Effect of troglitazone on the desaturases in a rat model of insulin-resistance induced by a sucrose-rich diet.

A sucrose-rich diet generates time-dependent metabolic disorders similar to those found in diabetes type 2. After 8 month (mo) this diet evoked in the rat an increase of blood glucose, free fatty acids (FFA) and triacylycerides (TG) without insulin modification, an interruption of liver stearoyl-CoA desaturase-1 (SCD-1) mRNA and activity increase found at 6 mo, and an enhacement of Delta6 and Delta5 desaturase mRNA and Delta6 activity. We found that the administration of troglitazone (TRO), a peroxisome-proliferator-activated receptors gamma (PPAR-gamma) agonist, for 2 mo normalized plasma FFA, TG, and glucose without altering the insulinemia. It depressed liver SCD-1 mRNA in both control and sucrose-fed rats, decreasing the 18:1n-9/18:0 ratio in serum and liver lipids, and eliminated the increasing effect on mRNA and activity of Delta6 and Delta5 desaturases. These findings evidence again that desaturases are not affected through an insulin resistant effect evoked by the sucrose-rich diet and TRO recovers the altered metabolic plasma parameters as it corresponds to a PPAR-gamma agonist, but its effect on hepatic desaturases can not be attributed to a direct action on liver by PPAR-gamma, insulin, and even by an insulin sensitizing mechanism, suggesting it would be evoked indirectly through hepatic PPAR-alpha deactivation induced by the FFA decrease.

Phosphatidyl choline fatty acid remodeling in the hepatic cell nuclei.

This study was performed to determine whether fatty acids incorporated into liver cell nuclei phosphatidylcholine (PtdCho) could be remodeled in the isolated nuclear. For this reason, rat liver cell nuclei were incubated in vitro with [1-14C]20:4n-6-CoA. PtdCho molecular species with the highest specific activity had an unsaturated fatty acid at sn-1 and sn-2 positions (20:4-20:4>18:2-20:4>18:1-20:4). 16:0-20:4 and 18:0-20:4 PtdChos showed a minor specific activity. When labeled nuclei were reincubated in the absence of labeled substrate with the addition of cytosol, ATP and CoA, the specific activity of 20:4-20:4, 18:2-20:4 and 18:1-20:4 species decreased, while that of 16:0-20:4 and 18:0-20:4 increased. In conclusion, the asymmetric fatty acid distribution of saturated fatty acids at sn-1 position, and unsaturated fatty acids at sn-2 position of nuclear PtdCho molecular species was re-established by an acyl-CoA-dependent remodeling process.

Immunohistochemical detection of a very high density lipoprotein (VHDL) in ovarian follicles of Triatoma infestans.

The ability of Triatoma infestans ovarian follicles to synthesize a very high-density lipoprotein (VHDL) has been examined by immunohistochemical methods. This kind of lipoprotein can be envisaged as a storage hexameric protein present in the hemolymph of some insect species. VHDL immunoreactivity is observed in oocytes at different stages of maturation. The antigen is present in the oocyte cytoplasm as well as in the follicular epithelial cells. The immunopositive reaction in the apical surface of follicle cells suggests both a VHDL synthesis and a secretion process. Furthermore, VHDL seems to be stored into oocyte in yolk granules. On the contrary, no immunopositive reaction is observed in the intracellular spaces between follicle cells, suggesting that VHDL is not incorporated from hemolymph into the oocyte.

Incorporation and distribution of saturated and unsaturated fatty acids into nuclear lipids of hepatic cells.

Liver nuclear incorporation of stearic (18:0), linoleic (18:2n-6), and arachidonic (20:4n-6) acids was studied by incubation in vitro of the [1-14C] fatty acids with nuclei, with or without the cytosol fraction at different times. The [1-14C] fatty acids were incorporated into the nuclei as free fatty acids in the following order: 18:0 > 20:4n-6 >> 18:2n-6, and esterified into nuclear lipids by an acyl-CoA pathway. All [1-14C] fatty acids were esterified mainly to phospholipids and triacylglycerols and in a minor proportion to diacylglycerols. Only [1-14C]18:2n-6-CoA was incorporated into cholesterol esters. The incorporation was not modified by cytosol addition. The incorporation of 20:4n-6 into nuclear phosphatidylcholine (PC) pools was also studied by incubation of liver nuclei in vitro with [1-14C]20:4n-6-CoA, and nuclear labeled PC molecular species were determined. From the 15 PC nuclear molecular species determined, five were labeled with [1-14C]20:4n-6-CoA: 18:0-20:4, 16:0-20:4, 18:1-20:4, 18:2-20:4, and 20:4-20:4. The highest specific radioactivity was found in 20:4-20:4 PC, which is a minor species. In conclusion, liver cell nuclei possess the necessary enzymes to incorporate exogenous saturated and unsaturated fatty acids into lipids by an acyl-CoA pathway, showing specificity for each fatty acid. Liver cell nuclei also utilize exogenous 20:4n-6-CoA to synthesize the major molecular species of PC with 20:4n-6 at the sn-2 position. However, the most actively synthesized is 20:4-20:4 PC, which is a quantitatively minor component. The labeling pattern of 20:4-20:4 PC would indicate that this molecular species is synthesized mainly by the de novo pathway.

Effects of diabetes and insulin on hepatic delta6 desaturase gene expression.

It has been recognized that rat liver microsomal Delta6 desaturation activity is defective in experimental diabetes, a fact that may be reverted by means of insulin treatment. In the present study, we used streptozotocin-induced diabetic rats in order to determine the regulatory role of insulin on the expression of hepatic Delta6 desaturase gene. The abundance of hepatic Delta6 desaturase mRNA in the diabetic rats is sevenfold lower than in the control. Insulin administration to diabetic rats induces Delta6 desaturase mRNA eightfold within 24 h. The effect of insulin on the Delta6 desaturase mRNA was inhibited 70% with dibutyryl-cAMP and theophylline administration and 90% by cycloheximide administration. Therefore, our data demonstrate that the activity of hepatic Delta6 desaturase in response to insulin is, at least in part, regulated by pretranslational events that require the synthesis of an unknown protein(s).

Immunohistochemical detection of a very high density lipoprotein (VHDL) in ovarian follicles of Triatoma infestans

The ability of Triatoma infestans ovarian follicles to synthesize a very high-density lipoprotein (VHDL) has been examined by immunohistochemical methods. This kind of lipoprotein can be envisaged as a storage hexameric protein present in the hemolymph of some insect species. VHDL immunoreactivity is observed in oocytes at different stages of maturation. The antigen is present in the oocyte cytoplasm as well as in the follicular epithelial cells. The immunopositive reaction in the apical surface of follicle cells suggests both a VHDL synthesis and a secretion process. Furthermore, VHDL seems to be stored into oocyte in yolk granules. On the contrary, no immunopositive reaction is observed in the intracellular spaces between follicle cells, suggesting that VHDL is not incorporated from hemolymph into the oocyte.

Immunohistochemical detection of a very high density lipoprotein (VHDL) in ovarian follicles of Triatoma infestans

The ability of Triatoma infestans ovarian follicles to synthesize a very high-density lipoprotein (VHDL) has been examined by immunohistochemical methods. This kind of lipoprotein can be envisaged as a storage hexameric protein present in the hemolymph of some insect species. VHDL immunoreactivity is observed in oocytes at different stages of maturation. The antigen is present in the oocyte cytoplasm as well as in the follicular epithelial cells. The immunopositive reaction in the apical surface of follicle cells suggests both a VHDL synthesis and a secretion process. Furthermore, VHDL seems to be stored into oocyte in yolk granules. On the contrary, no immunopositive reaction is observed in the intracellular spaces between follicle cells, suggesting that VHDL is not incorporated from hemolymph into the oocyte.(AU)

Effect of dexamethasone on the fatty acid composition of total liver microsomal lipids and phosphatidylcholine molecular species.

Dexamethasone depresses delta6 and delta5 and increases delta9 desaturase and synthase activities. Therefore, we investigated the effect on the fatty acid composition of microsomal liver lipids and phosphatidylcholine (PtdCho) molecular species. After 15 d of treatment we found a notable decrease in arachidonic acid, a small decrease in stearic acid, and increases of linoleic, oleic, palmitoleic, and palmitic acids in liver microsomal total lipids and PtdCho. The study of the distribution of the PtdCho molecular species indicated that 18:0/20:4n-6, 16:0/20:4n-6, and 16:0/18:2n-6 predominated in the control animals. Dexamethasone, as expected because of its depressing effect on arachidonic acid synthesis and activation of oleic and palmitic acid synthesis, evoked a very significant decrease in 18:0/20:4n-6 PtdCho (P<0.001) and an important increase in 16:0/18:2n-6. The invariability of 16:0/20:4n-6 PtdCho could be related to the antagonistic effect of arachidonic and palmitic acid synthesis. PtdCho species containing oleic acid were not significant. The bulk fluidity and dynamic properties of the microsomal lipid bilayer measured by fluorometry using the probes 1,6-diphenyl-1,3,5-hexatriene and 4-trimethylammonium-phenyl-6-phenyl-1,3,5-hexatriene showed no significant modification, probably owing to a compensatory effect of the different molecular species, but changes of particular domains not detected by this technique are possible. However, the extremely sensitive Laurdan detected increased lipid packing in the less-fluid domains of the polar-nonpolar interphase of the bilayer, possibly evoked by the change of molecular species and cholesterol/phospholipid ratio. The most important effect found is the decrease of arachidonic acid pools in liver phospholipids as one of the corresponding causes of dexamethasone-dependent pharmacological effects.

Immunohistochemical detection of a very high density lipoprotein (VHDL) in ovarian follicles of Triatoma infestans.

The ability of Triatoma infestans ovarian follicles to synthesize a very high-density lipoprotein (VHDL) has been examined by immunohistochemical methods. This kind of lipoprotein can be envisaged as a storage hexameric protein present in the hemolymph of some insect species. VHDL immunoreactivity is observed in oocytes at different stages of maturation. The antigen is present in the oocyte cytoplasm as well as in the follicular epithelial cells. The immunopositive reaction in the apical surface of follicle cells suggests both a VHDL synthesis and a secretion process. Furthermore, VHDL seems to be stored into oocyte in yolk granules. On the contrary, no immunopositive reaction is observed in the intracellular spaces between follicle cells, suggesting that VHDL is not incorporated from hemolymph into the oocyte.

Effect of experimental diabetes on the fatty acid composition, molecular species of phosphatidyl-choline and physical properties of hepatic microsomal membranes.

Streptozotocin diabetes depresses delta 9, delta 6 and delta 5 fatty acid desaturases, decreasing arachidonic acid and increasing linoleic acid, but also unexpectedly increasing docosahexaenoic acid in the different phospholipids of liver microsomal lipids. 18:0/20:4n-6, 16:0/20:4n-6 and 16:0/18:2n-6 are the predominant phosphatidyl choline (PC) molecular species in control rats, determining mainly PC contribution to the dynamic and biochemical properties of this bilayer. Diabetes decreases 20:4n-6 containing species and increases 18:2n-6 and 22:6n-3 containing species, maintaining the bulk dynamic properties in the hydrophobic interior of the bilayer, but changing its biochemical properties. The different dynamic parameters were measured by fluorometry using the probes 1,6-diphenyl-1,3,5-hexatriene (DPH), (4-trimethylammonium phenyl) 6-phenyl-1,3,5 (TMA-DPH) and 6-lauroyl-2,4-dimethyl aminonaphtalene (Laurdan). In the surrounding of the hydrophobic/hydrophilic interphase lipid molecules were less ordered and tightly packed in the diabetic samples, allowing a higher mobility of incorporated water molecules. The fact that diabetes decreases highly polyunsaturated acid of n-6 family, but increases docosahexaenoic acid, indicates the necessity of re-evaluating its effect in human physiology.

Effect of temperature and salinity stress on growth and lipid composition of Shewanella gelidimarina.

The maximum growth temperature, the optimal growth temperature, and the estimated normal physiological range for growth of Shewanella gelidimarina are functions of water activity (a(w)), which can be manipulated by changing the concentration of sodium chloride. The growth temperatures at the boundaries of the normal physiological range for growth were characterized by increased variability in fatty acid composition. Under hyper- and hypoosmotic stress conditions at an a(w) of 0.993 (1.0% [wt/vol] NaCl) and at an a(w) of 0.977 (4.0% [wt/vol] NaCl) the proportion of certain fatty acids (monounsaturated and branched-chain fatty acids) was highly regulated and was inversely related to the growth rate over the entire temperature range. The physical states of lipids extracted from samples grown at stressful a(w) values at the boundaries of the normal physiological range exhibited no abrupt gel-liquid phase transitions when the lipids were analyzed as liposomes. Lipid packing and adaptational fatty acid composition responses are clearly influenced by differences in the temperature-salinity regime, which are reflected in overall cell function characteristics, such as the growth rate and the normal physiological range for growth.

Dietary cholesterol induces changes in molecular species of hepatic microsomal phosphatidylcholine.

After 21 days on a diet containing 1 g% cholesterol and 0.5 g% cholic acid, rats had an increased content of cholesterol in liver microsomal lipids. In liver, both cholesterol content and delta9 desaturase activity increased, whereas delta6 and delta5 desaturase activities decreased. These changes correlated with increases in oleic, palmitoleic, and linoleic acids and decreases in arachidonic and docosahexenoic acids in total microsomal lipids. Similar fatty acid changes were found in phosphatidylcholine (PC), the principal lipid of the microsomal membrane. In PC the predominant molecular fatty acid species (67% of the total) in the control rats were 18:0/20:4, 16:0/20:4, and 16:0/18:2; and they mainly determined the contribution of PC to the biophysical and biochemical properties of the phospholipid bilayer. The cholesterol diet decreased specifically the 18:0/20:4 species, and to a lesser extent, 16:0/20:4 and 18:0/22:6. The 18:1-containing species, especially 18:1/18:2 and less so 16:0/18:1 and 18:1/20:4, were increased. A new 18:1/18:1 species appeared. The independent effects of the presence of cholesterol and change of the fatty acid composition of the phospholipid bilayer of liver microsomes on the packing were studied by fluorescence methods using 6-lauroyl-2,4-dimethylaminonaphthalene, 1,6-diphenyl-1,3,5-hexatriene and 1-(4-trimethylammonium phenyl)-6-phenyl-1,3,5-hexatriene, which test different parameters and depths of the bilayer. Data showed that the increase of cholesterol in the membrane, and not the change of the fatty acid composition of phospholipids, was the main determinant of the increased bulk packing of the bilayer. The increase of fluid oleic- and linoleic-containing species almost compensated for the drop in 20:4- and 22:6-containing molecules. But the most important effect was that the general drop in essential n-6 and n-3 polyunsaturated fatty acids meant that this endogenous source for the needs of the animal decreased.

Fatty acid delta9-desaturation in the Triatoma infestans fat body: response to food and trehalose administrations.

The effects of food intake and carbohydrate administration on fatty acid delta9-desaturation were investigated in isolated microsomes from Triatoma infestans fat body. Fifth instar nymphs, which were solely blood-fed just after the molt and then fasted, were used as controls and for determination of optimal assay conditions. Both [1-14C]palmitic and [1-14C]stearic acids in a medium containing ATP, CoA, MgCl2, NADH, NaF, and O2 were tested. For the control group, optimal conditions were a pH of 6.8-7.2, an incubation temperature of 29 degrees C, and an incubation time of 10 min. delta9-Desaturation depended on the presence of reduced pyridine dinucleotides; NADH and NADPH were equally efficient. Stearic acid showed a higher apparent Vmax than palmitic acid, but the apparent Km were very similar. When fifth instar nymphs were blood-fed weekly, a marked increase of delta9-desaturation was observed for both acids. Higher desaturation activity was also induced by injection of the carbohydrate trehalose into the hemolymph of fasted nymphs. These results indicate that insect delta9-desaturation, like the mammalian counterpart, is sensitive to dietary changes and carbohydrate administration.

Partial characterization of mosquito larvae extract modulating mouse and human cell proliferation.

Mosquito larvae crude extract have been found to alter the mitotic rate of several mouse epithelial cell populations such as enterocytes and tongue keratinocytes. Also, the dialysed fraction inhibits hepatocyte proliferation in hepatectomized males. These experiments suggested an inhibitory effect on the G1/S interphase. Consequently, we suggested the presence of some molecule or molecules related to the TGF-beta superfamily. In the present paper, we have assayed the crude extract on human mononuclear cells and the dialysed fraction of the extract on tongue keratinocyte proliferation. Furthermore, different protein fractions obtained using a molecular exclusion chromatographic column were assayed on hepatocyte proliferation of hepatectomized mice. Three groups of proteins have been isolated. Results show a dose-dependent effect of crude extract on mononuclear cell proliferation and the dialysed extract caused an inhibitory effect on tongue keratinocyte proliferation. With regard to the hepatocyte mitotic rate, an inhibitory effect appeared only in animals receiving the fraction with lower molecular weight. These results suggest the presence in mosquito larvae of some peptidic molecule or molecules resembling the activity of members of the TGF-beta superfamily.

Purification and properties of a small lipid-binding protein from the hemolymph of Triatoma infestans.

A small lipid-binding protein (sLBP) was purified from the hemolymph of the blood-sucking bug Triatoma infestans. Its isolation involved size exclusion-high performance liquid chromatography (HPLC) followed by anion exchange chromatography-HPLC. The molecular weight of the protein, as determined by gel permeation chromatography, was 20 kDa. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) resolved the protein into a single polypeptide with M(r) approximately equal to 16 kDa. The sLBP contains 6% lipids. Diacylglycerols represent the major lipid class, whereas phosphatidyl-choline, phosphatidyl-ethanolamine, free fatty acids and hydrocarbons were found in minor amounts. The amino acid composition indicated a high content of aspartic and glutamic acids and non-polar aliphatic amino acids. The N-terminal sequence did not resemble the sequence of any other previously reported insect hemolymph protein. Far-UV circular dichroism suggested that sLBP adopts a conformation rich in beta-sheet structure. The presence of this protein in hemolymph, fat body and unfertilized eggs was explored throughout the last nymphal and adult stages of the insect by Western blot assays. These assays indicated that sLBP is particularly abundant in hemolymph. A high concentration of sLBP was also detected in the fat body of the nymphs.

Incorporation of delta 5 desaturase substrate (dihomogammalinolenic acid, 20:3 n-6) and product (arachidonic acid 20:4 n-6) into rat liver cell nuclei.

The incorporation of [1-(14)C]20:3 n-6 and its desaturation product, [1-(14)C]20:4 n-6 into nuclear lipids from rat liver cells were investigated during in vitro delta5 desaturation. [1-(14)C]20:3 n-6 activated as 20:3 n-6-CoA by nuclear long chain acyl-CoA synthetase was: (1) incorporated into nuclear lipids mainly esterified to phospholipids and in a lesser proportion, to triglycerides and diglycerides; and (2) desaturated to 20:4 n-6-CoA by the nuclear delta5 desaturase. The amount of [1-(14)C]20:4 n-6 acid synthesized in cell nuclei increased along with time and was stimulated by the cytosol fraction. The major proportion of 20:4 n-6 was found in phospholipids and in a lesser proportion it remained as free fatty acids and was esterified to triglycerides and diglycerides. 20:4 n-6-CoA was incorporated into nuclear lipids and hydrolyzed to free fatty acid. These results indicate that liver cell nuclei possess the necessary enzymes to incorporate the delta5 desaturase substrate (20:3 n-6) as well as the product of desaturation (20:4 n-6) into nuclear TG, DG and PL following an acyl-CoA dependent pathway.

Conformation of apolipoprotein AI in reconstituted lipoprotein particles and particle-membrane interaction: effect of cholesterol.

Discoidal recombinant high density lipoproteins (rHDL) of apolipoprotein AI (apoAI) and palmitoyloleoylphosphatidylcholine (POPC), with or without cholesterol, were prepared by cholate dialysis. By gel filtration, rHDL containing 2-4 (Lp2, Lp3 and Lp4) apoAI molecules/particle were obtained. The ApoAI conformation in these rHDL was investigated by tryptophan fluorescence, denaturation with guanidine HCl, and immunoreactivity with two monoclonal antibodies recognizing epitopes in the N-terminal and central domains. Data show that apoAI conformation is highly dependent on particle size as well as on cholesterol. The ability of rHDL to interact with lipid bilayer was studied by measuring leakage induction on POPC and POPC/cholesterol vesicles loaded with terbium/dipicolinic acid. Among the cholesterol-free rHDL, the most efficient ones were the smallest Lp2. Leakage induction on POPC vesicles is dramatically decreased by the presence of cholesterol in Lp2 and Lp3. All the rHDL, but specially those containing cholesterol, induced more leakage on the POPC/cholesterol than on the POPC vesicles. These results suggest that in small cholesterol-poor particles, apoAI could have a conformation determining a high affinity for membranes, which could facilitate cholesterol efflux. After cholesterol enrichment, a conformational change in apoAI could decrease the affinity for membranes allowing the lipoprotein release.

Temperature acclimation of Trypanosoma cruzi epimastigote and metacyclic trypomastigote lipids.

This study examines the changes in cellular lipids that take place when Trypanosoma cruzi epimastigotes and metacyclic trypomastigotes are transferred from 28 to 37 degrees C. We found a rise in the sterol to phospholipid ratio, as well as in the triacylglycerol and steryl ester cellular content in T. cruzi epimastigotes. In addition, saturated to unsaturated fatty acid ratios in phospholipids increase. This latter effect appears to be due to two concurrent processes. Firstly, fatty acyl delta9 and, especially, delta12 desaturations are significantly diminished at 37 degrees C. Secondly, triacylglycerols and steryl esters undergo changes in their fatty acyl composition opposite to those simultaneously observed in phospholipids, i.e. the ratio of saturated to unsaturated fatty acids markedly decreases. Similar alterations in each of the lipid classes and in the fatty acid composition of polar and neutral lipids were found in cultured metacyclic trypomastigotes on exposure to the same shift-up. These observations suggest that a global remodeling of cellular lipids that involves extensive fatty acid exchange between neutral and polar lipid pools represents a novel and important mechanism of adaptation of the parasites to the temperature changes they encounter in their life cycle.

Molecular species of phosphoglycerides in liver microsomes of rats fed a fat-free diet.

The influence of a fat-free diet on the molecular species composition of phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylinositol (PI) of rat liver microsomes was studied by using reversed-phase high-pressure liquid chromatography. In the three phosphoglyceride classes analyzed, the fat-free diet produced a large decrease in the 18:0/20:4n-6 species but less important changes were found in the 16:0/20:4n-6 species. In PC, the most abundant phosphoglyceride class of rat liver microsomes, the fall in the 18:0/20:4n-6 species was counterbalanced mainly by an enhancement in the 16:0/18:1n-9 species although it was not evident in PE. In PI, the decrease in the 18:0/20:4n-6 species was counterbalanced by an increase in the 18:0/20:3n-9 species. Fluorescence polarization measurements of 1,7-diphenyl-1,3,5-hexatriene in liposomes of 16:0/18:1n-9, 18:0/18:1n-9-, 16:0/20:4n-6-, and 18:0/20:4n-6-PC indicated that the change in the saturated fatty acid in the sn-1 position accompanying the replacement of 20:4n-6 by 18:1n-9 could be very important for a homeoviscous compensation, maintaining the membrane physical properties without large alterations in spite of the essential fatty acid deficiency due to the fat-free diet.

Effect of polyunsaturated fatty acid deficiency on dipole relaxation in the membrane interface of rat liver microsomes.

The influence of a fat-free diet on the lipid dynamics of rat liver microsomes and liposomes of microsomal lipids was studied by using different fluorescence methods. Lifetime distribution and rotational diffusion of probes with different localization in the lipid bilayer were measured using multifrequency fluorometry. Lateral mobility was studied by measuring excimer formation of pyrenedodecanoic acid. Dipolar relaxation in the interfacial region was studied using 2-dimethyl-amino-6-lauroylnaphthalene (Laurdan). In spite of large changes in the fatty acid composition of microsomal lipids, polyunsaturated fatty acid deficiency showed no effect on the lifetime distribution and rotational mobility of 1,6-diphenyl-1,3,5-hexatriene (DPH). l-(4-(trimethylamino)phenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH), 2- 7- and 12-(9-anthroiloxy)stearic acids. The treatment did not change the lateral diffusion of pyrenedodecanoic acid, either. However, generalized polarization of Laurdan fluorescence was higher in polyunsaturated fatty acid deficient microsomes as compared to the polyunsaturated fatty acid sufficient ones. This effect was also observed in liposomes of the total microsomal lipids, indicating that the changes in fatty acid composition resulting from polyunsaturated fatty acid deficiency produced a small but significant decrease in the rate of dipolar relaxation in the region of the lipid polar groups of the bilayer. The absence of lipid gel phase domains in rat liver microsomes was also indicated by Laurdan fluorescence features.