Roberta J. Pentney

Alcohol‐Induced Neurodegeneration: When, Where and Why?

This manuscript reviews the proceedings of a symposium organized by Drs. Antonio Noronha and Fulton Crews presented at the 2003 Research Society on Alcoholism meeting. The purpose of the symposium was to examine recent findings on when alcohol induced brain damage occurs, e.g., during intoxication and/or during alcohol withdrawal. Further studies investigate specific brain regions (where) and the mechanisms (why) of alcoholic neurodegeneration. The presentations were (1) Characterization of Synaptic Loss in Cerebella of Mature and Senescent Rats after Lengthy Chronic Ethanol Consumption, (2) Ethanol Withdrawal Both Causes Neurotoxicity and Inhibits Neuronal Recovery Processes in Rat Organotypic Hippocampal Cultures, (3) Binge Drinking-Induced Brain Damage: Genetic and Age Related Effects, (4) Binge Ethanol-Induced Brain Damage: Involvement of Edema, Arachidonic Acid and Tissue Necrosis Factor alpha (TNFalpha), and (5) Cyclic AMP Cascade, Stem Cells and Ethanol. Taken together these studies suggest that alcoholic neurodegeneration occurs through multiple mechanisms and in multiple brain regions both during intoxication and withdrawal.

Quantitative analysis of ethanol effects on Purkinje cell dendritic tree

Abstract An animals age at the time of ethanol treatment is related to dendritic vulnerability to ethanol damage. The dendritic networks of Purkinje neurons of 5-month-old rats treated with ethanol were quantitavely similar to those of age-matched pair-fed rats, but dendritic networks of 14-month-old rats treated with ethanol were significantly reduced compared with those of their age-matched pair-fed controls.

Ethanol enhances neurite outgrowth in primary cultures of rat cerebellar macroneurons

Effects of ethanol on neurite outgrowth and morphometry were investigated in primary cultures of rat cerebella. Cell cultures were prepared from cerebella on embryonic day 17 (E17) for treatment with a series of ethanol concentrations (50, 75, 100, 150 and 200 mM). Ethanol did not reduce neuronal survival or attachment to the substrate at any of the concentrations that were used. Treatment with 75 mM ethanol significantly enhanced neurite outgrowth. Measurements from dissociated cultures exposed to 75 mM ethanol immediately after plating showed a significant increase in the percentage of neurite-bearing cells after 8 and 24 h in vitro. Measurements of the area and perimeter of neuronal cell bodies in dissociated cell cultures showed that the cell bodies of ethanol-treated neurons were also larger than those of control neurons. Ethanol was also associated with significant increases in the total neuritic length per cell and in the length of the longest neurite in each cell. The mean number of neurite branches was also greater in the ethanol-treated neurons. Measurements from suspension cell cultures, in which dissociated cells were suspended overnight in the presence of 75 mM ethanol prior to plating, corroborated these results. These findings suggest that ethanol may have distinct effects on neurite initiation and outgrowth and branching. The cellular mechanisms involved and the functional significance of these effects are currently not known. The present results also indicated that high concentrations of ethanol (150-200 mM) and long periods of exposure (4-7 days) were required to produce toxic effects on neurons and glial cells in this system.

Morphometric analyses of purkinje and granule cells in aging F344 rats

In this study, the Purkinje neurons and granule neurons in the cerebellar cortex were studied in male Fischer 344 rats at 3, 9, 18, and 27 months of age. The numbers of Purkinje cells (PC) and granule cells (GC) in folia IV, VII, and X of the vermis were quantitated with the disector stereological technique. The numbers of PC and GC and the ratio of PC to GC were stable with advancing age. Measurements of the molecular layer, however, showed that this layer, the site of synaptic contact between GC and PC, decreased in volume with age.

Quantitative immunocytochemistry of glia in the cerebellar cortex of old ethanol-fed rats

It is clear from results of studies in this laboratory that chronic ethanol consumption causes regression of the extensive Purkinje neuron (PN) dendritic arbor. There are, however, a paucity of studies on the effects of chronic ethanol consumption on glia cells that reside in the molecular layer of the cerebellar cortex with PN dendrites. The purpose of the present study was to investigate the possibility that chronic ethanol consumption in old F344 rats results in gliosis within the molecular layer of the cerebellar cortex. Ten 12-month-old, male, F344 rats received a liquid diet containing 35% ethanol for 40 weeks. Pair-fed controls (n=10) received a liquid diet in which maltose dextrins were substituted for ethanol. Chow-fed rats (n=10) served as controls for age. At the end of the treatment period, rats were euthanized and perfused through the aorta, and cerebella were prepared for immunocytochemistry. Free floating sections were stained with (1) glial fibrillary acidic protein antibody for labeling of Bergmann glial cells and fibers, (2) OX-42 antibody for labeling of microglia, and (3) 0.5% cresyl violet for estimates of molecular layer volume. Results indicate that the densities of Bergmann glial cell processes and microglia within the cerebellar molecular layer are not altered by ethanol consumption.

Evidence for nonrandom regression of dendrites of Purkinje neurons during aging

This study examined dendritic networks of Purkinje neurons for randomness of age-related changes in cerebella from Fischer 344 rats at three ages. Terminal dendritic segments were studied in relation to their distance from the neuronal soma (path length). The data indicated that the terminal dendritic segments furthest from the cell body were preferentially affected during aging and there was a redistribution of the remaining terminal segments in cells from rats of the intermediate age group. The data also suggested that dendritic regrowth occurred in proximal regions of networks from rats in the oldest group.

Effects of chronic ethanol consumption on SER of Purkinje neurons in old F344 rats

The purpose of this study was to determine whether the observed swelling of smooth endoplasmic reticulum (SER) profiles in Purkinje dendrites in our old ethanol-fed F344 rats: (1) represented measurable dilatation, (2) was present in dendritic shafts and spines, and (3) was reversed following recovery from ethanol. Of the 45 rats in 3 treatment groups (chow-fed, pair-fed, and ethanol-fed), 30 rats were euthanized after 40 weeks, and 15 were maintained on rat chow for an additional 20-week recovery period. Electron microscopy of cerebellar preparations was used to analyze morphological alterations in SER profile size within the dendritic shafts and spines of Purkinje neurons. Results showed significant SER dilatation following 40 weeks of ethanol consumption, which disappeared after ethanol withdrawal.

The Number of Granule Cells and Spine Density on Purkinje Cells in Aged, Ethanol-fed Rats

The purpose of this study was to determine whether chronic intake of ethanol by aged F344 rats was associated with a reduction in parallel fiber input to cerebellar Purkinje neurons (PN). Previous results from this laboratory provided direct evidence that synaptic density in PN dendritic arbors was significantly decreased and indirect evidence that terminal dendritic segments of PN were deleted during chronic ethanol treatment. From these results, it was hypothesized that an ethanol-related deletion of PN terminal dendritic segments might result from 1) a reduction in parallel fiber input to PN from cerebellar granule neurons or 2) a reduction in dendritic spines, the postsynaptic sites for parallel fiber input to PN dendrites. Measurements of the total number of cerebellar granule neurons (GN) and the volume of the GN layer, and measurements of the density of spines on PN terminal dendritic segments were made in separate groups of aged, ethanol-treated and control rats. There were no significant ethanol-related changes in these parameters after 40-48 weeks of ethanol treatment.

The total numbers of cerebellar granule neurons in young and aged Fischer 344 and Wistar-Kyoto rats do not change as a result of lengthy ethanol treatment.

It is generally accepted that long term chronic ethanol consumption by young rats will lead to significant losses of cerebellar granule neurons (GN). A recent study in this laboratory showed, however, that 40 weeks of chronic ethanol consumption had no effect on the total numbers of GN in aged Fischer 344 rats (F344). The goals of the present study were to determine whether F344 GN were resistant to ethanol toxicity only in aged rats and whether resistance of GN in aged rats to ethanol toxicity occurred only in the F344 strain. To accomplish those goals, young and aged adult F344 and Wistar-Kyoto (WKY) rats were treated chronically with ethanol for 40 weeks during the first or second half of their life span. In each rat the total numbers of GN were estimated with the optical fractionator and the volumes of the GN layer were estimated according to Cavalieri’s theorem. After the 40 weeks of ethanol, there were significant age-related differences in the total numbers of GN in the F344 rats. There were also significant strainrelated differences in the total numbers of GN and volumes of the GN layer. There were no significant ethanol-related differences, however, in numbers of cerebellar GN or volumes of the GN layer in F344 rats or WKY rats. The results presented here show that consumption of ethanol over long periods of time had no effect on the total numbers of cerebellar GN or the granular layer volumes in young or aged F344 or WKY rats.

Biochemical and structural brain alterations in female mice with cerebral pyruvate dehydrogenase deficiency

Pyruvate dehydrogenase complex (PDC) deficiency is an inborn metabolic disorder associated with a variety of neurologic abnormalities. This report describes the development and initial characterization of a novel murine model system in which PDC deficiency has been introduced specifically into the developing nervous system. The absence of liveborn male and a roughly 50% reduction in female offspring following induction of the X‐linked mutation indicate that extensive deficiency of PDC in the nervous system leads to pre‐natal lethality. Brain tissue from surviving females at post‐natal days 15 and 35 was shown to have approximately 75% of wild‐type PDC activity, suggesting that a threshold of enzyme activity exists for post‐natal survival. Detailed histological analyses of brain tissue revealed structural defects such as disordered neuronal cytoarchitecture and neuropil fibers in grey matter, and reduced size of bundles and disorganization of fibers in white matter. Many of the histologic abnormalities resemble those found in human female patients who carry mutations in the X‐linked ortholog. These findings demonstrate a requirement for PDC activity within the nervous system for survival in utero and suggest that impaired pyruvate metabolism in the developing brain can affect neuronal migration, axonal growth and cell–cell interactions.