AMPA receptor (AMPAR)-mediated ionic currents that govern gene expression, synaptic strength, and plasticity also can trigger excitotoxicity. However, native AMPARs exhibit heterogeneous ...pharmacological, biochemical, and ionic permeability characteristics, which are governed partly by receptor subunit composition. Consequently, the mechanisms governing AMPAR-mediated excitotoxicity have been difficult to elucidate. The GluR2 subunit is of particular interest because it influences AMPAR pharmacology, Ca(2+) permeability, and AMPAR interactions with intracellular proteins. In this paper we used mutant mice lacking the AMPAR subunit GluR2 to study AMPAR-mediated excitotoxicity in cultured cortical neurons and in hippocampal neurons in vivo. We examined the hypothesis that in these mice the level of GluR2 expression governs the vulnerability of neurons to excitotoxicity and further examined the ionic mechanisms that are involved. In cortical neuronal cultures AMPAR-mediated neurotoxicity paralleled the magnitude of kainate-evoked AMPAR-mediated currents, which were increased in neurons lacking GluR2. Ca(2+) permeability, although elevated in GluR2-deficient neurons, did not correlate with excitotoxicity. However, toxicity was reduced by removal of extracellular Na(+), the main charge carrier of AMPAR-mediated currents. In vivo, the vulnerability of CA1 hippocampal neurons to stereotactic kainate injections and of CA3 neurons to intraperitoneal kainate administration was independent of GluR2 level. Neurons lacking the GluR2 subunit did not demonstrate compensatory changes in the distribution, expression, or function of AMPARs or of Ca(2+)-buffering proteins. Thus GluR2 level may influence excitotoxicity by effects additional to those on Ca(2+) permeability, such as effects on agonist potency, ionic currents, and synaptic reorganization.
Ligand-gated ion channels gated by glutamate constitute the major excitatory neurotransmitter system in the mammalian brain. The functional modulation of GluR6, a kainate-activated glutamate ...receptor, by adenosine 3′,5′-monophosphate-dependent protein kinase A (PKA) was examined with receptors expressed in human embryonic kidney cells. Kainate-evoked currents underwent a rapid desensitization that was blocked by lectins. Kainate currents were potentiated by intracellular perfusion of PKA, and this potentiation was blocked by co-application of an inhibitory peptide. Site-directed mutagenesis was used to identify the site or sites of phosphorylation on GluR6. Although mutagenesis of two serine residues, Ser$^{684}$ and Ser$^{666}$, was required for complete abolition of the PKA-induced potentiation, Ser$^{684}$ may be the preferred site of phosphorylation in native GluR6 receptor complexes. These results indicate that glutamate receptor function can be directly modulated by protein phosphorylation and suggest that a dynamic regulation of excitatory receptors could be associated with some forms of learning and memory in the mammalian brain.
There is growing evidence that NMDA receptor-dependent long-term potentiation (LTP) in the hippocampus mediates the synaptic
plasticity that underlies spatial learning and memory. LTP deficiencies ...correlate well with spatial memory deficits and LTP
enhancements may improve spatial memory. In addition, LTP deficiencies are associated with abnormal place cells as expected
from the spatial mapping hypothesis of hippocampal function. In contrast, nothing is known on how enhanced NMDA receptor-dependent
LTP affects place cells. To address this question we recorded place cells from mice lacking the nociceptin receptor (NOP 1 /ORL 1 /OP4) that have enhanced hippocampal LTP. We found that the enhanced LTP was mediated by NMDA receptors, did not require L-type
calcium channels, and occurred only when high frequency tetanizing stimulus trains were used. Place cells in nociceptin receptor
knockout mice were abnormal in several ways: they were less stable, had noisier positional firing patterns, larger firing
fields and higher discharge rates inside and outside the firing fields. Our results suggest that excessive LTP can cause subnormal
hippocampal place cell function. The effects of LTP enhancement on place cell function may therefore also depend on molecular
details of synaptic plasticity, including the relationship between stimulus frequency and synaptic strength, and not merely
on the magnitude of synaptic strength increases. The data have important clinical implications on development of strategies
to improve cognitive function.
Palmitoylation of the GluR6 Kainate Receptor Pickering, D S; Taverna, F A; Salter, M W ...
Proceedings of the National Academy of Sciences - PNAS,
12/1995, Letnik:
92, Številka:
26
Journal Article
Recenzirano
Odprti dostop
The G-protein-coupled metabotropic glutamate receptor mGluR1α and the ionotropic glutamate receptor GluR6 were examined for posttranslational palmitoylation. Recombinant receptors were expressed in ...baculovirus-infected insect cells or in human embryonic kidney cells and were metabolically labeled with 3Hpalmitic acid. The metabotropic mGluR1α receptor was not labeled whereas the GluR6 kainate receptor was labeled after incubation with 3Hpalmitate. The 3Hpalmitate labeling of GluR6 was eliminated by treatment with hydroxylamine, indicating that the labeling was due to palmitoylation at a cysteine residue via a thioester bond. Site-directed mutagenesis was used to demonstrate that palmitoylation of GluR6 occurs at two cysteine residues, C827 and C840, located in the carboxyl-terminal domain of the molecule. A comparison of the electrophysiological properties of the wild-type and unpalmitoylated mutant receptor (C827A, C840A) showed that the kainate-gated currents produced by the unpalmitoylated mutant receptor were indistinguishable from those of the wild-type GluR6. The unpalmitoylated mutant was a better substrate for protein kinase C than the wild-type GluR6 receptor. These data indicate that palmitoylation may not modulate kainate channel function directly but instead affect channel function indirectly by regulating the phosphorylation state of the receptor.
Male Long-Evans rats were injected with 32 ng/μl of the
N
-methyl-
D
-aspartate (NMDA) receptor antagonist 3-(2-carboxypiperazin-4-yl) propyl-1-phosphonic acid (CPP) or vehicle and trained to locate ...a hidden platform in a different location (reversal training) than used on the initial 4 days of training. Rats treated with vehicle or CPP into the dorsal hippocampus, basolateral amygdala, or mediodorsal striatum had similar latencies to locate the platform on the reversal day. Rats infused with CPP into the dorsal hippocampus or mediodorsal striatum failed to search preferentially in the novel location during a 24-hr, drug-free retention test, whereas all other groups searched preferentially in this location. Therefore, blocking dorsal hippocampal or mediodorsal striatal NMDA receptors selectively blocked long-term spatial retention without producing short-term performance deficits.
The objective of the present work was to identify the compositional parameters of raw milk that affected ethanol stability at natural pH when natural milk conditions were not modified. Heat ...stability, measured as coagulation time (CT), was included in the analysis to verify relation to alcohol test. Statistical models were proposed for alcohol and heat (CT) stabilities. Milk samples of good hygienic quality from dairy farms were classified in two groups according to their alcohol stability. Unstable samples to ethanol (72%, v/v) presented lower values of pH, somatic cells count, casein and non-fat-solids relative to ethanol stable samples (ethanol at 78%, v/v or more); whereas freezing point, chloride, sodium and potassium concentrations were higher in the unstable group. Logistic regression and multiple regression were applied to modelling alcohol and heat stability behaviour respectively. Chloride, potassium, ionic calcium and somatic cell count were included in the alcohol regression model, whereas calcium, phosphorous, urea, pH and ionic calcium were part of CT model. Ionic calcium was the only measured variable that contributed to both models; however coagulation time was noted to be more sensitive to ionic calcium than alcohol. The relation between ionic strength and casein was found to contribute to the alcohol model but not to the CT model. However, the interaction calcium plus magnesium plus phosphorous and casein contributed only to CT model.
Advanced prostate cancer is a very heterogeneous disease reflecting in diverse regulations of oncogenic signaling pathways. Aberrant spatial dynamics of epidermal growth factor receptor (EGFR) ...promote their dimerization and clustering, leading to constitutive activation in oncogenesis. The EphB2 and Src signaling pathways are associated with the reorganization of the cytoskeleton leading to malignancy, but their roles in regulating EGFR dynamics and activation are scarcely reported. Using single-particle tracking techniques, we found that highly phosphorylated EGFR in the advanced prostate cancer cell line, PC3, was associated with higher EGFR diffusivity, as compared with LNCaP and less aggressive DU145. The increased EGFR activation and biophysical dynamics were consistent with high proliferation, migration, and invasion. After performing single-cell RNA-seq on prostate cancer cell lines and circulating tumor cells from patients, we identified that upregulated gene expression in the EphB2 and Src pathways are associated with advanced malignancy. After dasatinib treatment or siRNA knockdowns of EphB2 or Src, the PC3 cells exhibited significantly lower EGFR dynamics, cell motility, and invasion. Partial inhibitory effects were also found in DU145 cells. The upregulation of parts of the EphB2 and Src pathways also predicts poor prognosis in the prostate cancer patient cohort of The Cancer Genome Atlas. Our results provide evidence that overexpression of the EphB2 and Src signaling pathways regulate EGFR dynamics and cellular aggressiveness in some advanced prostate cancer cells.
In contrast to anaemia, polycythaemia is a distinctly uncommon finding in patients with multiple myeloma. We describe the presence of otherwise unexplained polycythaemia in a 57-year-old Caucasian ...man who was found to have IgG κ multiple myeloma. After treatment of myeloma, the polycythaemia resolved. We reviewed previous reports of polycythaemia associated with multiple myeloma and discuss potential pathophysiological mechanisms that link these 2 conditions.
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