Improvement by nefiracetam of beta-amyloid-(1-42)-induced learning and memory impairments in rats.
(1/969)
1. We have previously demonstrated that continuous i.c.v. infusion of amyloid beta-peptide (A beta), the major constituent of senile plaques in the brains of patients with Alzheimer's disease, results in learning and memory deficits in rats. 2. In the present study, we investigated the effects of nefiracetam [N-(2,6-dimethylphenyl)-2-(2-oxo-1-pyrrolidinyl) acetamide, DM-9384] on A beta-(1-42)-induced learning and memory deficits in rats. 3. In the A beta-(1-42)-infused rats, spontaneous alternation behaviour in a Y-maze task, spatial reference and working memory in a water maze task, and retention of passive avoidance learning were significantly impaired as compared with A beta-(40-1)-infused control rats. 4. Nefiracetam, at a dose range of 1-10 mg kg(-1), improved learning and memory deficits in the A beta-(1-42)-infused rats when it was administered p.o. 1 h before the behavioural tests. 5. Nefiracetam at a dose of 3 mg kg(-1) p.o. increased the activity of choline acetyltransferase in the hippocampus of A beta-(1-42)-infused rats. 6. Nefiracetam increased dopamine turnover in the cerebral cortex and striatum of A beta-(1-42)-infused rats, but failed to affect the noradrenaline, serotonin and 5-hydroxyindoleacetic acid content. 7. These results suggest that nefiracetam may be useful for the treatment of patients with Alzheimer's disease. (+info)
Establishment and characterization of human neuroblastoma cell lines.
(2/969)
Three new tissue culture cell lines, CHP-100, CHP-126, and CHP-134, have been established from explant cultures of human neuroblastoma. The cell lines have been characterized with respect to morphology, chromosomes constitution, growth, neural enzyme content, and their ability to grow in nude mice. The cells grow as dense masses comprised of fibroblast-or neuroblast-like cells with small processes. The cell lines differ in their neural enzyme acitivity. The chromosomal content of the 3 cell lines is near diploid, and all are capable of forming tumors in nude mice. The morphological findings indicate that the cells in culture resemble those found in the tumor, and the enzyme activities are consistent with those of nervous tissue. This the morphological, biochemical, and tumorigenic properties confirm that the 3 cell lines are neoplastic cells of neural origin. (+info)
Immunohistological studies of metabotropic glutamate receptor subtype 6-deficient mice show no abnormality of retinal cell organization and ganglion cell maturation.
(3/969)
Immature retinal ganglion cells (RGCs) initially show a multistratified dendritic pattern, and, during the postnatal period, these dendrites gradually monostratify into ON and OFF sublaminae. The selective agonist of group III metabotropic glutamate receptors (mGluR), L-2-amino-4-phosphonobutyrate (L-AP-4), hyperpolarizes ON bipolar cells and reduces glutamate release. On the basis of L-AP-4-evoked inhibitory effects on ON-OFF segregation of developing RGCs, it has been hypothesized that glutamate-mediated synaptic activity is crucial for formation of the ON-OFF network. Gene-targeted ablation of mGluR6 specifically expressed in ON bipolar cells blocks normal ON responses but has been predicted to enhance glutamate release from ON bipolar cells. The mGluR6 knock-out mouse therefore provides a unique opportunity to investigate whether glutamate release and ON responses are important factors in the development of ON-OFF segregation. The combination of several different morphological analyses indicates that ON bipolar cells, as well as several distinct amacrine cells, in mGluR6 knock-out mice are normally distributed and correctly extend their terminals to defined retinal laminae. Importantly, both alpha and delta RGCs in adult mGluR6 knock-out mice are found monostratified into cell type-specific layers. Furthermore, no difference between wild-type and mGluR6 knock-out mice is observed in the maturation and dendritic stratification of developing RGCs. Hence, despite a deficit in normal ON responses, mGluR6 deficiency causes no abnormality in the retinal cellular organization nor in the stratifications of both ON bipolar cells and developing and mature RGCs. Based on these findings, we discuss several possible mechanisms that may underlie ON-OFF segregation of RGCs. (+info)
A nerve growth factor mimetic TrkA antagonist causes withdrawal of cortical cholinergic boutons in the adult rat.
(4/969)
Cholinergic neurons respond to the administration of nerve growth factor (NGF) in vivo with a prominent and selective increase of choline acetyl transferase activity. This suggests the possible involvement of endogenous NGF, acting through its receptor TrkA, in the maintenance of central nervous system cholinergic synapses in the adult rat brain. To test this hypothesis, a small peptide, C(92-96), that blocks NGF-TrkA interactions was delivered stereotactically into the rat cortex over a 2-week period, and its effect and potency were compared with those of an anti-NGF monoclonal antibody (mAb NGF30). Two presynaptic antigenic sites were studied by immunoreactivity, and the number of presynaptic sites was counted by using an image analysis system. Synaptophysin was used as a marker for overall cortical synapses, and the vesicular acetylcholine transporter was used as a marker for cortical cholinergic presynaptic sites. No significant variations in the number of synaptophysin-immunoreactive sites were observed. However, both mAb NGF30 and the TrkA antagonist C(92-96) provoked a significant decrease in the number and size of vesicular acetylcholine transporter-IR sites, with the losses being more marked in the C(92-96) treated rats. These observations support the notion that endogenously produced NGF acting through TrkA receptors is involved in the maintenance of the cholinergic phenotype in the normal, adult rat brain and supports the idea that NGF normally plays a role in the continual remodeling of neural circuits during adulthood. The development of neurotrophin mimetics with antagonistic and eventually agonist action may contribute to therapeutic strategies for central nervous system degeneration and trauma. (+info)
Mapping the active site in vasoactive intestinal peptide to a core of four amino acids: neuroprotective drug design.
(5/969)
The understanding of the molecular mechanisms leading to peptide action entails the identification of a core active site. The major 28-aa neuropeptide, vasoactive intestinal peptide (VIP), provides neuroprotection. A lipophilic derivative with a stearyl moiety at the N-terminal and norleucine residue replacing the Met-17 was 100-fold more potent than VIP in promoting neuronal survival, acting at femtomolar-picomolar concentration. To identify the active site in VIP, over 50 related fragments containing an N-terminal stearic acid attachment and an amidated C terminus were designed, synthesized, and tested for neuroprotective properties. Stearyl-Lys-Lys-Tyr-Leu-NH2 (derived from the C terminus of VIP and the related peptide, pituitary adenylate cyclase activating peptide) captured the neurotrophic effects offered by the entire 28-aa parent lipophilic derivative and protected against beta-amyloid toxicity in vitro. Furthermore, the 4-aa lipophilic peptide recognized VIP-binding sites and enhanced choline acetyltransferase activity as well as cognitive functions in Alzheimer's disease-related in vivo models. Biodistribution studies following intranasal administration of radiolabeled peptide demonstrated intact peptide in the brain 30 min after administration. Thus, lipophilic peptide fragments offer bioavailability and stability, providing lead compounds for drug design against neurodegenerative diseases. (+info)
Millimeter-scale positioning of a nerve-growth-factor source and biological activity in the brain.
(6/969)
Toxicity prevents the systemic administration of many therapeutic proteins, and attempts at protein targeting via the circulatory system (i.e., "magic bullets") have failed in all but a few special cases. Direct administration at the target site is a logical alternative, particularly in the central nervous system, but the limits of direct administration have not been defined clearly. Nerve growth factor (NGF) enhances survival of cholinergic neurons and, therefore, has generated considerable interest for the treatment of Alzheimer's disease. We tested the effectiveness of local delivery by implanting small polymer pellets that slowly released NGF into the central nervous system of adult rats at controlled distances from a target site containing transplanted fetal cholinergic cells. NGF-releasing implants placed within 1-2 mm of the treatment site enhanced the biological function of cellular targets, whereas identical implants placed approximately 3 mm from the target site of treatment produced no beneficial effect. Effective NGF therapy required millimeter-scale positioning of the NGF source, and efficacy correlated with the spatial distribution of NGF concentration in the tissue; this result suggests that NGF must be delivered within several millimeters of the target to be effective in treating Alzheimer's disease. Because the human brain is divided into functional regions that are typically several centimeters in diameter and often irregular in shape, new methods for sculpting larger-scale drug fields are needed. We illustrate a concept, called pharmacotectonics, in which drug-delivery systems are arranged spatially in tissues to shape concentration fields for potent agents. (+info)
The induction of apoptosis in HeLa cells by the loss of LBP-p40.
(7/969)
To analyze the function of the laminin-binding protein precursor p40 (LBP-p40) in higher eukaryotic cells, plasmid DNA expressing antisense or sense cDNA for p40 under the control of the LacSwitch system was introduced into HeLa cells. Stable transformants were isolated, and the expression of p40 was assayed by Western and Northern blotting. The expression level of p40 was not affected in HeLa cell transformants cultured in 10% serum-supplemented media with the induction of antisense (AS)-p40 with 5 mM IPTG. However, both the protein and message for endogenous p40 in serum-depleted media with 5 mM IPTG were reduced to about 30 - 10% of the expression level in serum-free media without 5 mM IPTG. Colony formation was inhibited with the suppression of p40. AS-p40 clones died in 7 days when cultured in serum-depleted media with 5 mM IPTG, while clones without 5 mM IPTG AS-p40 clones never died, even in serum-depleted media. Additionally, sense (S)-p40 clones and control CAT clones survived more than 2 weeks in serum-free media with 5 mM IPTG. DNA fragmentation assay revealed that cell death induced by the reduction of AS-p40 resulted from apoptosis. Both the inhibition of cell growth and apoptotic cell death were partially rescued by the transfer of the p40 cDNA expression vector to AS-p40 clones. Moreover, the introduction of a synthetic hammerhead ribozyme for LBP-p40 using a fusigenic viral liposome suppressed the message for LBP-p40 even in the presence of 10% serum, and it also induced apoptosis. (+info)
Egr-1 inhibits apoptosis during the UV response: correlation of cell survival with Egr-1 phosphorylation.
(8/969)
UV irradiation of normal or immortalized cells induces a rapid increase in the expression of several transcription factors and is thought to serve a protective function. The human fibrosarcoma cell line, HT1080 clone H4, expresses almost undetectable levels of Egr-1 and does not respond to UV-C irradiation by the induction of Egr-1. The H4 cells are hypersensitive to UV which induces apoptosis and reduces clonogenicity. The introduction of exogenous Egr-1 into H4 (H4E9 and H4E4 cell-lines) confers protection from UV damage as measured by a number of assays. In both NIH3T3 (with inducible Egr-1) and H4E9 (constitutive Egr-1) cells, UV irradiation gave enhanced transactivation of Egr-1 reporters that correlated with phosphorylated Egr-1. Studies using inhibitors indicated that protein kinase-C and tyrosine kinases are involved in the anti-apoptotic effects of Egr-1 after UV damage. This is the first description of a biological effect of phosphorylated Egr-1. (+info)