Ezio A. Moscatelli

Effects of chronic consumption of ethanol and low-thiamin, low-protein diets on the lipid composition of rat whole brain and brain membranes

Four groups of rats were used in a nutritionally-controlled study of effects of chronic ethanol consumption on brain membrane lipid composition. Rats chronically consuming ethanol were fed high-nutrient or low-thiamin, low-protein diets. After 4 months, lipid analyses were performed on brains, brain microsomes and myelin from each group and from pair-fed, non-ethanol controls. Among the effects of ethanol was an increase of the relative proportion of cholesterol in microsomal lipids while there was decrease of it in myelin. Ethanol also increased plasmenylethanolamine while decreasing phosphatidylethanolamine proportions in myelin and in whole brain lipids, decreased the total lipid phosphorus of whole brain, and elevated the proportion of phosphatidylserine in microsomal and whole brain lipids. Effects of poor diet generally did not interfere with ethanol effects except in the case of microsomal lipids, where it apparently prevented an ethanol-induced increase in proportion of cholesterol. These changes may be adaptive responses to the fluidizing effect of ethanol on membranes.

Methanolysis of cerebrosides with boron trifluoride-methanol

A time study was carried out to determine conditions under which BF3-methanol could be used reliably for methanolysis of cerebrosides. Results indicate that heating at 100 C in a sealed tube with 14% w/v BF3-methanol for 60 min yields satisfactory cleavage of cerebrosides to fatty acid methyl esters without significant destruction of normal or hydroxy fatty acids.

Synaptosomal and Brain Mitochondrial Lipids in Hibernating and Cold‐Acclimated Golden Hamsters

Synaptosomes and mitochondria were isolated from the brains of warm‐adapted, hibernating, and cold‐acclimated golden hamsters (Mesocricetus auratus). Lipid extracts of these subcellular fractions were prepared and assayed for plasmenylethanolamin (ethanolamine plasmalogen) and cholesterol levels. The ganglioside composition of synaptosomes was also determined. Samples from the hibernating animals showed characteristic changes in lipid composition. These changes include decreases in plasmenylethanolamine levels and a shift in the ganglioside composition toward a higher percentage of the more polar gangliosides. Those animals which were exposed to cold and did not hibernate (cold‐acclimated) showed no such changes. Fatty acid analyses of synaptosomal and mitochondrial ethanolamine glycerophospholipids demonstrated a similar trend. Samples from hibernators showed decreases in 16:0, 18:0, and 22:6 (n‐3), and increases in 16:1, 18:1, and 20:4 (n‐6) fatty acids. No changes were detectable in samples from cold‐acclimated animals, indicating that hibernating and cold‐acclimated hamsters represent chemically distinct populations.

Effects of plasmenylethanolamine on the dynamic properties of the hydrocarbon region of mixed phosphatidylcholine-phosphatidylethanolamine aqueous dispersions. A spin label study

Spin-labeled aqueous dispersions of total phospholipid extracts from whole brains of hibernating hamsters and rats chronically consuming ethanol were compared with dispersions from control animals. Order parameter values and approximate rotational correlation times for the nitroxide spin labels indicated that ethanol consumption results in an adaptive decrease in bilayer membrane fluidity, while hibernation produces increases in fluidity. Since it has been proposed that changes in plasmenylethanolamine such as those seen with hibernation play a role in the homeoviscous adaptation of brain membranes, electron spin resonance studies using aqueous phospholipid dispersions containing equimolar mixtures of rat brain phosphatidylethanolamine and phosphatidylcholine, or synthetic dioleylphosphatidylcholine and dioleylphosphatidylethanolamine, and brain plasmenylethanolamine were performed. The molar amount of plasmenylethanolamine was varied within the ethanolamineglycerophospholipid fraction of each dispersion. Order parameter values of spin labels in liposomes containing brain phosphatidylcholine and phosphatidylethanolamine increased in parallel with increases in plasmenylethanolamine concentrations, indicating that fluidity was decreasing. Liposomes composed of synthetic dioleyl phospholipids exhibited biphasic changes in order parameter (S) values as plasmenylethanolamine replaced the diacyl form. Below 30% (mol%) plasmenylethanolamine, S values decreased, while above 30%, S values were seen to increase; indicating an initial fluidization, followed by a decrease in fluidity.

The effect of hibernation on the positional distribution of ethanolamineglycerophospholipid fatty acids in hamster brain membranes

Microsomal and myelin membrane fractions were prepared from the brains of warm-adapted (room temperature) and hibernating Syrian Hamsters (Mesocricetus auratus). Ethanolamineglycerophospholipids were isolated and subjected to a fraction scheme to separate the fatty acids of the plasmenylethanolamine and the phosphatidylethanolamine 1 and 2 positions. The major changes in microsomal phosphatidylethanolamine with hibernation were relative increases in 18∶1 at the 1 position and 20∶4(n−6) in the 2 position. In myelin, 18∶1 increased and 18∶0 decreased at the 1 position while the 2 position showed an increase in 16∶0 and a decrease in 22∶6(n−3). Plasmenylethanolamine fatty acids also changed in microsomes and myelin although the magnitudes were not as great and confined to longer chain fatty acids. In both membranes, fatty acid alterations were position-specific, and no new types of fatty acids were introduced at any position.

EFFECTS OF CHRONIC CONSUMPTION OF ETHANOL AND SUCROSE ON RAT WHOLE BRAIN 5-HYDROXYTRYPTAMINE

Abstract— Whole brain 5‐hydroxytryptamine (5‐HT) levels were determined after 3 weeks in rats chronically consuming ethanol, in pair‐fed controls, and in pair‐fed controls consuming sucrose in quantities isocaloric to the ethanol of the first group. A cryogenic harvesting and storage technique was developed to insure accurate measurement of whole rat brain 5‐HT. It was found that whole brain 5‐HT is completely stable for at least 16 days after decapitation into liquid N2 followed by storage of frozen whole heads at – 70°C. Using this technique and spectrophotofluorometric 5‐HT assays, no differences were found in whole brain 5‐HT among the three groups. Chronic consumption of ethanol and sucrose apparently lead to no chronic change in rat whole brain 5‐HT.

Gas—liquid chromatographic analysis of phenolic and non-phenolic chlorpromazine metabolites in the urine of chronic schizophrenic patients

Abstract The distribution of chlorpromazine and its phenolic and non-phenolic metabolites was studied in the urine of schizophrenic patients by gas—liquid chromatography on OV-17, using a flame ionization detector. Unconjugated phenolic metabolites, 7-hydroxychlorpromazine and 7-hydroxydemethylchlorpromazine in the crude extract were determined as their trimethylsilyl derivatives using N-trimethylsilylimidazole in the presence of pyridine. Trimethylsilyl 7-hydroxychlorpromazine and trimethylsilyl 7-hydroxydemethylchlorpromazine have greatly improved chromatographic properties and enable the investigator both to separate and identify the 7-hydroxy derivatives easily by gas—liquid chromatography. In addition to the phenolic metabolites, the major non-phenolic derivatives identified in urine include chlorpromazine N-oxide, chlorpromazine, demethylchlorpromazine, chlorpromazine sulfoxide and demethylchlorpromazine sulfoxide.

Effects of storage conditions on rat brain ethanolamine glycerophospholipids, cerebrosides, and cholesterol

The effects of storage on rat brain lipid composition were studied in terms of ethanolamine glycerophospholipid, cerebrosides, and cholesterol. Rat brains were stored at several combinations of temperature and time. Storage conditions were: 2 hr at room temperature, 12 hr of refrigeration, and a sequence of both of these conditions. Two-dimensional thin layer chromatography followed by colorimetric analyses of eluted lipids were used to determine molar ratios of phosphatidylethanolamine, ethanolamine plasmalogen, lysophosphatidylethanolamine, and cerebrosides. Cholesterol was also determined. These studies revealed small but significant increases in lysophosphatidylethanolamine in all three cases. A slight increase was also noted in the apparent molar proportion of cholesterol.

An improved method for the determination of creatinine in urine containing chlorpromazine metabolites

Abstract A simple spectrophotometric method which uses the Jaffe reaction is described for the determination of creatinine in urine containing seven metabolites of chlorpromazine. The phenothiazines are preferentially adsorbed on a column of Amberlite XAD-2, while creatinine does not bind and is recovered quantitatively. This procedure enables the investigator to relate chlorpromazine excretion to creatinine levels and may be useful in correcting the errors so commonly encountered in obtaining a 24 hr urine sample for metabolic studies.