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Objective: Our purpose was to test the hypothesis that prenatal ethanol exposure alters the hippocampal muscarinic cholinergic neurochemistry of albino rats., Study Design: Ethanol was administered in a liquid diet to pregnant albino Sprague-Dawley rats. Liquid diet control animals received the same diet in which ethanol was replaced by an isocaloric amount of maltose-dextrin. Chow-fed control animals were fed laboratory chow as desired. Progeny were killed at 90 days of age, and their hippocampi were analyzed for muscarinic cholinergic receptors by use of tritiated quinuclidinyl benzilate., Results: Prenatal ethanol exposure produced a statistically significant decrease in the number of muscarinic receptors in males. Similar trends were noted in females, but the results were not statistically significant., Conclusion: Prenatal ethanol treatment caused long-lasting alterations in the muscarinic cholinergic receptors of the hippocampus in male rats.

By combining techniques designed to increase the amount of horseradish peroxidase (HRP) available for transport and to improve the sensitivity of the reaction for detecting HRP, we have produced an anterograde HRP technique suitable for labeling sparse efferent projections. The efficacy of this light, microscopic technique was demonstrated by lebeling the hippocampal commissural and crossed temporo-ammonic projections to the dentate gyrus of the rat.

The effect of ethanol on hippocampal axonal sprouting was studied with a histochemical technique for identifying acetylcholinesterase. Unilateral lesion of the entorhinal cortex in adult rats produced an increase in the density of acetylcholinesterase staining in the outer molecular layer and a concomitant increase in the width of the pale-staining commissural-associational zone of the dentate gyrus. Other rats were given ethanol (11.3 +/- 0.45 grams per kilogram) for 2 weeks before and 9 days after receiving the lesion. Ethanol abolished the expansion of the commissural-associated zone. The effect of ethanol on sprouting axons suggests that it may inhibit recovery of function after brain injury.

Using both the Timm's sulfide silver and anterograde horseradish peroxidase (HRP) histochemical techniques, we have demonstrated that granule cell in the dentate gyrus of the rat have topographical projections to both suprapyramidal and distal infrapyramidal terminal fields in the rostral one-third of the hippocampus. An anterograde HRP technique modified for visualization of axonal morphology indicated that intrapyramidal mossy fiber axons possess classic periodic swellings characteristic of suprapyramidal mossy fibers. Most distal infrapyramidal mossy fiber axons appear to be in position to make synaptic contact with deeper-lying neurons in the pyramidal cell layer.

Three separate groups of pregnant Sprague-Dawley rats were (1) fed a liquid diet containing 35% ethanol-derived calories, or (2) pair-fed this diet containing an isocaloric amount of maltose-dextrin instead of ethanol,or (3) fed laboratory chow ad libitum. Their offspring were killed after reaching at least 60 days of age, and their brains were processed with the Timm's or horseradish peroxidase histochemical techniques. Both procedures revealed that intrauterine exposure to ethanol produced a dramatic change in the topography of hippocampal mossy fibres. An aberrant distal infrapyramidal mossy fibre terminal band was present at mid-temporal levels (where it does not normally occur). Pair-fed controls did not show the aberrant band. Rats exposed to ethanol after birth (using an artificial rearing procedure) showed even greater aberrations in the mossy fibre terminal field than rats exposed to ethanol in utero. Thus, postnatal exposure to ethanol (equivalent to the third trimester in the human) produced more striking changes in the mossy fibre terminal field than exposure to ethanol during the entire length of gestation in the rat (equivalent to the first and second trimesters in the human).

Rats exposed to ethanol throughout their gestation were found to have abnormally distributed mossy fibers in temporal regions of the hippocampus. This demonstrates that prenatal exposure to ethanol causes alterations in neuronal circuitry that persist to maturity. Such defects may play a role in the mental retardation often observed in children with fetal alcohol syndrome.

Rats were exposed to ethanol from postnatal day 4 to day 10, using an artificial rearing procedure. One group received ethanol administered by a cyclic regimen, which resulted in high peak blood ethanol concentrations (BACs). Another group received the same daily dose of ethanol using a regimen which resulted in stable, moderate BACs. Two control groups consisted of rats reared artificially but not exposed to alcohol and rats reared normally by dams. All rats were raised to adulthood. Approximately half the rats were tested for behavioral activity levels and then sacrificed in order to assay the hippocampus for levels of cyclic GMP and protein after incubation with bethanechol (a muscarinic cholinergic agonist). The remaining rats were sacrificed and their hippocampi were assayed for muscarinic cholinergic receptors and protein content. Exposure to cyclic BACs with high peaks but not exposure to stable moderate BACs resulted in greater behavioral activity levels in both sexes and also a greater cyclic GMP responsiveness to a muscarinic cholinergic agonist. Furthermore, exposure to cyclic BACs with high peaks resulted in a significantly larger dissociation constant and a greater number of muscarinic cholinergic receptors. There were no differences among groups or sexes in protein content of the hippocampus. There were no interactions of the alcohol treatment with sex. Thus, in adult rats, exposure to alcohol during the brain growth spurt results in an increase in the dissociation constant and number of muscarinic cholinergic receptors in the hippocampus.