Accumulation of DHA (docosahexaenoic acid;

22:6n-3) in larval and juvenile fish brain

Gabriel Mourente

Departamento de Biología, Facultad de Ciencias del Mar y Ambientales

Universidad de Cádiz, E-11510 Puerto Real, Cádiz, Spain
E-mail: gabriel.mourente@uca.es

Key words: docosahexaenoic acid, brain, accumulation, metabolism, development, marine fish

Abstract
As in mammals, a critical functional role for n-3HUFA (highly unsaturated fatty acids), specifically
docosahexaenoic acid (DHA; 22:6n-3), in neural tissues has been established in larval and juvenile
fish. Accumulation of DHA in brain during development has been demonstrated in several marine
fish species. A very low rate of DHA biosynthesis was observed in turbot brain but a rapid accu-
mulation of DHA in brain of turbot and gilthead sea bream was observed during weaning from
live to pelleted food. The incorporation of [1-14C] linolenic acid (LNA; 18:3n-3) and [1-14C] DHA
in juvenile turbot brain cells showed no significant differences between the amounts of LNA and
DHA incorporated into brain phospholipids demonstrating no preferential uptake and incorpora-
tion of DHA into brain cells. However, during 24h incubation, 1.1% and 8.5% of radioactivity
from [1-14C] LNA and [1-14C] eicosapentaenoic acid (EPA; 20:5n-3), respectively, were recovered
in the DHA fraction of turbot brain lipids. Thus, LNA bioconversion cannot contribute significantly
to brain DHA, whereas EPA can to a greater extent. In a further study, the in vivo metabolism of in-
traperitoneally injected [1-14C] LNA in liver, brain and eyes of juvenile rainbow trout and gilthead
sea bream showed that, although the sea bream incorporated more LNA into its lipids, the biocon-
version of LNA was greater in the trout. The proportion of radioactivity recovered in desaturated/
elongated products was much lower in liver than in brain and eyes in both species, but the recovery
of radioactivity in DHA in brain was significantly higher in trout compared to sea bream. Overall,
although the results did not eliminate a role for liver in the biosynthesis and provision of DHA for
developing neural tissues in fish, they indicated that DHA can be synthesised in fish brain and eye
in vivo. However, they also suggested that the level of DHA in marine fish brain is largely due to
dietary DHA levels than to PUFA bioconversion capabilities. In conclusion, as the DHA in neural
tissues is mainly of dietary origin, irrespective of the metabolic capacity of PUFA bioconversion in
the different tissues and fish species, the adequate supply of n-3HUFA, particularly DHA, during
the early stages of fish is crucial for the normal development of the neural system and to avoid be-
havioural abnormalities (raptorial or schooling behaviour) due to visual and/or neural impairment.
Dietary deprivation of DHA can be particularly deleterious in larvae and juveniles from fast
growing large pelagic marine species such as carangids and tunids.

The Big Fish Bang. Proceedings of the 26th Annual Larval Fish Conference. 2003. Edited by Howard I. Browman and Anne Berit Skiftesvik
Published by the Institute of Marine Research, Postboks 1870 Nordnes, N-5817, Bergen, Norway. ISBN 82-7461-059-8
240 THE BIG FISH BANG

Introduction
The understanding of nutrition during early development in all species of interest is a major
prerequisite to counter the major challenge of marine fish larvae culture and, in particular,
lipid nutrition in marine fish is considered crucial to ensure successful culture. The importance
of lipid biochemistry and metabolism in marine fish has been extensively reviewed, as well as
the requirements and possible roles of particular lipid molecules in marine fish larvae during
development (Sargent et al. 1993b; Watanabe 1993). The abundance of long chain polyunsaturat-
ed fatty acids of the n-3 series (n-3 PUFA), mainly eicosapentaenoic acid (EPA; 20:5(n-3)) and
docosahexaenoic acid (DHA; 22:6(n-3)) in fish oils is only surpassed by the greater abundance
of these fatty acids in the phosphoglycerides of fish cellular membranes. The very high concen-
tration of DHA in fish neural tissues is also well established (Tocher and Harvie 1988; Bell and
Dick 1991).
Marine fish are not capable of bioconverting C18 PUFA to their C20 and C22 homologues,
probably due to a deficiency in the ∆5 fatty acid desaturase and/or the C18-20 elongase (Sargent et
al. 1993a), consequently, marine fish invariably obtain EPA and DHA from their natural diet which
is rich in these particular fatty acids (Sargent et al. 1995).
DHA has a critical role in neural tissues in mammals, and is accumulated by these tissues
during development; nutritional depletion of DHA in the brain and retinal membranes is accom-
panied by functional defects such as reduced visual acuity and impaired learning abilities (Lau-
ritzen et al. 2001). In fish, neural tissue, especially the eyes, can form a relatively large proportion of
the total body mass of the embryo and larvae, so that a large portion of the total DHA required for
embryonic and larval growth is directed towards the formation of cell membranes vital for normal
development and functioning of the visual and neural systems. Therefore, any deficiency in DHA
during embryonic and larval development will have serious consequences for the successful per-
formance of sophisticated predatory larvae, particularly during and immediately following first
feeding (Sargent 1995).
The present paper summarises a series of studies which show the specificity of DHA uptake by
the brain of larvae and juvenile fish, as well as the metabolism of (n-3) PUFA in the brain, eye and
liver clarifying the relative importance of these tissues in the biosynthesis and provision of DHA.

Variations in brain DHA at different stages of development in a wild marine fish species. A study was
carried out to determine the changes in composition of brain lipids and fatty acids in wild Atlantic
herring (Clupea harengus L.) at four different stages of development: larvae at the end of the yolk
sac stage, two different juvenile stages and sexually mature adults. In brain total lipids, saturated
fatty acids generally decreased and monounsaturated fatty acids and dimethyl acetals (derived
from plasmalogens) increased from larvae to adults. However, the proportions of polyunsaturated
fatty acids in total lipids and individual brain phosphoglycerides were generally highest in juvenile
stages, due mainly to increased DHA, and were lowest in adult fish (Figure 1). In conclusion, a sig-
nificant accretion of DHA in brain lipids of Atlantic herring was observed during larval and juve-
nile stages (Mourente and Tocher 1992a).
 A C C U M U L AT I O N OF DHA IN FISH BRAIN 241

Figure 1. Variations of selected total lipid

fatty acids in brain of Atlantic herring
(Clupea harengus L.), including DHA, at
different stages of development. T. DMA,
total dimethyl acetals; EPA, eicosapen-
taenoic acid; DHA, docosahexaenoic acid;
24:1, nervonic acid. Values not bearing the
same superscript are significantly different
(p<0.05). Data taken from Mourente and
Tocher (1992a).

Specific accumulation of DHA in brain lipids during development of juvenile turbot. Juvenile turbot
(Scophthalmus maximus L.) were sampled during a 10-week period immediately following
weaning from live feed to a pelleted diet and the changes in the lipid class and fatty acid composi-
tions of total lipids and individual glycerophospholipids in brain were investigated during these
early developmental stages. The percentage of DHA in brain lipids was low at the beginning of the
study period but a specific and significant accumulation of this fatty acid was observed during de-
velopment. The percentages of DHA increased in total brain lipids, phosphatidylcholine (PC),
phosphatidylethanolamine (PE), phosphatidylserine (PS) and phosphatidylinositol (PI) by 3.2-
fold, 11.7-fold, 2.7-fold, 2.0-fold and 2.1-fold, respectively, by the end of the 10-week period
(Figure 2). The percentages of other n-3 and n-6 PUFAs generally decreased during this period, as did
that of monoenes; the percentages of saturated fatty acids remained relatively constant. This result
clearly demonstrated that the n-3 PUFA composition of the brains of early developing turbot can be
rapidly and markedly influenced by the PUFA composition of the diet (Mourente et al. 1991).

Figure 2. Specific accumula-

tion of DHA in brain lipids
during development of juve-
nile turbot (Scophthalmus
maximus L.). A, variation of
DHA in brain total lipids; B,
variation of DHA in brain
diradyl glycerophosphocholine;
C, variation of DHA in brain
diradyl glycerophosphoetha-
nolamine; D, variation of
DHA in brain phosphati-
dylserine. Data taken from
Mourente et al. (1991).
242 THE BIG FISH BANG

Effects of weaning onto a pelleted diet on brain DHA of developing cultured marine fish. The results
of the above study prompted a further investigation of the changes that might occur in brain DHA
levels during the weaning period. The early larvae of marine fish are generally fed a mixture of live
feeds consisting initially of rotifers Brachionus spp. and consequently of nauplii of the brine shrimp
Artemia. Because both live preys are usually naturally deficient in EPA and specifically DHA, they
have to be enriched with microalgae and/or supplements rich in these fatty acids (Bell 1998). After
growth for a suitable period of time on these live feeds, the small juvenile fish are weaned on to dry
pelleted diets consisting mainly of fish meal and fish oil. In this context, we studied the influence of
the weaning diet alone on brain lipid and fatty acid composition of two marine fish species farmed
in Europe, the turbot Scophthalmus maximus L. and the gilthead sea bream Sparus aurata L. Each
species was maintained on the same dietary regime throughout early development, and fed en-
riched Artemia prior to weaning. At this point, one group of fish was fed a dry pelleted food whereas
the other group remained on enriched Artemia. Both populations were sampled 7 days later and
the brains dissected out and analyzed for lipid
content, lipid class and fatty acid composi-
tions. The brine shrimp metanauplii, although
enriched with fish oil, exhibited low levels of
DHA. As a result, phospholipids from the
brains of fish reared only with enriched
Artemia were rich in EPA but relatively defi-
cient in DHA. In contrast, in fish that had been
weaned on to a fish meal and oil-based diet
rich in DHA, the level of this particular fatty
acid increased rapidly and significantly in their
brain lipids, primarily in PC and PE (Figure 3).
The rapid and specific incorporation of DHA
into brain phosphoglycerides was accompanied
by significant decreases in EPA (Mourente and
Tocher 1992b; 1993).

Metabolism of [1-14C](n-3) PUFA in brain

Figure 3. Effects of weaning onto a pelleted diet on
brain total lipid DHA level in (A) turbot (Scopthalmus
maximus L.) and (B) gilthead sea bream (Sparus
aurata L.). Data taken from Mourente and Tocher
(1992b; 1993).
cells from juvenile turbot. The incorporation of
[1-14C] 18:3(n-3) (LNA) and [1-14C] 22:6(n-
3) (DHA), and the metabolism via the desat-
urase / elongase pathways of [1-14C] 18:3(n-3)
(LNA) and [1-14C] 20:5(n-3) (EPA) were
studied in brain cells from newly-weaned (1-
month-old) and 4-month-old turbot. In this
study we aimed to determine: i) if there was a
differential uptake and incorporation of DHA
into brain cells, ii) the extent to which LNA
and EPA contribute to brain DHA levels and
 A C C U M U L AT I O N OF DHA IN FISH BRAIN 243

iii) if these processes were affected by age and

development of the fish. The rank order of the
extent of net incorporation of both LNA and
DHA into glycerophospholipids was PC > PE
> PS > PI, and was independent of the PUFA
added, the age of the fish and the time of incu-
bation. However, the rate of incorporation of
LNA into total lipid, PC, PE and PS was signif-
icantly greater than the rate of incorporation
of DHA, and there was a significantly greater
amount of DHA incorporated into PE than
LNA. There was no significant difference
between the amounts of LNA and DHA incor-
porated into total lipids, PC, PS and PI. There-
fore, little preferential uptake and incorpora-
tion of DHA into brain cells was apparent.
During 24 hours incubation, on average 1.1%
and 8.5% of radioactivity from [1-14C] LNA
and [1-14C] EPA, respectively, were recovered
in the DHA fraction. Thus, LNA cannot con-
tribute significantly to brain DHA levels in the
turbot whereas EPA can to a greater extent.
There were no significant differences between
the amounts of radioactivity from either [1-
Figure 4. Metabolism of [1-14C](n-3) PUFA in brain
14C] LNA or [1-14C] EPA recovered in the in-
cells from juvenile turbot (Scophthalmus maximus L.).
dividual products / intermediates of the desat- Metabolism via the desaturase/elongase pathway 30
urase / elongase pathways in brain cells from days and 120 days after hatch of (A) [1-14C]18:3(n-3)
30-day-old and 120-day-old turbot (Figure 4). and (B) [1-14C]20:5(n-3). Data taken from Tocher et
These results strongly suggested that the accu- al. (1992).
mulation of DHA in turbot brain during de-
velopment is not due to specific uptake and incorporation of DHA and that endogenous desatura-
tion of LNA cannot contribute to DHA levels in the brain. This confirms our previous conclu-
sions, from studies on lipid and fatty acid compositions of developing turbot brain, that the level
of DHA in the brain was mainly dependent on the level of DHA in the diet (Tocher et al. 1992).
The in vivo incorporation and metabolism of [1-14C] linolenate (18:3n-3) in liver, brain and
eyes of juveniles of rainbow trout Oncorhynchus mykiss L and gilthead sea bream Sparus aurata L.
Accumulation of docosahexaenoic acid (DHA; 22:6n-3) in brain and eyes during development
had been demonstrated in marine fish and significant biosynthesis of DHA from dietary 18:3n-3
was not observed in either liver or neural tissues. However, it was not clear whether this was also
the case in freshwater fish and whether liver or neural tissues themselves are of greater importance in
the biosynthesis of DHA from dietary 18:3n-3. In this study, we investigated the in vivo metabolism of
intraperitoneally injected [1-14C] 18:3n-3 in liver, brain and eyes of young juvenile freshwater and
244 THE BIG FISH BANG

Figure 5. Metabolism via the desaturase elongase
pathway of [1-14C]18:3n-3 in brain of (A) gilthead
sea bream (Sparus aurata L.) and (B) rainbow trout
(Oncorhynchus mykiss L.). Data taken from
Mourente and Tocher (1998).
Figure 6. Metabolism via the desaturase elongase
pathway of [1-14C]18:3n-3 in eye of (A) gilthead sea
bream (Sparus aurata L.) and (B) rainbow trout
(Oncorhynchus mykiss L.). Data taken from
Mourente and Tocher (1998).

marine fish. Metabolism was followed over a 48h time course in order to obtain dynamic information
that could aid the elucidation of the roles of the different tissues in the biosynthesis and provision
of DHA from dietary 18:3n-3. The study was performed in rainbow trout Oncorhynchus mykiss L. as
an example of a freshwater fish, and gilthead sea bream Sparus aurata L., as an example of a marine
fish, to determine the effect that low or limiting ∆5-desaturase activity may have in this process. As
expected, the results showed that although sea bream incorporated more 18:3n-3 into its lipids,
metabolism of the incorporated fatty acid by desaturation and elongation was significantly greater
in the trout. In liver, the percentages of radioactivity recovered in tetraene and pentaene products
were greater in trout than in sea bream although there was no difference in hexaenes. In contrast, the
recovery of radioactivity in DHA was significantly greater in brain of trout compared to sea bream. In
both species, the percentage of radioactivity recovered in desaturated / elongated products was much
 A C C U M U L AT I O N OF DHA IN FISH BRAIN 245

Figure 8. Unmetabolised [1-14C]18:3n-3 recovered in

Figure 7. Metabolism via the desaturase elongase
pathway of [1-14C]18:3n-3 in liver of (A) gilthead sea
bream (Sparus aurata L.) and (B) rainbow trout
(Oncorhynchus mykiss L.). Data taken from
Mourente and Tocher (1998).
brain, eye and liver of (A) gilthead sea bream (Sparus
aurata L.) and (B) rainbow trout (Oncorhynchus
mykiss L.) Percentage of radioactivity from [1-
14C]18:3n-3 recovered unmetabolised as 18:3n-3 or
elongated to 20:3n-3. Data taken from Mourente and
Tocher (1998).

lower in liver than in brain and eyes, but the percentages increased over the 48h time course. In trout,
the highest percentages of desaturated products in brain and eye were observed after 12 and 24h,
respectively. However in sea bream the highest percentages of desaturated products in the neural
tissues were observed after 24-48h (Figures 5, 6, 7, and 8). Overall, although the results could eliminate
a role for liver in the biosynthesis and provision of DHA for developing neural tissues in fish, they
suggest that DHA can be synthesised in fish brain and eye in vivo (Mourente and Tocher 1998).
Role(s) of DHA in the formation of neural tissues and its consequences on performance and
health of marine fish during development and growth.
Marine fish occupy an environment where the ability to see in low light intensities is of consider-
able importance. For this reason they have highly developed systems of rods in their retina. The
abundance of DHA in the rods of higher vertebrates is well known and the fact that more than
246 THE BIG FISH BANG

50% of the total fatty acids in PE from rod outer segment membranes can be DHA has revealed the
presence of di-DHA molecular species of this phosphoglyceride. The molecular species composi-
tion of the phosphoglycerides from the brain and retinas of several fish species were determined
(Bell and Tocher 1989; Bell and Dick 1990; 1991) and results indicated that di-DHA species occurred
in large proportions in phosphoglycerides of brain and retina. Therefore, a correlation was demon-
strated between the appearance of rods in the eyes of herring (Clupea harengus L.) and the increasing
of di-DHA molecular species of phospholipids (Bell and Dick 1993). Furthermore, dietary defi-
ciency of DHA resulted in impaired vision at low light intensities in juvenile herring (Clupea
harengus L.) (Bell et al. 1995), deficits of di-DHA phospholipids in eyes of sea bass (Dicentrarchus
labrax L.) (Bell et al. 1996; Navarro et al. 1997), abnormal pigmentation in flatfish (Estevez and
Kanazawa 1996) and behavioural differences in herring larvae (Navarro and Sargent 1992).
It is also relevant and noteworthy that di-DHA molecular species of phosphoglycerides are
absent from eggs of marine fish (Bell 1989), so that the di-DHA species abundant in neural tissue
must be assembled from pre-existing lipids in the eggs or from dietary lipid ingested by the larvae
during embryogenesis and larval development, respectively (Sargent et al. 1993b).
More recent research has elucidated the critical role of dietary DHA during larval stages as es-
sential for the development of schooling behaviour in carangid fish such as yellowtail (Seriola qui-
queradiata) and the striped jack (Pseudocaranx dentex) (Masuda et al. 1998; Masuda and
Tsukamoto 1999; Masuda et al. 1999; Ishizaki et al. 2001). Furthermore, mortality of northern
bluefin tuna (Thunnus thynnus), resulting from trauma caused by collisions with culture tank
walls (Miyashita et al. 2000; Masuma et al. 2001), is possibly a result of visual disorientation related
to impaired retinomotor responses. Although not as yet demonstrated, it is possible that such
visual impairment is related to dietary DHA deficiency.

Conclusions and Perspectives

Marine fish larvae have an absolute requirement for (n-3) HUFA, particularly DHA; dietary defi-
ciencies of these induce a range of pathologies in fish including malpigmentation and visual and
behavioural abnormalities. Therefore, carnivorous (piscivorous) marine fish have little or no
ability to modify C18 PUFA assimilated from their diets through anabolic reactions in the liver
and other tissues, although they can modify assimilated C22 PUFA by catabolic (chain shortening)
reactions. This limitation might appear to place a requirement for selectivity of uptake of DHA
from blood lipids, enabling the brain and eyes to achieve a PUFA profile from that of circulating
blood lipids derived from the intestine and/or liver. Thus, the ease with which changes in brain PUFA
compositions of these species can be elicited by changing their dietary PUFA composition, leads to
the conclusion that the specificities of the brain fatty acyl transferases incorporating PUFA into
major brain phosphoglycerides are very low. Therefore, the neural and visual tissues of these fishes
are highly vulnerable to changes in circulating PUFA in the blood lipids resulting from changes in
dietary PUFA (Sargent et al. 1993b). These studies emphasize the importance of providing suffi-
ciently high dietary levels of DHA for adequate neural development in species or individuals with
limited ability to elongate and desaturate C18 and C20 PUFA to their C22 homologues.
 A C C U M U L AT I O N OF DHA IN FISH BRAIN 247

References
Bell, J.G. 1998. Current aspects of lipid nutrition in fish farming. p. 114-145. In K. D. Black and A. D. Picker-
ing (Eds.), Biolology of Farmed Fish, Sheffield Academic Press, England.
Bell, M.V. 1989. Molecular species analysis of phosphoglycerides from the ripe roes of cod (Gadus morhua).
Lipids 24: 585-588.
Bell, M.V., Batty, R.S., Dick, J.R., Fretwell, K., Navarro, J.C. and J.R. Sargent. 1995. Dietary deficiency of
docosahexaenoic acid impairs vision at low light intensities in juvenile herring (Clupea harengus L.).
Lipids 30: 443-449.
Bell, M.V. and J.R. Dick. 1990. Molecular species composition of phosphatidylinositol from the brain, retina,
liver and muscle of cod (Gadus morhua). Lipids 25(11): 691-694.
Bell, M.V. and J.R. Dick. 1991. Molecular species composition of the major diacyl glycerophospholipids from
muscle, liver, retina and brain of cod (Gadus morhua). Lipids, 26: 565-573.
Bell, M.V. and J.R. Dick. 1993. The appearance of rods in the eyes of herring and increased di-docosa-
hexaenoyl molecular species of phospholipids. J. Mar. Biol. Ass. U. K. 73: 679-688.
Bell, M.V., McEvoy, L.A. and J.C. Navarro. 1996. Deficit of didocosahexaenoyl phospholipid in the eyes of
larval sea bass fed an essential fatty acid deficient diet. Journal of Fish Biology 49: 941-952.
Bell, M.V. and D.R. Tocher. 1989. Molecular species composition of the major phospholipids in brain and
retina from rainbow trout (Salmo gairdneri). Biochem. J. 264: 909-915.
Estevez, A. and A. Kanazawa. 1996. Fatty acid composition of normally pigmented and unpigmented juveniles
of Japanese Flounder using rotifer and Artemia enriched in n-3 HUFA. Fisheries Science 62(1): 88-93.
Ishizaki, Y., Masuda, R., Uematsu, K., Shimizu, K., Arimoto, M. and T. Takeuchi. 2001. The effect od dietary
docosahexaenoic acid on schooling behaviour and brain development in larval yellwotail. Journal of Fish
Biology 58: 1691-1703.
Lauritzen, L., Hansen, H.S., Jrrgensen, M.H. and K.F. Michelsen. 2001. The essentiality of n-3 fatty acids in re-
lation to development and function of the brain and retina. Progress in Lipid Research 40: 1-94.
Masuda, R., Takeuchi, T., Tsukamoto, K., Ishizaki, Y., Kanematsu, M. and K. Imaizumi. 1998. Critical invol-
ment of dietary docosahexaenoic acid in the ontogeny of shooling behaviour in the yellowtail. Journal of
Fish Biology 53: 471-484.
Masuda, R. and K. Tsukamoto. 1999. School formation and concurrent developmental changes in carangid
fish with reference to dietary conditions. Environmental Biology of Fishes 56: 234-252.
Masuda, R., Takeuchi, T., Tsukamoto, K., Sato, H., Shimizu, K., and K. Imaizumi. 1999. Incorporation of
dietary docosaheaenoic acid in the central nervous system of the yellowtail Seriola quiqueradiata. Brain,
Behaviour and Evolution 53: 173-179.
Masuma, S., Kawamura, G., Tezuka, N., Koiso, M. and K. Namba. 2001. Retinomotor responses of juvenile
bluefin tuna Thunnus thynnus. Fisheries Science 67: 228-231.
Miyashita, S., Sawada, Y., Hattori, N., Nakatsukasa, H., Okada, T., Murata, O. and H. Kumai. 2000. Mortality
of norther bluefin tuna Thunnus thynnus due to trauma caused by collision during growout culture.
Journal of the World Aquaculture Society 31(4): 632-639.
Mourente, G. and D.R. Tocher. 1992a. Lipid class and fatty acid composition of brain lipids from Atlantic
herring (Clupea harengus) at different stages of development. Marine Biology, 112: 553-558.
Mourente, G. and D.R. Tocher. 1992b. Effects of weaning onto a pelleted diet on docosahexaenoic acid (22:6n-
3) levels in brain of developing turbot (Scophthalmus maximus L.). Aquaculture, 105: 363-377.
Mourente, G. and D.R. Tocher. 1993. The effects of weaning on to a pelleted diet on brain lipid and fatty acid
composition in post-larval gilthead sea bream (Sparus aurata L.). Comp. Biochem. Physiol., 104A(3):
605-611.
Mourente, G. and D.R. Tocher. 1998. The in vivo incorporation and metabolism of [1-14C] linolenate (18:3n-
3) in liver, brain and eyes of juveniles of rainbow trout Oncorhynchus mykiss L, and gilthead sea bream
Sparus aurata L. Fish Physiology and Biochemistry 18: 149-165.
248 THE BIG FISH BANG

Mourente, G., Tocher, D.R. and J.R. Sargent. 1991. Specific accumulation of docosahexaenoic acid (22:6n-3)
in brain lipids during development of juvenile turbot Scophthalmus maximus L. Lipids, 26(11): 871-877.
Navarro, J.C. and J.R. Sargent. 1992. Behavioural differences in starving herring Clupea harengus L. Larvae
correlate with body levels of essential fatty acids. Journal of Fish Biology 41: 509-513.
Navarro, J.C., McEvoy, L.A., Bell, M.V., Amat, F., Hontoria, F. and J.R. Sargent. 1997. Effect of different dietary
levels of docosahexaenoic acid (DHA, 22:6n-3) on the DHA composition of lipid classes in sea bass larvae
eyes. Aquaculture International 5: 509-516.
Sargent, J.R. 1995. Origins and functions of lipids in fish eggs: nutritional implications, p. 353-372. In N. R.
Bromage and R. R. Roberts (Eds.), Broodstock Management and Egg and Larval Quality, Blackwell,
Oxford.
Sargent, J.R., Bell, J.G., Bell, M.V., Henderson, R.J. and D.R. Tocher. 1993a. The metabolism of phospholipids
and polyunsaturated fatty acids in fish, v: 43, p. 103-124. In B. Lahlou and P. Vitiello (Eds.), Aquaculture:
Fundamental and Applied Research, Coastal and Estuarine Studies. American Geophysical Union, Wash-
ington DC.
Sargent, J.R., Bell, M.V. and D.R. Tocher. 1993b. Docosahexaenoic acid and the development of brain and
retina in marine fish. p. 139-149. In C. A. Drevon, I. Baksaas and H. E. Krokan (Eds.), Omega-3 Fatty
Acids: Metabolism and Biological Effects. Birkhäuser Verlag Basel, Switzerland.
Sargent, J.R., Bell, J.G., Bell, M.V., Henderson, R.J. and D.R. Tocher. 1995. Origin and functions of n-3 polyun-
saturated fatty acids in marine organisms. p. 248-259.In G. Ceve and F. Paltauf (Eds.), Phospholipids:
Characterization, Metabolism and Novel Biological Applications Am. Oil Chem. Soc. Press, Champaign,
Ill, USA.
Tocher, D.R. and D.G. Harvie. 1988. Fatty acid compositions of the major phosphoglycerides from fish neural
tissues: (n-3) and (n-6) polyunsaturated fatty acids in rainbow trout (Salmo gairdneri) and cod (Gadus
morhua) brains and retinas. Fish Physiology and Biochemistry 5: 229-239.
Tocher, D.R., Mourente, G. and J.R. Sargent. 1992. Metabolism of [1-14C] docosahexaenoate (22:6n-3), [1-14C]
eicosapentaenoate (20:5n-3) and [1-14C] linolenate (18:3n-3) in brain cells from juvenile turbot Scoph-
thalmus maximus. Lipids 27: 494-499.
Watanabe, T. 1993. Importance of Docosahexaenoic Acid in Marine Larval Fish. J. World Aquacult. Soc. 24(2):
152-161.