Brian L. G. Morgan

Effects of essential fatty acid deficiency during late gestation on brain N-acetylneuraminic acid metabolism and behaviour in the progeny.

1. Rat dams given a diet containing 100 g maize oil/kg for approximately two weeks before mating and during the first 14 d of gestation, were given the same diet or one containing 100 g hydrogenated coconut oil/kg (essential fatty acid (EFA)-deficient) in place of maize oil until parturition. After parturition the dams were given the same diets and all progeny were weaned to the maize oil diet at 21 d of age. Brain N-acetylneuraminic acid (NeuNAc) content as well as neuraminidase (sialidase; (EC 3.2.1.18), and cytidine monophosphate N-acetylneuraminic acid synthetase (CMP-NeuNAc synthetase) activities were measured at days, 7, 14, 21 and 168 in the progeny. Y-maze learning was measured at 168 d. 2. Brain weight was independent of dietary fat at all ages. 3. Lack of EFA in the maternal diet during gestation and lactation depressed ganglioside and glycoprotein NeuNAc levels and the activities of sialidase and CMP-NeuNAc synthetase. 4. Maternal dietary deprivation of EFA irreversibly impaired learning behaviour of the progeny. A relationship exists between early exposure to EFA deficiency and learning potential of the progeny.

Delta-9-tetrahydrocannabinol during pregnancy in the rat: I. Differential effects on maternal nutrition, embryotoxicity, and growth in the offspring☆

Either 15 or 50 mg/kg of delta-9-tetrahydrocannabinol (THC) in sesame oil was administered by gastric intubation to gravid rats during the last two weeks of gestation. A pair-fed control group was administered the vehicle alone and allowed to eat and drink only the amount consumed by the 50 mg/kg group on the same gestation days. A nontreated control group was left undisturbed during pregnancy. All treated and control litters were fostered at birth to untreated dams. Among the dams receiving 50 mg/kg of THC, food and water intake was initially reduced to 75-80% of nontreated controls but then recovered over 3-4 days to approximately a 15-20% reduction until term. Compared with the nontreated dams, both THC dose-level groups and pair-fed control dams gained significantly less body weight from conception to term. Offspring mortality did not differ between the nontreated and pair-fed controls but was significantly higher among both dose-level THC exposed groups. In addition, there was a dose-related increase in the sex-ratio of live male to female offspring as well as significant effects on rate of growth for both sexes. The results are discussed with respect to published animal and clinical studies of cannabinoid exposure during pregnancy.

The subcellular localization of administered N-acetylneuraminic acid in the brains of well-fed and protein restricted rats

1. This study investigated the subcellular localization of injected N-acetylneuraminic acid (NeuNAc) in brain. Forty pregnant rats were distributed into four groups. Two groups were given a 200 g casein/kg diet and the other two groups a 100 g casein/kg diet throughout gestation. One group from each of the low- and high-protein groups were given their respective diets for the first 11 d of lactation. On day 12 of lactation, 2.5 microCi [14]NeuNAc/kg body-weight were injected intraperitoneally into their pups. After 1 h the pups were killed, their brains removed and subjected to subcellular fractionation. On day 16 of lactation the other two groups were similarly treated. 2. In all groups of animals 80% of the [14C]NeuNAc incorporated into the brains was found in the synaptosomal fraction and the remainder distributed among the other subcellular fractions in proportion to their total NeuNAc content. 3. These results suggest that NeuNAc exerts its effects on behaviour via the synaptic membrane.

Delta-9-tetrahydrocannabinol during pregnancy in the rat: II. Effects on ontogeny of locomotor activity and nipple attachment in the offspring

Either 15 or 50 mg/kg delta-9-tetrahydrocannabinol (THC) in sesame oil was administered by gastric intubation to gravid rats during the last two weeks of gestation. A pair-fed control group was administered the vehicle alone and allowed to eat and drink only the amount consumed by the 50 mg/kg group on the same gestation days. A nontreated control group was left undisturbed during pregnancy. All treated and control litters were fostered at birth to untreated dams. Intact litters from the two THC treated and the two control groups were tested at 3-day intervals from birth to 32 days of age for differences in locomotor activity. In addition, pups were tested for nipple attachment on days 2, 5, 8, 11, and 14 of age. There were no differences in locomotor activity among any of the groups although activity level varied during development. Pups from dams exposed to 50 mg/kg of THC, as well as the pair-fed controls, displayed significantly longer latencies to attach to a nipple. These results suggest that the impaired nipple attachment observed among the high-dose offspring was not a primary effect of THC, but rather was secondary to the significant reduction of food and water intake among the dams. The behavioral findings are discussed with respect to other animal and clinical reports of prenatal cannabinoid exposure.

Delta-9-tetrahydrocannabinol during pregnancy in the rat: effects on development of RNA, DNA, and protein in offspring brain.

Either 15 or 50 mg/kg of delta-9-tetrahydrocannabinol (THC) was administered from Day 2 through Day 22 of gestation. Pair-fed and nontreated groups served as controls and all treated and control litters were fostered at birth to untreated dams. To determine the effects of THC on offspring brain development, DNA, RNA and protein values were determined at 7, 14, and 21 days of postnatal age. DNA and RNA levels appeared unaffected by THC but brain protein levels of the 50 mg/kg offspring were significantly lower than in the other groups at Day 7 and 14. This suggests that the high THC dose inhibited protein synthesis for at least the first 14 days of life. Subsequently, protein levels of the 50 mg/kg offspring increased rapidly so that there were no differences between any of the groups at 21 days of age. These findings for developing CNS parallel the delayed rate of somatic growth previously reported from our laboratory and suggest a transitory rather than a permanent effect of THC on both somatic and brain growth. We also found that THC produces a significant dose-related increase in the sex-ratio of live male-to-female offspring, a finding we have reported previously.

A Possible Relationship between Brain N-Acetylneuraminic Acid Content and Behavior

Summary Repeated intraperitoneal injections of N-acetylneuraminic acid (NANA) into malnourished and well-fed rats during the brain growth spurt were associated with a permanent increase in NANA concentrations in brain gangliosides and glycoproteins. Further, there was an alleviation of some expected behavioral abnormalities in the malnourished group and an above normal behavioral performance shown by the well-fed pups. The results suggest the existence of a relationship between NANA and behavior.

A possible control of food intake during pregnancy in the rat

1. Brain hypothalamic concentrations of serotonin and norepinephrine have been implicated in the control of food intake. During pregnancy and lactation a rat dams food consumption is increased and so a study was performed to ascertain whether this was associated with changes in the hypothalamic content of serotonin and norepinephrine. 2. In the first experiment, forty-eight Sprague Dawley rat dams were given a diet containing 250 g casein/kg adlib. After 2 weeks, eight were killed and their hypothalmic analysed for the previously-mentioned neurotransmitters. The rest were mated and continued on the diet. On each of days 7, 14 and 20 of gestation, day 14 of lactation and 2 weeks after weaning of their pups a further eight dams were killed and their hypothalami assayed as described previously. Food intake was monitored throughout the experimental period. 3. The increase food intake of the dams during gestation and lactation increased to the same extent as elevation in hypothalamic norepinephrine content and depression in serotonin content. After lactation food intake returned to pre-pregnancy levels as did the hypothalamic levels of norepinephrine and serotonin. 4. By using the same experimental design but limiting the increase in food intake in pregnancy and lactation to half the expected amount the same changes were found in hypothalamic norepinephrine and serotonin contents. 5. The possibility of hypothalamic neurotransmitter contents controlling food intake is discussed.

The effects of folic acid supplementation during pregnancy in the rat.

1. Twenty-four Sprague-Dawley female rats were randomly assigned to three groups (groups A, B, C). Group A was given a folic acid-free diet and groups B and C received 0.0018 g folic acid/kg diet. Rats in group C were also given a supplement of 1 mg folic acid/d by intraperitoneal injection. 2. After 14 d of feeding the rats were mated. The diets were continued throughout gestation. On day 21, of gestation the dams were killed and their livers and products of conception assayed for RNA, DNA, protein and tetrahydrofolate dehydrogenase (5,6,7,8-tetrahydrofolate dehydrogenase; EC 1.5.1.3) activity. 3. The foetuses, placentas and livers from supplemented rats (group C) were significantly larger than those from groups A and B and had a higher content of RNA, DNA and protein. Those tissues from group A dams were smaller than those from the other groups and had a correspondingly reduced nucleic acid and protein content. 4. The activity of tetrahydrofolate dehydrogenase, the first and rate-limiting enzyme in the metabolism of folate, was increased in the folate supplemented rats (group C) and reduced in the rats given a folic acid-free diet (group A). These changes in enzyme activity could explain the differences in nucleic acid biosynthesis and growth shown by the different groups.

Nutrition and Cellular Growth of the Brain

The most common method employed in altering the nutritional status of neonatal rats is to vary the number of pups nursing from a single mother. The normal rat litter consists of from 8 to 12 pups; a nursing group of 10 animals has arbitrarily been considered normal. Malnutrition is imposed by increasing the size of the nursing group to 18 animals and overnutrition by decreasing the size to 3 animals. But in addition to altering the nutrition, this changes the amount of maternal stimulation available to each pup. More recently, other methods of inducing undernutrition have been employed. Protein restriction in the lactating mother reduces the quantity of milk produced without altering its composition. Allowing the animals to nurse for only a single 8-hr period per day also reduces the quantity of milk consumed. In a combination of these approaches, protein-restricted mothers have been given an increased number of animals to nurse. All these methods produce a total caloric restriction as well as a restriction in individual nutrients, the most important of which is probably protein. So far, all the methods have had comparable effects on brain growth, and we will therefore examine them together.

Brain β-Hydroxybutyrate Utilization in Neonatal Hypothyroidism in Rats

The in vitro and in vivo utilization of 14C-β-hydroxybutyrate (BOHB) by the brain in neonatal hypothyroidism was studied in rat pups made hypothyroid by feeding dams a diet containing 0.02% propylthiouracil during pregnancy and lactation. In the in vitro studies, the oxidation of BOHB to CO2 was lower at 14 and 21 days of age in hypothyroid pups than in pair-fed controls but higher at 35 days. The incorporation of BOHB into brain lipid was significantly higher in hypothyroid pups at 14, 21 and 35 days. In contrast, in the in vivo studies, incorporation of BOHB into lipid was significantly lower in hypothyroid pups at 14 and 21 days. These results suggest that hypothyroid pups may have a normal or increased capability to use BOHB for lipid synthesis, but were unable to achieve normal rates of utilization in vivo, possibly due to a decreased substrate availability. These findings may have implications for the decreased brain lipid content characteristic of neonatal hypothyroidism.