Synaptic membrane phospholipids: effects of maternal ethanol consumption.

The cholesterol and phospholipid content and phospholipid composition were determined in synaptic membranes from the 17- to 31-day-old offspring of rats that were pair-fed either a control or 6.6% (v/v) ethanol liquid diet on a chronic basis prior to parturition. At all ages examined, the major synaptic membrane phospholipid was phosphatidyl choline (greater than 40%). Other prominent synaptic membrane phospholipids included phosphatidyl ethanolamine (approximately 17 to 21%), ethanolamine plasmalogen (approximately 5 to 16%), and phosphatidyl serine (approximately 13%). Smaller proportions of sphingomyelin (4 to 7%), phosphatidyl inositol (approximately 1%), and phosphatidic acid (approximately 1%) were detected. Between 17 and 31 days of age, there was a significant decrease in the proportion of phosphatidyl choline and a significant increase in the proportion of ethanolamine plasmalogen. When the offspring of control and ethanol-treated rats were compared, no significant differences were found in either the yield of synaptic membrane protein, or in the concentration of synaptic membrane cholesterol and total phospholipid. However, the proportion of ethanolamine plasmalogen was significantly decreased in the 24-day-old offspring of ethanol-treated rats, suggestive of a delay in the normal development-related increase of this lipid. In addition, there was a small increase in the proportion of sphingomyelin in the 31-day-old offspring of ethanol-treated rats.

Biosynthesis and transport of gangliosides in peripheral nerve.

Radiolabelled glucosamine was injected into L-7 dorsal root ganglion (DRG) of rabbits. At several different times after injection DRG, lumbosacral trunks (LST) and sciatic nerves (SN) removed and gangliosides extracted. Two and 3 weeks after injection the amounts of radioactivity in the ganglioside fractions of LST SN were significantly higher than at days 1 and 2. The TCA soluble radioactivity decreased dramatically over the same time period. Colchicine prevented the appearance of radiolabelled lipid in LST and SN. From these experiments we conclude that some ganglioside is synthesized in the neuronal cell bodies of DRG and transported in the axons of the sciatic nerve. In another experiment the sciatic nerve was transected and ends separated to prevent regeneration. Ganglioside synthesis and transport were studied in these animals the same way as the previous experiment. There was no difference the amount of radiolabelled ganglioside that was isolated from DRG or LST of transected compared with control nerves. The behavior of several potential acid soluble contaminants was studied in several steps used to isolate gangliosides. Of those studied only CMP-NeuAc could cause significant contamination of the final ganglioside preparation.

Rabbit sciatic nerve as a model for studying ganglioside metabolism in regenerating nerve.

The rabbit sciatic nerve contains a sufficient amount of gangliosides for chemical analyses. We have developed a method by which the dorsal root ganglion of L-7 can easily be injected with a radiolabeled precursor of ganglioside biosynthesis. We have used this to demonstrate that gangliosides are synthesized in dorsal root ganglia and then transported into peripheral nerve. Sialyltransferase-I activity is present in dorsal root ganglia, lumbosacral trunk, and sciatic nerve. This model is now well characterized for studies on ganglioside metabolism in degenerating and regenerating peripheral nerve.

Peripheral nerve phospholipid composition: development in normal nerve and age-dependent changes in Wallerian degenerated nerve.

The phospholipid composition of normal peripheral nerve as a function of developmental age as well as that of Wallerian-degenerated nerve as a function of age at nerve transection and duration of Wallerian degeneration have been quantitated in rabbit sciatic nerve. During development, increases in the proportions of ethanolamine plasmalogen, sphingomyelin, and combined phosphatidyl serine plus phosphatidyl inositol and decreases in the proportions of phosphatidyl choline and phosphatidyl ethanolamine correlated well with the concurrent myelin accretion. During Wallerian degeneration, age-dependent changes in phospholipid composition were observed. The large and statistically significant increase in the proportion of phosphatidyl choline and decrease in the proportion of ethanolamine plasmalogen were manifest promptly in nerves transected at 2 weeks of age but in a delayed manner in nerves transected at 8, 12, and 20 weeks of age. The rate of loss of individual phospholipids was greater in nerves transected at younger ages. The findings from normal developing peripheral nerve may well serve as baseline data for subsequent studies of phospholipid composition in pathological peripheral nerve. The findings from Wallerian-degenerated peripheral nerve provide additional evidence for age-dependent chemical changes occurring in Wallerian-degenerated peripheral nerve that may be of significance in explaining the superior functional recovery from peripheral nerve injury observed in younger compared with older subjects.

Chemical and morphological studies on garfish peripheral nerves.

Gangliosides were extracted, separated by thin layer chromatography, and quantitated in three cranial nerves of the garfish (Lepisosteus osseus): the completely unmyelinated olfactory nerve (OLF), and two nerves composed of both myelinated and unmyelinated fibers, viz., the main trunk of the maxillary nerve (MAX) and a branch of the maxillary nerve (BR-MAX). Morphological studies on each of these nerves were done to verify that the OLF had been excised free of any contamination from the accompanying myelinated BR-MAX, to aid in the interpretation of the biochemical findings, and to clarify the nature of the OLF supporting cell. The chief chemical findings were (1) documentation of the presence of gangliosides in nerves previously thought not to contain them, (2) demonstration that gangliosides can be associated with unmyelinated nerves, (3) demonstration of a greater proportion of one simple ganglioside (G-6) in the OLF but greater proportions of two complex gangliosides (G-2 and G-0) in the MAX and BR-MAX, and (4) that either GM4 or a variant of the GM3 is present in OLF. The morphological findings with respect to the difficulty of ascribing only peripheral nervous system characteristics to the OLF supporting cell are discussed in relation to the ganglioside band chromatographing slightly ahead of GM4 in the OLF.

Wallerian degeneration of peripheral nerve. Age-dependent loss of nerve lipids.

The age-dependent loss of the major peripheral nerve lipids (cholesterol, phospholipid, and total galactolipid) was quantitated over a period of 9 weeks of Wallerian degeneration induced by surgical transection of rabbit sciatic nerves in animals of several ages. Proportionate losses of these lipids were determined by calculating the content of each lipid on a per nerve and on a per gram fresh weight basis remaining after a given period of Wallerian degeneration as a percent of original normal values at several time following surgery. The proportionate loss of each lipid from the distal stump was the most prompt and the most complete in nerves transected at 2 weeks of age, and the least in nerves transected at 20 weeks of age. The prompter clearance of these lipids from younger than older degenerating nerve gives convincing evidence that the suggestion from light-microscopic studies of faster clearance of neural debris in younger than in older animals is correct. A possible relationship between these biochemical findings and the phenomenon of greater functional recovery from peripheral nerve injury in younger than in older subjects is discussed.

Central nervous system myelination in rats exposed to ethanol in utero.

Pregnant rats were pair-fed using isocaloric control or 6.6% (v/v) ethanol liquid diets. At 18, 25, and 53 days, the in vivo incorporation of [3H] leucine and [14C] glucose into proteins and lipids of three central nervous system (CNS) myelin subfractions was examined in the offspring (control and ethanol pups) of control and ethanol-treated females. With few exceptions, the pattern of CNS myelination appeared near-normal in the ethanol pups. The ethanol pups had brain weights and total myelin protein content within the normal range. In addition, the amount of protein in light, medium, and heavy myelin was normal at 18 and 25 days. However, the ethanol pups had an excess of the chemically and morphologically immature heavy fraction at 53 days. The incorporation of [3H] leucine and [14C] glucose into myelin subfractions and separated proteins and lipids was similar in control and ethanol pups. The findings of near-normal CNS myelination in the offspring of female rats fed an ethanol liquid diet during gestation differ from our previous findings of a premature onset and slowdown of active CNS myelination in the offspring of female rats that consumed an ethanol liquid diet for one month prior to conception as well as during gestation.