NMDA receptor dysfunction in Alzheimer's disease

The inherent neurotoxic potential ofthe endogenous excitatory amino acid glutamate, may be causally related to the pathogenesis ofAD neurodegeneration disorders. Neuronal excitotoxicity is conceivably mediated by the N-methyl-D-aspartate-(NMDA)-Ca2+- ionotropic receptor. NMDA receptors exist as multimeric complexes comprising proteins from two families – NR1 and NR2(A-D). The polyamines, spermine and spermidine bind to, and modulate NMDA receptor efficacy via interaction with exon 5, an alternatively-spliced, 21 amino acid, N-terminal cassette. ADassociated cognitive impairment may therefore occur via subunitspecific NMDA receptor dysfunction effecting regional selectivity ofneuronal degradation. Total RNA was prepared from pathologically spared and susceptible regions from AD cases and matched controls. Quantitation was performed using standard curve methodology in which a known amount ofa synthetic ribonucleic acid competitor deletion construct was co-amplified against total RNA. Expression profile analysis oftwo NR1 mRNA subsets has revealed significant differences in NR11XX mRNA levels in cingulate gyrus, P.

Novel, isoform-selective, cholecystokinin A receptor antagonist inhibits colon and pancreatic cancers in preclinical models through novel mechanism of action

Colon and pancreatic cancers contribute to 90,000 deaths each year in the USA. These cancers lack targeted therapeutics due to heterogeneity of the disease and multiple causative factors. One important factor that contributes to increased colon and pancreatic cancer risk is gastrin. Gastrin mediates its actions through two G-protein coupled receptors (GPCRs): cholecystokinin receptor A (CCK-A) and CCK-B/gastrin receptor. Previous studies have indicated that colon cancer predominantly expresses CCK-A and responds to CCK-A isoform antagonists. However, many CCK-A antagonists have failed in the clinic due to poor pharmacokinetic properties or lack of efficacy. In the present study, we synthesized a library of CCK-A isoform-selective antagonists and tested them in various colon and pancreatic cancer preclinical models. The lead CCK-A isoform, selective antagonist PNB-028, bound to CCK-A at 12 nM with a 60-fold selectivity towards CCK-A over CCK-B. Furthermore, it inhibited the proliferation of CCK-A-expressing colon and pancreatic cancer cells without affecting the proliferation of non-cancerous cells. PNB-028 was also extremely effective in inhibiting the growth of MAC-16 and LoVo colon cancer and MIA PaCa pancreatic cancer xenografts in immune-compromised mice. Genomewide microarray and kinase-array studies indicate that PNB-028 inhibited oncogenic kinases and angiogenic factors to inhibit the growth of colon cancer xenografts. Safety pharmacology and toxicology studies have indicated that PNB-028 is extremely safe and has a wide safety margin. These studies suggest that targeting CCK-A selectively renders promise to treat colon and pancreatic cancers and that PNB-028 could become the next-generation treatment option.

Computer assisted synthesis and in-vitro cytotoxic evaluation of new pyrazole-fused isoquinolinoquinones derivatives as PI3K receptor antagonist with promising antitumoral activity

The importance of pyrazole and isoquinoline-5,8-dione scaffolds in medical chemistry is underlined by the high number of drugs currently on trading that contains these active ingredients. Due to their cytotoxic capability, the interest of medicinal chemists in these heterocyclic rings has grown exponentially especially, for cancer therapy. In this project, the first synthesis of pyrazole-fused isoquinoline-5,8-diones has been developed. 1,3-Dipolar cycloaddition followed by oxidative aromatization, established by our research group, has been employed. Screening of reaction conditions and characterization studies about the regioselectivity have been successfully performed. A remote control of regioselectivity, to achieve the two possible regioisomers has been accomplished. Through Molecular Docking studies, Structure-Activity relationship of differently substituted scaffolds containing our central core proved that a family of PI3K inhibitors have been discovered. Finally, in order to verify the promising antitumor activity, a first test of cell viability in vitro on T98G cell line of a solid brain tumor, the Glioblastoma Multiforme, showed cytotoxic inhibition comparable to currently trade anticancer drugs.

Immune proteins in neurons : towards an understanding of the neuronal role of the immune protein CD3[Zeta] in NMDA receptor signaling and cytoskeleton remodeling

Otto-von-Guericke-Universität Magdeburg, Fakultät für Naturwissenschaften, Dissertation, 2016

First-episode psychosis in a 15 year-old female with clinical presentation of anti-NMDA receptor encephalitis: a case report and review of the literature

BACKGROUND: Anti-NMDA receptor encephalitis is an autoimmune disease that was identified in 2007, and manifests in a stepwise manner with psychiatric, neurological and autonomic symptoms. The disease is caused by autoantibodies against NMDA receptors. It can have a paraneoplastic origin, mainly secondary to ovarian teratomas, but it can also be unrelated to the tumor. This disease can affect both sexes and all ages. CASE PRESENTATION: Here, we present a case of a 15 year-old female adolescent with first-episode psychosis with anti-NMDA receptor encephalitis not related to tumor, which manifested with delusion, hallucinations, panic attacks, agitation, and neurological symptoms, and later with autonomic instability. She was treated with immunotherapy and psychiatric medication resulting in improvement of her main psychiatric and neurological symptoms. CONCLUSION: Our main objective in presenting this case is to alert clinicians to this challenging and recent disease that has a clinical presentation that might resemble a functional psychiatric condition and can be underdiagnosed in the context of child and adolescent psychiatry

Laminar and cellular distribution of AMPA, kainate, and NMDA receptor subunits in monkey sensory-motor cortex

In situ hybridization histochemistry and immunocytochemistry were used to examine lamina- and cell-specific expression of glutamate receptor (GluR) mRNAs and polypeptide subunits in motor and somatosensory cortex of macaque monkeys. Radioactive complementary RNA (cRNA) probes were prepared from cDNAs specific for α-amino-3-hydroxy-5-methylisoxozolepropionate (AMPA)/kainate (GluR1-GluR4), kainate (GluR5-GluR7), and N-methylD-aspartate (NMDA; NR1, NR2A-NR2D) receptor subunits. AMPA/kainate and NR1, NR2A, and NR2B receptor transcripts show higher expression than other transcripts. All transcripts show lamina-specific patterns of distribution. GluR2 and GluR4 mRNAs show higher expression than do GluR1 and GluR3 mRNAs. GluR6 transcript expression is higher than that of GluR5 and GluR7. NR1 mRNA expression is much higher than that of NR2 mRNAs. NR2C subunit expression is very low except for a very distinct band of high expression in layer IV of area 3b. Immunocytochemistry, using subunit-specific antisera and double labeling for calbindin, parvalbumin, or α type II Ca2+/calmodulin-dependent protein kinase (CaMKII-α), allowed identification of cell types expressing different subunit genes. GluR1 and GluR5/6/7 immunoreactivity is found in both pyramidal cells and gamma-amino butyric acid (GABA) cells; GluR2/3 immunoreactivity is preferentially found in pyramidal cells, whereas GluR4 immunoreactivity is largely restricted to GABA cells; NMDA receptor subunit immunoreactivity is far greater in excitatory cells than in GABA cells. The density of expression of AMPA/kainate, kainate, and NMDA receptor subunit mRNAs differed within and across the architectonic fields of sensory-motor cortex. This finding and the lamina- and cell-specific patterns of expression suggest assembly of functional receptors from different arrangements of available subunits in specific neuronal populations.

Stress enables synaptic depression in CA1 synapses by acute and chronic morphine: Possible mechanisms for corticosterone on opiate addiction

The hippocampus, being sensitive to stress and glucocorticoids, plays significant roles in certain types of learning and memory. Therefore, the hippocampus is probably involved in the increasing drug use, drug seeking, and relapse caused by stress. We have studied the effect of stress with morphine on synaptic plasticity in the CA1 region of the hippocampus in vivo and on a delayed-escape paradigm of the Morris water maze. Our results reveal that acute stress enables long-term depression (LTD) induction by low-frequency stimulation (LFS) but acute morphine causes synaptic potentiation. Remarkably, exposure to an acute stressor reverses the effect of morphine from synaptic potentiation ( similar to 20%) to synaptic depression ( similar to 40%), precluding further LTD induction by LFS. The synaptic depression caused by stress with morphine is blocked either by the glucocorticoid receptor antagonist RU38486 or by the NMDA-receptor antagonist D-APV. Chronic morphine attenuates the ability of acute morphine to cause synaptic potentiation, and stress to enable LTD induction, but not the ability of stress in tandem with morphine to cause synaptic depression. Furthermore, corticosterone with morphine during the initial phase of drug use promotes later delayed-escape behavior, as indicated by the morphine-reinforced longer latencies to escape, leading to persistent morphine-seeking after withdrawal. These results suggest that hippocampal synaptic plasticity may play a significant role in the effects of stress or glucocorticoids on opiate addiction.

DE-71-induced apoptosis involving intracellular calcium and the Bax-mitochondria-caspase protease pathway in human neuroblastoma cells In vitro

Polybrominated diphenyl ethers (PBDEs) are used extensively as flame-retardants and are ubiquitous in the environment and in wildlife and human tissue. Recent studies have shown that PBDEs induce neurotoxic effects in vivo and apoptosis in vitro. However, the signaling mechanisms responsible for these events are still unclear. In this study, we investigated the action of a commercial mixture of PBDEs (pentabrominated diphenyl ether, DE-71) on a human neuroblastoma cell line, SK-N-SH. A cell viability test showed a dose-dependent increase in lactate dehydrogenase leakage and 3-(4,5-dimethylthia-zol-2-yl)-2,5-diphenyl-tetrazolium bromide reduction. Cell apoptosis was observed through morphological examination, and DNA degradation in the cell cycle and cell apoptosis were demonstrated using flow cytometry and DNA laddering. The formation of reactive oxygen species was not observed, but DE-71 was found to significantly induce caspase-3, -8, and -9 activity, which suggests that apoptosis is not induced by oxidative stress but via a caspase-dependent pathway. We further investigated the intracellular calcium ([Ca2+](i)) levels using flow cytometry and observed an increase in the intracellular Ca2+ concentration with a time-dependent trend. We also found that the N-methyl d-aspartate (NMDA) receptor antagonist MK801 (3 mu M) significantly reduced DE-71-induced cell apoptosis. The results of a Western blotting test demonstrated that DE-71 treatment increases the level of Bax translocation to the mitochondria in a dose-dependent fashion and stimulates the release of cytochrome c (Cyt c) from the mitochondria into the cytoplasm. Overall, our results indicate that DE-71 induces the apoptosis of ([Ca2+](i)) in SK-N-SH cells via Bax insertion, Cyt c release in the mitochondria, and the caspase activation pathway.

Dopamine affects parvalbumin expression during cortical development in vitro.

This study was undertaken to determine how dopamine influences cortical development. It focused on morphogenesis of GABAergic neurons that contained the calcium-binding protein parvalbumin (PV). Organotypic slices of frontoparietal cortex were taken from neonatal rats, cultured with or without dopamine, harvested daily (4-30 d), and immunostained for parvalbumin. Expression of parvalbumin occurred in the same regional and laminar sequence as in vivo. Expression in cingulate and entorhinal preceded that in lateral frontoparietal cortices. Laminar expression progressed from layer V to VI and finally II-IV. Somal labeling preceded fiber labeling by 2 d. Dopamine accelerated PV expression. In treated slices, a dense band of PV-immunoreactive neurons appeared in layer V at 7 d in vitro (DIV), and in all layers of frontoparietal cortex at 14 DIV, whereas in control slices such labeling did not appear until 14 and 21 DIV, respectively. The laminar distribution and dendritic branching of PV-immunoreactive neurons were quantified. More labeled neurons were in the superficial layers, and their dendritic arborizations were significantly increased by dopamine. Treatment with a D1 receptor agonist had little effect, whereas a D2 agonist mimicked dopamine's effects. Likewise, the D2 but not the D1 antagonist blocked dopamine-induced changes, indicating that they were mediated primarily by D2 receptors. Parvalbumin expression was accelerated by dopaminergic reinnervation of cortical slices that were cocultured with mesencephalic slices. Coapplication of the glutamate NMDA receptor antagonist MK801 or AP5 blocked dopamine-induced increases in dendritic branching, suggesting that changes were mediated partly by interaction with glutamate to alter cortical excitability.

Acquisition of Pavlovian Fear Conditioning Using beta-Adrenoceptor Activation of the Dorsal Premammillary Nucleus as an Unconditioned Stimulus to Mimic Live Predator-Threat Exposure

In the present work, we sought to mimic the internal state changes in response to a predator threat by pharmacologically stimulating the brain circuit involved in mediating predator fear responses, and explored whether this stimulation would be a valuable unconditioned stimulus (US) in an olfactory fear conditioning paradigm (OFC). The dorsal premammillary nucleus (PMd) is a key brain structure in the neural processing of anti-predatory defensive behavior and has also been shown to mediate the acquisition and expression of anti-predatory contextual conditioning fear responses. Rats were conditioned by pairing the US, which was an intra-PMd microinjection of isoproterenol (ISO; beta-adrenoceptor agonist), with amyl acetate odor-the conditioned stimulus (CS). ISO (10 and 40 nmol) induced the acquisition of the OFC and the second-order association by activation of beta-1 receptors in the PMd. Furthermore, similar to what had been found for contextual conditioning to a predator threat, atenolol (beta-1 receptor antagonist) in the PMd also impaired the acquisition and expression of OFC promoted by ISO. Considering the strong glutamatergic projections from the PMd to the dorsal periaqueductal gray (dPAG), we tested how the glutamatergic blockade of the dPAG would interfere with the OFC induced by ISO. Accordingly, microinjections of NMDA receptor antagonist (AP5, 6 nmol) into the dPAG were able to block both the acquisition, and partially, the expression of the OFC. In conclusion, we have found that PMd beta-1 adrenergic stimulation is a good model to mimic predatory threat-induced internal state changes, and works as a US able to mobilize the same systems involved in the acquisition and expression of predator-related contextual conditioning. Neuropsychopharmacology (2011) 36, 926-939; doi:10.1038/npp.2010.231; published online 5 January 2011