The phosphoglycerides that constitute cell membrane bilayers in fish generally contain 16:0, 18:1n-9, 20:5n-3, and 22:6n-3 as their principal fatty acids. The former two are located preferentially in thesn-1 position of the glycerol backbone of the phosphoglycerides, the latter two preferentially in thesn-2 position. Additionally, 22:6n-3 is generally present at about twice the level of 20:5n-3 in fish phospholipids. However, that generalization must be qualified with respect to phosphoglyceride classes and also with respect to
particular tissues. The following account rests mainly on extensive studies, es- pecially of molecular species compositions of phosphoglycerides, in our own laboratory (Bell and Tocher, 1989; Bell and Dick, 1990, 1991, 1993a, b; M. V.
Bellet al.,1997). First, with respect to phosphoglyceride classes, the highest levels of 22:6n-3 are usually contained in phosphatidylethanolamine (which includes the plasmalogens, i.e., alk-1-enyl/acyl as distinct from acyl/acyl phosphoglycerides), with 16:0/22:6n-3, 18:0/22:6n-3, and 18:1n-9/22:6n-3 molecular species all being present. Phosphatidylserine is also rich in 22:6 n-3, with 18:0/22:6n-3 being an abundant molecular species. Phosphatidyl- choline commonly has the lowest levels of 22:6n-3, in fish tissues, being rich instead in 16:0 and 18:1n-9, and appears to be more easily influenced by dietary fatty acids than the other phosphoglycerides. A special situation exists for phosphatidylinositol, which has the highest level of arachidonic acid, 20:4n-6, of all the fish phosphoglycerides and is also rich in 18:0.
Indeed thesn-2 position of fish phosphoinositides has a distinct preference for C20PUFA. Thus, high levels of 20:5n-3 can also be found in phosphatidyli- nositol, its fatty acid composition can be readily influenced by changes in dietary levels of both 20:4n-6 and 20:5n-3, and it can also accumulate 20:3n-6.
Second, with respect to variations in the fatty acid compositions of phospho- glycerides in fish tissues, particularly high levels of 22:6n-3 exist in neural tissues, whether brain or eyes, to the extent that di-22:6n-3 species of both phosphatidylserine and phosphatidylethanolamine can account for up to 60 and 72%, respectively, of the total molecular species of these phospho- glycerides. Fish sperm can also contain high levels of dipolyunsaturated phosphoglycerides (M. V. Bellet al.,1997). As discussed in Section 4.5, the abundance of 22:6n-3 in neural tissue has particular implications for fish larval nutrition.
The precise reason(s) for the preponderance of long-chainn-3 PUFA, specifically 20:5n-3 and, especially, 22:6n-3, in fish phosphoglycerides and, above all, the remarkable preponderance of 22:6n-3 in neural tissue, which occurs in vertebrates generally, albeit not to the same extent as in fish, remains elusive. Traditionally it was believed that the preponderance of 22:6n-3 in fish tissues was an adaptation to low temperatures to maintain membrane fluidity (homeoviscous regulation). However, the abundance of 22:6n-3 in both tropical species such as the warm-blooded tuna and polar species such as the cod belies that notion. Indeed, it is now recognized that the main determinant of fluidity in phosphoglycerides is the ratio of mo- nounsaturated to saturated fatty acids, which is increased and decreased, respectively, in response to lowered and elevated environmental tempera- tures (Wodtke and Cossins, 1991). More sophisticated and credible expla- nations for the abundance of 22:6n-3 in the phosphoglycerides of fish (and indeed of marine organisms in general) start with the recognition that
22:6n-3 is unique among fatty acids in having the maximum number of cis-methylene-interrupted double bonds possible in a C22fatty acid. As such it has a unique conformation dictated by a helical twist, or an “angle iron”
shape, induced by its cis-methylene interrupted dienes that results in the molecule assuming a squat, compact form, with an overall length similar to that of 16:0 (Applegate and Glomsett, 1986). Such a structure favors the formation of the hexagonal (inverted micellar) phase rather than the conventional bilayer phase in phosphoglycerides, above all in di-22:6n-3- phosphoglycerides containing small head groups such as phosphatidylser- ine and phosphatidyl ethanolamine. Therefore, bilayers containing 22:6 n-3 phosphoglycerides, and above all di-22:6n-3-phosphatidylserine and di- 22:6n-3-phosphatidylethanolamine, will be “energized” by the tendency of these molecules to form hexagonal phases, and this will facilitate conforma- tional changes, especially very fast conformational changes, in membrane proteins that might otherwise be energetically unfavorable (Brown, 1994).
Such changes are particularly important in visual and neuromuscular pro- cesses where di-22:6n-3-phosphoglycerides are particularly abundant. More- over, the intrinsic structure of 22:6n-3 is inherently resistant to temperature and pressure changes, such that its effects continue to be exerted essentially independently of these environmental variables (Rabinovich and Ripatti, 1991). The advantages of such properties in the aquatic and, especially, the marine environment are self-evident and may well have been exploited evolutionary by the single-cell, motile and phototactic organisms (the flag- ellates) that produce 22:6n-3 at the base of the marine food web and from which the bulk, if not all, of the 22:6n-3 in the marine ecosystem is ulti- mately derived (Sargentet al.,1995c). In this context, the sperm cell may be considered as a chemotactic rather than a phototactic flagellate. Such considerations, though speculative, are well consistent with more metaboli- cally specific roles of 22:6n-3, e.g., its particular role in Ca2+-ATPase activity in the sarcoplasmic reticulum of heart and skeletal muscle, which has long been established in higher mammals and more recently established in fish (Ushioet al.,1997).
As described in previous reviews (Henderson and Tocher, 1987; Sargent et al.,1993b, 1995b; Sargent, 1995a; Tocher, 1995), a major role for the C20
PUFA, especially 20:4n-6, is as precursors for the group of highly biologically active compounds, the eicosanoids which are C20derivatives of C20PUFA.
These include cyclic compounds, comprised inter alia of prostaglandins, prostacyclins, and thromboxanes and formed initially by the action of cy- clooxygenase on C20PUFA, and linear compounds, including the leukotri- enes and lipoxins, formed initially by the action of lipoxygenases on C20
PUFA. These enzymes act on free C20PUFA released from plasma membrane phosphoglycerides by the action of phospholipase A2. The eicosanoids are
autocrines, i.e., hormone-like compounds produced by cells to act in their immediate vicinity with a short half-life. Virtually every tissue in the body produces eicosanoids and they have a wide range of physiological actions, e.g., in blood clotting, the immune response, the inflammatory response, cardiovascular tone, renal function, neural function, and reproduction.
Eicosanoid production is associated very broadly with stressful situations and is a normal physiological process, with excess eicosanoid production often occurring in pathological conditions. In higher terrestrial mammals 20:4n-6 is the chief precursor of the eicosanoids, generating 2-series pros- tanoids and 4-series leukotrienes. However, 20:5n-3 competitively interferes with eicosanoid production from 20:4n-6 catalyzed by cyclooxygenase and lipoxygenases, and is itself converted to 3-series prostanoids and 5-series leukotrienes, which are generally much less biologically active than the cor- responding 2-series prostanoids and 4-series leukotrienes produced from 20:4n-6 (Fig. 4.9). Thus, eicosanoid actions are determined by the ratio of 20:4n-6 to 20:5n-3 in cellular membranes, this in turn being determined by the dietary intake ofn-6 andn-3 PUFA. There is now a large body of evidence supporting the notion that high incidences of cardiovascular and inflammatory conditions and cancers in developed societies are associated
20:4n-6 20:5n-3
Cyclo-oxygenase or Lipoxygenase
2-series prostanoids 3-series prostanoids
4-series leukotrienes 5-series leukotrienes
HIGH biological activity LOW biological activity
FIG. 4.9
Arachidonic acid, 20:4n-6, and eicosapentaenoic acid, 20:5n-3, compete with the same cyclooxygenase or the same lipoxygenases to produce 2-series prostanoids/4-series leukotrienes and 3-series prostanoids/5-series leukotrienes, respectively. Therefore, the ratio of 20:4n-6 to 20:5n-3 determines the ratio of high-activity to low-activity eicosanoids.
with an over high dietary intake of 18:2n-6 relative to 18:3n-3, which gen- erates over high levels of 20:4n-6 in cells and thence over high levels of eicosanoids (Anonymous 1992, 1994a; Okoyumaet al.,1997). Dietary sup- plementation with 20:5n-3, e.g., as fish oil or fish oil concentrates, can be beneficial under these conditions by damping down excess eicosanoid production from 20:4n-6 (Anonymous, 1992, 1994a, 1999).
With the exception of the phosphoinositides, the cellular phosphogly- cerides of most species of fish, especially marine fish, have a large excess of 20:5n-3 relative to 20:4n-6. Despite this, 20:4n-6 is the chief source of biologically active eicosanoids in fish, where it is concentrated in phospho- inositides. It is tempting to conclude that the phosphoinositides are the origin of the 20:4n-6 used to produce eicosanoids in fish but there is no clear-cut experimental evidence for this conclusion. It has been established in fish, as in mammals, that 20:5n-3 and also 20:3n-6 (dihomo-γ-linolenic acid) competitively depress the production of eicosanoids from 20:4n-6 (Bell et al.,1994a), as does 20:4n-3 (Ghioni, Tocher, and Sargent, unpublished data), which can be produced by elongation of stearidonic acid, 18:4n-3, which is a significant constituent of many fish oils. Therefore, in fish, as in mammals, eicosanoid production is influenced by the cellular ratio of 20:4n-6 to 20:5n-3, although, clearly, the optimal ratio of 20:4n-6 to 20:5n-3 for eicosanoid production is much lower in fish than in mammals. It remains the case, however, that an imbalanced ratio of 20:4n-6 to 20:5n-3 can be as damaging in fish as in mammals, as discussed later for marine fish larvae (Section 4.5).
The importance of 22:6n-3 in fish neural tissue, evidenced by the abun- dance of di-22:6n-3 species of phosphatidylethanolamine and phos- phatidylserine in brain and eye, has been noted above, and, as discussed later, provision of an adequate dietary supply of 22:6n-3 is particularly im- portant in marine fish larval nutrition for normal development of neural and visual functions. The fatty acid 22:6n-3 is no less important in this re- spect in human neural tissue and, therefore, in prenatal and early postnatal life when development of the neural and visual systems take place. There is now clear cut evidence that suboptimal provision of 22:6n-3 during prena- tal and early postnatal life can generate visual and mental subnormalities in infants (Anonymous, 1994b). Indeed, there is accumulating evidence that a range of psychiatric disorders including schizophrenia may involve ab- normalities in the metabolism of brain phospholipids and their associated PUFA, 22:6n-3, 20:5n-3 and 20:4n-6 (Horrobinet al.,1999; Horrocks and Yeo, 1999; Okuyamaet al.,1997; Puriet al.,1999; Peetet al.,1999). There is also accumulating evidence that dietary supplementation with fish oils and fish oil concentrates can be beneficial in various psychiatric disorders (Horrocks and Yeo, 1999; Puriet al.,1999).
These beneficial effects of long-chainn-3 PUFA in a range of important human disorders stem basically from a dietary excess ofn-6 PUFA, specifi- cally 18:2n-6, which is a major component of the vegetable seed oils whose production is rapidly increasing on a global scale, relative to 18:3n-3, whose chief dietary source for man is green leaf vegetables. The problem can be expressed alternatively as a relative deficiency of 18:3n-3, which can most effectively be reversed by providing the biologically active end productn-3 PUFA, 20:5n-3 and 22:6n-3. Fish are far and away the greatest providers of 20:5n-3 and 22:6n-3 in the human diet, so that fish have a critical role to play in human nutrition. Fish nutrition is concerned with both optimizing the health and welfare of the farmed animal and optimizing its nutritional value for the consumer. Therefore, a major concern of fish nutrition is to generate an end product with high levels of health-promoting 20:5n-3 and 22:6n-3 for the consumer. These HUFA have unique, important, and paral- lel roles to play in both fish and human nutrition and it is natural that they feature prominently in fish lipid nutrition.
4.4