Characterization of a Na∶K∶2Cl cotransport system in the apical membrane of a renal epithelial cell line (LLC-PK1)

Summary ⁸⁶Rb uptake into LLC-PK₁ cells (an established renal epithelial cell line) was found to be comprised of an active ouabain-sensitive component, a loop diuretic-sensitive component which was passive and strictly dependent upon the presence of extracellular Na⁺ and Cl^– for activity, and a leak component. The diuretic-sensitive component of influx was investigated further in apical membrane vesicles derived from these cells. A large fraction of⁸⁶Rb,²²Na and³⁶Cl flux into these vesicles was sensitive to inhibition by furosemide and dependent upon the presence of the other two co-ions, in keeping with the presence of a loop diuretic-sensitive Na⁺K⁺Cl^– cotransport system. The kinetic parameters for Na⁺ and K⁺ interaction have been analyzed under initial linear zerotrans conditions. The following values were obtained:K _mNa+=0.42±0.05 mmol/liter,V _max=303±24 pmol/mg/6 sec;K _mK+=11.9±1.0 mmol/liter,V _maxK+=307±27 pmol/mg/6 sec. For Cl^– interaction evidence for two cooperative binding sites with different affinities and different specificities were obtained. Thus, a stoichiometry of 1Na⁺1K⁺2Cl^– can be calculated. It is concluded that the apical membrane of LLC-PK₁ cells contains a Na⁺K⁺2Cl^– cotransport system with properties similar to those described for the thick ascending limb of the loop of Henle.     

Cellular fatty acid composition of six fastidious,gramnegative, xylem-limited bacteria from plants

Composition of cellular fatty acids was determined for strains of fastidious, Gramnegative, xylem-limited bacteria causing or associated with Pierce's disease of grapevine, phony disease of peach, plum leaf scorch, stunt of ragweed, elm leaf scorch, and periwinkle wilt. The most abundant fatty acids were straight-chain 150, 160, 170, and 180, unsaturated 161, 181, and unsaturated 17-and 19=carbon homologs. Minor fatty acids included straightchain 120, 140, 190, and 200; an unsaturated 15-carbon homolog; hydroxy-substituted 2-OH 120, 3-OH 120, and 3-OH 140; and branched chain iso-140 and iso-200. Cyclopropane acids were not detected. Physiological age had no effect on fatty acid composition. Class analysis of data indicated relative uniformity within the group. Saturated even-carbon chains comprised 31%–42%, unsaturated acids 41%–52%, saturated odd-carbon chains 10%–18%, hydroxysubstituted chains 2%–7%, and branched-chains 1%–4% of total fatty acids. The ratio of saturated-unsaturated acids ranged from 0.8 to 1.2.     

Conformational dynamics in mixed alpha/beta-oligonucleotides containing polarity reversals: a molecular dynamics study using time-averaged restraints

Nucleic acid duplexes featuring a single alpha-anomeric thymidine inserted into each DNA strand via 3-3 and 5-5 phosphodiester linkages exhibit local conformational dynamics that are not adequately depicted by conventional restrained molecular dynamics (rMD) methods. We have used molecular dynamics with time-averaged NMR restraints (MDtar) to explore its applicability to describing the conformational dynamics of two -containing duplexes – d(GCGAAT-3-3-T-5-5-CGC)₂ and d(ATGG-3-3-T-5-5-GCTC)r(gagcaccau). In contrast to rMD, enforcing NOE-based distance restraints over a period of time in MDtar rather than instantaneously results in better agreement with the experimental NOE and J-data. This conclusion is based on the dramatic decreases in average distance and coupling constant violations (d _av, J _rms, and J _av) and improvements in sixth-root R-factors (R ^x). In both duplexes, the deoxyribose ring puckering behavior predicted independently by pseudorotation analysis is portrayed remarkably well using this approach compared to rMD. This indicates that the local dynamic behavior is encoded within the NOE data, although this is not obvious from the local R ^x values. In both systems, the backbone torsion angles comprising the 3-3 linkage as well as the (high S-) sugars of the -nucleotide and preceding residue (–1) are relatively static, while the conformations of the 5-5 linkage and the sugar in the neighboring -nucleotide (+1) show enhanced flexibility. To reduce the large ensembles generated by MDtar to more manageable clusters we utilized the PDQPRO program. The resulting PDQPRO clusters (in both cases, 13 structures and associated probabilities extracted from a pool of 300 structures) adequately represent the structural and dynamic characteristics predicted by the experimental data.     

Nutrient proportions in foliage of semi-mature loblolly pine

Summary Nitrogen and phosphorus limitations to growth are common in many loblolly pine (Pinus taeda) stands. Interactions of these nutrients may complicate interpretation of foliar nutrient analysis for predicting response to forest fertilization. Proportions of foliar nutrient concentrations (and the changes in these proportions following fertilization) were examined in 36 semi-mature loblolly pine plantations in the southeastern United States. Mean proportions of nutrient concentrations (NPKCaMg) for non-fertilized stands were 1009.336.517.29.2. Potassium and phosphorus were higher. Nitrogen fertilization generally decreased the PN ratio and enhanced growth, indicating a nitrogen deficiency in most stands under study. Additions of nitrogen and phosphorus together yielded a significant increase in the PN ratio. Effects of fertilization effects on other nutrient concentration ratios were also examined.Paper No. 9401 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh, NC 27695.     

Physiologic,morphologic and behavioral responses of perpetual cultures ofCaulobacter crescentus to carbon,nitrogen and phosphorus limitations

Perpetual cultures ofCaulobacter crescentus were maintained in a chemostat and provided with media that provided glucose, glutamate, ammonium chloride, and Na/K phosphate as sources, respectively, of C, of C and N, of N and of P. All cultures were maintained at the same relative flow rate, f/V, so that reproductive rate was constant. As the CNP ratio was shifted from a balanced ratio of 100 g C9–10 g N1 g P, the cells modulated their rates of uptake, reducing the rates of uptake of excess element-sources and accelerating the uptake of potential sources of the limiting element. The change in N-limited cells was the greatest, resulting in a nearly 150-fold increase in rate of uptake of amino acids. Poly--hydroxybutyrate or polyphosphate was stored whenever C or P, respectively, was not the limiting element. Direct measurement of cell and stalk surface areas on purified peptidoglycan sacculi of P-limited cells revealed that the surface of both swarmer and stalked cells, as well as the stalk surface, participated in the accelerated rate of phosphate uptake. Swarmers from N-limited populations were the only cells that exhibited chemotactic responsiveness-to methionine, NH₄Cl and glutamate, but not to glucose or phosphate-sufficient to be detectable in a microcapillary assay. In populations that were C-limited and provided with ammonium ions as the principal source of N, morphogenesis and reproduction deteriorated and a steady state could not be maintained. Generally, the responses ofC. crescentus were appropriate to manage unbalanced nutrient supplies, but this oligoheterotrophic organism did not tolerate excesses of inorganic nutrients when limited for its sources of C, of energy, and of ammonium acceptor.     

Mutation spectra of N-ethyl-N''-nitro-N-nitrosoguanidine and 1-(2-hydroxyethyl)-1-nitrosourea in Escherichia coli

Summary DNA sequencing was used to determine the specific types of DNA base changes induced following in vivo exposure of Escherichia coli to the ethylating agent N-ethyl-N-nitro-N-nitrosoguanidine (ENNG) and the hydroxyethylating agent 1-(2-hydroxyethyl)-1-nitrosourea (HENU) using the xanthine guanine phosphoribosyltransferase (gpt) gene as the genetic target. We observed that 22/30 of the ENNG-induced mutations were GCAT transitions, 4/30 were ATGC transitions, 3/30 were ATTA transversions, and 1/30 was an ATCG transversion. We observed that 37/40 HENU-induced mutations were GCAT transitions and that the remaining 3/40 were ATGC transitions. A majority of the GCAT transitions induced by ENNG and HENU (68% and 73%, respectively) occurred at the second guanine of the sequence 5-GG(A or T)-3; this sequence specificity was similar to that previously seen with the alkylating agents N-methyl- and N-ethyl-N-nitrosourea (MNU and ENU) and N-methyl-N-nitro-N-nitrosoguanidine (MNNG). A DNA strand preference for the GA changes (antisense strand), previously noted for MNU, ENU, and MNNG, was observed following exposure to HENU and ENNG. The ATGC transitions induced by ENNG, HENU, and ENU also exhibit a sequence specificity with 13/13 mutations occurring at the T of the sequence 5-NTC-3. A strand preference was not apparent for these mutations.     

Organization of α-chain genes among Hb G-Philadelphia heterozygotes in association with Hb S, β-thalassemia,and α-thalassemia-2

The percentages of the -chain variant Hb G-Philadelphia (Hb G) or ₂ 68 AsnLys₂ were evaluated in 84 adult and 18 newborn heterozygotes. These included members of three families who were studied in more detail by nucleic acid hybridization techniques. The adult heterozygotes fell in two categories, one with a higher proportion of Hb G [46.5±1.0% (SD), N=21] and another with lower values (33.9±3.4%, N=63). Among the newborn heterozygotes, two babies fell in the category with the higher proportion of Hb G while 16 babies gave values between 25 and 34%. Studies of -chain gene organization on the parents of one neonate with a Hb G level of 27% at birth and 37% at 8 months excluded the presence of chromosomes with triplicated -chain genes which could lead to the ^0G/ genotype. Rather, these studies on five Hb G heterozygotes from three families confirmed the linkage between Hb G and a specific type of -thalassemia-2 associated with the presence of a 16-kbp Bgl II fragment which most probably carries the ^G locus since it has been found in 19 Hb G heterozygotes studied to date. The presence of an -thal-2 heterozygosity and three -chain genes (^0G/) was confirmed among Hb G heterozygotes with lower proportions of this variant. It is likely that the even lower values found in some newborn could arise through defective assembly of ^G- dimers. The presence of an -thal-2 homozygosity and two active -chain genes, one on each chromosome (^0G/⁰), was confirmed among heterozygotes with the higher proportion of Hb G. One of each of these categories was present in each of the three families investigated. This type of variability in the number of active -chain genes due to a heterozygosity or a homozygosity for -thalassemia-2 explains the trimodality of Hb S percentages among heterozygotes and the atypical hematological or biosynthetic features among patients with -thalassemia and sickle-cell syndromes.This research was supported by USPHS Research Grants HLB-05168 and HLB-15158 and by designated research funds of the Veterans Administration. This is Contribution No. 0693 of the Department of Cell and Molecular Biology, Medical College of Georgia, Augusta.     

Inhibition of NF-κ B activity and cFLIP expression contribute to viral-induced apoptosis

Virus-induced activation of nuclear factor-kappa B (NF-B) is required for Type 3 (T3) reovirus-induced apoptosis. We now show that NF-B is also activated by the prototypic Type 1 reovirus strain Lang (T1L), which induces significantly less apoptosis than T3 viruses, indicating that NF-B activation alone is not sufficient for apoptosis in reovirus-infected cells. A second phase of virus-induced NF-B regulation, where NF-B activation is inhibited at later times following infection with T3 Abney (T3A), is absent in T1L-infected cells. This suggests that inhibition of NF-B activation at later times post infection also contributes to reovirus-induced apoptosis. Reovirus-induced inhibition of stimulus-induced activation of NF-B is significantly associated with apoptosis following infection of HEK293 cells with reassortant reoviruses and is determined by the T3 S1 gene segment, which is also the primary determinant of reovirus-induced apoptosis. Inhibition of stimulus-induced activation of NF-B also occurs following infection of primary cardiac myocytes with apoptotic (8B) but not non-apoptotic (T1L) reoviruses. Expression levels of the NF-B-regulated cellular FLICE inhibitory protein (cFLIP) reflect NF-B activation in reovirus-infected cells. Further, inhibition of NF-B activity and cFLIP expression promote T1L-induced apoptosis. These results demonstrate that inhibition of stimulus-induced activation of NF-B and the resulting decrease in cFLIP expression promote reovirus-induced apoptosis.     

Inhibition of protein synthesis enhances the lytic effects of tumor necrosis factor α and interferon γ in cell lines derived from gynecological malignancies

Summary Few clinical responses have occurred in preliminary studies using the cytokines tumor necrosis factor (TNF) or interferon (IFN) in cancer patients. This may be related to the observation that many malignant cell lines are resistant to lysis by these cytokinesin vitro. Resistance to lysis by TNF or IFN in many cells is controlled by a protein-synthesis-dependent mechanism, such that when protein synthesis is inhibited cells become sensitive to lysis by these cytokines. Because there is some evidence that TNF and IFN act through different lytic mechanisms and are opposed by different resistance mechanisms, we treated a panel of eight cell lines, five derived from human cervical carcinomas (ME-180, MS751, SiHa, HT-3, and C-33A) and three derived from ovarian carcinomas (Caov-3, SK-OV-3, and NIH: OVCAR-3) with both TNF and IFN to determine whether such combination treatment might maximizein vitro cell lysis. Our results showed that pretreatment with IFN followed by exposure to TNF in the presence of protein synthesis inhibitors increased lysis of seven of the eight cell lines above that seen with either TNF or IFN and inhibitors of protein synthesis. Only the cell line C-33A was resistant to lysis by TNF and IFN, when exposed to these agents both alone and in combination with protein synthesis inhibitors. Clinically, combining the cytokines TNF and IFN with protein synthesis inhibitors may maximize thein vivo lytic effects of these cytokines.Supported by American Cancer Society Career Development Award 90-221     

Bacterial regeneration of ammonium and phosphate as affected by the carbon:nitrogen:phosphorus ratio of organic substrates

The effect of carbonnitrogenphosphorus (CNP) ratio of organic substrates on the regeneration of ammonium and phosphate was investigated by growing natural assemblages of freshwater bacteria in mineral media supplemented with the simple organic C, N, and P sources (glucose, asparagine, and sodium glycerophosphate, respectively) to give 25 different substrate CNP ratios. Both ammonium and phosphate were regenerated when CN and NP atomic ratios of organic substrates were 101 and 161, respectively. Only ammonium was regenerated when CN and NP ratios were 101 and 10–201, respectively. On the other hand, neither ammonium nor phosphate was regenerated when CN and NP ratios were 151 and 51, respectively. In no case was phosphate alone regenerated. As bacteria were able to alter widely the CNP ratio of their biomass, the growth yield of bacteria appeared primarily dependent on the substrate carbon concentration, irrespective of a wide variation in the substrate CNP ratio.     

Variant forms ofα-2-l-fucosyltransferase in human submaxillary glands from blood group ABH “secretor” and “non-secretor” individuals

The properties of the -2-l-fucosyltransferases in submaxillary gland preparations from blood group ABH secrefors and non-secretors were compared. The level of activity in the non-secretor gland homogenates amounted to about 5% only of that found in the secretor gland preparations. The enzymes from the two sources differed in solubility properties, charge and affinities for donor and acceptor substrates. The enzyme from secretor glands showed a preference for acceptors with Type 1 [d-galactosyl(1–3)-N-acetyl-d-glucosamine] structures whereas the enzyme from non-secretor glands had a preference for Type 2 [d-galactosyl(1–4)-N-acetyl-d-glucosamine] structures.These results demonstrate that expression of the secretor gene (Se) is associated with a molecular form of the -2-l-fucosyltransferase that is different from the species present in the same tissue when theSe gene is not expressed.     

Refinement of the protein backbone angle ψ in NMR structure calculations

Cross-correlated relaxation rates involving the C-H dipolar interaction and the carbonyl (C) chemical shift anisotropy (CSA) have been measured using two complementary 3D experiments. We show that the protein backbone angle can be directly refined against such cross-correlated relaxation rates (^{H C,C}) and the three-bond H/D isotope effect on the C chemical shifts (³C _(ND)). By simultaneously using both experimental parameters as restraints during NMR structure calculations, a unique value for the backbone angle is defined. We have applied the new refinement method to the -Spectrin SH3 domain (a -sheet protein) and to the Sgs1p HRDC domain (an -helical protein) and show that the quality of the NMR structures is substantially improved, judging from the atomic coordinate precision and the Ramachandran map. In addition, the -refined NMR structures of the SH3 domain deviate less from the 1.8 Å crystal structure, suggesting an improved accuracy. The proposed refinement method can be used to significantly improve the quality of NMR structures and will be applicable to larger proteins.     

Design and synthesis of thrombin inhibitors

Summary As part of our programme directed at the development of enzyme inhibitors based on transition-state mimics, we discovered in the early 1980s that P3-P3 fragments of human fibrinogen A, containing the ketomethylene isostere Arg--[COCH₂]Gly at P1-P1, were potent inhibitors of thrombin. Such low-molecular-weight inhibitors are expected to be clinically useful as anticoagultant drugs. In our more recent investigations, the P1-P1 moiety has been replaced with various arginine or lysine ketones. The resulting compounds showed the following order of thrombin inhibitory potency: -ketoesters > fluoroketones >alkoxymethylketones > difluoro--ketoamides >-ketoesters >alkyl ketones. In contrast to all other lysine/arginine pairs studied previously, the inhibitor based on a lysine -ketoester proved superior to the corresponding arginine analogue. A possible explanation for this finding is discussed. All the highly electrophilic ketones (e.g., fluoroketones) were found to exhibit slow-binding kinetics with thrombin, which is likely to be a disadvantage in clinical use. Alkoxymethyl ketones were devoid of such behaviour and have been developed further to yield nanomolar inhibitors of low molecular weight and good selectivity for thrombin. One of these ketones was found to compare favourably with known thrombin inhibitors in anticoagulant assays. The synthesis of various types of inhibitor mentioned above is described, together with structure-activity correlations for inhibition of thrombin.     

Effects of D‐mannoheptulose upon D‐glucose metabolism in tumoral pancreatic islet cells

D[³H]mannoheptulose was recently reported to be poorly taken up by tumoral pancreatic islet cells of the RINm5F and INS1 lines. We have now investigated the effects of Dmannoheptulose upon Dglucose metabolism in these two cell lines. Dmannoheptulose (1.0–10.0 mM) only caused a minor decrease of Dglucose metabolism in RINm5F cells, whether at low (1.1 mM) or higher (8.3 mM) Dglucose concentration. A comparable situation was found in INS1 cells examined after more than 20 passages. In both cases, however, the hexaacetate ester of Dmannoheptulose (5.0 mM) efficiently inhibited Dglucose metabolism. In the INS1 cells, the relative extent of the inhibitory action of Dmannoheptulose upon Dglucose metabolism increased from 12.4 ± 2.6 to 38.3 ± 3.8% as the number of passages was decreased from more than 20 to 13–15 passages, the latter percentage remaining lower, however, than that recorded in INS1 cells also examined after 13–15 passages but exposed to Dmannoheptulose hexaacetate (66.9 ± 2.2%). These findings when compared to our recent measurements of D[³H]mannoheptulose uptake, reinforce the view that the entry of the heptose into cells and, hence, its inhibitory action on Dglucose metabolism are dictated by expression of the GLUT2 gene.     

Synthesis of aspartame precursor by solid thermolysin in organic solvent

Summary The enzymatic synthesis of a peptide compound was carried out successfully in homogeneous organic solvent.Solid Thermolysin was found to catalyze the synthetic reaction of N-benzyloxycarbonyl-L-aspartyl-L-phenylalanine methyl ester (Z-APM; a precursor of sweetner Aspartame) from N-benzyloxycarbonyl-L-aspartic acid (Z-L-Asp) and L-phenylalanine methyl ester (L-PheOMe) in a 98 percent organic medium (ethylacetatebenzenemethanolwater=5029192). The dissolution of enzyme was not observed. The optimal pH shifted to acidic side by 1.0 pH unit, compared with that in aqueous medium. The enzymatic activity of solid thermolysin with an average size of 3.4×9.5 m was determined to be 0.18 moles-product/(mg-solid)·h under the initial concentrations of L-PheOMe of 0.1M and Z-L-Asp of 0.05M, and at pH 6.0 and 40°C.     

Modification of the properties of a translocation in the German cockroach by selection

Summary A stock of Blattella germanica bearing the interchange T(3; 12)/3;12 was subjected to close inbreeding with selection for random disjunction at metaphase I. After 3–4 generations of selection, interchange quadrivalent chiasma frequency decreased, variability in free bivalent chiasma frequency increased sharply, and individuals with either random or directed disjunction were present in the stock. Random disjunction was modified from a ratio of 2112 (adj.-1; alt.-1; adj.-2; alt.-2) to a ratio of 1111. After 7–8 generations of selection, chiasma frequency appeared to stabilize at lower than normal levels and variability decreased for both quadrivalents and free bivalents. Directed disjunction was modified from a ratio of 2114 to 1112, and no individuals with the original high level of directed disjunction were detected. Chains-of-four tended to orient randomly, especially in individuals where the ring quadrivalents showed directed disjunction. Relaxation of inbreeding, but not selection, produced an increase in chiasma frequency and variability in both free bivalents and quadrivalents, but the modified ratios for both random and directed disjunction were retained. These results are discussed with respect to inbreeding effects and genetic control of chiasma frequency and metaphase I disjunction in interchange quadrivalents.     

Sex determination in Sarotherodon (tilapia)

Summary The simplest possible model of the sex determination process adding autosomal influence to a minimal number of sex chromosomes was developed to explain matings of Tilapia (Sarotherodon) species. Eighteen different genotypes, each having two autosomes (AA, Aa, or aa) and two sex chromosomes (WX, WY, WW, XY, XX or YY) involved in sex determination, are predicted by the theory. Their sex (10 males and 8 females) were determined using a series of directed graphs, showing the relative strength of the chromosome pairs, developed on the basis of Chen's sex ratio results (Chen 1969). This theoretical model predicts eight different sex ratios (01, 13, 35, 11, 97, 53, 31, 10 ); three of them are not predicted by the WXYZ theory. The greatest part of these sex ratios have been obtained experimentally in extensive series of crosses between related species of Tilapia and their hybrids, carried out by several authors. The theory succeeds in explaining all of Chen's results, including those ratios 53 and 01 seen in certain crosses but not predicted by the WXYZ theory. The importance of the autosomes is seen in comparisons of the genotype pairs (AaWY, aaWY), (AaXY, aaXY) and (AAWW, AaWW) in which the first genotype in each case is male while the second is female as proven by the sex ratio results. The members of the pair differ only in the substitution of one autosome for the other. To test the theory, experiments consisting of hormonal sex reversion and a series of crosses are proposed. Finally, theoretical and practical implications of the theory are discussed.     

Desensitization of prostaglandin F2α receptor-mediated phosphoinositide hydrolysis in cultured rat astrocytes

Desensitization of prostaglandin (PG) F₂ receptor-mediated phosphoinositide (PI) hydrolysis was investigated in cultured rat astrocytes. Prolonged exposure of astrocytes differentiated by dibutyryl cyclic AMP-treatment to PGF₂ caused the desensitization of subsequent PGF₂-induced PI hydrolysis. The desensitization was time- and PGF₂ dose-dependent; maximal decrease in the PI hydrolysis was observed after exposure to 10 M PGF₂ for 4 h and the degree of the desensitization was 31.7±2.7% of control. Pretreatment with either PGD₂ or PGE₂ also induced the desensitization of subsequent PGF₂-stimulated PI hydrolysis and conversely pretreatment of PGF₂ decreased the PI responses to PGD₂ and PGE₂. The desensitization prevented by phloretin and was reversible upon removal of the agonist. Protein synthesis inhibitors blocked the recovery of the desensitization. Treatment of the cells with phorbol 12-myristate 13-acetate had no effect on the desensitization. These results suggest that prolonged exposure of the astrocytes to PGF₂ caused the desensitization of the receptors.     

Evidence of β-carotene 7,8(7′,8′) oxygenase (β-cyclocitral,crocetindial generating) in Microcystis

A -carotene oxygenase is described which occurs in the Cyanobacterium Microcystis. It cleaves -carotene and zeaxanthin specifically at the positions 7,8 and 7,8, while echinenone and myxoxanthophyll are not affected. The oxidative cleavage of -carotene leads to the formation of -cyclocitral and crocetindial and that of zeaxanthin to hydroxy--cyclocitral and crocetindial in nearly stoichiometric amounts. Oxidant is dioxygen as has been demonstrated by high incroporation (86%) of ¹⁸O₂ into -cyclocitral. -Carotene oxygenase is membrane bound, sensitive to sulfhydryl reagents, antioxidants and chelating agents. Iron seems to be an essential part of the enzyme activity. Cofactors necessary for the reaction could not be detected.Abbreviations TLC thin layer-chromatography - PIPES piperazine-N,N-bis-(2-ethanesulfonate) Na - TES 2{[tris-(hydroxymethyl)-methyl]-amino} ethanesulfonic acid Dedicated to Professor G. Drews on occasion of his 60th birthday     

Developmental changes in the fatty acids of synaptic membrane phospholipids: Effect of protein malnutrition

The acyl-linked fatty acid composition of the major phospholipid species in rat cortical synaptic membranes was determined at various stages of development. For most species there was a decrease during development in the short chain saturated fatty acids, 140 and 160, an increase in 180 and 226 (n-3) and an increase in the ratio of 226 (n-3)/225 (n-6). Pups were protein deprived by feeding the dams a 12% casein diet as compared to the 24% casein control diet. Protein malnutrition markedly affected the composition of acyl-linked fatty acids in the synaptic membranes. The increases in the ratio of 226 (n-3)/225 (n-6) fatty acids were especially compromised.Abbreviations used (PC) Phosphatidylcholine - (PE) phosphatidylethanolamine - (EP) ethanolamine plasmalogen - (PS) phosphatidylserine - (PI) phosphatidylinositol - (SP) sphingomyelin - (t.l.c.) thin layer chromatography - (BHT) butylated-hydroxytoluene