Rat Dentate Gyrus Research Articles

Zuogui Jiangtang Jieyu Decoction promotes neural stem cell self-renewal and activates Shh signaling in the hippocampal dentate gyrus of diabetic rats with depression

To investigate the effect of Zuogui Jiangtang Jieyu Decoction (ZJJ) on Shh signaling and self-renewal of neural stem cells in the hippocampal dentate gyrus of diabetic rats with depression. Diabetic rat models with depression were randomly divided into model group, positive drug (metformin + fluoxetine) group, and low-, medium-, and high-dose ZJJ groups (n=16), with normal SD rats as the control group. The positive drugs and ZJJ were administered by gavage, and the rats in the control and model groups were given distilled water. After the treatment, blood glucose level was detected using test strips, and behavioral changes of the rats were assessed by forced swimming test and water maze test. ELISA was used to examine the serum level of leptin; The expressions of nestin and Brdu proteins in the dentate gyrus of the rats were detected using immunofluorescence assay, and the expressions of self-renewal marker proteins and Shh signaling proteins were detected using Western blotting. The diabetic rats with depression showed significantly increased levels of blood glucose and leptin (P < 0.01) and prolonged immobility time in forced swimming test (P < 0.01) and increased stage climbing time with reduced stage seeking time and stage crossings in water maze test (P < 0.01). The expressions of nestin and Brdu in the dentate gyrus, the expressions of cyclin D1, SOX2, Shh, Ptch1, Smo in the hippocampus and the nuclear expression of Gli-1 were decreased (P < 0.01) while hippocampal Gli-3 expression was increased significantly (P < 0.01) in the rat models. Treatment of rat models with high-dose ZJJ significantly reduced the blood glucose (P < 0.01) and leptin level (P < 0.05) and improved their performance in behavioral tests (P < 0.01). The treatment also obviously increased the expressions of nestin, Brdu, cyclin D1, SOX2, Shh, Ptch1, and Smo and the nuclear expression of Gli-1 in the dentate gyrus (P < 0.01) and reduced hippocampal expression of Gli-3 (P < 0.05) in the rat models. ZJJ can significantly improve the self-renewal ability of neural stem cells and activate Shh signaling in dentate gyrus of diabetic rats with depression.

Efeitos dos exercícios físicos sobre a neurogênese do cérebro de ratos adolescentes

Introduction. The neurogenesis of the brain of young rats can be affected by several factors in their development, with the practice of physical exercises being an important factor in the neurogenesis of nervous tissue. Objective. To investigate the effects of different physical exercise protocols of medium and high intensity on neurogenesis in the brain of adolescent rats. Method. The experimental study, where three groups of animals were used: a control group (RC), a group that performed medium-intensity physical exercise (R1) and a group that performed high-intensity physical exercise (R2). Each group with 8 animals, totaling 24 animals. The progressive resisted physical exercise program on the vertical ladder was carried out for 4 weeks, 5 days a week with a duration varying between 30 and 45 minutes. After the end of the experiments, the animals were euthanized for removal of the brain and subsequent processing, staining using the Nissl method and histomorphometric analysis by counting neurons in the dentate gyrus of the hippocampus and subventricular zone. Results. The data demonstrated that there was no significant difference in the area and height of the apex of the hippocampal dentate gyrus between the groups (p=0.2474 and p=0.3337; respectively). The results of the length of the apex of the hippocampal dentate gyrus showed significant differences (p=0.0172). Conclusion. The practice of medium and high intensity progressive resisted physical exercises did not modify the morphology and neuronal density in the hippocampal dentate gyrus and subventricular zone of young rats.

Moderate Copper Deprivation during Gestation and Lactation Affects Dentate Gyrus and Hippocampal Maturation in Immature Male Rats

The hippocampal formation (HF) is involved in higher brain functions including learning and declarative memory. The possibility that dietary copper has a role in the morphological development, and therefore the function of the HF, has received little attention. A rat model of tiered copper deficiency, initiated during gestation, was employed to determine the susceptibility of the HF, regions of which develop postnatally, to copper deficiency. At postnatal 23, pups whose dams had received either 1.8 or 1.4 mg Cu/kg diet during both gestation and lactation, compared with offspring of a group that had received 4.3 mg Cu/kg diet during both periods had, significantly more cell nuclei in the infrapyrimidal arm of the dentate gyrus. Offspring of rats fed 1.4 mg Cu/kg diet, but not those fed 1.8 mg/kg, compared with those fed 4.3 mg/kg, exhibited smaller, shorter, and narrower cell nuclei in the infrapyrimidal and suprapyrimidal arms of the dentate gyrus and smaller cell nuclei in region CA3c of the hippocampus. A fourth group (gestation, 1.8 mg Cu/kg diet; lactation, 0.9 mg Cu/kg diet) exhibited alterations less marked than those exhibited by the group fed 1.4 mg Cu/kg diet. All alterations in the groups fed low copper diets were consistent with slowed cell nuclear maturation. The findings indicate that copper is required for maturation of the dentate gyrus and hippocampus. Also, copper supplied at or below 1.8 mg/kg is insufficient for morphological maturation of the dentate gyrus and hippocampus. Because the HF is important for higher brain functions, further research is needed to determine whether the copper deficiency-induced alterations in dentate gyrus and hippocampus development are transient or permanent.

Repeated administration of acrylamide for 28 days suppresses adult neurogenesis of the olfactory bulb in young-adult rats

Acrylamide (AA) is a neurotoxicant that inhibits synaptic function in distal axons. We previously found that AA decreased neural cell lineages during late-stage differentiation of adult hippocampal neurogenesis and downregulated genes related to neurotrophic factor, neuronal migration, neurite outgrowth, and synapse formation in the hippocampal dentate gyrus in rats. To investigate whether olfactory bulb (OB)-subventricular zone (SVZ) neurogenesis is similarly affected by AA exposure, AA was administered to 7-week-old male rats via oral gavage at doses of 0, 5, 10, and 20 mg/kg for 28 days. Immunohistochemical analysis revealed that AA decreased the numbers of doublecortin-positive (+) cells and polysialic acid-neural cell adhesion molecule+ cells in the OB. On the other hand, the numbers of doublecortin+ cells and polysialic acid-neural cell adhesion molecule+ cells in the SVZ did not change with AA exposure, suggesting that AA impaired neuroblasts migrating in the rostral migratory stream and OB. Gene expression analysis in the OB revealed that AA downregulated Bdnf and Ncam2, which are related to neuronal differentiation and migration. These results suggest that AA decreased neuroblasts in the OB by suppressing neuronal migration. Thus, AA decreased neuronal cell lineages during late-stage differentiation of adult neurogenesis in the OB-SVZ, similar to the effect on adult hippocampal neurogenesis.

Lesion-induced sprouting promotes neurophysiological integration of septal and entorhinal inputs to granule cells in the dentate gyrus of rats

Axonal sprouting of dentate gyrus (DG) afferents after entorhinal cortex (EC) lesion is a model preparation to assess lesion-induced functional reorganization in a denervated target structure. Following a unilateral EC lesion, the surviving contralateral entorhinal projection, termed the crossed temporodentate pathway (CTD), and the heterotypic septal input to the DG, the septodentate pathway (SD), undergo extensive axonal sprouting. We explored whether EC lesion alters the capacity of the SD pathway to influence CTD-evoked granule cell excitability in the DG. We recorded extracellular field excitatory postsynaptic potentials (fEPSPs) after CTD stimulation alone and paired SD-CTD stimulation. Male rats were given unilateral EC lesions or sham operations; evoked fEPSPs in the DG were recorded at 4-, 15-, and 90-days post-entorhinal lesion to assess functional reorganization of the CTD and SD pathways. We found significantly increased fEPSP amplitudes in cases with unilateral lesions compared to sham-operates at 15- and 90-days post lesion. Within each time point, paired SD-CTD stimulation resulted in significantly depressed fEPSP amplitudes compared to amplitudes evoked after CTD stimulation alone and this effect was solely seen in cases with EC lesion. In cases where granule cell discharge was observed, SD stimulation increased discharge amplitude elicited by the CTD stimulation at 90-days postlesion. These findings demonstrate that synaptic remodeling following unilateral cortical lesion results in a synergistic interaction between two established hippocampal afferents that is not seen in uninjured brains. This work may be important for models of neurodegenerative disease and neural injury that target these structures and associated hippocampal circuitry.

Isolation and Culture of Neural Stem/Progenitor Cells from the Hippocampal Dentate Gyrus of Young Adult and Aged Rats.

Adult neural stem/progenitor cells (NSPCs) in two neurogenic areas of the brain, the dentate gyrus and the subventricular zone, are major players in adult neurogenesis. Addressing specific questions regarding NSPCs outside of their niche entails in vitro studies through isolation and culture of these cells. As there is heterogeneity in their morphology, proliferation, and differentiation capacity between these two neurogenic areas, NSPCs should be isolated from each area through specific procedures and media. Identifying region-specific NPSCs provides an accurate pathway for assessing the effects of extrinsic factors and drugs on these cells and investigating the mechanisms of neurogenesis in both healthy and pathologic conditions. A great number of isolation and expansion techniques for NSPCs have been reported. The growth and expansion of NSPCs obtained from the dentate gyrus of aged rats are generally difficult. There are relatively limited data and protocols about NSPCs isolation and their culture from aged rats. Our approach is an efficient and reliable strategy to isolate and expand NSPCs obtained from young adult and aged rats. NSPCs isolated by this method maintain their self-renewal and multipotency. Key features • NSPCs isolated from the hippocampal dentate gyrus of young adult and aged rats, based on Kempermann et al. (2014) and Aligholi et al. (2014). • Maintenance of NSPCs isolated from the dentate gyrus of aged rats (20-24 months) in our culture condition is feasible. • According to our protocol, maximum growth of primary neurospheres obtained from isolated NSPCs of young and aged rats took 15 and 35 days, respectively.

Adult-born neurons inhibit developmentally-born neurons during spatial learning

Ongoing neurogenesis in the dentate gyrus (DG) subregion of the hippocampus results in a heterogenous population of neurons. Immature adult-born neurons (ABNs) have physiological and anatomical properties that may give them a unique role in learning. For example, compared to older granule neurons, they have greater somatic excitability, which could facilitate their recruitment into memory traces. However, recruitment is also likely to depend on interactions with other DG neurons through processes such as lateral inhibition. Immature ABNs target inhibitory interneurons and, compared to older neurons, they receive less GABAergic inhibition. Thus, they may induce lateral inhibition of mature DG neurons while being less susceptible to inhibition themselves. To test this we used a chemogenetic approach to silence immature ABNs as rats learned a spatial water maze task, and measured activity (Fos expression) in ABNs and developmentally-born neurons (DBNs). A retrovirus expressing the inhibitory DREADD receptor, hM4Di, was injected into the dorsal DG of male rats at 6w to infect neurons born in adulthood. Animals were also injected with BrdU to label DBNs or ABNs. DBNs were significantly more active than immature 4-week-old ABNs. Silencing 4-week-old ABNs did not alter learning but it increased activity in DBNs. However, silencing ABNs did not affect activation in other ABNs within the DG. Silencing ABNs also did not alter Fos expression in parvalbumin- and somatostatin-expressing interneurons. Collectively, these results suggest that ABNs may directly inhibit DBN activity during hippocampal-dependent learning, which may be relevant for maintaining sparse hippocampal representations of experienced events.

Sucrose Consumption during Late Adolescence Impairs Adult Neurogenesis of the Ventral Dentate Gyrus without Inducing an Anxiety-like Behavior.

Sucrose consumption impairs behavioral and cognitive functions that correlate with decreased neurogenesis in animal models. When consumed during early adolescence, this disaccharide promotes anxious and depressive behaviors, along with a reduction in the generation of new neurons in the dentate gyrus of the hippocampus. Data concerning sucrose consumption during late adolescence are lacking, and the effect of sucrose intake on the ventral dentate gyrus of the hippocampus (which modulates anxiety and depression) remains elusive. Here, we tested whether sucrose intake during late adolescence causes anxiety or impaired neurogenesis in the ventral dentate gyrus. Rats did not display anxiety-like behaviors neither at the light-dark box test nor at the open field exploration. However, there was a significant increase in proliferative cells in the subgranular zone of the ventral dentate gyrus in rats exposed to sucrose (p &lt; 0.05). This increased proliferation corresponded to neural stem cells (Radial Type 1 cells) in the group exposed to sucrose until adulthood but was not present in rats exposed to sucrose only during late adolescence. Remarkably, the phosphorylation of ERK1/2 kinases was increased in the hippocampi of rats exposed to sucrose only during late adolescence, suggesting that the increased proliferation in this group could be mediated by the MAPK pathway. On the other hand, although no differences were found in the number of immature granular neurons, we observed more immature granular neurons with impaired dendritic orientation in both groups exposed to sucrose. Finally, GAD65/67 and BCL2 levels did not change between groups, suggesting an unaltered hippocampal GABAergic system and similar apoptosis, respectively. This information provides the first piece of evidence of how sucrose intake, starting in late adolescence, impacts ventral dentate gyrus neurogenesis and contributes to a better understanding of the effects of this carbohydrate on the brain at postnatal stages.

Daily Consumption of High-Polyphenol Olive Oil Enhances Hippocampal Neurogenesis in Old Female Rats

Objective The aim of this study is to evaluate the effect of daily consumption of high-polyphenol (HP) olive oil on neurogenesis by investigating neuronal cell proliferation and maturation in the hippocampus of old rats, and to evaluate the relationship between neurogenesis, spatial memory, and anxiety-like behavior. Methods A total of 34 female, 20–22-month-old Sprague Dawley rats were divided into three groups: control group, low-polyphenol (LP) group, and high-polyphenol (HP) group. The animals were fed distilled water, LP olive oil and HP-extra virgin olive oil, respectively for 6 weeks using an oral gavage. At 43 days, animals were tested using the Morris Water Maze to evaluate spatial memory, and the Open-field test to evaluate anxiety-like behavior. Neural cell proliferation in the dentate gyrus (DG) was determined by BrdU labeling and Nestin protein expression. Neuronal maturation was determined by NeuN labeling. Synaptic density in the hippocampus and prefrontal cortex was examined by measuring Synaptophysin (SYN) levels. Hippocampal Calbindin levels were measured to assess cellular calcium metabolism. Results Daily consumption of HP olive oil significantly improved cell proliferation and neuronal maturation in the DG of old rats. HP-olive oil significantly increased SYN levels in the prefrontal cortex, and nestin and calbindin levels in the hippocampus (p < 0.05). LP olive oil diet has shown no effect on any parameter (p > 0.05). We also did not find any statistically significant difference between the groups in terms of spatial memory and anxiety-like behavior (p > 0.05). Conclusion Our study is first to show that daily consumption of HP-olive oil enhances hippocampal neurogenesis in old rats, which has been confirmed by proliferation and maturation biomarkers. In addition, increased SYN and calbindin levels showed that the generated cells were also functionally developed in the HP group. We suggest that daily consumption of HP olive oil may have beneficial effects on brain aging by triggering neurogenesis.

Gadolinium Deposition in the Rat Brain Measured with Quantitative MRI versus Elemental Mass Spectrometry.

Background T1-weighted MRI and quantitative longitudinal relaxation rate (R1) mapping have been used to evaluate gadolinium retention in the brain after gadolinium-based contrast agent (GBCA) administration. Whether MRI measures accurately reflect gadolinium regional distribution and concentration in the brain remains unclear. Purpose To compare gadolinium retention in rat forebrain measured with in vivo quantitative MRI R1 and ex vivo laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) mapping after gadobenate, gadopentetate, gadodiamide, or gadobutrol administration. Materials and Methods Adult female Sprague-Dawley rats were randomly assigned to one of five groups (eight per group) and administered gadobenate, gadopentetate, gadodiamide, gadobutrol (2.4 mmol/kg per week for 5 weeks), or saline (4.8 mL/kg per week for 5 weeks). MRI R1 mapping was performed at baseline and 1 week after the final injection to determine R1 and ΔR1. Postmortem brains from the same rats were analyzed with LA-ICP-MS elemental mapping to determine regional gadolinium concentrations. Student t tests were performed to compare results between GBCA and saline groups. Results Rats that were administered gadobenate showed gadolinium-related MRI ΔR1 in 39.5% of brain volume (ΔR1 = 0.087 second-1 ± 0.051); gadopentetate, 20.6% (ΔR1 = 0.069 second-1 ± 0.018); gadodiamide, 5.4% (ΔR1 = 0.055 second-1 ± 0.019); and gadobutrol, 2.2% (ΔR1 = 0.052 second-1 ± 0.041). Agent-specific gadolinium-related ΔR1 was detected in multiple forebrain regions (neocortex, hippocampus, dentate gyrus, thalamus, and caudate-putamen) in rats treated with gadobenate or gadopentetate, whereas rats treated with gadodiamide showed gadolinium-related ΔR1 in caudate-putamen. By contrast, LA-ICP-MS elemental mapping showed a similar regional distribution pattern of heterogeneous retained gadolinium in the forebrain of rats treated with gadobenate, gadopentetate, or gadodiamide, with the average gadolinium concentration of 0.45 μg · g-1 ± 0.07, 0.50 μg · g-1 ± 0.10, and 0.60 μg · g-1 ± 0.11, respectively. Low levels (0.01 μg · g-1 ± 0.00) of retained gadolinium were detected in the forebrain of gadobutrol-treated rats. Conclusion Differences in in vivo MRI longitudinal relaxation rate versus ex vivo elemental mass spectrometry measures of retained gadolinium in rat forebrains suggest that some forms of retained gadolinium may escape detection with MRI. © RSNA, 2022 Online supplemental material is available for this article.

Oral Exposure to Lead Acetate for 28 Days Reduces the Number of Neural Progenitor Cells but Increases the Number and Synaptic Plasticity of Newborn Granule Cells in Adult Hippocampal Neurogenesis of Young-Adult Rats.

Lead (Pb) causes developmental neurotoxicity. Developmental exposure to Pb acetate (PbAc) induces aberrant hippocampal neurogenesis by increasing or decreasing neural progenitor cell (NPC) subpopulations in the dentate gyrus (DG) of rats. To investigate whether hippocampal neurogenesis is similarly affected by PbAc exposure in a general toxicity study, 5-week-old Sprague-Dawley rats were orally administered PbAc at 0, 4000, and 8000ppm (w/v) in drinking water for 28days. After exposure to 4000 or 8000ppm PbAc, Pb had accumulated in the brains. Neurogenesis was suppressed by 8000ppm PbAc, which was related to decreased number of type-2b NPCs, although number of mature granule cells were increased by both PbAc doses. Gene expression in the 8000ppm PbAc group suggested suppressed NPC proliferation and increased apoptosis resulting in suppressed neurogenesis. PbAc exposure increased numbers of metallothionein-I/II+ cells and GFAP+ astrocytes in the DG hilus, and upregulated Mt1, antioxidant genes (Hmox1 and Gsta5), and Il6 in the DG, suggesting the induction of oxidative stress and neuroinflammation related to Pb accumulation resulting in suppressed neurogenesis. PbAc at 8000ppm also upregulated Ntrk2 and increased the number of CALB2+ interneurons, suggesting the activation of BDNF-TrkB signaling and CALB2+ interneuron-mediated signals to ameliorate suppressed neurogenesis resulting in increased number of newborn granule cells. PbAc at both doses increased the number of ARC+ granule cells, suggesting the facilitation of synaptic plasticity of newborn granule cells through the activation of BDNF-TrkB signaling. These results suggest that PbAc exposure during the young-adult stage disrupted hippocampal neurogenesis, which had a different pattern from developmental exposure to PbAc. However, the induction of oxidative stress/neuroinflammation and activation of identical cellular signals occurred irrespective of the life stage at PbAc exposure.

Effect of gabapentin on neurogenesis in hippocampal dentate gyrus of adult rats with co-disease of chronic pain and depression.

Chronic pain lasts for more than 3 months and is often associated with negative emotions such as depression and anxiety. Long-term chronic pain stress can lead to plastic changes in hippocampal structure and function. In addition to its analgesic effect, gabapentin also has certain cerebral protective effects. This study aims to observe the effect of gabapentin on neurogenesis in hippocampal dentate gyrus (DG) of the adult rats with co-disease of chronic pain and depression. The adult rats were randomly divided into 4 groups: A sham operation (Sham) group, a comorbidity model+normal saline (CCI+Veh) group (1 mL saline), a comorbidity model+low-dose gabapentin (CCI+LG) group (diluting gabapentin with normal saline to 1 mL at the dose of 30 mg/kg), and a comorbidity model+high-dose gabapentin (CCI+HG) group (diluting gabapentin with normal saline to 1 mL at the dose of 100 mg/kg) (8 rats per group). The comorbidity model was established by sciatic nerve encirclement. On the 30th day after operation, normal saline, low-dose gabapentin, and high-dose gabapentin were given intraperitoneally, respetively, for 7 consecutive days. The paw withdrawal mechanical threshold (PWMT) of the right hindlimb was measured before the operation and on the 7th, 14th, 21th, 28th, and 40th day after the operation. The time of immobility and sugar water preference rate were measured by forced swimming test and sugar water preference test, respectively, on the 28th and 40th day after the operation. The number of doublecortin (DCX) positive neurons and the expression of brain-derived neurotrophic factor (BDNF) in hippocampal dentate gyrus were observed by immunohistochemical staining, and the morphological changes of the hippocampal neurons were observed by Golgi staining. Compared with the Sham group, the PWMT of the CCI comorbidity model rats reached the lowest level on the 7th day after the operation and lasted until the 28th day after the operation, and remained at a low level on the 40th day after the operation (all P<0.05). Compared with the CCI+Veh group, the PWMT in the CCI+LG group and the CCI+HG group was increased on the 40th day after the operation (all P<0.05). Compared with the Sham group, the time of immobility in the CCI comorbidity model rats was increased significantly (all P<0.01) and the sugar water preference rate was decreased significantly (all P<0.01) on the 28th day after the operation. Compared with the CCI+Veh group, the time of immobility in the CCI+HG group was shortened (P<0.05) and the sugar water preference rate was significantly increased (P<0.01) on the 40th day after the operation. Compared with the CCI+Veh group, the number of DCX positive cells in hippocampal DG of the CCI+LG group and the CCI+HG group was increased, and that in the CCI+HG group was increased more significantly (P<0.05). Compared with the Sham group, the expression of BDNF in hippocampal DG was decreased in the CCI+Veh group (P<0.05). Compared with the CCI+Veh group, the expression of BDNF in hippocampal DG and the length of dendritic spines of the hippocampal neurons were increased in the CCI+HG group (all P<0.05). Gabapentin can relieve chronic pain and depression-like behavior in rats with chronic pain and depression, and promote neurogenesis of hippocampal dentate gyrus neurons.

Immunophenotypic Characteristics of the Expression of Inducible NO Synthase in the Dentate Gyrus in Mature Rats and Its Pharmacological Correction

Immunophenotypic Characteristics of the Expression of Inducible NO Synthase in the Dentate Gyrus in Mature Rats and Its Pharmacological Correction

P-Coumaric acid mitigates passive avoidance memory and hippocampal synaptic plasticity impairments in aluminum chloride-induced Alzheimer's disease rat model

The present study was designed to determine the effects of p-coumaric acid (p-CA), a phenolic compound, on passive avoidance memory function and hippocampal long-term potentiation (LTP) in an Alzheimer's disease (AD) rat model induced by aluminum chloride (AlCl3). p-CA (100 mg/kg; P.O.) was administered concomitantly with AlCl3 (100 mg/kg; P.O.) for 42 consecutive days. Results of this study revealed an attenuation of memory capacity, which was accompanied by a reduction in both components of LTP (field excitatory postsynaptic potentials slope and population spike amplitude) and an increase in amyloid-beta (Aβ) plaques production in the AlCl3-exposed rats. Intriguingly, treatment with p-CA improved passive avoidance memory dysfunction, ameliorated hippocampal LTP impairment, and hindered Aβ plaque accumulation in the hippocampal dentate gyrus of AlCl3-treated rats. This data provides evidence that p-CA improves AlCl3-induced passive avoidance memory decline, which may be partly related to amelioration of synaptic dysfunction and suppression of Aβ plaque formation.

Differential effects of levetiracetam on hippocampal CA1 synaptic plasticity and molecular changes in the dentate gyrus in epileptic rats

Temporal lobe epilepsy (TLE) is the most common form of focal epilepsies. Pharmacological treatment with anti-seizure drugs (ASDs) remains the mainstay in epilepsy management. Levetiracetam (LEV) is a second-generation ASD with a novel SV2A protein target and is indicated for treating focal epilepsies. While there is considerable literature in acute models, its effect in chronic epilepsy is less clear. Particularly, its effects on neuronal excitability, synaptic plasticity, adult hippocampal neurogenesis, and histological changes in chronic epilepsy have not been evaluated thus far, which formed the basis of the present study. Six weeks post-lithium-pilocarpine-induced status epilepticus (SE), epileptic rats were injected with levetiracetam (54 mg/kg b.w. i.p.) once daily for two weeks. Following LEV treatment, Schaffer collateral - CA1 (CA3-CA1) synaptic plasticity and structural changes in hippocampal subregions CA3 and CA1 were evaluated. The number of doublecortin (DCX+) and reelin (RLN+) positive neurons was estimated. Further, mossy fiber sprouting was evaluated in DG by Timm staining, and splash test was performed to assess the anxiety-like behavior. Chronic epilepsy resulted in decreased basal synaptic transmission and increased paired-pulse facilitation without affecting post-tetanic potentiation and long-term potentiation. Moreover, chronic epilepsy decreased hippocampal subfields volume, adult hippocampal neurogenesis, and increased reelin expression and mossy fiber sprouting with increased anxiety-like behavior. LEV treatment restored basal synaptic transmission and paired-pulse facilitation ratio in CA3-CA1 synapses. LEV also restored the CA1 subfield volume in chronic epilepsy. LEV did not affect epilepsy-induced abnormal adult hippocampal neurogenesis, ectopic migration of newborn granule cells, mossy fiber sprouting in DG, and anxiety-like behavior. Our results indicate that in addition to reducing seizures, LEV has favorable effects on synaptic transmission and structural plasticity in chronic epilepsy. These findings add new dimensions to the use of LEV in chronic epilepsy and paves way for further research into its effects on cognition and affective behavior.

Enhanced Non-Associative Long-Term Potentiation in Immature Granule Cells in the Dentate Gyrus of Adult Rats.

The dentate gyrus is one of the few sites of neurogenesis in the adult brain. Integration of new-generated granule cells into the hippocampal circuitry provides a substrate for structural plasticity, fundamental for normal function of adult hippocampus. However, mechanisms of synaptic plasticity that mediate integration of new-generated granule cells into the existing circuitry remain poorly understood. Especially mechanisms of plasticity at GABA-ergic synapses remain elusive. Here, we show that postsynaptic spiking without presynaptic activation can induce heterosynaptic, non-associative plasticity at GABA-ergic inputs to both immature and mature granule cells. In both immature and mature neurons, plastic changes were bidirectional and individual inputs could express long-term potentiation (LTP) or long-term depression (LTD), or do not change. However, properties of non-associative plasticity dramatically change with maturation of newly generated granule cells: while in immature cells there was a clear predominance of non-associative LTP and net potentiation across the inputs, in mature neurons, potentiation and depression were balanced with no net change on average. We conclude that GABA-ergic inputs to granule cells are plastic, and that the rules for induction of non-associative plasticity change with maturation. We propose that potentiation-biased non-associative plasticity of GABA-ergic transmission might help to counter-balance an increase of excitatory drive that is facilitated by enhanced LTP at glutamatergic synapses in maturating granule cells. Such mechanism might help to build a strong GABA-ergic input to surviving active new cells, necessary for normal function of mature granule cells, which operate under a tight inhibitory control and generate sparse spiking activity.

Glutamate released by Cajal-Retzius cells impacts specific hippocampal circuits and behaviors.

SUMMARYThe impact of Cajal-Retzius cells on the regulation of hippocampal circuits and related behaviors is unresolved. Here, we directly address this issue by impairing the glutamatergic output of Cajal-Retzius cells with the conditional ablation of vGluT2, which is their main vesicular glutamate transporter. Although two distinct conditional knockout lines do not reveal major alterations in hippocampal-layer organization and dendritic length of principal neurons or GABAergic cells, we find parallel deficits in specific hippocampal-dependent behaviors and in their putative underlying microcircuits. First, conditional knockout animals show increased innate anxiety and decreased feedforward GABAergic inhibition on dentate gyrus granule cells. Second, we observe impaired spatial memory processing, which is associated with decreased spine density and reduced AMPA/NMDA ratio of postsynaptic responses at the perforant- and entorhino-hippocampal pathways. We conclude that glutamate synaptically released by Cajal-Retzius cells is critical for the regulation of hippocampal microcircuits and specific types of behaviors.

Sex-specific effects of adolescent intermittent ethanol exposure-induced dysregulation of hippocampal glial cells in adulthood.

Adolescent alcohol abuse is a significant public health concern, with approximately 4.3 million U.S. adolescents reporting monthly binge drinking. Excessive ethanol consumption during adolescence has been linked to dysregulation of the neuroimmune system, particularly in the hippocampus. Because there are sex differences in both neuroimmune responses and ethanol's pharmacologic actions, this study tested whether there were disparate effects based on sex in glial cells and neurodegeneration in adulthood after the adolescent intermittent ethanol (AIE) model. Male and female adolescent Sprague-Dawley rats underwent AIE. In adulthood, immunohistochemical techniques were utilized to determine the effects of AIE on astrocytes and microglia, and Fluoro-Jade C (FJC) was used to assess neurodegeneration in the hippocampus. AIE exposure significantly increased astrocyte activation in the cornu ammonis 1 (CA1), CA2/3, and dentate gyrus (DG) in both male and female rats with no discernible sex differences in immunoreactivity. Likewise, the number of GFAP+cells was significantly increased by AIE across the hippocampus. In our microglial assessment, AIE only led to increased Iba1 immunoreactivity in the CA1 but not CA2/3 or DG regions. However, the number of Iba1+ cells was increased by AIE in both the CA1 and DG subregions. In the DG, the ethanol effect was observed in both sexes, but in the CA1, AIE-induced increased Iba1 cells were only observed in females. In regard to neurodegeneration, there were no persisting AIE effects on FJC+cells. These findings indicate that AIE alters hippocampal glial cells in adulthood, in the absence of active neurodegeneration. However, while AIE induced long-term elevation of astroglial measures in both males and females, persisting AIE-induced microglial activation was more sparse and sex-dependent. While the majority of these findings suggest that AIE has similar effects on glial morphology and number between males and females, additional work should determine whether there are molecular differences as well as innate sex differences in glial interaction with AIE's influence on glial functions in behavior.

Luteolin Enhances Choroid Plexus 5-MTHF Brain Transport to Promote Hippocampal Neurogenesis in LOD Rats.

Folates, provided by food, are commonly used antidepressant synergists in late-onset depression (LOD). However, increased intake of folic acid in the elderly population might lead to the accumulation of unmetabolized folic acid in the systemic circulation, leading to enhanced deterioration of the central nervous system function. In addition, folates cannot access the brain directly because of the blood–brain barrier. Choroid plexus (CP) 5-methyltetrahydrofolate (5-MTHF) brain transport plays a critical role in regulating the cerebrospinal fluid (CSF) 5-MTHF content. Luteolin is a natural flavonoid that has antidepressant effects and is involved in the anti-folate resistance pathway. It remains unclear whether the antidepressant effects of luteolin are associated with the CP 5-MTHF brain transport. In this study, 20–21-month-old Wistar rats were exposed to the chronic unpredictable mild stress (CUMS) protocol for 6 consecutive weeks to explore the long-term effects of luteolin on behavior, 5-MTHF levels, hippocampal neurogenesis, and folate brain transport of the CP. In vitro primary hippocampal neural stem cells (NSCs) cultured in media containing 10% CSF from each group of rats and choroid plexus epithelial cells (CPECs) cultured in media containing 20 μM luteolin were treated with 100 μM corticosterone and 40 mg/ml D-galactose. We found that aged rats exposed to CUMS showed a significantly reduced sucrose preference, decreased locomotion activity in the open field test and accuracy of the Morris water maze test, increased immobility time in the forced swimming test, accelerated dysfunctional neurogenesis and neuronal loss in the dentate gyrus of LOD rats, as well as decreased CSF and hippocampus 5-MTHF levels, and zona occludens protein 1 (ZO-1), proton-coupled folate transporter (PCFT), and reduced folate carrier (RFC) protein levels. In vitro assays showed media containing 10% aged CSF or LOD+ Luteolin-CSF significantly increased the viability of CORT + D-gal-injured NSCs and alleviated dysfunctional neurogenesis and neuronal loss compared with the CORT + D-gal medium. However, media containing 10% LOD-CSF had no such effect. In the meantime, induction of CORT + D-gal significantly decreased the ZO-1, PCFT, RFC, and folate receptor alpha (FR-α) protein levels and transepithelial electrical resistance in rat CPECs. As expected, luteolin treatment was effective in improving these abnormal changes. These findings suggested that luteolin could ameliorate CUMS-induced LOD-like behaviors by enhancing the folate brain transport.

Development and Validation of Arc Nanobodies: New Tools for Probing Arc Dynamics and Function

Activity-regulated cytoskeleton-associated (Arc) protein plays key roles in long-term synaptic plasticity, memory, and cognitive flexibility. However, an integral understanding of Arc mechanisms is lacking. Arc is proposed to function as an interaction hub in neuronal dendrites and the nucleus, yet Arc can also form retrovirus-like capsids with proposed roles in intercellular communication. Here, we sought to develop anti-Arc nanobodies (ArcNbs) as new tools for probing Arc dynamics and function. Six ArcNbs representing different clonal lines were selected from immunized alpaca. Immunoblotting with recombinant ArcNbs fused to a small ALFA-epitope tag demonstrated binding to recombinant Arc as well as endogenous Arc from rat cortical tissue. ALFA-tagged ArcNb also provided efficient immunoprecipitation of stimulus-induced Arc after carbachol-treatment of SH-SY5Y neuroblastoma cells and induction of long-term potentiation in the rat dentate gyrus in vivo. Epitope mapping showed that all Nbs recognize the Arc C-terminal region containing the retroviral Gag capsid homology domain, comprised of tandem N- and C-lobes. ArcNbs E5 and H11 selectively bound the N-lobe, which harbors a peptide ligand binding pocket specific to mammals. Four additional ArcNbs bound the region containing the C-lobe and C-terminal tail. For use as genetically encoded fluorescent intrabodies, we show that ArcNbs fused to mScarlet-I are uniformly expressed, without aggregation, in the cytoplasm and nucleus of HEK293FT cells. Finally, mScarlet-I-ArcNb H11 expressed as intrabody selectively bound the N-lobe and enabled co-immunoprecipitation of full-length intracellular Arc. ArcNbs are versatile tools for live-cell labeling and purification of Arc, and interrogation of Arc capsid domain specific functions.