Biomedical subjects

N Sousa

Publications and source records attributed to N Sousa.

33 records · Page 2Linked to original sources

PubMed1998

Effects of corticosterone treatment and rehabilitation on the hippocampal formation of neonatal and adult rats. An unbiased stereological study.

Elevations in the plasma levels of glucocorticoids are associated with cognitive impairments that have been ascribed to loss of neurons in the hippocampal formation. However, recent studies have strongly challenged this view. In order to clarify this issue, we have employed for the first time the optical fractionator and the Cavalieri principle, two unbiased stereological tools, to estimate respectively the total number of neurons and the volumes of the main subdivisions of the hippocampal formation of rats submitted to corticosterone treatment for different periods, either neonatally or in adulthood. A significant reduction in the number of neurons and in the volumes of the layers of the dentate gyrus and CA3 hippocampal field was found in rats exposed to glucocorticoids in the neonatal period; furthermore, animals treated with corticosterone from birth until 180 days of age had also a reduction in the volume of the stratum radiatum of the CA1 hippocampal field. Conversely, when the exposure occurred only during adulthood, no significant neuronal loss was observed, but there were significant reductions in the volume of layers in the dentate gyrus and CA3 hippocampal field. To search for signs of structural recovery, we incorporated a group of rats submitted to corticosterone treatment during the neonatal period in which the hormonal conditions were restored thenceforth. In this group we found a significant increase in the volume of the molecular layer of the dentate gyrus when compared with rats that were kept under corticosteroid treatment. In conclusion, these data provide a sound structural basis for the cognitive deficits observed during, and following, exposure to increased levels of glucocorticoids.

Aging↗

PubMed1998

Maintenance of hippocampal cell numbers in young and aged rats submitted to chronic unpredictable stress. Comparison with the effects of corticosterone treatment.

Exposure of rats to sustained stress has been associated with behavioural impairments, the degree of impairment being greater with increasing age of the subject. Although the behavioural deficits have been frequently attributed to stress-induced neuronal loss in the hippocampus, the validity of that view may be disputed since it is based on data collected using conventional morphometric methods which are subject to bias. The question of whether stress per se does indeed induce hippocampal cell losses was therefore re-examined using unbiased stereological tools in the present work. Specifically, we used the optical fractionator and the Cavalieri principle, to respectively estimate the total number of neurons and volumes of the main divisions of the hippocampal formation of young and old rats which had been exposed for 1 month to an unpredictable stress paradigm. The efficacy of the treatment was confirmed by elevated serum corticosterone levels measured at various intervals during the experimental period. In order to evaluate whether any deleterious effects might have occurred merely due to the stress-induced elevations in corticosterone secretion, we conducted a parallel study on animals that were injected with corticosterone over a similar duration. Neither stress nor treatment with corticosterone was found to result in significant cell losses in any division of the hippocampal formation; likewise, neither treatment produced significant volumetric differences. Further, these results were not influenced by age of the experimental subjects. The present findings therefore call for a reappraisal of the hypothesis that hippocampal cell loss accounts for the behavioural impairments observed by others following prolonged stress and/or chronic elevation of serum corticosterone levels.

Aging↗

PubMed1997

Chronic alcohol consumption and withdrawal do not induce cell death in the suprachiasmatic nucleus, but lead to irreversible depression of peptide immunoreactivity and mRNA levels.

There is evidence that chronic ethanol treatment (CET) disrupts the biological rhythms of various brain functions and behaviors. Because the suprachiasmatic nucleus (SCN) is widely recognized as the dominant pacemaker of the circadian system, we have examined the effects of CET and withdrawal on the main morphological features and chemoarchitecture of this hypothalamic nucleus. Groups of rats ethanol-treated for 6 and 12 months were compared with withdrawn rats (ethanol-treated for 6 months and then switched to a normal diet for an additional 6 months) and with groups of age-matched control and pair-fed control rats. The volume and the total number of neurons of the SCN were estimated from conventionally stained material, whereas the total number of astrocytes and of neurons containing vasopressin (AVP), vasoactive intestinal polypeptide (VIP), gastrin-releasing peptide (GRP), and somatostatin (SS) were estimated from immunostained sections. The estimates were obtained using unbiased stereological methods, based on Cavalieri's principle and the optical fractionator. The volume of the SCN and the total number of SCN neurons and astrocytes did not vary among groups. We found, however, that CET induced a significant reduction in the total number of AVP-, VIP-, GRP-, and SS-containing neurons. Withdrawal from alcohol did not reduce but rather augmented the loss of VIP- and GRP-immunoreactive neurons. The CET-induced neurochemical alterations seem to result from a decrease in neuropeptide synthesis, as revealed by the reduction in AVP and VIP mRNA levels demonstrated by in situ hybridization with radioactively labeled 48-mer AVP and 30-mer VIP probes. It is thus possible to conclude that the irreversible CET-induced changes in the neurochemistry of the SCN might underpin the disturbances in circadian rhythms observed after long-term alcohol consumption.

Alcohol Drinking↗

PubMed1997

Structural alterations of the hippocampal formation of adrenalectomized rats: an unbiased stereological study.

Previous studies have demonstrated that adrenalectomy rapidly induces cell death in hippocampal formation. However, these previous studies have involved only qualitative observations or biased estimates. Therefore, the selectivity of the effects of adrenalectomy and the magnitude of changes occurring, remain controversial. The present work employed unbiased stereological tools to examine the effects of adrenalectomy on the number of neurons in, and the volume of, the hippocampal formation. Male rats were adrenalectomized 15, 30 or 120 days before sacrifice at 180 days of age. The total number of neurons in the somal layers and hilus of the hippocampal formation was estimated using the optical fractionator. The volume of the different layers of each subdivision in the hippocampal formation was determined according to the Cavalieri principle. A progressive reduction, reaching 43%, was found in the total number of granule cells. Adrenalectomized animals exhibited a reduction in the volume of all layers of the dentate gyrus. No other region of the hippocampal formation displayed significant cell loss or a reduction in volume. In addition, the main neuronal subpopulations of the dentate gyrus were also evaluated, and a reduction in the total number of GABA- and neuropeptide Y-immunoreactive neurons in the molecular and granule cell layers of adrenalectomized rats was found. No quantitative changes were observed in the hilus. To characterize the glial response to the neuronal degeneration, we estimated the total number of cells immunoreactive for glial fibrillary acidic protein in the dentate gyrus. Although no variation in the total number of glial cells was found, signs of astroglial activation were observed in the adrenalectomized group. The present data strengthen the evidence pointing to the critical role of corticosteroids in maintaining the structural integrity of dentate gyrus.

Adrenalectomy↗

PubMed1995

Age and sex do not affect the volume, cell numbers, or cell size of the suprachiasmatic nucleus of the rat: an unbiased stereological study.

The circadian rhythms displayed by numerous biological functions are known to be sex specific and affected by aging. It has not been settled yet whether the sex- and age-related characteristics of circadian rhythms derive from changes in the anatomy of the suprachiasmatic nucleus. To shed light on these issues, we applied unbiased stereological techniques to estimate the volume of the suprachiasmatic nucleus as well as the total number of its cells and the mean volume of their somata and nuclei in progressively older groups of male and female Wistar rats (aged 1, 6, 12, 18, 24, and 30 months). The volume of the nucleus was estimated with the Cavalieri principle on serial sections. The total numbers of neurons and astrocytes were estimated by applying the optical fractionator, and the mean somatic and nuclear volumes of cells were estimated by using isotropic, uniform random sections and the nucleator method. On average, the volume of the suprachiasmatic nucleus was 0.044 mm3, and the total number of neurons and astrocytes was 17,400. Cells of the dorsomedial and ventrolateral components of the nucleus, which are morphologically different, have identical mean perikaryal and nuclear volumes, which we estimated to be 750 microns3 and 400 microns3, respectively. We further demonstrated that, at all ages analysed, the volume of the suprachiasmatic nucleus, the total cell number, and the mean somatic and nuclear volumes of its cells are affected neither by the age nor by the sex of the animal, regardless of the presence of sex- and age-related variations in circadian rhythms. However, the possibility that females may display changes in the volume of the suprachiasmatic nucleus at older ages cannot be ruled out. No effect of aging was observed in the total number of neurons or in the total number of astrocytes.

Age Factors↗

PubMed1995

Effects of chronic alcohol consumption on the cholinergic innervation of the rat hippocampal formation as revealed by choline acetyltransferase immunocytochemistry.

The specific aim of this study was to evaluate whether the cholingeric innervation of the hippocampal formation is affected by chronic alcohol consumption in the rat. Choline acetyltransferase-immunoreactive fibres and neurons were analysed in both alcohol-fed and control rats using a monoclonal antibody against choline acetyltransferase and quantitative methods. We found a global reduction in the cholinergic plexus, which was more pronounced in the hippocampus proper than in the dentate gyrus. The areal density of choline acetyltransferase immunoreactive neurons was also reduced. Differences from controls in neuronal number were particularly striking in the stratum lacunosum moleculare of the regio superior, which is precisely the zone of the hippocampal formation where choline acetyltransferase immunoreactive neurons are more abundant in controls. In conclusion, our results show that prolonged ethanol consumption leads to a substantial reduction in the cholinergic innervation of the hippocampal formation, as there was a loss of cholinergic fibres and also an apparent loss of hippocampal cholingeric neurons. These findings may help to explain the cognitive dysfunctions observed after chronic alcohol consumption.

Alcoholism↗

PubMed1995

Structural reorganization in the supraoptic nucleus of withdrawn rats following long-term alcohol consumption.

We have recently shown in the supraoptic nucleus (SON) of the rat that prolonged ethanol consumption induces cell degeneration and enlargement of the surviving neurons and of their subcellular organelles. We analyzed the SON of withdrawn rats to evaluate whether it displays any evidence of morphological reorganization following abstinence from ethanol, inasmuch as in this condition the ethanol-induced changes in the plasma levels of neurohormones and plasma osmolality are no longer detectable. A group of 18-month-old withdrawn rats was compared with age-matched, pair-fed control and ethanol-treated rats. To differentiate between the effects of withdrawal and the effects of rehydration, a group of 18-month-old rehydrated rats was also included in this study and compared with age-matched, pair-fed control and dehydrated rats. We estimated the volume of SON, and the total number and mean volume of its neurons. The cross-sectional areas of the vasopressinergic and oxytocinergic populations were also evaluated. At the ultrastructural level, we determined the volumes and surface areas of the rough endoplasmic reticulum and Golgi apparatus, and the volumes of neurosecretory granules and nucleoli. In withdrawn animals, the total number of SON neurons was smaller than in controls, although the neuronal volume was greater. The number of SON neurons did not differ between withdrawn and ethanol-treated rats, despite the reduced volume of SON in the former animals. The decrease of SON volume correlated with and was caused by a reduction in the volume of SON neurons and in the size of the organelles involved in neuro-hormone synthesis.(ABSTRACT TRUNCATED AT 250 WORDS)

Alcoholism↗

PubMed1994

The vasopressinergic innervation of the lateral septum of the rat after chronic alcohol consumption and withdrawal.

We have recently reported that ethanol ingestion induces morphological changes in the vasopressinergic neurons of the supraoptic nucleus and that withdrawal from alcohol partially reverses these alterations. Since the production of vasopressin is not restricted to the magnocellular neurons of the hypothalamus, we investigated the effects of long-term ethanol intake and withdrawal on the lateral septum, an area heavily innervated by vasopressinergic fibers. Besides, as ethanol leads to a decrease of the plasma levels of testosterone, a hormone which plays a pivotal role in the development and maintenance of the vasopressinergic innervation of the lateral septum, we included groups of alcohol-fed animals submitted to testosterone replacement both in physiological and supraphysiological doses. In ethanol-treated rats there was a marked reduction in the number of vasopressin-immunoreactive fibers in the lateral septum. Following ethanol withdrawal a partial recovery in the number of vasopressin-immunoreactive fibers was observed. In both groups of ethanol + testosterone-treated animals the vasopressinergic innervation was increased when compared to the alcohol-fed group, although a complete reversal was not achieved. Therefore, two mechanisms might be regarded as underlying the impoverishment of the vasopressinergic innervation in the lateral septum after prolonged alcohol consumption: alcohol-induced cell death in the bed nucleus of the stria terminalis, from where these fibers arise, and/or alcohol-induced decrease in testosterone plasma levels.

Animals↗

PubMed1994

Stereological study of the ultrastructural changes induced by chronic alcohol consumption and dehydration in the supraoptic nucleus of the rat hypothalamus.

We have previously shown that prolonged alcohol ingestion leads to neuronal loss in the supraoptic nucleus of the rat and that the surviving neurons, mainly the vasopressinergic ones, display marked increase in volume. In an attempt to establish correlates for the volumetric alterations we have studied the organelles of supraoptic nucleus neurons in three groups of rats--ethanol-fed, pair-fed, and dehydrated, in all cases treated from 2 to 12 months of age. The volume and surface area of the rough endoplasmic reticulum and Golgi apparatus, and the volume of nucleoli and neurosecretory granules were estimated on the basis of the respective volume and surface densities. The volumes and surface areas of all quantified organelles were increased in both alcohol-fed and dehydrated animals, although the increases were greater in the former group. Changes in the organelles studied are commonly regarded as reliable indicators of the neurosecretory activity of magnocellular neurons. Thus, our results suggest that under conditions of chronic alcohol exposure, the synthesizing activity of the surviving supra-optic neurons is augmented to compensate for the alcohol-induced neuronal loss and/or as a consequence of the alcohol-induced hyperosmolality. Changes in the transport and release of the neurosecretory material cannot, however, be ruled out as an additional cause of neuronal enlargement.

Adaptation, Physiological↗

PubMed1993

Ultrastructural evidence of sexual dimorphism in supraoptic neurons: a morphometric study.

We have recently shown that in spite of the absence of receptors for gonadal steroids in the supraoptic nucleus (SON) of the rat hypothalamus, the volume of the nucleus and the size of its neurons are larger in males than in females, and that these differences between male and female rats are correlated with body weight and dependent on the vasopressinergic neurons. As supraoptic neurons and their organelles enlarge when they are engaged in active peptidergic secretion we have carried out a morphometric ultrastructural analysis to determine if cell structures involved in the synthesis and storage of neurosecretory material also display weight-dependent sex dimorphism. Groups of six male and six female rats aged 30, 60 and 180 days were used. Nucleoli, rough endoplasmic reticulum and neurosecretory granules were analysed and we estimated their volume or surface densities and the total volume of nucleoli and rough endoplasmic reticulum, and total surface area of rough endoplasmic reticulum. We found that, with the exception of neurosecretory granules, the densities of the organelles did not differ among the groups studied, but total values were higher in males. These differences were found to be weight-dependent. Since the organelles studied are regarded as reliable indicators of the neurosecretory activity of supraoptic neurons, our data fully support the view that the weight-dependent sexual dimorphism observed in this nucleus reflects greater synthetic activity of its vasopressinergic neurons associated with the need to maintain water balance in larger bodies.

Animals↗

PubMed1993

Effects of chronic alcohol consumption and of dehydration on the supraoptic nucleus of adult male and female rats.

Ethanol ingestion affects the hypothalamo-neurohypophysial system resulting in increased diuresis, dehydration and hyperosmolality. We studied the supraoptic nucleus, of the hypothalamus, in ethanol-treated rats, to determine if ethanol alone and/or the associated disturbances of water metabolism lead to structural alterations in a nucleus known to play a central role in fluid homeostasis. Groups of male and female rats were ethanol-treated until 12 and 18 months of age and compared with age-matched pair-fed controls. Twelve and 18-month-old control groups and 12-month-old water control groups (rats submitted to chronic dehydration) were also included in this study in an attempt to differentiate between the effects of undernutrition and dehydration/hyperosmolality, and the specific neurotoxic effects of ethanol. We estimated the volume of the supraoptic nucleus and the numerical density of its neurons and calculated the total number of supraoptic neurons. The volume of both supraoptic neurons and neuropil were also estimated. In immunostained material the ratio of vasopressin to oxytocin neurons and the cross-sectional areas of the two neuronal types were evaluated. There was marked neuronal loss in alcohol-treated rats, but the volume of the supraoptic nucleus was increased. The increase in the volume of the supraoptic nucleus correlated with and was due to increases in the volume was particularly marked for vasopressin neurons. No significant differences were found between controls and pair-fed controls in any of the parameters investigated. In water control rats, the volume of the supraoptic nucleus and of the supraoptic neurons and neuropil was also greater than in pair-fed controls. However, the variations found were not as marked as in ethanol-treated rats and there was no cell loss. These findings reveal, for the first time, that chronic ethanol consumption affects the morphology of supraoptic neurons and neuropil and, consequently, the structure of the entire supraoptic nucleus. Moreover, this study supports the view that ethanol has direct neurotoxic effects on supraoptic neurons because the alterations that occur are not mimicked in animals in which water metabolism alone is disturbed.

Animals↗

PubMed1993

The supraoptic nucleus of the adult rat hypothalamus displays marked sexual dimorphism which is dependent on body weight.

The neurons of the supraoptic nucleus in the rat hypothalamus are reported not to possess receptors for gonadal steroids and sexual dimorphism has not previously been described in this nucleus. We have analysed this nucleus in groups of Sprague-Dawley rats (six males or six females per group), one, two, six, 12 and 18 months after birth. Body and brain weights were recorded, the volume of the nucleus was determined from the right hemisphere and all other quantitative parameters were determined from the left nucleus. In addition, different groups of four male and four female rats aged two and 18 months were analysed after immunocytochemical staining to distinguish between vasopressin and oxytocin neurons. The total number of neurons was constant in all groups studied, despite which the volume of the supraoptic nucleus increased progressively with age in both males and females. The cross-sectional areas and volumes of supraoptic neurons also increased with age. The volume density of the neuropil remained constant in all groups and there was a progressive decrease with age in the numerical density of neurons. Immunocytochemistry revealed that the age-dependent increases in the size of the neurons involved primarily the vasopressin neurons. The age-related changes were much greater in males than in females, resulting in significant differences between the sexes at two, six, 12 and 18 months with respect to the volume of the supraoptic nucleus, the cross-sectional areas of neuronal somata and nuclei, and the volume of supraoptic neurons. Thus the supraoptic nucleus and its vasopressin neurons are larger in adult males than in age-matched females. Since we have also shown that body weight is very closely correlated with changes in the size of supraoptic neurons, and adult male rats are heavier than females of the same age, we suggest that these size changes reflect adaptation of the vasopressin neurons of the supraoptic nucleus to increasing functional demands associated with the regulation of water balance in bodies of increasing size.

Aging↗

PubMed1992

Selective vulnerability of the hippocampal pyramidal neurons to hypothyroidism in male and female rats.

Thyroid hormone deficiency has long been considered to affect profoundly such cognitive functions as learning and memory, which are known to depend on the structural integrity of the hippocampal formation. Since we previously found that the number of granule cells of the dentate gyrus is reduced in hypothyroid animals, we decided to extend our observations to the pyramidal cells of the hippocampus in order to gain further insight into the effects of hypothyroidism upon the other neuronal links of the hippocampal trisynaptic circuitry, inasmuch as CA1 neurons are known to be particularly vulnerable to aggressive agents. Groups of 6 male and 6 female rats aged 30 and 180 days were analysed separately after being treated as follows: (1) hypothyroid from day 0 until day 30 (30-day-old hypothyroid group); (2) respective 30-day-old control; (3) hypothyroid from day 0 until day 180 (180-day-old hypothyroid group); (4) hypothyroid until day 30 and thenceforth maintained euthyroid (recovery group); (5) hypothyroid since day 30 (adult hypothyroid group); and (6) respective 180-day-old control. The volume of the pyramidal cell layer of the CA1 and CA3 regions and the numerical density of the respective neurons were evaluated, thereby allowing us to estimate the total number of pyramidal cells in each hippocampal region. The areal density and the mean nuclear volume of CA1 and CA3 pyramidal cells were also estimated. In the CA3 region, we found that hypothyroidism, whatever its duration and time of onset, induces a reduction in the volume of the pyramidal cell layer and a parallel increase in the numerical density of its neurons, without interfering with the total number of pyramidal cells. Conversely, in the CA1 region, thyroid hormone deficiency started either neonatally or during maturity was found to lead to a decrease in the total number of pyramidal cells. Reductions ranging between 14.2 and 22.5% were found in 30 and 180-day-old hypothyroid groups. The reestablishment of a euthyroid state did not ameliorate the referred neuronal loss. The present results support the view that hypothyroidism induces small alterations in the structural organization of the hippocampal CA3 region, contrary to what happens in CA1 in which neuronal death occurs. Furthermore, the data presented herein demonstrate that the total number of CA1 pyramidal cells displays sexual dimorphism that is not affected by thyroid hormone manipulations.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1991

Sexual dimorphism in the mossy fiber synapses of the rat hippocampus.

The presence of sexual dimorphism in the hippocampal formation has long been recognized. Differences between male and female rats have been detected with respect to the number of dentate granule cells and branching patterns of dentate granule and hippocampal pyramidal cell dendrites. Groups of 6 male and 6 female Sprague-Dawley rats were studied at 180 days of age. Based on light microscopical Timm-staining and Golgi-impregnation and electron microscopy, and applying morphometric techniques, we now report that the total number of synapses between mossy fibers and the apical dendritic excrescences of CA3 pyramidal cells is the same in male and female rats, despite a higher numerical density in the latter. Moreover, the volume of the mossy fiber system was found to be smaller in females. Because the number of dentate granule cells is smaller in females than in males, the increased numerical density of synapses may be thought of as a compensatory mechanism to equalize the number of synaptic contacts between dentate granule and CA3 pyramidal cells in the two sexes. We demonstrate that an increase in the number of mossy fiber boutons in female rats is a determining factor for the sexual differences found.

Animals↗

PubMed1991

The supraoptic nucleus in hypothyroid and undernourished rats: an experimental morphometric study.

The supraoptic nuclei of both male and female 30-day-old rats rendered hypothyroid by daily subcutaneous injection of propylthiouracil were studied and the results were compared with age- and sex-matched rats fed ad libitum (control rats) and with undernourished rats. Morphometric methods were used to evaluate the volume of the supraoptic nucleus and the areal and numerical densities of its neurons. These parameters allowed us to estimate the total number of neurons of this nucleus. In addition, the mean cross-sectioned area and mean nuclear diameter of the same neurons were also evaluated. The volume of the supraoptic nucleus was reduced both in hypothyroid and undernourished animals when compared with normal controls. The areal and numerical densities of neurons from the former groups were increased and the volume density of the neuropil was reduced. As a consequence, the total number of neurons was found to be identical in all groups studied. Furthermore, the mean nuclear diameters and cross-sectioned areas of the supraoptic neurons were similar in all groups. The volumetric differences and the increased packing found were always more marked for hypothyroid than for undernourished rats. Differences were not detected between male and female groups. The present results support the view that the increased cell packing in hypothyroid animals depends upon a reduction in the neuropil of the nucleus, as has been described under similar conditions in other central nervous system areas displaying identical patterns of neurogenesis. In addition, it was found that the effects of undernourishment cannot be discriminated from those dependent on neonatal hypothyroidism.

Analysis of Variance↗