Sofía Díaz-Cintra

Effects of prtein undernutrition on the dentate gyrus in rats of three] age groups

Abstract The effect on an 8% and a control 25% casein diet on the granule cells in the dorsal blade of the dentate gyrus of the rat hippocampal formation was studied at 30, 90 ans 220 days of age. Female rats were fed wither an 8% or 25% casein deit 5 weeks prior to conception and the letters were maintained on these respective deits until killed. In rapid-Golgi-impregnated cells, we measured major and minor axes of the soma of the dentate granule neurons, the number of spines on 50-μm segments of proximal, middle and terminal regions of the largest dendrite per granule cell and the number of te dendrites intersecting 8 concentric rings 38 μm apart. At all 3 ages undernourished rats showed, when compared to controls, significant reductions of the major and minor axes od the somata and significant reductions in the number of spines on dendrites in the middle ans terminal denderitic segments. Dendritic branching was significantly reduced in undernounrished rats compared to control in all but the 4th concentric rings, with the greartest effect being seen on the outer 3 concentric rings at 90 and 220 days of age. The location of the deficit in dendric synaptic spines and the greatest dificit in dendritic branching correspond to the sites termination of the lateral and medial perfonant pathway projection to the dentate gyrus on the terminal and middle dendritic segments of the granule cells. The deficits noted in the granule cells of the detate gyrus in this study were more severe than those found in our previous studies on the effect of the low protein diet in these same rats on visual cortical pyramidal cells and on the 3 cell types nucleus raphe dorsalis and nucleus locus coerules.

Effects of prenatal protein malnutrition on hippocampal CA1 pyramidal cells in rats of four age groups

The present study was undertaken to investigate the effect of prenatal protein deprivation on area CA1 hippocampal pyramidal cells on postnatal (P) days 15, 30, 90 and 220 using Golgi techniques. Age related changes in both groups and diet related changes between groups were assessed. There were significant diet effects at all four ages, with one of 12 different measurements showing a significant diet effect on P15, five on P30, one on P90, and seven on P220. The most marked effect of the diet was on pyramidal cell dendrite spine density in the stratum moleculare and stratum radiatum, with a different pattern of diet effects in the two strata. In pyramidal cell dendrites in the stratum moleculare, there was a deficit in spine density that was significant at three of the four ages and there were similar age‐related changes in the two diet groups. Spines on pyramidal cell dendrites in the stratum radiatum showed a lack of synchrony of age‐related changes in the two diet groups, with an increased spine density in the malnourished rats on P30 and a widening deficit in this parameter on P90 and P220. The bimodal distribution to these changes, with most marked deficits occurring on P30 and P220, with an intervening period of apparent “catch‐up” on P90, is of interest and may be a significant brain adaptation to malnutrition.

Effects of prenatal malnutrition and postnatal nutritional rehabilitation on CA3 hippocampal pyramidal cells in rats of four ages.

The effects of prenatal protein malnutrition and postnatal nutritional rehabilitation on CA3 hippocampal pyramidal cells were investigated in rats of 15, 30, 90 and 220 days of age. Female rats were fed either 6% or 25% casein diet 5 weeks before conception. Following delivery, litters born the same day to 6% and 25% casein diet rats were randomly cross-fostered to 25% casein diet dams and maintained on that diet until sacrificed. In 288 rapid-Golgi impregnated cells, we measured somal size, length of the longest apical dendrite, number of apical and basal dendrites intersecting 10 concentric rings 38 microns apart, synaptic spine density in three 50 microns segments of the largest apical dendrite and the thorny excrescence area. Prenatal protein malnutrition produced differential morphological changes on CA3 pyramidal cells. We observed significant decreases of somal size (at 90 and 220 days of age), of length of apical dendrites (at 15 days old), of apical (in 15 day animals) and basal (in 15, 90 and 220 day animals) dendritic branching and of spine density (in 30, 90 and 220 day animals). We also found significant increases of apical dendritic branching in 90 and 220 day old rats. These results indicate that prenatal protein malnutrition affects normal development and produces long-term effects on CA3 pyramidal cells.

Effects of Prenatal Protein Malnutrition on Mossy Fibers of the Hippocampal Formation in Rats of Four Age Groups

The present study was undertaken to investigate the effect of prenatal protein deprivation on the postnatal development of the mossy fiber plexus of the hippocampal formation on postnatal (P) days 15, 30, 90, and 220. Although there is extensive information about the effects of malnutrition on cell body and dendrite morphology, little attention has been paid to axons or axon plexuses. The mossy fiber plexus represents the dentate gyrus granule cell axonal projection to areas CA4 and CA3 of the hippocampal formation and is readily demonstrated with Timms heavy metal stain. With the use of this stain, the plexus was measured at 13 levels throughout the hippocampal complex. There was no effect of the diet on the anatomical distribution of the plexus. The current study, however, does show significant effects of prenatal protein malnutrition on postnatal development of the mossy fiber plexus that are age dependent. The prenatally malnourished rats show significant deficits in the total rostro‐caudal extent and volume of the plexus on P15, P90, and P220, with the most marked dietary effect on P220. There was no significant diet effect on P30 in either extent or volume. Hippocampus 7:184–191, 1997.

Effect of protein malnutrition on CA3 hippocampal pyramidal cells in rats of three ages

Prenatal and postnatal protein deprivation effects on CA3-hippocampal pyramidal cells were investigated in 30-, 90- and 220-day-old rats. Female rats were fed either a 6% or a 25% casein diet 5 wk before conception and the litters were maintained on their respective diet until sacrificed. In 216 rapid Golgi-impregnated cells, we measured somal size, length and diameter of apical dendrite, number of apical dendrites intersecting 10 concentric rings 38 microns apart, thorny excrescence area and length, head diameter and density of synaptic spines on 50-microns segments of apical dendrite. The present experiments showed that malnutrition produced significant reductions of somal size in animals at 220 days of age. There were significant reductions of apical dendrite diameters in animals of 30 and 90 days, and of density and head diameter of synaptic spines at the three ages studied, and significant decrease of the thorny excrescence area at 220 days of age. At this latter age, dendritic branching was significantly decreased in the last four rings representing the area into which the perforant pathway projects. In 30-day malnourished rats, dendritic branching showed a significant increase in rings 4-6 representing the area in which the Schaffer collaterals synapse. The location of the deficit in dendritic spines corresponds to the sites where mossy fibers synapse on the apical dendrites of CA3 neurons. Age-related changes normally observed in control rats (e.g., the 30-day-old control group showed the smallest somal size and 220-day-old controls the largest size) failed to occur in the malnourished rats. The deficits in spine density and dendritic branching (in animals of 220 days old) were similar to those found in our previous studies on fascia dentata.

The effects of protein deprivation on the nucleus locus coeruleus: A morphometric golgi study in rats of three age groups

In a previous study we identified 3 cell types in the nucleus raphe dorsalis (NRD): fusiform, multipolar and ovoid. In the present study, we have investigated the effect of an 8% casein diet on these 3 cell types using quantitative techniques on rapid Golgi-impregnated neurons from rats of 3 different ages: 30, 90 and 220 days. Major and minor axes of the cell body and dendritic diameter were unaffected and primary dendritic linear extent was only slightly affected by the diet. All 3 cell types in control rats showed an increase in synaptic spines on both primary and secondary dendrites between 30 and 90 days followed by a decrease for all 3 of the cell types at 220 days. Protein-deprived rats failed to show these age-related changes. Other parameters of comparison showed clear differences between the 3 cell types. These differences could be readily seen when total synaptic spine input to the primary and secondary dendrites was calculated from the data on dendritic number, linear extent and spine density. When viewed in this way the fusiform and ovoid cells show either little change or a decreased synaptic input at all ages, while the presumed serotonergic multipolar cells showed an increase. This is in agreement with neurochemical studies in these rats showing increased levels of this biogenic amine in protein malnourished rats.

Increase of mushroom spine density in CA1 apical dendrites produced by water maze training is prevented by ovariectomy.

Dendritic spine density increases after spatial learning in hippocampal CA1 pyramidal neurons. Gonadal activity also regulates spine density, and abnormally low levels of circulating estrogens are associated with deficits in hippocampus-dependent tasks. To determine if gonadal activity influences behaviorally induced structural changes in CA1, we performed a morphometric analysis on rapid Golgi-stained tissue from ovariectomized (Ovx) and sham-operated (Sham) female rats 7 days after they were given a single water maze (WM) training session (hidden platform procedure) or a swimming session in the tank containing no platform (SC). We evaluated the density of different dendritic spine types (stubby, thin, and mushroom) in three segments (distal, medial, and proximal) of the principal apical dendrite from hippocampal CA1 pyramidal neurons. Performance in the WM task was impaired in Ovx animals compared to Sham controls. Total spine density increased after WM in Sham animals in the proximal and distal CA1 apical dendrite segments but not in the medial. Interestingly, mushroom spine density consistently increased in all CA1 segments after WM. As compared to the Sham group, SC-Ovx rats showed spine pruning in all the segments, but mushroom spine density did not change significantly. In Ovx rats, WM training increased the density of stubby and thin, but not mushroom spines. Thus, ovariectomy alone produces spine pruning, while spatial learning increases spine density in spite of ovariectomy. Finally, the results suggest that mushroom spine production in CA1 after spatial learning requires gonadal activity, whereas this activity is not required for mushroom spine maintenance.

Protein malnutrition differentially alters the number of glutamic acid decarboxylase-67 interneurons in dentate gyrus and CA1-3 subfields of the dorsal hippocampus

In 30- and 90-day-old rats, using immunohistochemistry for glutamic acid decarboxylase 67 (GAD-67), we have tested whether malnutrition during different periods of hippocampal development produces deleterious effects on the population of GABA neurons in the dentate gyrus (DG) and cornu Ammonis (CA1-3) of the dorsal hippocampus. Animals were under one of four nutritional conditions: well-nourished controls (Con), prenatal protein malnourished (PreM), postnatal protein malnourished (PostM), and chronic protein malnourished (ChroM). We found that the number of GAD-67-positive (GAD-67+) interneurons was higher in the DG than in the CA1-3 areas of both Con and malnourished groups. Regarding the DG, the number of GAD-67+ interneurons was increased in PreM and PostM and decreased in ChroM at 30 days. At 90 days of age the number of GAD-67+ interneurons was increased in PostM and ChroM and remained unchanged in PreM. With respect to CA1-3, the number of labeled interneurons was decreased in PostM and ChroM at 30 days of age, but no change was found in PreM. At 90 days no changes in the number of these interneurons were found in any of the groups. These observations suggest that 1) the cell death program starting point is delayed in DG GAD-67+ interneurons, and 2) protein malnutrition differentially affects GAD-67+ interneuron development throughout the dorsal hippocampus. Thus, these changes in the number of GAD-67+ interneurons may partly explain the alterations in modulation of dentate granule cell excitability, as well as in the emotional, motivational, and memory disturbances commonly observed in malnourished rats.

Prenatal protein malnutrition decreases mossy fibers-CA3 thorny excrescences asymmetrical synapses in adult rats

Prenatal protein malnutrition has deleterious effects on hippocampal structure and function that likely result from decreased synapse number. We thus evaluated long-term effects of prenatal protein malnutrition on the mossy fibers-CA3 thorny excrescences asymmetrical synapses in 220-day-old rats. Protein malnourished rats born from pregnant dams fed with 6% casein diet were cross-fostered to lactating control rats at birth. Control animals were fed with a 25% casein diet. Timms stained material was used to estimate the total reference volume of the mossy fiber system suprapyramidal bundle by means of stereology. The mossy fiber-CA3 asymmetrical synapse numerical density was obtained by electron microscopy, using the physical disector method. The total number of mossy fiber-CA3 asymmetrical synapses was determined on the basis of the total reference volume of the mossy fiber system suprapyramidal bundle and the mossy fiber-CA3 asymmetrical synapse numerical density. Prenatal protein malnutrition produced long-lasting, significant decreases in the volume of the mossy fiber system suprapyramidal bundle and in the numerical density of mossy fiber-CA3 asymmetrical synapse, suggesting a reduction in the total number of this synapse type. Hence, prenatal protein malnutrition induces long lasting deleterious effects on the progression of developmental programs controlling synaptogenesis and/or synaptic consolidation, likely by affecting a myriad of cellular processes.

Effects of Lesions of Hippocampal Fields CA1 and CA3 on Acquisition of Inhibitory Avoidance

Performance decrements of inhibitory avoidance (IA) induced by lesions in either the dorsal or ventral hippocampus have been interpreted as a deficiency in acquisition. Alternative interpretations are that short-term learning occurs despite the lesions and the long-term performance decrements reflect a failure of consolidation or retrieval. To assess the alternative explanations of the performance decrements, rats received lesions in either CA1 or CA3 fields of dorsal and ventral hippocampus, respectively, 8 days before IA training. Retention was tested at 30 min or 24 h after training. Kainic acid lesions were also produced in either hippocampal field 1 day after training and retention measured 8 days later. The group assessed 30 min after IA training showed little or no performance decrements, whereas the remaining groups did show marked performance decrements. These results do not support the conclusion that the hippocampus is essential for acquisition and support the idea that the hippocampus is highly involved in the consolidation or retrieval of information germane to these procedures.