Rodrigo Pascual

Effects of prenatal stress and exercise on dentate granule cells maturation and spatial memory in adolescent mice

Exposure to prenatal stress (PS) increases the risk of developing neurobehavioral disturbances later in life. Previous work has shown that exercise can exert beneficial effects on brain damage; however, it is unknown whether voluntary wheel running (VWR) can ameliorate the neurobehavioral impairments induced by PS in adolescent offspring. Pregnant CF‐1 mice were randomly assigned to control (n = 5) or stressed (n = 5) groups. Pregnant dams were subjected to restraint stress between gestational days 14 and 21 (G14–21), whereas controls remained undisturbed in their home cages. On postnatal day 21 (P21), male pups were randomly assigned to the following experimental groups: control (n = 5), stressed (n = 5), and stressed mice + daily submitted to VWR (n = 4). At P52, all groups were behaviorally evaluated in the Morris water maze. Animals were then sacrificed, and Golgi‐impregnated granule cells were morphometrically analyzed. The results indicate that PS produced significant behavioral and neuronal impairments in adolescent offspring and that VWR significantly offset these deleterious effects.

EARLY SOCIAL ISOLATION DECREASES THE EXPRESSION OF CALBINDIN D-28K AND DENDRITIC BRANCHING IN THE MEDIAL PREFRONTAL CORTEX OF THE RAT

Social isolation in rodents is the most well characterized animal model for early stressful experiences and their neurobehavioral consequences. The present study analyzed the effects of early social isolation on the expression of the calcium binding protein calbindin-D28k (CAD) and dendritic arborization in the medial prefrontal cortex (mPFC) of the rat. Sprague-Dawley male rats were reared either under isolation or social conditions from 21 to 51 postnatal days. At the end of this period the animals were behaviorally evaluated in the open-field test, sacrificed, and mPFC serial sections were processed either for immunocytochemical labeling against CAD or Golgi-Cox-Sholl staining. Isolated-reared rats exhibited a dramatic decrease in the number of CAD immunoreactive neurons and a significant dendritic atrophy of layer II/III pyramidal cells in association with a reduced exploratory behavior.

Maternal exercise during pregnancy ameliorates the postnatal neuronal impairments induced by prenatal restraint stress in mice

Clinical and preclinical studies have demonstrated that prenatal stress (PS) induces neuronal and behavioral disturbances in the offspring. In the present study, we determined whether maternal voluntary wheel running (VWR) during pregnancy could reverse the putative deleterious effects of PS on the neurodevelopment and behavior of the offspring. Pregnant CF‐1 mice were randomly assigned to control, restraint stressed or restraint stressed + VWR groups. Dams of the stressed group were subjected to restraint stress between gestational days 14 and delivery, while control pregnant dams remained undisturbed in their home cages. Dams of the restraint stressed + VWR group were subjected to exercise between gestational days 1 and 17. On postnatal day 23 (P23), male pups were assigned to one of the following experimental groups: mice born from control dams, stressed dams or stressed + VWR dams. Locomotor behavior and pyramidal neuronal morphology were evaluated at P23. Animals were then sacrificed, and Golgi‐impregnated pyramidal neurons of the parietal cortex were morphometrically analyzed. Here, we present two major findings: first, PS produced significantly diminished dendritic growth of parietal neurons without altered locomotor behavior of the offspring; and second, maternal VWR significantly offset morphological impairments.

Prenatal stress alters the behavior and dendritic morphology of the medial orbitofrontal cortex in mouse offspring during lactation.

Several preclinical and clinical studies have shown that prenatal stress alters neuronal dendritic development in the prefrontal cortex, together with behavioral disturbances (anxiety). Nevertheless, neither whether these alterations are present during the lactation period, nor whether such findings may reflect the onset of anxiety disorders observed in childhood and adulthood has been studied. The central aim of the present study was to determine the effects of prenatal stress on the neuronal development and behavior of mice offspring during lactation (postnatal days 14 and 21). We studied 24 CF‐1 male mice, grouped as follows: (i) control P14 (n = 6), (ii) stressed P14 (n = 6), (iii) control P21 (n = 6) and (iv) stressed P21 (n = 6). On the corresponding days, animals were evaluated with the open field test and sacrificed. Their brains were then stained in Golgi‐Cox solution for 30 days. The morphological analysis dealt with the study of 96 pyramidal neurons. The results showed, first, that prenatal stress resulted in a significant (i) decrease in the apical dendritic length of pyramidal neurons in the orbitofrontal cortex at postnatal day 14, (ii) increase in the apical dendritic length of pyramidal neurons in the orbitofrontal cortex at postnatal day 21, and (iii) reduction in exploratory behavior at postnatal day 14 and 21.

Maternal stress induces long-lasting Purkinje cell developmental impairments in mouse offspring

A number of clinical studies suggest that prenatal stress can be a risk factor in the development of various psychopathologies, including schizophrenia, depression, anxiety, and autism. The cerebellar vermis has been shown to be involved in most of these disorders. In the present study, therefore, we evaluate the effect of maternal stress on long-term alterations in vermal Purkinje cell morphology. Furthermore, to discern whether these structural changes are associated with anxious behavior, the exploratory drive in the elevated plus maze was evaluated. Pregnant CF-1 mice were randomly assigned to control (n = 14) or stressed (n = 16) groups. Dams of the stressed group were subjected to restraint stress between gestational days 14 and 20, while control pregnant dams remained undisturbed in their home cages. Anxious behavior and Purkinje cell morphology were evaluated in three ontogenetic stages: postweaning, adolescence, and adulthood. Although exploratory behavior in the elevated plus maze was unaffected by prenatal stress, the Purkinje cell morphology showed a transient period of abnormal growth (at postweaning and juvenile stages) followed by dramatic dendritic atrophy in adulthood. In conclusion, prenatal stress induced significant long-lasting bimodal changes in the morphology of vermal Purkinje cells. These structural alterations, however, were not accompanied by anxious behaviors in the elevated plus maze.

Effects of a single course of prenatal betamethasone on dendritic development in dentate gyrus granular neurons and on spatial memory in rat offspring.

BACKGROUND Preterm babies treated with synthetic glucocorticoids (sGC) in utero exhibit behavioral alterations and disturbances in brain maturation during infancy. However, the effects on dentate granule cell morphology and spatial memory in rats that were given clinically equivalent doses of antenatal betamethasone remain unclear. METHODS Pregnant rats were randomly divided into the following two experimental groups: control (CON) and betamethasone-treated (BET) groups. At gestational day 20 (G20), BET dams were subcutaneously injected with a 0.17 mg/kg betamethasone solution, and CON animals received a similar volume of saline. At postnatal days 22 (P22) and P52, BET and CON offsprings were behaviorally evaluated in the Y-Maze test, and the dentate gyrus granular neurons were histologically analyzed. RESULTS Animals prenatally treated with a single course of betamethasone exhibit a significant decrement in the dendritic outgrowth of dentate granule cells along with impaired spatial memory when evaluated at P52. Moreover, the body and brain weight of the BET group was significantly lower than the CON group at P0, P22, and P52. CONCLUSION The current results indicate that a single course of betamethasone in pregnant rats produces significant neuronal and behavioral impairments of the offspring at adolescence along with a decrement in somatic and brain weights at each of the three ages evaluated.

Melatonin ameliorates neocortical neuronal dendritic impairment induced by toluene inhalation in the rat

The present study investigated the effects of toluene inhalation and the restorative effects of melatonin on branching and basal dendritic outgrowth of superficial pyramidal neurons in rats frontal, parietal, and occipital cortices. At postnatal day 21 (P21), Sprague-Dawley male rats were randomly assigned to either an air-only group or a toluene group. From P22 to P32 the animals were exposed to either clean air or toluene vapors (5000-6000 ppm) for 10 min/day. This strategy simulated common toluene abuse in humans, which consists of 15-20 rapid inhalations of highly concentrated solvent. Once the inhalation period was over (P32), toluene exposed animals were randomly reassigned to one of following experimental groups: (i) air-control/saline; (ii) toluene/saline; (iii) toluene/melatonin 0.5mg/kg; (iv) toluene/melatonin 1.0mg/kg; (v) toluene/melatonin 5.0mg/kg; and (vi) toluene/melatonin 10mg/kg. Seven days after the last inhalation (P39), all the animals were sacrificed under deep anesthesia; brains were dissected out and stained according to the Golgi-Cox-Sholl procedure. Layer II/III pyramidal neurons were morphologically analyzed by measuring their basilar dendritic length and the number of branches. The results obtained revealed that (i) toluene inhalation significantly reduced dendritic outgrowth and branching in all cortical areas studied, and (ii) intraperitoneal administration of melatonin (0.5-10mg/kg) was able to restore the dendritic impairment induced by toluene exposure.

Purkinje cell dendritic atrophy induced by prenatal stress is mitigated by early environmental enrichment.

BACKGROUND Prenatal stress (PS) in experimental animals causes long-lasting changes in Purkinje cell dendritic morphology. Furthermore, these structural changes are associated with an increase in anxiogenic behaviors in the elevated plus maze (EPM) and open-field (OF) test. OBJECTIVES As environmental enrichment (EE) has significant restorative effects on brain neurons and behavior, the aim of this study was to evaluate if postweaning EE mitigates the decrease in Purkinje cell dendritic expansion and exploratory behavior induced by PS in mice. MATERIALS AND METHODS Restraint stress was induced from gestational day 14 (G14) to G21. Approximately 50% of the PS animals were submitted to the EE paradigm between postnatal days 22 (P22) and P52. At P52 and P82, male animals were behaviorally evaluated, and then the morphology of the cerebellar vermal Purkinje cells was analyzed. RESULTS We found that EE significantly ameliorates the Purkinje cell dendritic atrophy and anxiety-like behavior in the EPM. CONCLUSION Our data show that long-lasting Purkinje cell dendritic impairments and anxiety-like behavior can be mitigated by postweaning EE.

Melatonin promotes distal dendritic ramifications in layer II/III cortical pyramidal cells of rats exposed to toluene vapors

We have previously shown that toluene inhalation produces significant impairments in the basilar dendritic outgrowth of pyramidal cortical cells. This neurotoxic effect was markedly inhibited by melatonin administration at a dose of 5mg kg(-1). The present study was designed to determine whether toluene and melatonin equally affect all basilar dendritic segments or if a differential response exists between the segments. Twenty-eight male mice were weaned at postnatal day 21 (P21) and randomly assigned to either the control (C; n=10,) or toluene (T; n=18) group. Between P22-P32, male rats were placed into a glass chamber and exposed to either toluene vapors (5-000-6000 ppm) or clean air for 10 min a day. When toluene exposure ended (P32), animals were further assigned to the following experimental groups: (a) control/saline (C/S; n=10), (b) toluene/saline (T/S; n=10), or (c) toluene/melatonin 5mg kg(-1) (T/M; n=8). Melatonin or vehicle solutions were administered daily between P32 and P38. Forty-eight hours after the final toluene exposure, the animals were sacrificed, and the pyramidal cortical cells were stained using the Golgi-Cox-Sholl procedure. The number of basilar dendritic branches/order was counted using the centrifugal ordering method. The results indicate that (i) toluene inhalation significantly impairs both proximal and distal basilar dendritic ramifications (in the parietal and frontal/occipital cortices, respectively) and (ii) melatonin both protects neurons from toluene neurotoxicity in all cortical areas studied and increases the complexity of the dendritic tree above control values.

Antenatal betamethasone produces protracted changes in anxiety-like behaviors and in the expression of microtubule-associated protein 2, brain-derived neurotrophic factor and the tyrosine kinase B receptor in the rat cerebellar cortex

Using classic Golgi staining methods, we previously showed that the administration of synthetic glucocorticoid betamethasone in equivalent doses to those given in cases of human premature birth generates long‐term alterations in Purkinje cell dendritic development in the cerebellar cortex. In the present study, we evaluated whether betamethasone alters the immunohistochemical expression of proteins that participate in cerebellar Purkinje cell dendritic development and maintenance, including microtubule‐associated protein 2 (MAP2), brain‐derived neurotrophic factor (BDNF) and the tyrosine kinase B receptor (TrkB), which are located predominantly in the cerebellar molecular layer where Purkinje cell dendritogenesis occurs. Consistent with our previous Golgi stain studies, we observed that animals prenatally exposed to a single course of betamethasone showed long‐term alterations in the expression of MAP2, BDNF and TrkB. Additionally, these protracted molecular changes were accompanied by anxiety‐like behaviors in the elevated plus maze and marble burying tests.