nerve cell membranes.

Nothing can be more important than an adequate supply of DHA at the beginning of life, since it is essential for the growth and functional development of the brain in infants. DHA deficiencies in infancy have been associated with visual impairment and the later development of disorders including attention deficit hyperactivity disorder (ADHD), learning disabilities and aggressive behavior. DHA is also required for the maintenance of normal brain function in adults, for learning and for memory, and low levels have been shown to be a risk factor for Alzheimer’s disease (Horrocks et al., 1999).

Many experimental studies on mice and rats have been conducted to clarify the effects of DHA on learning and memory. These studies clearly indicate that DHA deficiency is associated with a loss of discriminative learning ability (Greiner et al., 1999), while omega-3 enriched diets increase learning ability in elderly animals.

The Japanese research team Lim and Suzuki demonstrated superior maze-learning ability in old mice fed a DHA supplemented diet. After four months on the diet the mice made significantly fewer mistakes and spent less time in the maze than the control group. They even performed better than the young rats on the control diet (Lim & Suzuki, 2000). When the re-searchers studied the relationship between the time of DHA intake and maze behavior, they found that an improved maze-learning ability was evident at one month after the feeding started, whereas increased DHA levels in the brain were apparent as early as two weeks. These results suggest that improvement in learning ability may take some time after the incorporation of DHA into the brain (Lim & Suzuki, 2001).

Dementia

As we have seen, aging is often connected to a decreased meta-bolism of EFAs. Changes in the fatty aid composition of brain lipids during aging appear to be correlated with a deterioration of the central nervous system. Knowing that DHA constitutes a major portion of the fatty acids in the brain, it may not be surprising that low DHA levels are shown to be a significant risk factor for the development of Alzheimer’s disease.
In a recent study tracking DHA levels in 1188 elderly American subjects for 10 years, Alzheimer’s disease was 67% more likely to develop in individuals with DHA levels in the lower half of the distribution (Kyle et al., 1999).

Brain cholinergic systems are generally thought to be critical for memory function. Dysfunction of the central cholinergic system has been seen both in patients with vascular dementia and with senile dementia of Alzheimer’s type. In a study on stroke-prone spontaneously hypertensive rats Minami et al. (1997) demonstrated that DHA increased choline and acetylcholine levels in the brain, while improving passive avoidance performance.

Interesting results from a Japanese clinical trial on DHA and dementia provide encouragement for further research. This pilot study involved 20 elderly people (average 83 years) with moderately severe dementia from thrombotic cerebrovascular disorder (stroke) (Terano et al., 1999). The participants all lived in the same home for the elderly and ate the same food. They were divided into two groups according to age and baseline scores on psychometric tests. The individuals in the treatment group received 720 mg of DHA daily for one year. Significant improvement in the dementia scores was noticeable after three to six months of DHA supplementation. The control group showed no improvement.

Safety

With all these benefits of GLA and DHA in mind it is important to remember that too much of a good thing is not always good. Balance is the key, in this case between omega-6 and omega-3 fatty acids. The easiest and safest way to accomplish this balance is by taking a high quality combination supplement (ideally in the 2:1 range), while reducing dietary intake of saturated and hydrogenated fats.

Through the simple and safe procedure of supplementing our diet with a balanced combination of GLA and DHA it seems evident from current research that we have the chance to prevent a significant portion of the age-related degenerative diseases that plague our society today. It will ease our bodies’ response to stress and may even help us to escape dementia.

RELATED ARTICLE

1.

Administration of docosahexaenoic acid influences behavior and plasma catecholamine levels at times of psychological stress.
Lipids 1999;34 Suppl:S33-7
Hamazaki T, Sawazaki S, Nagasawa T, Nagao Y, Kanagawa Y, Yazawa K.

Department of Clinical Application, Institute of Natural Medicine, School of Medicine, Toyama Medical and Pharmaceutical University, Japan. hamazaki@ms.toyama-mpu.ac.jp

The purpose of the present research was to clarify the effect of docosahexaenoic acid (DHA) intake on behavior and plasma catecholamines (CA). In Study 1, 42 students took either DHA-rich oil capsules containing 1.5-1.8 g DHA/d or control oil capsules containing 97% soybean oil plus 3% of another fish oil for 3 mon in a double-blind fashion. They took a psychological test (PF Study) at the start and end of the study. This study started at the end of summer vacation and ended just before the final exams. In the control group, external aggression (aggression against others) in PF Study was significantly increased at the end of the study as compared with that measured at the start (+8.9%), whereas it was not significantly changed in the DHA group (-1.0%). In a similar double-blind study (Study 2), we measured external aggression under nonstressful conditions. External aggression slightly decreased in the control group, whereas there were no significant changes in the DHA group. In Study 3 with 14 students, plasma CA were measured at the start and end of capsule administration period of 2 mon. Subjects were under continuous stress of the final exams that lasted throughout the whole study period. The ratio of plasma epinephrine to norepinephrine concentrations was significantly increased in the DHA group (78%), whereas it stayed at the same level in the control group. In Study 4, mice were fed either DHA-deficient diet or -sufficient diet for 4 wk, and their rearing frequency (an anxiety index) was measured. In the DHA-sufficient group, the rearing frequency was significantly less than in the other group. These effects of DHA intake may be applied to people in an attempt to ameliorate stress-related diseases.

Publication Types:

• Clinical Trial
• Multicenter Study
• Randomized Controlled Trial

2.

Anti-stress effects of DHA.
Biofactors 2000;13(1-4):41-5
Hamazaki T, Itomura M, Sawazaki S, Nagao Y.

Department of Clinical Application, Institute of Natural Medicine, Toyama Medical and Pharmaceutical University, Japan. hamazaki@ms.toyama-mpu.ac.jp

DHA is abundant in the brain. Deficiency of DHA changes behavior in animals. The purpose of the present studies was to clarify the effect of DHA intake on hostility and plasma catecholamines. In study 1, forty-one students took either DHA-rich oil capsules containing 1.5-1.8 g DHA/d (17 females and 5 males) or control oil capsules containing 97% soybean oil plus 3% fish oil (12 females and 7 males) for 3 mon in a double blind fashion. They took a psychological test (P-F Study) at the start and end of the study. Study I started at the end of summer vacation and ended in the middle of mental stress of final exams. In the control group, hostility measured by P-F Study was significantly increased at the end of the study as compared with that measured at the start (+58%), whereas it was not significantly changed in the DHA group (-14%). In a similar double blind two-mon study (study 2), we measured plasma catecholamines and cortisol of students (3 females and 4 males for the DHA group and the same numbers for the control) at the start and end of the study. In study 2 the students were under a continuous stress of final exams that lasted for two mon throughout the whole study period. The plasma cortisol did not change in either group, but the norepinephrine concentration was significantly decreased in the DHA group (-31%), whereas it stayed at the same level in the control group. These effects of DHA intake may be applied to people under psychological stress.

PMID: 11237197 [PubMed - indexed for MEDLINE]

Fish diet, fish oil and docosahexaenoic acid rich oil lower fasting and postprandial plasma lipid levels.

Agren JJ, Hanninen O, Julkunen A, Fogelholm L, Vidgren H, Schwab U, Pynnonen O, Uusitupa M.

Department of Physiology, University of Kuopio, Finland.

OBJECTIVE: The present study was carried out to clarify the effects of fish diet, fish oil and docosahexaenoic acid (DHA) rich oil on fasting and postprandial lipid levels in healthy male students. DESIGN: The study was a randomized single-blind study with a control and three study groups. SETTING: The study was carried out in the Departments of Physiology and Clinical Nutrition of University of Kuopio. SUBJECTS: Healthy male volunteers were recruited for the study from the university student population. Fifty-nine subjects entered and 55 completed the study. INTERVENTIONS: For 15 weeks the subjects in the fish diet group ate 4.3 +/- 0.5 fish containing meals per week and those in the fish oil and DHA-oil groups ate 4 g oil per day. Fish diet provided 0.38 +/- 0.04 g eicosapentaenoic acid (EPA) and 0.67 +/- 0.09 g DHA, fish oil 1.33 g EPA and 0.95 g DHA and DHA-oil (EPA-free) 1.68 g DHA per day. RESULTS: Fasting plasma triglyceride levels decreased in all test groups in 14 weeks when compared to the control group (P < 0.05). Total plasma cholesterol levels did not change but the HDL2/HDL3-cholesterol ratio increased in all test groups by over 50% (P < 0.05). The postprandial total and chylomicron triglyceride responses, measured as areas under the response curve, were lowered in 15 weeks by the fish diet and fish oil (P < 0.05), the same tendency (P < 0.1) being seen in DHA-oil group. CONCLUSIONS: These results show that both fasting and postprandial triglyceride concentrations can be decreased with moderate intakes of long-chain n-3 fatty acids either from a fish diet or fish oil and that also pure DHA has a hypotriglyceridemic effect.

Publication Types:

• Clinical Trial
• Randomized Controlled Trial

4.

Incorporation of dietary docosahexaenoic acid into the central nervous system of the yellowtail Seriola quinqueradiata.
Brain Behav Evol 1999;53(4):173-9
Masuda R, Takeuchi T, Tsukamoto K, Sato H, Shimizu K, Imaizumi K.

Ocean Research Institute, University of Tokyo, Nakano, Tokyo, Japan. reiji@compuserve.com

In order to show the involvement of docosahexaenoic acid (DHA) in the development of the central nervous system (CNS) in carangid fish, we conducted tracer experiments by feeding radioactive DHA to larval yellowtail (Seriola quinqueradiata). Artemia nauplii were enriched with 14C-labeled DHA and fed to larval yellowtail for eight or ten days. Autoradiography of frozen sections, using both electric imaging plates and X-ray sensitive film, clearly showed that DHA was incorporated into and retained in the brain, spinal cord, and eyes. The brain, eyes, gill raker, liver, guts, and other muscle and bone structures were dissected, and radioactivity was measured in each organ by liquid scintillation counter. The results of this study suggest the incorporation of DHA into the brain. Considering our previous results indicating that DHA-free fish cannot form schools, we conclude that the incorporation of DHA into the brain might be a critical factor in the ontogeny of schooling behavior