Early post-natal, low-level lead exposure increases the number of PSA-NCAM expressing cells in the dentate gyrus of adult rat hippocampus

Although lead is widely known as a potent neurotoxin, the effect of lead exposure on the expression of the polysialic acid linked neural cell adhesion molecule (PSA-NCAM) remains unclear. We exposed Wistar rat pups to 0.2% lead acetate from postnatal day (PND) 1 to PND 30. This exposure protocol resulted in pup blood lead levels, which increased to 29.3+/-5.0 mg/dl on PND 15, and subsequently rose to 34.2+/-5.8 mg/dl at weaning. Corresponding brain tissue lead levels were 456+/-23 ng/g on PND 15 and 781+/-87 ng/g on PND 30. Animals were sacrificed on PND 80, when the blood and brain lead concentrations did not differ from those of the control group. Lead exposure induced a significant increase in the total number of PSA-NCAM expressing cells, compared to the control group (p<0.01), and did not change the proportion of cells co-expressing PSA-NCAM with glial or neuronal markers (calbindin, TuJ1, GFAP). These results suggest that early post-natal lead exposure induces persistent changes in the number of PSA-NCAM expressing cells, which could be, at least, partly the basis of impairments in the learning and memory formation, which follows low-level lead exposure.     

Time of neuron origin in the hippocampus and dentate gyrus of normal and reeler mutant mice: an autoradiographic analysis

The hippocampus and dentate gyrus of reeler mutant mice contain the normal neuronal classes. The majority of cells are included in laminar aggregates reminiscent of normal structure though these are irregular in outline, interrupted at points, and less densely packed. Many hippocampal neurons and granule cells of the dentate gyrus are scattered in the external plexiform zone (strata radiatum and lacunosum-moleculare) of the hippocampus rather than being included in the laminar aggregates. Neurons in the reeler are generated at the same time as their normal homologs. However, the relationship of adult cell position to sequence of cell generation is abnormal in two general ways in the mutant. First, although in both the mutant and normal, cells in the external plexiform zone of the hippocampus are among the earliest formed, a relatively greater proportion of cells generated through a relatively longer embryonic time are destined to be scattered in this distal location in the mutant. Secondly, within the laminar aggregates of the hippocampus in reeler, the more superficial cells tend to be formed earlier, and the more deeply located ones are the latest formed. A reversed relationship exists in the normal in which the deepest cells are formed earliest, the more superficial ones last. The observations are consistent with the view that a mechanism which provides for orderly lamination of cells in the normal is defective in reeler.     

Quantitative analysis of postnatal neurogenesis and neuron number in the macaque monkey dentate gyrus

The dentate gyrus is one of only two regions of the mammalian brain where substantial neurogenesis occurs postnatally. However, detailed quantitative information about the postnatal structural maturation of the primate dentate gyrus is meager. We performed design‐based, stereological studies of neuron number and size, and volume of the dentate gyrus layers in rhesus macaque monkeys (Macaca mulatta) of different postnatal ages. We found that about 40% of the total number of granule cells observed in mature 5–10‐year‐old macaque monkeys are added to the granule cell layer postnatally; 25% of these neurons are added within the first three postnatal months. Accordingly, cell proliferation and neurogenesis within the dentate gyrus peak within the first 3 months after birth and remain at an intermediate level between 3 months and at least 1 year of age. Although granule cell bodies undergo their largest increase in size during the first year of life, cell size and the volume of the three layers of the dentate gyrus (i.e. the molecular, granule cell and polymorphic layers) continue to increase beyond 1 year of age. Moreover, the different layers of the dentate gyrus exhibit distinct volumetric changes during postnatal development. Finally, we observe significant levels of cell proliferation, neurogenesis and cell death in the context of an overall stable number of granule cells in mature 5–10‐year‐old monkeys. These data identify an extended developmental period during which neurogenesis might be modulated to significantly impact the structure and function of the dentate gyrus in adulthood.     

Effect of low-frequency stimulation on kindling induced changes in rat dentate gyrus: an ultrastructural study

It has been shown that low-frequency stimulation (LFS) can induce anticonvulsant effects. In this study, the effect of different LFS frequencies on kindling induced behavioral and ultrastructural changes was investigated. For induction of kindled seizures in rats, stimulating and recording electrodes were implanted in perforant path and dentate gyrus, respectively. Animals were stimulated in a rapid kindling manner. Different groups of animals received LFS at different frequencies (0.5, 1 and 5 Hz) following kindling stimulations and their effects on kindling rate were determined using behavioral and ultrastructural studies. Kindling stimulations were applied for 7 days. Then, the animals were sacrificed and their dentate gyrus was sampled for ultrastructural studies under electron microscopy. All three used LFS frequencies (0.5, 1 and 5 Hz) had a significant inhibitory effect on kindling rate and decreased afterdischarge duration and the number of stimulations to achieve stage 4 and 5 seizures significantly. In addition, application of LFS prevented the increase in the post-synaptic density and induction of concave synaptic vesicles following kindling. There was no significant change between anticonvulsant effects of LFS at different frequencies. Obtained results show that LFS application can prevent the neuronal hyperexcitability by preventing the ultrastructural changes during kindling and this may be one of the mechanisms of LFS anticonvulsant effects.     

Seizures increasetrkC mRNA expression in the dentate gyrus of rat hippocampus

In this study we have shown, byin situ hybridization and RNase protection assay, a significanttrkC mRNA increase confined to the dentate gyrus of hippocampus, both after seizures induced by intracerebroventricular injection of kainic acid and bicuculline. Moreover, after bicuculline treatment we observed an earlier increase oftrkC mRNA level, which peaked after 3 h and returned back to normal levels by 12 h. In contrast, the kainic acid treatment produced a delayed increase oftrkC mRNA, which initiated after 6 h, peaked at 12 h, and returned to normal levels at 24 h. This increase, which involves alsotrkC mRNA receptor with tyrosine kinase activity, was mediated by non-NMDA receptors and counteracted by GABA potentiating agent diazepam. Using embryonic neuronal cultures from cerebral hemispheres, including hippocampus, we found that glutamate receptor agonists, including glutamate, kainate, NMDA, and t-ACPD, increasetrkC mRNA levels with the following rank order of efficacy: NMDA>t-ACPD>kainic acid>glutamate. In conclusion, our data show thattrkC mRNA expression in granule cells of the hippocampus dentate gyrus is increased during seizure activity and that it is mediated by non-NMDA receptors.     

Prenatal exposure to a non-steroidal anti-inflammatory drug or saline solution impairs sciatic nerve morphology: a stereological and histological study

The toxic effect of non-steroidal anti-inflammatory drugs (NSAIDs) during development has been widely investigated. While it has been shown that these drugs impair central nervous development and compromise the neural activity, the effects of these substances on the development of peripheral nerves are still not clarified. In the present study, sciatic nerves withdrawn from three experimental groups of 4-week-old rats, prenatally exposed to either saline solution, or diclofenac sodium, and controls not exposed to any substance, were evaluated in terms of axon number, cross-sectional area of axon and myelin sheet thickness as well as of the ultrastructure of nerve fibers. Comparisons of stereological estimations among these three groups showed that axon number and mean axon cross-sectional area, but not average myelin sheet thickness, were significantly decreased in rats that were exposed to both diclofenac sodium and also to the saline solution, in comparison of the control group. Electron microscope analysis revealed, in both treated groups, deterioration of myelin sheaths that was more pronounced in rats that were exposed to diclofenac sodium. Altogether, these findings show that the prenatal administration of both diclofenac sodium and saline solution impairs peripheral nervous system development, thus suggesting that this potential teratogenic effect should be also taken into consideration in the clinical use of these substances in pregnant patients.     

On the number of neurons in the dentate gyrus of the rat

We have estimated the number of dentate granule cells in Sprague-Dawley and Wistar rats at 1, 4 and 12 months of age. In Sprague-Dawley rats the number of granule cells is relatively constant throughout this period at about 1 million. In Wistar rats, on the other hand, there is a progressive increase in the number from about 700,000 at 1 month to 1 million at 4 months; thereafter the number declines to about 800,000 at 1 year. Estimates of the numbers of cells in the polymorphic zone that can be stained immunohistochemically for somatostatin, cholecystokinin, vasoactive-intestinal peptide, and glutamic acid decarboxylase show no appreciable differences in the two strains.     

Cholinergic innervation of the rat dentate gyrus: an immunocytochemical and electron microscopical study

Immunocytochemical studies using a monoclonal antibody to choline acetyltransferase (ChAT) were performed on sections of rat dentate gyrus. Light microscopical analysis of the immunoreactivity revealed dense fiber networks and many punctate structures predominantly located at the interface of the granule cell layer and molecular layer. In the elctron microscope, the immunostained punctate structures were identified as synaptic boutons which formed mainly symmetrical contacts onto dendritic elements. Few ChAT-immunoreactive boutons formed axosomatic contacts.     

The histochemical localization of cytochrome oxidase in the dentate gyrus of the rat hippocampus

In the dentate gyrus of the rat's hippocampal formation, the activity of an oxidative enzyme, cytochrome oxidase, has been localized mostly to the molecular layer with histochemical methods that utilize diaminobenzidine. The electron microscopic localization of cytochrome oxidase indicated that mitochondria within granule cell dendrites were very reactive while those within the somata and mossy fiber terminals of this neuronal type were less reactive. Caution must be used when predicting the relative physiological activities of neurons with this method because differential activities of this enzyme occur within separate parts of the same neuronal population.     

The effects of chronic lead exposure on long-term depression in area CA1 and dentate gyrus of rat hippocampus in vitro

Long-term potentiation (LTP) and long-term depression (LTD), two forms of synaptic plasticity, are believed to underlie the mechanisms of learning and memory. Previous studies have demonstrated that low-level lead exposure can impair the induction and maintenance of LTP in vivo and in vitro. The present study was carried out to investigate whether the low-level lead exposure affected the induction and maintenance of LTD. Neonatal Wistar rats were exposed to lead from parturition to weaning via milk of dams drinking 0.2% lead acetate solution. Field excitatory postsynaptic potentials (EPSPs) were recorded in hippocampal slices in adult rats (50–65 days) to study the alterations of LTD in area CA1 and dentate gyrus (DG) of hippocampus following chronic lead exposure. The input–output (I/O) curves before conditioning in both areas showed no evident alterations in basic synaptic transmission between the control and lead exposure groups. In area CA1, the mean amplitude of EPSP slope in control rats (61±11%, n=15) decreased significantly greater than that in lead-exposed rats (78±8%, n=8, P<0.05) following low frequency stimulation (LFS, 1 Hz, 15 min), which lasted at least 45 min. In area DG, with application of the same LFS, the LTD was induced in control rats (72±22%, n=8), while the LFS failed to induce LTD in lead-exposed rats (100±26%, n=8). These results showed that chronic lead exposure affected the induction of LTD in both area CA1 and DG. The effect of lead on synaptic plasticity in area CA1 was also investigated. The alteration of the amplitude of LTP in hippocampal slices caused by lead was reexamined in order to compare with that on LTD (control: 189±23, n=5; lead-exposed: 122±12, n=10). The result demonstrated that low-level lead exposure could reduce the range of synaptic plasticity, which might underlie the dysfunction of learning and memory caused by chronic lead exposure.     

Developmental changes of MAP2 immunoreactivity in the hippocampus proper and dentate gyrus of the rat

Microtubules are present in high concentration in the nervous system and are a prominent component of the neuronal cytoskeleton. Microtubules are composed of tubulin and variety of microtubule-associated proteins (MAPs), which have been implicated in the regulation of microtubule assembly and function. MAP2 is the most abundant of these proteins, and it has been extensively characterized in various functional and pathological conditions. In the present study the distribution of MAP2 was examined in each layer of the CA1 and CA3 regions of the hippocampus proper and dentate gyrus in rat development. A total of 40 brains at various ages starting from postnatal day (P) 0 to P90 were examined. After perfusional fixation the brains were frozen and cut on the coronal plane and stained with either cresyl violet or standard immunohistochemical methods using the anti-MAP2 antibody. MAP2 exhibited a somatodendritic pattern of localization in cells of the hippocampus. Staining was most prominent in dendrites and perikarya as well as granules surrounding cell bodies. In a newborn rat's brain immunostaining was intense in granules and faint in perikarya. Between P4 and P21 immunostaining density for MAP2 was stronger and appeared in perikarya, granules, and dendritic trees. After P21 the perikarya and dendrites of the pyramidal layer and stratum radiatum of the hippocampus proper, as well as the molecular and granular layer of dentate gyrus, showed reduced immunoreactivity. In the stratum oriens of the hippocampus and polymorphic layer of the dentate gyrus immunoreactivity was still strong until P90.     

Activation of the hippocampus and dentate gyrus by working-memory: a 2-deoxyglucose study of behaving rhesus monkeys

The 2-deoxyglucose method was used to examine metabolic activity in the hippocampus, dentate gyrus, and amygdala of rhesus monkeys performing working-memory and control tasks. A working-memory group was tested on 1 of 3 tasks requiring trial-by-trial updating of information: delayed spatial response, delayed spatial alternation, or delayed object alternation. A control group was tested either on an associative memory problem, visual pattern discrimination, or a sensory-motor task that did not have an explicit mnemonic component. Local cerebral glucose utilization (LCGU) in specific layers of the dentate gyrus and the CA1 and CA3 sectors of the hippocampus, as well as in 7 distinct nuclei of the amygdala, was measured and compared across groups. Metabolic rate in specific layers of the dentate gyrus and the CA3 and CA1 fields of the hippocampus was enhanced in the working-memory compared with the control group: LCGU was between 18 and 24% higher in the granule cell and molecular layers of the dentate gyrus and in the molecular and radiatum layers of CA1 and CA3 in the hippocampus. In contrast, no significant group differences in LCGU were found for any of the 7 amygdaloid nuclei examined: the lateral, lateral basal, medial basal, accessory basal, cortical, central, and medial nuclei. These results are consistent with previous evidence showing that lesions of the hippocampus affect memory selectively, producing deficits on some memory problems while sparing others. Our findings further suggest that working-memory may be a common denominator among those tasks that are sensitive to hippocampal damage in monkeys. The contribution of the amygdala to performance on memory tasks, on the other hand, appears to be independent of the specific type of memory process that is engaged.     

The seasonal pattern of cell proliferation and neuron number in the dentate gyrus of wild adult eastern grey squirrels

The dentate gyrus is one of two areas in the mammalian brain that produces neurons in adulthood. Neurogenesis (proliferation, survival, and differentiation of new neurons) is regulated by experience, and increased neurogenesis appears to be correlated with improved spatial learning in mammals and birds. We tested the hypothesis that in long-lived mammals that scatter-hoard food, seasonal variations in spatial memory processing (i.e. increased processing during caching season in the autumn) might correlate with changes in neurogenesis and neuron number in the granule cell layer of the dentate gyrus (gcl DG). We investigated the rate of cell proliferation and the total number of neurons in the granule cell layer of wild adult eastern grey squirrels (Sciurus carolinensis) at three different times of the year (October, January and June). We found no seasonal differences in cell proliferation rate or in total neuron number in the granule cell layer. Our findings are in agreement with those of previous studies in laboratory mice and rats, and in free-ranging, food-caching, black-capped chickadees, as well as with current hypotheses regarding the relationship between neurogenesis and learning. Our results, however, are also in agreement with the hypothesis that neurogenesis in the dentate gyrus represents a maintenance system that may be regulated by environmental factors, and that changes in total neuron number previously reported in rodents represent developmental changes rather than adult plasticity. The patterns observed in mature wild rodents, such as free-ranging squirrels, may represent more accurately the extent of hippocampal plasticity in adult mammals.     

Maternal administration of magnesium sulfate promotes cell proliferation in hippocampus dentate gyrus in offspring mice after exposing to prenatal stress

Prenatal stress (PS) inhibits cell proliferation in the hippocampal dentate gyrus (DG), which is related to hippocampal anatomy and function abnormality. The aim of the study was to investigate the effects of magnesium sulfate (MgSO₄) on PS-induced cell proliferation suppression in offspring during embryonic stage and postnatal spatial learning. MgSO₄ administration was performed after PS treatment on pregnant mice. Mice were randomly divided into four groups: non-PS or PS maternal mice injected with MgSO₄ or saline (P + NS, P + MG, C + MG and C + NS group). Corticosterone was collected from amniotic fluid of mother mice on day 17 of embryonic stage (E17). The ability for spatial learning and memory of pups postnatal 3 week was evaluated using water maze assay. Additionally, cell proliferation was detected by assessing the expression of Ki67 using immunohistochemistry in mice fetuses or pups. PS significantly increased corticosterone level in amniotic fluid (P < 0.05) and impaired the spatial learning and memory (P + NS vs C + NS of latency time and track path length: P < 0.05) of offspring on postnatal day 21. However, MgSO₄ administration could reverse PS-induced spatial learning and memory disability (P + MG vs P + NS, P < 0.05). Additionally, PS reduced the number of Ki67-positive cell in hippocampal DG on E17, E19 and postnatal day 21 (P + NS vs C + NS, P < 0.05), which were also abrogated by maternal administration of MgSO₄ (P + MG vs P + NS, P < 0.05). Collectively, prenatal administration of MgSO₄ can reverse PS-induced reduction of cell proliferation in hippocampal DG during embryonic stage and postnatal spatial learning.     

Synapse formation and morphological differentiation of neuron types in embryonic rat dentate gyrus explants in vitro

Cultured explants obtained from the dentate gyrus of rat embryos (embryonic day 19–20) were used to investigate synapse formation and morphological differentiation of neuron types in the absence of extrinsic afferents. Synaptogenesis was studied by whole-cell recordings of postsynaptic currents and by ultrastructural analysis. Neurons were visualized using Lucifer Yellow filling or staining with DiI. In short-term (3–5 days) cultured explants postsynaptic currents were rarely evoked by extracellular stimulation and synapses were almost completely absent at the ultrastructural level. After 6–10 days in vitro, the incidence of evoking postsynaptic currents mediated by glutamate and GABA_A receptors was strongly increased. At the ultrastructural level, the density of synapses increased more than 20-fold. These results demonstrate de novo formation of synapses in cultured embryonic dentate gyrus explants. Neuron types could be discriminated by their dendritic arborizations and by their electrophysiological properties. After 6–10 days in vitro, mossy-like cells exhibited 3–4 primary dendrites branching in a characteristic pattern and showed moderate spike-frequency adaptation. Application of serotonin (5-HT) to cultured explants elicited GABA_A-receptor-mediated postsynaptic currents in mossy-like cells, indicating synaptic GABA release from local interneurons. Comparison to 5-HT evoked GABA release in mossy cells in age-matched, acute slices revealed only slight quantitative differences. In contrast to mossy cells, granule cells showing several primary dendrites originating at one cell pole were almost completely absent in cultured explants, suggesting an involvement of extrinsic afferents in the differentiation of granule cells.     

Effect of electroacupuncture on synaptic transmission in dentate gyrus of the hippocampus in cerebral ischemic injured rats

IN T R O D U C T IO N Long-term potentiation (LTP ) is an essentialphenom enon concern- ing cerebral plasticity. It w as routinely used as a basic cellular m odel for the study of m em ory. LTP could be induced by m ul- ti-factors constructing the externa…     

GABAergic neurons containing the Ca2+-binding protein parvalbumin in the rat hippocampus and dentate gyrus

The distribution of Ca2+-binding protein, parvalbumin (PV), containing neurons and their colocalization with glutamic acid decarboxylase (GAD) were studied in the rat hippocampus and dentate gyrus using immunohistochemistry. PV immunoreactive (PV-I) perikarya were concentrated in the granule cell layer and hilus in the dentate gyrus and in the stratum pyramidale and stratum oriens in the CA3 and CA1 regions of the hippocampus. They were rare in the molecular layer of the dentate gyrus, in the stratum radiatum and in the stratum lacunosum-moleculare of the hippocampus. PV-I axon terminals were restricted to the granule cell layer, the stratum pyramidale and the immediately adjoining zones of these layers. Almost all PV-I neurons were also GAD immunoreactive (GAD-I), whereas only about 20% of GAD-I neurons also contained PV. The percentages of GAD-I neurons which were also immunoreactive for PV were dependent on the layer in which they were found; i.e. 40-50% in the stratum pyramidale, 20-30% in the dentate granule cell layer and in the stratum oriens of the CA3 and CA1 regions, 15-20% in the hilus and in the stratum lucidum of CA3 region and only 1-4% in the dentate molecular layer and in the stratum radiatum and the stratum lacunosum-moleculare of the CA3 and CA1 regions. PV-I neurons are a particular subpopulation of GABAergic neurons in the hippocampal formation. Based on their morphology and laminar distribution, they probably include basket cells and axo-axonic cells.     

Factors specifying the development of synapse number in the rat dentate gyrus: effects of partial target loss

The development of the dentate gyrus has been studied under conditions of partial reduction of granule cell number. Neonatal rats were subjected to X-irradiation, a procedure which reduces the number of granule cells to 20% of control values. In X-irradiated rats, quantitative analyses were performed on cells in the entorhinal cortex which give rise to the perforant path projection to the dentate granule cells, and on the remaining, undamaged dentate granule cells. These residual cells were examined morphologically for possible hyperdevelopment in comparison to granule cells from control animals. Granule cells in X-irradiated animals were similar to granule cells in control animals with respect to dendritic structure and synaptic density. The number of neurons in both the medial and lateral entorhinal cortices in X-irradiated animals appeared normal until day 12, at which time a selective reduction in cell numbers became apparent. By day 30, 25–55% of the cells of origin of the perforant path were absent in X-irradiated animals. It is hypothesized that these cells are subject to retrograde transynaptic degeneration as a result of target removal. Further, it appears that granule cells play an important role in determining the density of their innervation.     

Kainic acid-induced sprouting of dynorphin- and enkephalin- containing mossy fibers in the dentate gyrus of the rat hippocampus

This study utilized Timm histochemistry and immunocytochemistry to determine the prolonged effects of kainic acid on the distribution of dynorphin- and enkephalin-containing mossy fibers in the rat dentate gyrus at progressive time points following kainic acid injection. Beginning 1–2 weeks after kainic acid administration, a progressive increase in the distribution and intensity of staining for supragranular zinc, dynorphin and enkephalin was observed in the dentate gyrus. The kainic acid-induced sprouting of mossy fibers containing dynorphin and enkephalin strongly resembles the pattern observed in the dentate gyrus of humans with temporal lobe epilepsy.