K. S. Bedi

Early life undernutrition in rats

1. Male rats were undernourished either during the gestational and suckling periods or for a period of time immediately following weaning. Some rats were killed at the end of the period of undernutrition; others were nutritionally rehabilitated for lengthy periods of time before examination. Two muscles, the extensor digitorum longus (EDL) and soleus (SOL) were studied from each rat. Histochemically-stained transverse sections of these muscles were used to determine total number of fibres, the fibre cross-sectional areas and the relative frequency of the various fibre types. 2. All rats killed immediately following undernutrition showed significant deficits in body-weight, muscle weight and fibre cross-sectional area compared to age-matched controls. 3. Animals undernourished during gestation and suckling and then fed normally for 5 months showed persistent and significant deficits in body-weight, muscle weight, muscle weight and total fibre number. There were also significant deficits in mean fibre cross-sectional area of each fibre type except for red fibres in the EDL. No difference in the volume proportion of connective tissue was found. 4. Rats undernourished after weaning and then fed ad lib. for approximately 7 months had normal body-and muscle weights. Their muscles showed no significant differences in total fibre number, relative frequency of the various fibre types, fibre size or volume proportion of connective tissue. 5. These results indicate that, although the effects on rat skeletal muscle of a period of undernutrition after weaning can be rectified, undernutrition before weaning causes lasting deficits.

Inter-animal variation and its influence on the overall precision of morphometric estimates based on nested sampling designs

This paper provides additional experimental evidence that biological variation between individuals is likely to be the major factor influencing the overall precision and efficiency of nested sampling schemes for morphometric analysis of thin sections.

Spatial learning ability of rats undernourished during early postnatal life

Experiments to determine whether undernutrition during early life results in deficits in spatial learning behavior has produced conflicting results. It was hypothesized that this may be due to the differing degrees of undernutrition used in the various studies, and/or to the timing of the testing procedures with respect to the period of food deprivation. These possibilities were tested by undernourishing rats between birth and 30 days of age to two different levels (i.e., level-1 and level-2). The degree of undernutrition was greater at level-2 than level-1. Behavioral testing of these rats and well-fed controls was carried out in the Morris water maze when they were between either 35 and 65 or 170 and 200 days of age. Statistical analyses of the escape latency data from these experiments revealed that rats tested almost immediately after the period of undernutrition have alterations in their spatial learning behavior compared with controls. However, even a short period of nutritional rehabilitation removed any differences between control and previously undernourished groups, irrespective of the level of undernutrition.

Early maternal deprivation induces alterations in brain-derived neurotrophic factor expression in the developing rat hippocampus

The effects of maternal deprivation (MD) during early postnatal life on the brain-derived neurotrophic factor (BDNF) level were investigated in the present study. Wistar rats were assigned to either maternal deprivation or mother-reared control (MRC) groups. MD manipulation was achieved by separating rat pups from their mothers for 3h a day during postnatal days (PND) 10-15. At 16, 20, 30, and 60 days of age, the level of BDNF mRNA in the hippocampal formation of each group was determined using real-time PCR analysis. Early postnatal maternal deprivation of rat pups resulted in a significant increase in body weight at 60 days of age. The expression of BDNF mRNA in the hippocampus was significantly decreased at 16 days of age, and increased at 30 and 60 days of age. These data indicate that even a brief period of maternal deprivation during early postnatal life can affect hippocampal BDNF expression.

Spatial learning ability of rats following differing levels of exposure to alcohol during early postnatal life

Rats exposed to a relatively high dose (7.5 g/kg body weight) of alcohol on either the fifth or tenth postnatal day of age have been reported to have long-lasting deficits in spatial learning ability as tested on the Morris water maze task. The question arises concerning the level of alcohol required to achieve this effect. Wistar rats were exposed to either 2, 4 or 6 g/kg body weight of ethanol administered as a 10% solution. This ethanol was given over an 8-h period on the fifth postnatal day of age by means of an intragastric cannula. Gastrostomy controls received a 5% sucrose solution substituted isocalorically for the ethanol. Another set of pups raised by their mother were used as suckle controls. All surgical procedures were carried out under halothane vapour anaesthesia. After the artificial feeding regimes all pups were returned to lactating dams and weaned at 21 days of age. The spatial learning ability of these rats was tested in the Morris water maze when they were between 61-64 days of age. This task requires the rats to swim in a pool containing water made opaque and locate and climb onto a submerged platform. The time taken to accomplish this is known as the escape latency. Each rat was subjected to 24 trials over 3 days of the test period. Statistical analysis of the escape latency data revealed that the rats given 6 g/kg body weight of ethanol had significant deficits in their spatial learning ability compared with their control groups. However, there was no significant difference in spatial learning ability for the rats given either 2 or 4 g/kg body weight of ethanol compared with their respective gastrostomy or suckle control animals. We concluded that ethanol exposure greater than 4 g/kg over an 8-h period to 5-day-old rats is required for them to develop long-term deficits in spatial learning behaviour.

Effects of environmental diversity on brain morphology

This article provides a review of the literature on the effects of early environmental enrichment or impoverishment on the morphological structure of the brain. A critical re-appraisal of this literature shows that many of these effects have not been consistently observed in all experiments. In many experiments where effects have been observed the morphological methods adopted have not always been appropriate for the features being studied and thus the conclusions are brought into question. The review has also examined recent research concerned with the question of studying whether environmental enrichment in later life can ameliorate some of the morphological deficits known to be produced by undernutrition during early life. A consistent finding in this research has been that there is no statistically significant interaction between nutrition and environment, even for those few features which showed a main effect of environment. This indicates that undernutrition during early life does not preclude animals from interacting with their environment in a manner which can produce similar changes in brain morphology to those observed in normally nourished animals.

Neurons in the hilus region of the rat hippocampus are depleted in number by exposure to alcohol during early postnatal life

We have previously shown that exposing rats to a relatively high dose of ethanol during early postnatal life resulted in a deficit in spatial learning ability. This ability is controlled, at least in part, by the hippocampal formation. The purpose of the present study was to determine whether exposure of rats to ethanol during early postnatal life affected the number of specific neurons in the hippocampus. Wistar rats were exposed to a relatively high daily dose of ethanol between postnatal days 10 and 15 by placing them for 3 h each day in a chamber containing ethanol vapor. The blood ethanol concentration was about 430 mg/dl at the end of the exposure period. Groups of ethanol‐treated (ET) rats, separation controls (SC), and mother‐reared controls (MRC) were anesthetized and killed at 16 days of age by perfusion with phosphate‐buffered glutaraldehyde (2.5%). The Cavalieri principle was used to determine the volume of various subdivisions of the hippocampal formation (CA1, CA2+CA3, hilus, and granule cell layer), and the physical disector method was used to estimate the numerical densities of neurons within each subdivision. The total number of neurons was calculated by multiplying estimates of the numerical density with the volume. There were, on average, about 441,000 granule cells in the granule cell layer and 153,000 to 177,000 pyramidal cells in both the CA1 and CA2+CA3 regions in all three treatment groups. In the hilus region, ET rats had about 27,000 neuronal cells. This was significantly fewer than the average of 38,000 such neurons estimated to be present in both MRC and SC animals. Thus, neurons in the hilus region may be particularly vulnerable to the effects of a high dose of ethanol exposure during early postnatal life. Hippocampus 10:284–295, 2000

Ethanol exposure during the third trimester equivalent results in long-lasting decreased synaptic efficacy but not plasticity in the CA1 region of the rat hippocampus.

Fetal alcohol syndrome is a major cause of mental retardation. We investigated possible long‐lasting effects of alcohol on the hippocampus using a model for human third trimester brain development. Treatment of neonatal rats with an ethanol vapor atmosphere of 39.4 ± 2.6 mg ethanol/liter of air for 3 h a day from postnatal day 4 through 9 produced daily blood ethanol levels of 351 ± 14 mg/dL. Separation control animals were removed from their mothers in parallel with the ethanol vapor treatment, while suckle controls were left to develop normally. We prepared hippocampal slices from these animals between postnatal days 45 and 60 and recorded extracellular responses to Schaffer collateral stimulation. The maximum population spike in the CA1 pyramidal region and population excitatory postsynaptic potentials in the stratum radiatum did not differ significantly between groups. However, slices prepared from ethanol‐treated rats as opposed to separation and suckle controls required larger stimulus currents to produce normal postsynaptic responses. In addition, the ratio of the population excitatory postsynaptic potential (pEPSP) slope to the presynaptic volley was significantly reduced in ethanol‐treated rats. Ethanol vapor‐treated rats and separation control rats did not exhibit any significant changes in long‐term potentiation or paired‐pulse potentiation compared with normal suckle controls. These results suggest that early postnatal ethanol treatment produces a long‐lasting reduction in synaptic efficacy but not plasticity. Synapse 31:51–58, 1999.

Neonatal ethanol exposure reduces AMPA but not NMDA receptor levels in the rat neocortex.

Fetal alcohol syndrome (FAS) is the leading cause of mental retardation in western society. We investigated possible changes in glutamate receptor levels in neonatal animals following ethanol exposure using radioligand binding and western blot analysis. We used a vapor chamber to administer ethanol to neonatal Wistar rats 3 h a day from postnatal day (PND) 4-9. A separation control group was separated from their mothers for the same time and duration as the vapor treatment, while a normal control group was left to develop normally. Daily ethanol administrations resulted in decreased brain weight and body weight, as well as microencephaly (decreased brain:body weight ratio). Neither the affinity nor maximum binding of [(3)H]MK-801 (dizoclipine maleate) in the cortex of PND10 rats differed between treatment groups. Western blot analysis also failed to reveal any changes in NMDAR1, NMDAR2A, or NMDAR2B receptor levels. In contrast, the AMPA receptor subunit GluR1 was greatly reduced in vapor-treated pups compared with control pups, as revealed by western blot analysis. A similar reduction was found in westerns with an antibody recognizing the GluR2 and 4 subunits. These results indicate that ethanol reduces AMPA rather than NMDA receptors in the developing neocortex, possibly by blocking NMDA receptors during development.

The effects of undernutrition during early life on spatial learning

Recent research has shown that a lengthy period of undernutrition during early postnatal life can cause alterations in the morphological structure of the dentate gyrus. As this region is involved in the control of spatial memory, we decided to investigate whether undernourished rats also showed any deficits in this aspect of behaviour. Rats were undernourished from about birth until either 30 or 60 days of age and then nutritionally rehabilitated for a lengthy period before testing. There were significant differences in the body weight of control and undernourished rats in each experiment. The testing procedure involved rats being placed in a large pool of opaque water. They were required to swim to find a hidden platform located just below the water surface onto which they could escape. Each rat had to perform this test 20 times over a period of 3 days. The time taken and the total distance swum by each rat during each trial was measured. In Experiment 1, rats were familiarised with the water before testing took place, whereas in Experiment 2 they were not. There were no significant differences in the latency to find the platform or the distance swum between well-fed control and previously undernourished rats in either experiment. In conclusion, in our experiments we could not demonstrate that undernutrition during early life caused any deficits in spatial memory performance.