Propofol exposure during early gestation impairs learning and memory in rat offspring by inhibiting the acetylation of histone

Propofol is widely used in clinical practice, including non‐obstetric surgery in pregnant women. Previously, we found that propofol anaesthesia in maternal rats during the third trimester (E18) caused learning and memory impairment to the offspring rats, but how about the exposure during early pregnancy and the underlying mechanisms? Histone acetylation plays an important role in synaptic plasticity. In this study, propofol was administered to the pregnant rats in the early pregnancy (E7). The learning and memory function of the offspring were tested by Morris water maze (MWM) test on post‐natal day 30. Two hours before each MWM trial, histone deacetylase 2 (HDAC2) inhibitor, suberoylanilide hydroxamic acid (SAHA), Senegenin (SEN, traditional Chinese medicine), hippyragranin (HGN) antisense oligonucleotide (HGNA) or vehicle were given to the offspring. The protein levels of HDAC2, acetylated histone 3 (H3) and 4 (H4), cyclic adenosine monophosphate (cAMP) response element‐binding protein (CREB), N‐methyl‐D‐aspartate receptor (NMDAR) 2 subunit B (NR2B), HGN and synaptophysin in offspring's hippocampus were determined by Western blot or immunofluorescence test. It was discovered that infusion with propofol in maternal rats on E7 leads to impairment of learning and memory in offspring, increased the protein levels of HDAC2 and HGN, decreased the levels of acetylated H3 and H4 and phosphorylated CREB, NR2B and synaptophysin. HDAC2 inhibitor SAHA, Senegenin or HGN antisense oligonucleotide reversed all the changes. Thus, present results indicate exposure to propofol during the early gestation impairs offspring's learning and memory via inhibiting histone acetylation. SAHA, Senegenin and HGN antisense oligonucleotide might have therapeutic value for the adverse effect of propofol.     

7,8-dihydroxyflavone rescues spatial memory and synaptic plasticity in cognitively impaired aged rats

7,8-dihydroxyflavone (7,8-DHF) has recently been identified as a potential TrkB agonist that crosses the blood-brain barrier after i.p. administration. We previously demonstrated that 7,8-DHF in vitro rescues long-term synaptic plasticity in the hippocampus of aged rats. This study assessed the rescue effect of 7,8-DHF in vivo on aging-related cognitive impairment in rats, and further determined whether the effect of 7,8-DHF is age dependent. Aged rats at 22 and 30 months of age were pretested for spatial memory in Morris water maze. The aged-impaired rats were retested twice during 7,8-DHF or vehicle treatment, which started 3 weeks after the completion of the pretest. In the 22-month-old rats, daily i.p. administration of 7,8-DHF for 2 weeks improved spatial memory. The improvement in behavioral tests was associated with increases in synapse formation and facilitation of synaptic plasticity in the hippocampus, as well as the activation of several proteins crucial to synaptic plasticity and memory. A more extended treatment paradigm with 7,8-DHF was required to achieve a significant memory improvement in the severely impaired 30-month-old rats. Moreover, 7,8-DHF moderately facilitated the synaptic plasticity, modified the density but not number of spines in the hippocampus of the oldest rats. Taken together, our results suggest that 7,8-DHF can act in vivo to counteract aging-induced declines in spatial memory and synaptic plasticity and morphological changes of hippocampal neurons. The effect of 7,8-DHF is more pronounced in relatively younger impaired rats than in those of more advanced age. These findings demonstrate the reversal of age-dependent memory impairment by in vivo 7,8-DHF application and support the benefit of early treatment for cognitive aging.     

Structure of the neuronal protein calexcitin suggests a mode of interaction in signalling pathways of learning and memory

The three-dimensional structure of the neuronal calcium-sensor protein calexcitin from Loligo pealei has been determined by X-ray analysis at a resolution of 1.8A. Calexcitin is up-regulated following Pavlovian conditioning and has been shown to regulate potassium channels and the ryanodine receptor. Thus, calexcitin is implicated in neuronal excitation and plasticity. The overall structure is predominantly helical and compact with a pronounced hydrophobic core between the N and C-terminal domains of the molecule. The structure consists of four EF-hand motifs although only the first three EF hands are involved in binding calcium ions; the C-terminal EF-hand lacks the amino acids required for calcium binding. The overall structure is quite similar to that of the sarcoplasmic calcium-binding protein from Amphioxus although the sequence identity is very low at 31%. The structure shows that the two amino acids of calexcitin phosphorylated by protein kinase C are close to the domain interface in three dimensions and thus phosphorylation is likely to regulate the opening of the domains that is probably required for binding to target proteins. There is evidence that calexcitin is a GTPase and the residues, which have been implicated by mutagenesis in its GTPase activity, are in a short but highly conserved region of 3(10) helix close to the C terminus. This helix resides in a large loop that is partly sandwiched between the N and C-terminal domains suggesting that GTP binding may also require or may cause domain opening. The structure possesses a pronounced electropositive crevice in the vicinity of the 3(10) helix, that might provide an initial docking site for the triphosphate group of GTP. These findings elucidate a number of the reported functions of calexcitin with implications for neuronal signalling.     

孕期饮用酒精对子代大鼠学习记忆及海马cdk5表达的影响

目的:研究孕期饮用酒精对子代大鼠学习记忆及海马细胞周期依赖性蛋白激酶5(cdk5)表达的影响。方法:建立大鼠孕期饮用酒精模型,子代成年后,Y-型迷宫测试学习记忆成绩;聚合酶链式反应(RT-PCR)分析海马组织cdk5mRNA的表达;免疫荧光法检测子鼠海马区cdk5蛋白表达。结果:学习记忆测试结果显示孕期饮用酒精组子鼠学习记忆成绩比正常对照组和饮酒对照组明显下降;RT-PCR结果表明孕期饮用酒精组子鼠海马组织cdk5mRNA表达较正常对照组和饮酒对照组明显上升;免疫荧光结果显示孕期饮用酒精组子鼠海马区cdk5蛋白表达明显增加。结论:孕期饮用酒精对子代大鼠的神经损伤可能与cdk5蛋白表达的上调有关。     

Overexpression of the NR2A subunit in the forebrain impairs long‐term social recognition and non‐social olfactory memory

Animals must recognize and remember conspecifics and potential mates, and distinguish these animals from potential heterospecific competitors and predators. Despite its necessity, aged animals are known to exhibit impaired social recognition memory. As the brain ages, the ratio of NR2A:NR2B in the brain increases over time and has been postulated to underlie the cognitive decline observed during the aging process. Here, we test the hypothesis that an increased NR2A:NR2B subunit ratio underlies long‐term social recognition memory. Using transgenic overexpression of NR2A in the forebrain regions, we investigated the ability of these mice to learn and remember male and female conspecifics, mice of another strain and animals of another rodent species, the rat. Furthermore, due to the importance of olfaction in social recognition, we tested the olfactory memory in the NR2A transgenic mice. Our series of behavioral experiments revealed significant impairments in the NR2A transgenic mice in long‐term social memory of both male and female conspecifics. Additionally, the NR2A transgenic mice are unable to recognize mice of another strain or rats. The NR2A transgenic mice also exhibited long‐term memory impairments in the olfactory recognition task. Taken together, our results provide evidence that an increased NR2A:NR2B ratio in the forebrain leads to reduced long‐term memory function, including the ethologically important memories such as social recognition and olfactory memory .     

O‐GlcNAcylation of protein kinase A catalytic subunits enhances its activity: a mechanism linked to learning and memory deficits in Alzheimer's disease

Alzheimer's disease (AD) is characterized clinically by memory loss and cognitive decline. Protein kinase A (PKA)‐CREB signaling plays a critical role in learning and memory. It is known that glucose uptake and O‐GlcNAcylation are reduced in AD brain. In this study, we found that PKA catalytic subunits (PKAcs) were posttranslationally modified by O‐linked N‐acetylglucosamine (O‐GlcNAc). O‐GlcNAcylation regulated the subcellular location of PKAcα and PKAcβ and enhanced their kinase activity. Upregulation of O‐GlcNAcylation in metabolically active rat brain slices by O‐(2‐acetamido‐2‐deoxy‐d ‐glucopyranosylidenamino) N‐phenylcarbamate (PUGNAc), an inhibitor of N‐acetylglucosaminidase, increased the phosphorylation of tau at the PKA site, Ser214, but not at the non‐PKA site, Thr205. In contrast, in rat and mouse brains, downregulation of O‐GlcNAcylation caused decreases in the phosphorylation of CREB at Ser133 and of tau at Ser214, but not at Thr205. Reduction in O‐GlcNAcylation through intracerebroventricular injection of 6‐diazo‐5‐oxo‐l ‐norleucine (DON), the inhibitor of glutamine fructose‐6‐phosphate amidotransferase, suppressed PKA‐CREB signaling and impaired learning and memory in mice. These results indicate that in addition to cAMP and phosphorylation, O‐GlcNAcylation is a novel mechanism that regulates PKA‐CREB signaling. Downregulation of O‐GlcNAcylation suppresses PKA‐CREB signaling and consequently causes learning and memory deficits in AD.     

Expression of mRNA encoding insulin‐like growth factors I and II by uterine tissues and placenta during pregnancy in the rat

The uterus and the placenta synthesize insulin‐like growth factors (IGFs) and insulin‐like binding proteins (IGFBPs). These growth factors are implicated in processes of proliferation and differentiation that occur in the uterus. To determine the patterns of expression of IGFs during rat pregnancy we used in situ hybridization with digoxigenin labeled probes on uterus from day 7 to day 16 of pregnancy. In early gestation days (7–8) both IGF mRNAs showed similar tissue distribution with relative abundance in the stroma and circular muscle layer. On days 11 and 12 expression for IGF‐I mRNA was found in the mesometrial decidua and metrial gland and in the ectoplacental cone while clear expression of IGF‐II mRNA could only be found in the latter. On days 13 and 14, expression for IGF‐I mRNA could be detected in the mesometrial decidua and metrial gland but no expression was observed for IGF‐II mRNA. A gradient of IGF‐I mRNA expression could be observed in the placenta on day 16, with the trophoblastic cells of the basal zone expressing the signal with stronger intensity than in the labyrinthine zone. For IGF‐II mRNA the highest expression was associated with the labyrinthine zone. Endovascular trophoblast was positive for both mRNAs. The spatial and temporal patterns of expression suggests a role for IGFs in the process of decidualization as well as in the establishment, growth and differentiation of the various trophoblast cells of the placenta. Mol. Reprod. Dev. 53:294–305, 1999. © 1999 Wiley‐Liss, Inc.     

Up‐regulated CST5 inhibits bone resorption and activation of osteoclasts in rat models of osteoporosis via suppression of the NF‐κB pathway

Here, we aim at exploring the effect of CST5 on bone resorption and activation of osteoclasts in osteoporosis (OP) rats through the NF‐κB pathway. Microarray analysis was used to screen the OP‐related differentially expressed genes. Osteoporosis was induced in rats by intragastric retinoic acid administration. The serum levels of tartrate‐resistant acid phosphatase (TRAP), bone alkaline phosphatase (BALP) and osteocalcin (OC) and the expression of CD61 on the surface of osteoclasts were examined. The number of osteoclasts and the number and area of resorption pits were detected. Besides, the pathological changes and bone mineral density in bone tissues of rats were assessed. Also, the relationship between CST5 and the NF‐κB pathway was identified through determining the expression of CST5, RANKL, RANK, OPG, p65 and IKB. Poorly expressed CST5 was indicated to affect the OP. CST5 elevation and inhibition of the NF‐κB pathway decreased serum levels of TRAP, BALP and OC and expression of CD61 in vivo and in vitro. In OP rats, CST5 overexpression increased trabecular bones and bone mineral density of bone tissues, but decreased trabecular separation, fat within the bone marrow cavities and the number of osteoclasts through inhibiting the NF‐κB pathway. In vivo experiments showed that CST5 elevation inhibited growth in number and area of osteoclastic resorption pits and restrained osteoclastic bone absorption by inhibiting the NF‐κB pathway. In summary, overexpression of CST5 suppresses the activation and bone resorption of osteoclasts by inhibiting the activation of the NF‐κB pathway.     

Wnt5a regulates the cell proliferation and adipogenesis via MAPK‐independent pathway in early stage of obesity

The early stage of obesity is an important stage in the development of obesity. However, there are few studies which explored the property or changes in obesity at early stage especially involving Wnt5a. The associated gene expression of Wnt5a on cell regeneration and the effect of Wnt5a on rat adipose‐derived stem cell (rASC) proliferation and adipogenesis need additional study. Here, we investigated the changes in obesity at early stage and how Wnt5a regulates rASC regeneration, proliferation, and adipogenesis. Our data revealed that obesity at early stage measured by Lee index presented a state with impaired adipogenesis and more infiltrated inflammatory cells but without significant changes in adipocyte sizes and inflammatory factors. The process might be associated with anti‐canonical Wnt pathway and a reciprocal Wnt5a/JNK pathway. Besides the gene expression of Wnt5a decreased from cell passage 1 to passage 3. The cell proliferation was regulated by increasing dose of Wnt5a with the maximal effect at 50 ng/mL and 50 ng/mL Wnt5a suppressed adipogenic differentiation at middle‐late stage of adipogenesis via anti‐β‐catenin and a mitogen‐activated protein kinase (MAPK) signaling‐independent manner. Accordingly, the research helps to gain further insights into the early stage of obesity and its associated changes on a cellular and molecular level.     

Anti‐invasion effect of rosmarinic acid via the extracellular signal‐regulated kinase and oxidation–reduction pathway in Ls174‐T cells

Rosmarinic acid is a major phenylpropanoid isolated from Prunella vulgaris L., which is a composition of herbal tea for centuries in China. However, the anti‐invasion activity on Ls174‐T human colon carcinoma cells has not been studied. In this study, we investigated the anti‐metastasis functions according to wound healing assay, adhesion assay, and Transwell assay and found that rosmarinic acid could inhibit migration, adhesion, and invasion dose‐dependently. Rosmarinic acid also could decrease the level of reactive oxygen species by enhancing the level of reduced glutathione hormone. In addition, rosmarinic acid repressed the activity and expression of matrix metalloproteinase‐2,9. According to Western blot and quantitative real‐time PCR assay, rosmarinic acid may inhibit metastasis from colorectal carcinoma mainly via the pathway of extracellular signal‐regulated kinase. In animal experiment, intraperitoneal administration of 2 mg of rosmarinic acid reduced weight of tumors and the number of lung nodules significantly compared with those of control group. Therefore, these results demonstrated that rosmarinic acid can effectively inhibit tumor metastasis in vitro and in vivo. J. Cell. Biochem. 111: 370–379, 2010. © 2010 Wiley‐Liss, Inc.     

Expression profiles of interferon‐stimulated gene 15 and prostaglandin synthases in the ovine lymph nodes during early pregnancy

Lymph nodes are distributed all over the body and are part of the lymphatic system. The interferon‐stimulated gene 15 kDa protein (ISG15) and prostaglandins (PGs) are involved in the establishment of pregnancy and are expressed in the uterus during early pregnancy in sheep. In this study, the ovine lymph nodes were obtained on Day 16 of the estrous cycle, and Days 13, 16, and 25 of pregnancy, and the expression of ISG15 and PG synthases, including cyclooxygenase 1 (COX‐1), COX‐2, prostaglandin E (PGE) synthase (PTGES), and a PGF synthase (aldo‐keto reductase family 1, member B1, AKR1B1) were detected by quantitative real‐time polymerase chain reaction, western blot analysis, and immunohistochemistry analysis. Our results showed that there were peaks in the expression of mRNAs and the proteins of ISG15, COX‐1, COX‐2, PTGES, and AKR1B1 in the lymph nodes during early pregnancy and that the COX‐2 and AKR1B1 proteins were limited to the subcapsular sinus and lymph sinuses. In conclusion, the ISG15, COX‐1, COX‐2, PTGES, and AKR1B1 were upregulated in the maternal lymph nodes, which may be beneficial for the development of conceptus, maternal systemic immunoregulation, and anti‐luteolysis during early pregnancy in sheep.