Environmental enrichment restores neurogenesis and rapid acquisition in aged rats

Strategies combatting cognitive decline among the growing aging population are vital. We tested whether environmental enrichment could reverse age-impaired rapid spatial search strategy acquisition concomitantly with hippocampal neurogenesis in rats. Young (5–8 months) and aged (20–22 months) male Fischer 344 rats were pair-housed and exposed to environmental enrichment (n = 7 young, 9 aged) or housed individually (n = 7 young, 7 aged) for 10 weeks. After 5 weeks, hidden platform trials (5 blocks of 3 trials; 15 m inter-block interval), a probe trial, and then visible platform trials (5 blocks of 3 trials; 15 m inter-block interval) commenced in the water maze. One week after testing, rats were given 5 daily intraperitoneal bromodeoxyuridine (50 mg/kg) injections and perfused 4 weeks later to quantify neurogenesis. Although young rats outperformed aged rats, aged enriched rats outperformed aged individually housed rats on all behavioral measures. Neurogenesis decreased with age but enrichment enhanced new cell survival, regardless of age. The novel correlation between new neuron number and behavioral measures obtained in a rapid water maze task among aged rats, suggests that environmental enrichment increases their ability to rapidly acquire and flexibly use spatial information along with neurogenesis.     

Aging impairs amygdala-hippocampus interactions involved in hippocampal LTP.

Aging impairs amygdala-hippocampus interactions involved in hippocampal LTP. NEUROBIOL. AGING. We have recently shown that the stimulation of the basolateral nucleus of the amygdala (BLA) is able to prolong early-LTP (<4h) into late-LTP (>4h) in the dentate gyrus. To study whether aging affects this interaction, aged (24-27 months) rats were used, classified as cognitively impaired (I), or non-impaired (N) by means of their results in the Morris water maze. Paired pulses (30-90 ms interval) showed no differences among age groups. Among young controls, the early-LTP induced in the dentate gyrus by stimulation of the perforant path (PP) was prolonged in a late-LTP when the BLA was stimulated 15 min later. In aged-impaired rats the stimulation of the PP induced a reduced LTP, decaying to baseline in less than 2 h. BLA stimulation was without effect. Aged non-impaired rats showed an early-LTP identical to that of young animals; however, stimulation of the BLA showed no effect. These results suggest that deficient synaptic plasticity and memory functions in aged animals might be caused, in part by impaired mechanisms of heterosynaptic reinforcement.     

Running reduces stress and enhances cell genesis in aged mice

Cell proliferation and neurogenesis are diminished in the aging mouse dentate gyrus. However, it is not known whether isolated or social living affects cell genesis and stress levels in old animals. To address this question, aged (17–18 months old) female C57Bl/6 mice were single or group housed, under sedentary or running conditions. We demonstrate that both individual and socially housed aged C57Bl/6 mice have comparable basal cell proliferation levels and demonstrate increased running-induced cell genesis. To assess stress levels in young and aged mice, corticosterone (CORT) was measured at the onset of the active/dark cycle and 4 h later. In young mice, no differences in CORT levels were observed as a result of physical activity or housing conditions. However, a significant increase in stress in socially housed, aged sedentary animals was observed at the onset of the dark cycle; CORT returned to basal levels 4 h later. Together, these results indicate that voluntary exercise reduces stress in group housed aged animals and enhances hippocampal cell proliferation.     

Estrogen effects on pilocarpine-induced temporal lobe epilepsy in rats

Objetive

To evaluate the effects of conjugated equine estrogens (CEE) on the pilocarpine-induced epilepsy in rats.

Study design

40 female rats were divided into: GPC (positive control) presented “status epilepticus” (SE) induced by pilocarpine; GOC (ovariectomized control) only castrated; GNC (negative control) received only saline solution; GPE received pilocarpine, presented SE, castrated and received 50 μg/kg CEE treatment; GPV received pilocarpine, castrated and received propylene glycol (vehicle). The animals were monitored by a video system. At the end of observation, the brains removed for later histologic analysis using Neo-Timm and Nissl methods.

Results

The GPE presented a reduction in number of seizures compared to GPV. The Neo-Timm analysis showed that GPV had greater sprouting of mossy fibers, with a denser band in the area of the dentate gyrus hilum compared to GPE. On Nissl staining, GPE showed evident neuronal loss in the CA3 area. GPV presented loss in CA1 and dentate gyrus.

Conclusion

Estrogen may have a protecting effect on the central nervous system.     

Influence of aging on chondroitin sulfate proteoglycan expression and neural stem/progenitor cells in rat brain and improving effects of a herbal medicine, yokukansan

There is evidence of structural and functional deterioration in the brain, including the prefrontal cortex (PFC) and hippocampus, during the normal aging process in animals and humans. Extracellular matrix-associated glycoproteins, such as chondroitin sulfate proteoglycans (CSPGs), are involved in not only maintaining the structures and functions of adult neurons, but also regulating the proliferation, migration, and neurite outgrowth of neural stem cells in the brain. On the other hand, a herbal medicine, yokukansan (YKS), is used in a variety of clinical situations for treating symptoms associated with age-related neurodegenerative disorders such as Alzheimer's disease, but its pharmacological properties have not been fully understood. The present study was designed to clarify the influence of aging and the improving effects of YKS on the expression of aggrecan, a major molecule of CSPGs, and on the proliferation and migration of neural stem/progenitor cells identified by bromodeoxyuridine (BrdU) incorporation in the PFC and hippocampus including the dentate gyrus. Aged rats (24 months old) showed a significant increase in aggrecan expression throughout the PFC and in the hippocampus particularly in the CA3 subfield, but not the dentate gyrus compared to young rats (5 months old), evaluated by the immunohistochemical method. YKS treatment decreased the age-related increase in aggrecan expression as well as normal expression in young rats. Aged rats also showed a decreased number of BrdU-labeled cells in the PFC and hippocampus, and these decreases were improved by YKS treatment, which also increased the numbers in young rats. These results suggest that aging influences the microenvironment for adult and immature neurons in the brain, which may affect the proliferation and migration of neural stem/progenitor cells, and YKS has pharmacological potency for these age-related events. These findings help to understand the physiology and pathology of the aged brain and provide an anti-aging strategy for the brain.     

Deficits in radial-arm maze learning in aged rats

The present study was designed to investigate the possible deficits in the place learning on the 8-arm radial maze in aged rats. In this task, reward was given in the 4 predetermined arms. Aged rats (27 months old, N = 7) acquired this task more slowly than young rats (12 months old, N = 11), and didn't reach to the performance level of the young rats within 80 training trials. Analysis of error choices revealed that the aged animals first entered in the unbaited arms more often than the young rats, whereas there was no difference in the number of re-entered choices to the baited and unbaited arms between the aged and young animals. Therefore, it was concluded that learning deficits in aged rats were attributed to deficits in the reference memory but not in the working memory.     

Neuroinflammation not associated with cholinergic degeneration in aged-impaired brain

Degeneration of the cholinergic neurons in the basal forebrain and elevation of inflammatory markers are well-established hallmarks of Alzheimer's disease; however, the interplay of these processes in normal aging is not extensively studied. Consequently, we conducted a neuroanatomical investigation to quantify cholinergic neurons and activated microglia in the medial septum/vertical diagonal band (MS/VDB) of young (6 months) and aged (28 months) Fisher 344 × Brown Norway F₁ rats. Aged rats in this study were impaired relative to the young animals in spatial learning ability as assessed in the Morris water maze. Stereological analysis revealed no difference between aged and young rats in the total numbers of cholinergic neurons, demonstrating that loss of cholinergic neurons is not a necessary condition to observe impaired spatial learning in aged rats. In this same region, the total number of activated microglia was substantially greater in aged rats relative to young rats. Jointly, these data demonstrate that aging is characterized by an increase in the basal inflammatory state within the MS/VDB, but this inflammation is not associated with cholinergic neuron death.     

Investigation of oxidative stress involvement in hippocampus in epilepsy model induced by pilocarpine

The relationship between free radical and scavenger enzymes has been found in the epileptic phenomena and reactive oxygen species have been implicated in seizure-induced neurodegeneration. Using the epilepsy model obtained by systemic administration of pilocarpine in rats, we investigated the lipid peroxidation, nitrite content, superoxide dismutase (SOD) and catalase activities in the hippocampus of rats during chronic period. The enzyme activities as well as the lipid peroxidation and nitrite concentrations were measured using spectrophotometric methods and the results compared to values obtained from saline-treated animals. The superoxide dismutase and catalase activities increased during the chronic phase. In addition, lipid peroxidation and nitrite levels increased in same period in the hippocampus of animals observed during spontaneous recurrent seizures. Previous studies showed that animals presenting seizures and submitted to 24 h of status epilepticus showed normal levels of superoxide dismutase and increased in catalase activities as well as an increase in hippocampal lipid peroxidation and nitrite concentrations. These results show a direct evidence of lipid peroxidation and nitrite during seizure activity that could be responsible for neuronal damage in the hippocampus of rats, during the establishment of pilocarpine model of epilepsy.     

Delayed peripheral administration of the N-terminal tripeptide of IGF-1 (GPE) reduces brain damage following microsphere induced embolic damage in young adult and aged rats

We have previously reported that peripheral administration of GPE prevents neuronal injury after ischemic reperfusion injury in young adult rats. This study examined the ameliorating effects of GPE-treatment after embolic injury induced by microsphere injection in young adult and aged male rats. Unilateral injury was induced by injecting microspheres into the right internal carotid artery in both young adult (3–4 months) and aged (16–17 months) male rats. Either GPE (12 mg/kg) or the vehicle was infused intravenously over 1 h starting 3 h after embolic injury and the degree of brain injury, astrocytosis and vascular remodeling were examined using histological and immunohistochemical analysis 8 days later. Changes in core temperature, blood glucose concentration, oxygen saturation and heart rate were monitored. Microsphere injection induced multiple sites of focal damage in the ipsilateral subcortical regions. Massive numbers of microglia accumulated within the core of the tissue damage whereas astrocytes were located in the penumbra. There was no difference in the degree of brain injury between the young and aged control rats. However the aged rats showed less injury-induced astrocytosis and greater vascular remodeling. Intravenous infusion of GPE 3 h after the injury reduced overall damage scores in both young (p < 0.01) and aged rats (p < 0.05). GPE-treatment reduced astrocytosis in young, but not aged animals and did not significantly alter the vascular remodeling in either age group. The data suggested that the neuroprotection of the tripeptide is independent of cerebral reperfusion and is not age selective.     

Altered dendritic integration in hippocampal granule cells of spatial learning-impaired aged rats

Glutamatergic transmission at central synapses undergoes activity-dependent and developmental changes. In the hippocampal dentate gyrus, the non-N-methyl d-aspartate (NMDA) receptor component of field excitatory postsynaptic potentials (fEPSPs) increases with age in Fischer-344 rats. This effect may not depend on the animal's activity or experience but could be part of the developmental process. Age-dependent differences in synaptic transmission at the perforant path-granule cell synapse may be caused by changes in non-NMDA and NMDA receptor-mediated currents. To test this hypothesis, we compared whole cell excitatory postsynaptic currents (EPSCs) in dentate granule cells evoked by perforant path stimulation in young (3-4 mo) and aged (22-27 mo) Fischer-344 rats using a Cs+-based intracellular solution. Aged animals as a group showed spatial learning and memory deficits in the Morris water maze. Using whole cell recordings, slope conductances of both non-NMDA and NMDA EPSCs at holding potentials -10 to +50 mV were significantly reduced in aged animals and the non-NMDA/NMDA ratio in aged animals was found to be significantly smaller than in young animals. In contrast, we detected no differences in basic electrophysiological parameters, or absolute amplitudes of non-NMDA and NMDA EPSCs. Extracellular Cs+ increased the fEPSP in young slices to a greater degree than was found in the aged slices, while it increased population spikes to a greater degree in the aged rats. Our results not only provide evidence for reduced glutamatergic synaptic responses in Fischer-344 rats but also point to differential changes in Cs+-sensitive dendritic conductances, such as Ih or inwardly rectifying potassium currents, during aging.     

Efficacy of fish liver oil and propolis as neuroprotective agents in pilocarpine epileptic rats treated with valproate

Objective: To evaluate the action of fish liver oil and propolis in pilocarpine epileptic rats treated with the anticonvulsant drug valproate. Methods: Seven groups of rats were treated daily for six months: control; fish liver oil (0.4 ml/kg b.w); propolis (50 mg/kg b.w); pilocarpine-treated rats (epileptic control); epileptic rats treated with valproate (400 mg/kg b.w); groups 6 and 7, epileptic rats treated with valproate plus fish liver oil or propolis. Results: Pilocarpine administration caused a significant increase in hippocampal dopamine and serotonin levels accompanied with a significant decrease in their levels in serum. Lipid peroxidation level and LDH activity in hippocampus were significantly increased after pilocarpine treatment whereas Na⁺/K⁺-ATPase activity and total antioxidant capacity were significantly decreased compared to the controls. Animals treated with the combined treatments showed a significant improvement in tested parameters towards the normal values of the control. Conclusion: Fish liver oil and propolis when given in combination with valproate, neuroprotected against the neurophysiological disorders induced by pilocarpine epilepsy in rats.     

Protective effects of ascorbic acid and garlic extract against neurogenesis inhibition caused by developmental lead exposure in the dentate gyrus of rat

Lead is a well-known neurotoxin that affects the developing central nervous system and may potentially inhibit neurogenesis in adults. We investigated the effect of ascorbic acid and garlic extract against lead-induced neurotoxicity in developing rat dentate gyrus. Female Wistar rats were divided randomly into five groups: lead-treated (L; 1,500 ppm lead acetate in drinking water) group, lead plus ascorbic acid-treated (L?+?AA; 500 mg/kg, ip) group, lead plus garlic juice-treated (L?+?G; 1 ml /100 g BW, gavage) group, sham group (sh), and controls. All treatments were administered to female rats during pregnancy and lactation. At the end of the treatment, dentate gyrus neurogenesis were determined using Doublecortin (DCX) immunohistochemistry in the hippocampus of 50-day-old male pups. DCX-positive cells in the dentate gyrus were counted and compared between the groups. Lead exposure caused a significant increase in blood and brain lead concentration vs. control (P?<?0.001); whereas, co-administration of ascorbic acid or garlic?+?lead was effective in reducing blood and brain lead levels (P?<?0.01). The number of DCX-positive cells in the dentate gyrus of the lead-exposed group was significantly lower, when compared with controls. A statistically significant increase in number of DCX-positive cells in ascorbic acid and garlic groups compared with lead-exposed rats was noted (P?<?0.05). This study provides evidence of the beneficial role of ascorbic acid and garlic in early status of dentate gyrus neurogenesis in rats against lead exposure.     

Morphometric analysis of mitochondria and boutons in the dentate gyrus molecular layer of aged rats

Summary An electron microscopic morphometric analysis of the volume density of mitochondria and boutons in the dentate gyrus molecular layer was carried out in young (3–4 months old) and aged (26–27 months old) Wistar rats. This study showed that the volume fraction of mitochondria per unit volume of the neuropil in aged rats did not differ significantly from that in young animals. The comparison of different zones of the molecular layer (supragranular, inner, middle and outer zone) showed a significant increase in the mitochondrial volume density from the supragranular to the outer zone in both animal groups. These stereologic results are discussed in relation to the histochemical pattern of the mitochondrial enzymes in young and aged rats. Only in the supragranular zone was there a statistically significant difference in the volume density of boutons, i.e. aged rats showed about a 20% higher volume density than did young rats. It is suspected that this increase in bouton volume density could be due to the agerelated atrophy of smaller dendritic shafts previously reported in senescent Fischer rats.     

The effects of alcohol intake and withdrawal on the seizures frequency and hippocampal morphology in rats with epilepsy

The aim of our study, using the pilocarpine model of epilepsy, was to investigate the effects of alcohol administration and withdrawal on the spontaneous recurrent seizures (SRSs). Four groups of adult, male Wistar rats were studied: (A). control rats (n=10), received neither pilocarpine nor alcohol, (B). alcohol-treated rats (n=10), received a daily dose of 3.0 g x kg(-1) of a 30% alcohol solution via an oesophagic probe for 30 days, (C). rats with epilepsy (n=10), (D). rats with epilepsy with alcohol intake (n=10). SRSs were induced by a single dose of pilocarpine (i.p.) and the basal frequency of SRSs was video monitored (24h per day) for 30 days. Following this period, the animals of group D received a daily dose of alcohol solution as described above and at the end of this period, alcohol administration was stopped and the seizure frequency was assessed for more 30 days. The basal seizure frequency observed in groups C and D during the first 30 days was 2.2+/-1.8 seizures per week per animal. In group D, it was observed an increase to 12.2+/-5.8 during the first 2 weeks of alcohol administration. During the last 2 weeks of alcohol administration, the number of SRSs returned to the previous basal level. During alcohol withdrawal the seizure frequency increased to 14.3+/-7.4 seizures per week per animal for the first 2 weeks, and returned to the basal level in the remaining period of observation. The Neo-Timm and Nissl staining of hippocampal formation and of the dentate gyrus in rats with epilepsy showed a cell loss in the hippocampal subfield CA1 and in the hillus of dentate gyrus. In rats with epilepsy with alcohol intake, we observed a cell loss in hippocampal subfields CA3 and hillus of the dentate gyrus, with significant neuronal death in subfield CA1, when compared with control animals. The alcohol withdrawal syndrome is a crucial event for the development of functional and neuropathological alterations associated with epilepsy.     

Hippocampal CA field neurogenesis after pilocarpine insult: The hippocampal fissure as a neurogenic niche

Pilocarpine model for temporal lobe epilepsy has shown aberrant neurogenesis, but mainly restricted to the dentate gyrus (DG). Herein, by using a modified protocol, combining pilocarpine with ipratropium bromide, we unexpectedly observed a heretofore-unrecognized distinct cellular population expressing the neuroprogenitor marker doublecortin (DCX) on post insult days (PID) 10, 14 and 18, mainly located in the temporal segment of the hippocampal fissure (hf). Some of these DCX+ cells possessed high morphological complexity and seemed to disperse toward the CA fields. Next, we injected bromodeoxyuridine (BrdU) in early (PID 2–4) and delayed (PID 5–7) fashions and killed the rats 7–35 days later for immunohistochemical and anatomical analysis. Massive increase of BrdU labeling was found in the delayed group and the neural stem cell-specific marker nestin was highly expressed in the same narrow band on PID7, so was glial fibrillary acidic protein (GFAP). Using double labeling with BrdU and a mature neuron marker NeuN, we found discrete but clear BrdU+/NeuN+ double labeled cells in the Cornu Ammonis (CA) pyramidal cell layer on PID35. Based on immunohistochemical and anatomical observations, as well as time-course analysis of BrdU, nestin, GFAP, DCX and NeuN expressions in this population of cells located in/near hf, we wish to suggest that this structure harbors neurogenic niches, in addition of the possible dispersion of neuroprogenitors from subgranular niches to CA fields also revealed by this study. Our results support the few previous reports demonstrating hippocampal CA field neurogenesis in adult rats. Mechanistic basis of the phenomenon is discussed.     

Dendritic spines in the posterodorsal medial amygdala after restraint stress and ageing in rats

Several evidences suggest that the posterodorsal medial amygdala (MePD) can be a relevant part of the rat neural circuitry for the regulation of hypothalamic neuroendocrine secretion and for ontogenetically different behavioral displays. The dendritic spine density of Golgi-impregnated neurons from the MePD was evaluated in young rats following acute or chronic restraint stress and in aged animals (24 months old). Compared to the control group, a single 1 h restraint stress session promoted a decreased spine density (p < 0.01) whereas a single 6 h restraint stress session or daily 6-h restraint sessions for 28 consecutive days did not lead to the same effect (p > 0.05). Aged rats showed no difference in this dendritic spine parameter when compared to young adults (p > 0.05). These results indicate that short-term stress (1 h) can affect MePD dendritic spines and that neural plasticity is involved with adaptive responses onwards in restrained rats. On the other hand, brain structural modifications related with ageing appear not to influence the number of certain postsynaptic sites in the MePD of rats.     

Age-dependent alterations in hippocampal synaptic plasticity: relation to memory disorders

In this paper, we review the evidence indicating that the common disturbance in recent memory associated with aging is a consequence of functional and structural impairment in the hippocampal formation. In the Fischer 344 rat, an experimental model of the human age-related memory disorder was developed. The majority of aged rats of this strain show impaired performance in the 8-arm radial maze in a manner typical of young rats with bilateral hippocampal lesions. Aged animals also exhibit rapid decay of LTP and slower kindling of the perforant path-dentate synapse. Furthermore, quantitative morphometric analysis of the hippocampal synaptic architecture revealed that aged, memory-impaired rats had a specific loss of perforated axospinous synapses in the middle third of the dentate gyrus molecular layer; the extent of loss was directly related to the degree of memory dysfunction. Most important was the fact that the electrophysiological and morphological abnormalities did not appear in equally old animals with good memory.     

Postural control strategy during standing is altered in patients with multiple sclerosis

It was previously shown that the ictogenic potential of 4-aminopyridine (4-AP) was reduced in the parahippocampal region of kainate treated chronic epileptic rats. In the actual study we investigated the potential of 4-aminopyridine (50 and 100μM) to induce seizure like events (SLEs) in combined entorhinal cortex hippocampal slices from Wistar rats following pilocarpine induced status epilepticus. The potential of 4-AP to induce SLEs in the entorhinal cortex was reduced in the latent period and in slices of chronic epileptic animals with a high seizure incidence in vivo (>2seizures/24h). 4-AP induced SLEs in slices from animals with a low incidence of seizures in vivo (<2seizures/24h) in a similar manner as compared to controls. The hippocampal formation displayed no SLEs, instead short recurrent epileptiform discharges (REDs) were evoked by application of 4-AP in areas CA3 and CA1. The incidence of REDs was largest in slices from control animals. This study shows that the reduced ictogenic potential of 4-AP is not restricted to kainate treated chronic epileptic animals as it can be found in the pilocarpine model as well. The underlying mechanisms may relate to altered expression and editing of voltage gated potassium channels.     

Memory enhancement in aged rats: the differential outcomes effect

Aged (23 months) and young (3 months) rats were trained on an operant Matching-To-Position (MTP) task that had either (a) specific outcomes (reinforcers) correlated (differential groups), or (b) outcomes uncorrelated (nondifferential groups) for each correct sample-choice sequence. The traditional version of MTP uses a common outcome and is thought to assess spatial working memory. Aged rats are impaired on the traditional version of MTP. However, aged animals trained with the Differential Outcomes Procedure (DOP) did not display the typical age-related decline in spatial working memory. Differences in choice accuracy between old and young rats reached significance only if the subjects were trained with a nondifferential outcomes procedure (NOP)-similar to when a common outcome is used. These data demonstrate that employing behavioral procedures to tap intact cognitive functions is an effective means of enhancing spatial working memory in normal as well as aged subjects.     

Resistance training improves femoral artery endothelial dysfunction in aged rats

Although endurance exercise improves age-associated endothelial dysfunction, few studies have examined the effects of resistance training and the potential molecular mechanisms involved in altering vascular reactivity with age. Young (9 months) and aged (20 months) male, Fisher 344 rats were divided into four groups: Young Sedentary (YS, n = 14), Young Trained (YT, n = 10), Aged Sedentary (AS, n = 12), and Aged Trained (AT, n = 10). Resistance training consisted of climbing a 1 m wire ladder, at an 85° angle, 3 days/week for 6 weeks with increasing weight added to the tail. Endothelial function in femoral arteries was determined by constructing acetylcholine dose–response curves on a wire myograph. Femoral artery phospho-Ser1179-eNOS, eNOS and Hsp90 expression were evaluated by Western blot. Acetylcholine-induced vasorelaxation was significantly (P < 0.05) impaired in AS compared to YS and YT but not AT compared to YS and YT. Phospho-Ser1179-eNOS and eNOS were elevated (P < 0.05) in aged animals but not changed with resistance training. Resistance training increased Hsp90 levels in both young and old animals. Therefore, resistance training improves age-associated endothelial dysfunction in femoral arteries without changes in eNOS phosphorylation and expression. Increased Hsp90 expression, a regulator of eNOS activity and coupling, suggests a potential mechanism for this improvement.