Marta I. Aveldaño

Rapid production of diacylglycerols enriched in arachidonate and stearate during early brain ischemia

AMONG the rapid neurochemical reactions triggered in the mammalian brain at the onset of. ischemia, a strikingly active production of free ratty acids (FFA) has been described in the rat (BAZAN, 1970; BAZAN et al., 1971), monkey (BAZAN, 1971a) and mouse brain (AVELDAI~O & BAZAN, 1975) which was ascribed to polar lipid deacylation. This phcnomcnon was found to be unique to the mature homeothcrmic brain (BAZAN, 1971b; BAZAN et ul., 1971 ; AVELDAGO & BAZAN, 1975) being accompanied by a partial triacylglycerol breakdown. The measurement of diacylglycerols during ischemia was undertaken to estimate the actual contribution of triglycerides to the FFA, since the decreases undcrgoing these neutral lipids might be accompanied by the lost, not only of FFA but also of lower glycerides. The present communication demonstrate that within a short interval after decapitation, there occurs in the mouse brain a rapid and concomitant production of diacylglycerols and FFA, both surpassing in magnitude the decreases undcrgoing triacylglyccrols. Moreover it is shown that free arachidonic acid and arachidonoyl-diglycerides, the most rapidly released lipids, do not originate in triacylglyccrols.

Apoptosis of Retinal Photoreceptors During Development In Vitro: Protective Effect of Docosahexaenoic Acid

Abstract: When rat retinal cells are cultured in a serum‐free medium, the photoreceptor cells start dying after 7 days. The addition of docosahexaenoic acid (DHA) to the cultures prevents the selective death of photoreceptors. Here it is shown that, unlike other retinal neurons, photoreceptors die through an apoptotic pathway. Hallmarks of apoptosis, such as nuclear fragmentation and condensation and DNA cleavage forming a ladder pattern on an agarose gel, were observed. The timing and high selectivity of the triggering of photoreceptor cell apoptosis suggest the existence of a programmed cell death. Compared with other fatty acids, DHA not only was the most effective in promoting photoreceptor survival, but also the only one to decrease the number of apoptotic nuclei. The results suggest that DHA is important among the factors preventing apoptosis of photoreceptors in the developing retina. A limitation in the availability of this fatty acid might trigger apoptosis as a result of the failure to develop functional photoreceptor outer segments.

Docosahexaenoic acid is required for the survival of rat retinal photoreceptors in vitro

Abstract: The effect of docosahexaenoic acid (DHA) on neuronal survival was studied in cultured cells isolated from newborn rat retina. In vivo, the content of DHA in the retina increased nearly fourfold from days 2 to 12 after birth, whereas in retinal cells in culture it remained constant. Unlike amacrine cells, the photoreceptor cells in control cultures underwent a selective degeneration, starting at day 7, that led to their massive death by day 11. The addition of DHA at day 7 led to its active incorporation by the cultures, increasing from 6 to 21% of total fatty acids in cell lipids, and completely prevented photo‐receptor cell death. When other fatty acids were tested, both neuronal fatty acid composition and photoreceptor death were the same as in control cultures. These results indicate that DHA is specifically required for the survival of retinal photoreceptors.

Very Long-chain Polyunsaturated Fatty Acids Are the Major Acyl Groups of Sphingomyelins and Ceramides in the Head of Mammalian Spermatozoa

Very long-chain (C24 to C34) polyunsaturated fatty acids (VLCPUFA) are important constituents of sphingomyelin (SM) and ceramide (Cer) in testicular germ cells. In the present paper we focused on the SM and Cer and their fatty acids in spermatozoa and their main regions, heads and tails. In bull and ram spermatozoa, SM was the third most abundant phospholipid and VLCPUFA were the major acyl groups (∼70%) of SM and Cer. In rat epididymal spermatozoa the SM/Cer ratio was low in the absence of and could be maintained high in the presence of the cation chelator EDTA, added to the medium used for sperm isolation. This fact points to the occurrence of an active divalent cation-dependent sphingomyelinase. Bull and rat sperm had an uneven head-tail distribution of phospholipid, with virtually all the VLCPUFA-rich SM located at the head, the lower SM content in the rat being determined by the lower sperm head/tail size ratio. Most of the SM from bull sperm heads was readily solubilized with 1% Triton X-100 at 4 °C. The detergent-soluble SM fraction was richer in VLCPUFA than the nonsoluble fraction and richer in saturated fatty acids. Cer was produced at the expense of SM, thus decreasing severalfold the SM/Cer ratio in rat spermatozoa incubated for 2 h in presence of the sperm-capacitating agents, calcium, bicarbonate, and albumin. The generation of Cer from SM in the sperm head surface may be an early step among the biochemical and biophysical changes known to take place in the spermatozoon in the physiological events preceding fertilization.

Ceramides and sphingomyelins with high proportions of very long-chain polyunsaturated fatty acids in mammalian germ cells.

Very long-chain polyunsaturated fatty acids (VLCPUFA) have previously been shown to be components of sphingomyelin (SM) of mammalian testis and spermatozoa. Here we examined the fatty acids of testicular ceramide (Cer) in comparison with those of SM in some mammals with a special focus on the rat testis. In bull, cat, dog, rabbit, mouse, and rat, VLCPUFA were found in both testicular lipids, Cer having a higher percentage of VLCPUFA than SM. Rat testis had the highest percentage of VLCPUFA in both lipids, the major ones being 28:4n-6 and 30:5n-6. VLCPUFA-containing SM and Cer occurred in cells located in the seminiferous tubules, where germ cells had a higher percentage of these species than Sertoli cells. Seminiferous tubule fractionation showed that SM and Cer of mitochondria and lysosomes had mostly saturates and negligible VLCPUFA, the latter being important in the SM and Cer of microsomes and other membrane fractions. VLCPUFA were absent from the SM and Cer of rat prepuberal testis, increased with the onset of spermatogenesis to account for nearly 15 and 40% of the total fatty acids of testicular SM and Cer, respectively, remained at those levels throughout the adult life of fertile rats and tended to decrease at advanced ages. Four conditions that lead to selective death of germ cells in vivo, namely experimental cryptorchidism, post-ischemic reperfusion, focalized x-ray irradiation and treatments with the antineoplasic drug doxorubicin, caused the VLCPUFA to disappear from the testicular SM and Cer of adult fertile rats, showing that these lipids are specific traits of spermatogenic cells.

Lipid and fatty acid composition of canine lipoproteins.

Lipid classes and their fatty acids were studied in the major lipoprotein fractions from canine, in comparison with human, plasma. In dogs, high-density-lipoprotein (HDL), the main carrier of plasma phospholipid (PL), cholesterol ester (CE) and free cholesterol, was the most abundant lipoprotein, followed by low and very-low density lipoproteins (LDL and VLDL). Notably, LDL and VLDL contributed similarly to the total dog plasma triacylglycerol (TG). The PL composition was similar in all three lipoproteins, dominated by phosphatidylcholine (PC). Even though the content and composition of lipids within and among lipoproteins differed markedly between dog and man, the total amount of circulating lipid was similar. All canine lipoproteins were relatively richer than those from humans in long-chain (C20-C22) n-6 and n-3 polyunsaturated fatty acids (PUFA) but had comparable proportions of total saturated and monoenoic fatty acids, with 18:2n-6 being the main PUFA in both mammals. The fatty acid profile of canine and human lipoproteins differed because they had distinct proportions of their major lipids. There were more n-3 and n-6 long-chain PUFA in canine than in human plasma, because dogs had more HDL, their HDL had more PC and CE, and both these lipids were richer in such PUFA.

Effects of aging on the composition and metabolism of docosahexaenoate-containing lipids of retina

The amount of docosahexaenoate (22∶6n−3)-containing phospholipid species decreases with aging in the rat retina. Most lipids, but especially choline and serine glycerophospholipids, show a significant fall in 22∶6n−3, which is not compensated by increases in other polyenoic fatty acids. The decrease not only affects 22∶6 but also various very long chain n−3 hexaenoic fatty acids which, in phosphatidylcholine, have up to 36 carbon atoms, and which are probably synthesized by successive elongations of 22∶6n−3. The in vitro incorporation of [2-3H] glycerol into retinal lipids indicates that the de novo biosynthetic pathways are not impaired by aging. The incorporation of [1-14C]docosahexaenoate is significantly stimulated into all lipids of aged retinas, but to the largest extent in those showing the largest decreases in 22∶6, especially in choline glycerophospholipids. The results indicate that the decreased levels of 22∶6 with aging are due not to an impaired activity of the enzymes involved in the synthesis and turnover of phospholipids but to a decreased availability of this polyene in the retina. It is suggested that this may stem from a defect in some of the enzymatic steps that lead to the synthesis of 22∶6n−3, probably that catalyzed by Δ4 desaturase, the effect on longer hexaenes being secondary to the decreased synthesis of 22∶6.

Fatty acid composition and level of diacylglycerols and phosphoglycerides in brain and retina

Abstract The fatty acid composition of diacylglycerols of toad brain and retina is reported. In addition, the fatty acid composition of brain and retina phospholipids and of diacylglycerols and phospholipids from choroid is also presented. Retina diacylglycerol content is 7-fold that of the brain; fatty acid acyl groups are also more unsaturated in the former tissue. The diacylglycerols of the retina contain an unusually high amount of docosahexaenoate (42%), while arachidonate represents about 7%. Other major fatty acids are palmitate (19%) and stearate (20%). Brain diacylglycerols contain only 3% docosahexaenoate, while arachidonate represents about 27%. Other components are stearate (24%), palmitate (17%) and oleate (14%). In the retina, the fatty acid composition of phospholipids resembled that of diacylglycerols, and the total unsaturated components were greater than in diacylglycerols. The ratio docosahexaenoate to arachidonate in retina phospholipids and diacylglycerols was similar. On the contrary, the fatty acid composition in the phospholipids from brain and choroid was unlike that observed in their diacylglycerols. It is concluded that the unique characteristics of retina diacylglycerols may be related to a lipid metabolic differentiation developed to sustain the polar lipids of the photoreceptor cell membranes during their renewal process.

Displacement into incubation medium by albumin of highly unsaturated retina free fatty acids arising from membrane lipids

The concentration, composition and metabolic regulation of the free acid (FFA) pool in tissues other than the adipose are not well known. However, it has been reported that many cellular processes are affected by or are dependent upon the components of the FFA pool. FFAs, among other effects are potent uncouplers of the oxidative phosphorylation from the respirat0.v chain (for references see [ 1 and 21; they are necessary as precursors of prostaglandins [3,4] ; they may be involved in the onset of irreversible brain damage caused by ischemia [S, 61; they inhibit Na-K-dependent ATP-ase in brain [7] and they are inhibitors of key glycolytic enzymes (for references see [l] . Certain aspects of the intrinsic FFAs have been explored but only in a few tissues, e.g., in Ehrlich ascites tumor cells [S, 93 , human platelets [lo], cultured cells [ 1 l] and bran [ 1, 5, 6, 12, 131. Bovine retina neutral [ 141 and polar-lipids [ 151 are richly in long-chain polyenoic fatty acids. FFAs are a relatively small and labile pool, also composed of a large proportion of polyunsaturated constituents. This letter reports remarkable features of endogenously produced retina FFAs, showing that they arise from membrane lipids, and that a different behavior is apparent in the rate of release of individual FFA from the tissue in vitro. The present data may be of

Differentiation-related changes in lipid classes with long-chain and very long-chain polyenoic fatty acids in rat spermatogenic cells

In rat seminiferous tubules (ST), cells that contain polar and neutral lipids with long-chain polyenoic fatty acids (PUFA) and sphingomyelins (SM) and ceramides (Cer) with very long chain (VLC) PUFA of the n-6 series coexist. In this study, pachytene spermatocytes and round spermatids were isolated to determine how these lipids change during spermatogenesis. As the amount per cell of PUFA-rich glycerophospholipids (GPL) decreased with cell size, the 22:5/20:4 ratio increased with cell differentiation. The elovl2 and elovl5 genes, required for 22:5 formation, were expressed (mRNA) in both cell types. Residual bodies- particles with compacted organelles and materials discarded from late spermatids-concentrated cholesterol, 22:5-rich triacylglycerols, and GPL, including plasmalogens and phosphatidylserine. Species of SM and Cer with nonhydroxylated (n-) VLCPUFA (28:4, 30:5, and 32:5) predominated in pachytene spermatocytes, whereas species with the corresponding 2-hydroxy (2-OH) VLCPUFA prevailed in round spermatids. Thus, a dramatic increase in the 2-OH/n-VLCPUFA ratio in SM and Cer was a hallmark of differentiation. A substantial decrease of 2-OH SM occurred between spermatids and mature spermatozoa and 2-OH SM species were collected in residual bodies “en route” to Sertoli cells. Notably, spermatids and spermatozoa gained a significant amount of ceramides devoid of n-VLCPUFA but having 2-OH VLCPUFA as their main fatty acids.