The protective effect of trihexyphenidyl on the beta-amyloid peptide 25–35-induced cytotoxicity in PC12 cells

In the development and progression of Alzheimer’s disease (AD), β-amyloid peptide (Aβ) that induced cytotoxicity containing apoptosis and excess production of reactive oxygen species (ROS) is considered as a causal role. The aim of present study is to investigate the protective effect of Trihexyphenidyl (THY) on Aβ_25–35-induced cytotoxicity in cultured rat pheochromocytoma (PC12) cells. In this report, the cell survival was measured by MTT assay, the enzyme activity of superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX), the contents of lipid peroxidation products malondialdehyde (MDA) and ROS in the cells were determined. Acridine orange (AO) was used to observe the morphological characteristic of apoptotic cells. Mitochondrial membrane potential in PC12 cells were monitored by fluorospectrophotometer combining with Rh123. As a cell permeable fluorescent probe, Fura-2/AM was employed to detect intracellular [Ca²⁺]. The results showed that after incubation with Aβ_25–35 (10 μM) for 24 h, there were decreased changes in cell viability, SOD, and GSH-PX activity as well as mitochondrial membrane potential, in contrast, the levels of [Ca²⁺]_i, ROS, and MDA were increased, THY significantly attenuated all the changes induced by Aβ_25–35, indicating that THY exhibited protective effect against Aβ_25–35-induced cytotoxicity, which may represent the cellular mechanisms of the action.     

Effects of β-amyloid (25–35) on learning in the common snail

The effects of neurotoxic β-amyloid fragment (25–35) on the formation of behavioral sensitization and a conditioned defensive reflex to food were studied. Administration of β-amyloid (25–35) to common snails before the start of training led to a significant reduction in sensitization of the defensive reaction, weakening of the formation of the conditioned defensive reflex to food, and impairment of memory. These impairments to behavioral plasticity may be mediated by changes in synaptic plasticity previously observed in the presence of β-amyloid. __________ Translated from Zhurnal Vysshei Nervnoi Deyatel’nosti imeni I. P. Pavlova, Vol. 57, No. 2, pp. 229–236, March–April, 2007.     

Neuroprotective effects of cyanidin 3-O-glucopyranoside on amyloid beta (25–35) oligomer-induced toxicity

Recent studies suggest that the oligomers of short amyloid beta (Aβ) peptides such as Aβ_25–35 as well as full-length Aβ peptides (i.e. Aβ_1–40 and Aβ_1–42 peptides) are responsible for synaptic dysfunction and/or neuronal loss in Alzheimer's disease (AD). Among antioxidant phytochemicals derived from fruit and vegetables, cyanidin 3-O-glucoside (Cy-3G) has recently gained attention for its neuroprotective properties. In this in vitro study, we demonstrated that Cy-3G can inhibit Aβ_25–35 spontaneous aggregation into oligomers and their neurotoxicity in human neuronal SH-SY5Y cells. In particular, the pre- and co-treatment of SH-SY5Y cells with Cy-3G reduced the neuronal death, in terms of apoptosis and necrosis, elicited by Aβ_25–35 oligomers. Cy-3G also shows the interesting ability to prevent the early events leading to neuronal death such as the Aβ_25–35 oligomer binding to plasma membrane and the subsequent membrane integrity loss. Taken together, these findings suggest that Cy-3G may be considered a phytochemical with neuroprotective properties useful in finding potential drug or food supplements for the therapy of AD.     

Effects of corticoliberin fragment CRF4–6 on sexual behavior in male mice

Centrally administered doses of the tripeptide corticoliberin fragment CRF_4–6 (Pro-Pro-Ile) suppressed mating behavior in male rats. Doses of 1 and 2 μg of the tripeptide produced dose-dependent increases in the latent periods of mounting, intromission, and ejaculation. Changes in measures of sexual behavior demonstrated that the corticoliberin fragment CRF_4–6 suppressed both sexual motivation and sexual performance. __________ Translated from Rossiiskii Fiziologicheskii Zhurnal imeni I. M. Sechenova, Vol. 91, No. 7, pp. 785–790, July, 2005. An erratum to this article is available at.     

The stimulatory effects of caffeine with oseltamivir (Tamiflu) on light–dark behavior and open-field behavior in mice

Abnormal behaviors and death associated with the use of oseltamivir (Tamiflu^®) have emerged as a major issue in influenza patients taking the drug. Here, we investigated the mechanisms underlying the effects of oseltamivir on the behavior of mice using light–dark and open-field preference tests. Oseltamivir (75 and 150 mg/kg, intraperitoneally (i.p.)) alone affected neither time spent in the open area in the light–dark preference test nor ambulation in the open-field test at 2 h post-injection. However, a non-selective adenosine A₁/A₂ receptor antagonist, caffeine (10 mg/kg, i.p.) in combination with oseltamivir (150 mg/kg, i.p.) increased time spent in the open area in the light–dark preference test. This enhancement was not inhibited by a benzodiazepine receptor antagonist, flumazenil (10–20 mg/kg, subcutaneously (s.c.)). Enhancement of ambulation in the open-field test was also observed when caffeine (10 mg/kg, i.p.) was combined with oseltamivir (150 mg/kg, i.p.). This enhancement was inhibited by a dopamine D₂ receptor antagonist, haloperidol (0.1 mg/kg, s.c.). Furthermore, an adenosine A₂ receptor antagonist, SCH58261 (3 mg/kg, i.p.) in combination with oseltamivir (150 mg/kg, i.p.) increased ambulation in the open-field test, while an adenosine A₁ receptor antagonist, DPCPX (1–3 mg/kg, i.p.) did not. These findings suggest that the actions of oseltamivir may involve the dopamine and adenosine systems. Our findings suggest that due to the interaction between central blockade of adenosine A₂ receptors by caffeine, and oseltamivir-induced behavioral changes, patients being treated with oseltamivir should be closely monitored.     

Impairments of hippocampal synaptic plasticity induced by aggregated beta-amyloid (25–35) are dependent on stimulation-protocol and genetic background

The aggregation of beta-amyloid to plaques in the brain is one of the hallmarks of Alzheimer disease (AD). Numerous studies have tried to elucidate to what degree amyloid peptides play a role in the neurodegenerative developments seen in AD. While most studies report an effect of amyloid on neural activity and cognitive abilities of rodents, there have been many inconsistencies in the results. This study investigated to what degree the different genetic backgrounds affect the outcome of beta-amyloid fragment (25–35) on synaptic plasticity in vivo in the rat hippocampus. Two strains, Wistar and Lister hooded rats, were tested. In addition, the effects of a strong (600 stimuli) and a weak stimulation protocol (100 stimuli) on impairments of LTP were analysed. Furthermore, since the state of amyloid aggregation appears to play a role in the induction of toxic processes, it was tested by dual polarisation interferometry to what degree and at what speed beta-amyloid (25–35) can aggregate in vitro. It was found that 100 nmol beta-amyloid (25–35) injected icv did impair LTP in Wistar rats when using the weak but not the strong stimulation protocol (P < 0.001). One-hundred nano mole of the reverse sequence amyloid (35–25) had no effect. LTP in Lister Hooded rats was not impaired by amyloid at any stimulation protocol. The aggregation studies showed that amyloid (25–35) aggregated within hours, while amyloid (35–25) did not. These results show that the genetic background and the stimulation protocol are important variables that greatly influence the experimental outcome. The fact that amyloid (25–35) aggregated quickly and showed neurophysiological effects, while amyloid (35–25) did not aggregate and did not show any effects indicates that the state of aggregation plays an important role in the physiological effects.     

Studies of the Effects of Central Administration of β-Amyloid Peptide (25–35): Pathomorphological Changes in the Hippocampus and Impairment of Spatial Memory

The possible link between amnesia induced by central administration of β-amyloid (25–35) (Aβ(25–35)) and neurodegenerative changes in the hippocampus was studied. Male Wistar rats received single intracerebroventricular injections of Aβ(25–35) at a dose of 15 nmoles and one month later were trained in an eight-arm radial maze. Training was followed by histological assessment of the state of the hippocampus on brain sections stained with hematoxylin and eosin. Aβ(25–35) induced impairments in long-term (reference) and working memory on testing in the maze. There was a moderate reduction in the number of neurons in hippocampal field CA1; there was no change in the number of cells in field CA3. The numbers of errors made by the animals on testing in the maze were found to correlate negatively with the numbers of nerve cells in hippocampal field CA1. Thus, this is the first demonstration that impairments of learning and memory induced by single doses of Aβ(25–35) are specifically associated with neurodegenerative changes in hippocampal field CA1 in rats. __________ Translated from Zhurnal Vysshei Nervnoi Deyatel'nosti imeni I. P. Pavlova, Vol. 54, No. 5, pp. 705–711, September–October, 2004.     

Prolonged effects of DPP-4 inhibitors on steato-hepatitic changes in Sprague–Dawley rats fed a high-cholesterol diet

Objective

Sitagliptin and other dipeptidyl peptidase (DPP)‐4 inhibitors/gliptins are antidiabetic drugs known to improve lipid profile, and confer anti‐inflammatory and anti‐fibrotic effects, which are independent of their hypoglycemic effects. However, in our previous short-term (35 days) studies, we showed that sitagliptin accentuates the hepato-inflammatory effects of high dietary cholesterol (Cho) in male Sprague–Dawley rats. Since most type 2 diabetics also present with lipid abnormalities and use DPP-4 inhibitors for glucose management, the present study was conducted to assess the impact of sitagliptin during long-term (98 days) feeding of a high Cho diet. An additional component of the present investigation was the inclusion of other gliptins to determine if hepatic steatosis, necro‐inflammation, and fibrosis were specific to sitagliptin or are class effects.

Methods

Adult male Sprague–Dawley rats were fed control or high Cho (2.0%) diets, and gavaged daily (from day 30 through 98) with vehicle or DPP-4 inhibitors (sitagliptin or alogliptin or saxagliptin). On day 99 after a 4 h fast, rats were euthanized. Blood and liver samples were collected to measure lipids and cytokines, and for histopathological evaluation, determination of hepatic lesions (steatosis, necrosis, inflammation, and fibrosis) using specific staining and immunohistochemical methods.

Results

Compared to controls, the high Cho diet produced a robust increase in NASH like phenotype that included increased expression of hepatic (Tnfa, Il1b, and Mcp1) and circulatory (TNFα and IL-1β) markers of inflammation, steatosis, necrosis, fibrosis, and mononuclear cell infiltration. These mononuclear cells were identified as macrophages and T cells, and their recruitment in the liver was facilitated by marked increases in endothelium‐expressed cell adhesion molecules. Importantly, treatment with DPP‐4 inhibitors (3 tested) neither alleviated the pathologic responses induced by high Cho diet nor improved lipid profile.

Conclusions

The potential lipid lowering effects of DPP-4 inhibitors were diminished by high Cho (a significant risk factor for inducing liver damage). The robust inflammatory responses induced by high Cho feeding in long-term experiment were not exacerbated by DPP-4 inhibitors and a consistent hepatic inflammatory environment persisted, implying a prospective physiological adaptation.

     

Administration of Aggregated Beta-Amyloid Peptide (25–35) Induces Changes in Long-Term Potentiation in the Hippocampus in Vivo

Intracerebroventricular administration of aggregated -amyloid protein fragment (25–35) (7.5 nmol/ventricle) was followed one month later by significant changes in the dynamics of long-term potentiation in the hippocampus in vivo, expressed as powerful and stable increases in the amplitude of evoked potentials. This phenomenon may be associated with oxidative stress in the hippocampus, which has previously been demonstrated in this model, and, thus, with disturbances in ion homeostasis.     

Hexapeptides HLDF-6 and PEDF-6 restore memory in rats after chronic intracerebroventricular treatment with β-amyloid peptide Aβ(25–35)

Effects of homologous peptides HLDF-6 and PEDF-6 on behavior of animals with experimental Alzheimer’s disease induced by chronic intracerebroventricular administration of β-amyloid peptide Aβ(25–35) were studied in the zoosocial recognition test and Morris water maze. Peptides HLDF-6 and PEDF-6 possessed neuroprotective activity and counteracted the toxic effect of Aβ(25–35). Peptides HLDF-6 and PEDF-6 mainly improved long-term memory and working memory, respectively. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 141, No. 3, pp. 292–296, March, 2006     

Changes in Cell Proliferation in the Subventricular Zone of the Brain in Adult Rats Given β-Amyloid Peptide (25–35)

The effects of intracerebroventricular administration of fragment (25–35) of β-amyloid peptide [Aβ(25–35)] on cell proliferation in the subventricular zone of the dentate gyrus of the hippocampus in adult rats were analyzed. Animals received doses of 15 nmol of pre-aggregated Aβ(25–35) or the Aβ(35–25) control peptide, or solvent (sterile water) into the lateral ventricles. On post-injection days 1–5, rats received intraperitoneal injections of the thymidine analog 5-bromo-2’-deoxyuridine (BrdU). BrdU incorporated into DNA was detected immunohistochemically on frontal brain sections six and 12 days after peptide administration. At six days, the numbers of BrdU-containing cells in the subventricular zone showed no differences between the study groups. At 12 days, the total number of BrdU-positive cells decreased significantly in all study groups. At the same time, the number of labeled cells in rats given Aβ(25–35) was significantly greater in this brain zone than in animals given water or the control peptide. Thus, Aβ(25–35) significantly increased cell proliferation in the subventricular zone after intracerebroventricular administration.     

Protective effects of luteolin against cognitive impairment induced by infusion of Aβ peptide in rats

Luteolin can be found in many traditional Chinese medicines, it’s a falconoid compound derived from Lonicera japonica Thunb. This study aims to investigate the neuroprotective effects of luteolin against cognitive impairment induced by amyloid-β (Aβ) peptide and the underlying mechanisms in rats. The animal behavioral tests showed that luteolin could ameliorate Aβ-induced learning and memory impairment. In hippocampal tissue, the activity of choline acetyl transferase (ChAT), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) increased after treated by luteolin. Luteolin also reversed the increased activity of acetylcholine esterase (AchE). In hippocampi homogenate, the content of acetylcholine (Ach) increased, but malondialdehyde (MDA) reduced. Moreover, luteolin can increase Bcl-2/Bax ratio. This study demonstrated that luteolin could protect Alzheimer’s disease (AD) rats against Aβ-induced cognitive impairment through regulating the cholinergic system and inhibiting oxidative injuries. The results suggesting that luteolin may have potential as a therapy for AD.     

The effects of early isolation on sexual behavior and c-fos expression in naïve male Long-Evans rats

Previous findings have demonstrated that the maternal environment is important for the development of male sexual behavior. The present study examined the effects of complete early life isolation and replacement 'stroking' stimulation on male sexual behavior and neural activation as seen by Fos immunoreactivity (Fos-IR). Animals were either artificially reared (AR) with minimal (AR-MIN) or maximal (AR-MAX) body simulation, or maternally reared (MR). In adulthood, animals were either given an exposure to an estrous female (EXP) or left undisturbed (NoEXP). No significant effects of early development were found in sexual behavior; however differences in activation in response to this exposure were observed. AR-MIN animals showed lower Fos-IR in the medial preoptic area and the ventromedial hypothalamus compared to MR animals. AR-MAX animals were not significantly different from either condition. These findings demonstrate that although there are no differences in the quality of the first copulatory exposure between AR and MR animals, the brain's response to this exposure differs in sites within the brain that subserve sexual behavior.     

Alteration of the sialylation pattern and memory deficits by injection of Aβ(25–35) into the hippocampus of rats

Sialic acid in glycoconjugates participates in important cellular functions associated with normal development, growth, and communication. Therefore we evaluated the sialylation pattern and memory deficits caused by the injection of Aβ_(25–35) into the hippocampus (Hp) of rats. The eight-arm maze spatial-learning and memory test indicated that the injection of Aβ_(25–35) into subfield CA1 of the Hp impaired both learning and memory. The sialylation pattern was examined using sialic acid-specific lectins. Our results showed that Maackia amurensis agglutinin (MAA, specific for Neu5Acα2,3Gal) showed reactivity in the CA1 and dentate gyrus (DG) subfields of the Hp mainly in the group injected with vehicle, whereas Macrobrachium rosenbergii lectin (MRL, specific for Neu5,9,7Ac) and Sambucus nigra agglutinin (SNA, specific for Neu5Acα2,6Gal-GalNAc) had increased reactivity in the CA1 and DG subfields of the Hp in the Aβ_(25–35)-injected group. The staining pattern of the antibody specific for polysialic acid (a linear homopolymer of α-2,8-linked sialic acid) increased in the CA1 and DG subfields of the Hp of the Aβ_(25–35) group compared to the control group. Our results suggest that injection of Aβ_(25–35) causes impairment in spatial memory and alters the sialylation pattern in response to compensatory reorganization and-or sprouting of dendrites and axons of the surviving neurons.     

Neuroprotective effect of alpha-asarone on spatial memory and nitric oxide levels in rats injected with amyloid-β(25–35)

The chemical α-asarone is an important active substance of the Acori graminei rhizome (AGR). It has pharmacological effects that include antihyperlipidemic, antiinflammatory, and antioxidant activity. Our aim was to study the effects α-asarone on nitric oxide (NO) levels in the hippocampus and temporal cortex of the rat after injection of the fraction 25–35 from amyloid-β (Aβ_(25–35)). In addition we examined the working spatial memory in an eight-arm radial maze. Our results showed a significant increase of nitrites in the hippocampus and temporal cortex of Aβ_(25–35)-treated rats. Other evidence of neuronal damage was the expression of a glial-fibrillar-acid protein and a silver staining. There were impairments in the spatial memory evaluated in the eight-arm radial maze. We wanted to determine whether α-asarone improves the memory correlated with NO overproduction and neuronal damage caused by the injection of Aβ_(25–35) into rats. Then animals received a 16-day treatment of α-asarone before the Aβ_(25–35) injection. Our results show a significant decrease of nitrite levels in the hippocampus and temporal cortex, without astrocytosis and silver-staining cells, which correlates with memory improvement in the α-asarone-treated group. Our results suggest that α-asarone may protect neurons against Aβ_(25–35)–caused neurotoxicity by inhibiting the effects of NO overproduction in the hippocampus and temporal cortex.     

Alteration of the sialylation pattern and memory deficits by injection of Aβ(25-35) into the hippocampus of rats

Previous studies in Parkinson's disease (PD) models suggest that early events along the path to neurodegeneration involve activation of the ubiquitin-proteasome system (UPS), endoplasmic reticulum-associated degradation (ERAD), and the unfolded protein response (UPR) pathways, in both the sporadic and familial forms of the disease, and thus ER stress may be a common feature. Furthermore, impairments in protein degradation have been linked to oxidative stress as well as pathways associated with ER stress. We hypothesize that oxidative stress is a primary initiator in a multi-factorial cascade driving dopaminergic (DA) neurons towards death in the early stages of the disease. We now report results from proteomic analysis of a rotenone-induced oxidative stress model of PD in the human neuroblastoma cell line, SH-SY5Y. Cells were exposed to sub-micromolar concentrations of rotenone for 48h prior to whole cell protein extraction and shotgun proteomic analysis. Evidence for activation of the UPR comes from our observation of up-regulated binding immunoglobulin protein (BiP), heat shock proteins, and foldases. We also observed up-regulation of proteins that contribute to the degradation of misfolded or unfolded proteins controlled by the UPS and ERAD pathways. Activation of the UPR may allow neurons to maintain protein homeostasis in the cytosol and ER despite an increase in reactive oxygen species due to oxidative stress, and activation of the UPS and ERAD may further augment clean-up and quality control in the cell.     

Studies on a Novel Multi-sensitive Hydrogel: Influence of the Biomimetic Phosphorylcholine End-Groups on the PEO–PPO–PEO Tri-block Co-polymers

In the present study, a biomimetic phosphorylcholine group was employed in the end-capping modification of PEO–PPO–PEO tri-block co-polymers (Pluronic^®). The structures of the resulting materials were characterized by ¹H-NMR and GPC. The effects of the additional phosphorylcholine end-groups to the thermo-sensitive sol–gel transition behaviors of the aqueous solutions of the resulting polymers were studied by rheology test in neutral (0.1 M NaCl) aqueous solutions and in acidic solutions (pH 3). It was found that the phosphorylcholine-end-capped Pluronic hydrogels still kept their thermo-sensitive mechanical properties with a slight change on the sol–gel transition behaviors. The phosphorylcholine-modified Pluronics exhibited a response to the change of the pH value, which made this kind of material a multi-sensitive hydrogel system. Also, the resulting polymers showed improved hemocompatibilities in the blood coagulation test using full human blood.