Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ischemia results in increased phosphorylation of NMDA receptors. To investigate the possible role of lipid rafts in this increase, lipid rafts and post-synaptic densities (PSDs) were isolated by the extraction of rat brain synaptosomes with Triton X-100 followed by sucrose density gradient centrifugation. Lipid rafts accounted for the majority of PSD-95, whereas SAP102 was predominantly located in PSDs. Between 50 and 60% of NMDA receptors were associated with lipid rafts. Greater than 85-90% of Src and Fyn were present in lipid rafts, whereas Pyk2 was mainly associated with PSDs. Lipid rafts and PSDs were isolated from animals subjected to 15 min of global ischemia followed by 6 h of recovery. Ischemia did not affect the yield, density, flotillin-1 or cholesterol content of lipid rafts. Following ischemia, the phosphorylation of NR1 by protein kinase C and tyrosine phosphorylation of NR2A and NR2B was increased in both lipid rafts and PSDs, with a greater increase in tyrosine phosphorylation occurring in the raft fraction. Following ischemia, NR1, NR2A and NR2B levels were elevated in PSDs and reduced in lipid rafts. The findings are consistent with a model involving close interaction between lipid rafts and PSDs and a role for lipid rafts in ischemia-induced signaling pathways.
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PMID:Increased phosphorylation and redistribution of NMDA receptors between synaptic lipid rafts and post-synaptic densities following transient global ischemia in the rat brain. 1577 18

NR2C-containing N-methyl-D-aspartate (NMDA) receptors are highly expressed in cerebellar granule cells where they mediate the majority of current in the adult. NMDA receptors composed of NR1/NR2C exhibit a low conductance and reduced sensitivity to Mg(2+), compared with the more commonly studied NR2A- and NR2B-containing receptors. Despite these interesting features, very little is known about the regulation of NR2C function. Here we investigate the role of phosphorylation of NR2C in regulating NMDA receptor trafficking and ion channel properties. We identify a phosphorylation site, serine 1244 (Ser(1244)), near the extreme COOH terminus of NR2C, which is phosphorylated by both cAMP-dependent protein kinase and protein kinase C. This residue is located adjacent to the consensus PDZ ligand, a region that regulates protein-protein interactions and receptor trafficking in NR2A and NR2B. We show that Ser(1244) on NR2C is phosphorylated in vitro, in heterologous cells, and in neurons. Moreover, we demonstrate for the first time that NR2C interacts with the PSD-95 family of PDZ domain-containing proteins but that phosphorylation of Ser(1244) does not influence this PDZ interaction. Furthermore, Ser(1244) phosphorylation does not regulate surface expression of NR1/NR2C receptors. However, we find that this site does regulate the kinetics of the ion channel: a phosphomimetic mutation at Ser(1244) accelerates both the rise and decay of NMDA-evoked currents in excised patches from HEK-293 cells. Therefore, phosphorylation of Ser(1244) does not regulate trafficking but unexpectedly affects ion channel function, suggesting that phosphorylation of Ser(1244) on NR2C may be important in defining the functional properties of NMDA receptor-mediated currents in the cerebellum.
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PMID:Regulation of NR1/NR2C N-methyl-D-aspartate (NMDA) receptors by phosphorylation. 1660 16

The present study characterizes the involvement of the N-methyl-D-aspartic acid receptors (NMDARs) in mediating thermal hyperalgesia induced by activation of group I metabotropic glutamate receptors (mGluRs). Intrathecal administration of the mGluR1/5 agonist (S)-3,5-DHPG [(S)-3,5-dihydroxyphenylglycine] to mice resulted in significant hyperalgesia as assessed by the tail immersion test. The pretreatment of mice i.t. with CGS 19755 (selective antagonist of the NMDAR), CGP 78608 [[(1S)-1-[[(7-bromo-1,2,3,4-tetrahydro-2,3-dioxo-5-quinoxalinyl)methyl]amino]ethyl]phosphonic acid] (selective antagonist at the glycine-binding site of the NMDAR), ifenprodil and Ro 25-6981 (selective antagonists of the NR2B subunit of the NMDAR), bisindolylmaleimide I and Go-7874 [12-(2-cyanoethyl)-6,7,12,13-tetrahydro-13-methyl-5-oxo-5H-indolo(2,3-a)pyrrolo(3,4-c)-carbazole] (inhibitors of protein kinase C), or PKI-(14-22)-amide [Myr-N-Gly-Arg-Thr-Gly-Arg-Arg-Asn-Ala-Ile-NH(2)] (inhibitor of protein kinase A) dose-dependently inhibited the hyperalgesia induced by i.t. administration of the mGluR1/5 receptor agonist (S)-3,5-DHPG. In contrast, i.t. pretreatment of mice with NVP-AAM077 [[(R)-[(S)-1-(4-bromophenyl)-ethylamino]-(2,3-dioxo-1,2,3,4-tetrahydroquinoxalin-5-yl)-methyl]-phosphonic acid] (selective antagonist of the NR2A subunit of the NMDAR) or DT-3 [H-Arg-Gln-Ile-Lys-Ile-Trp-Phe-Gln-Asn-Arg-Arg-Met-Lys-Trp-Lys-Lys-Leu-Arg-Lys-Lys-Lys-Lys-Lys-His-OH] (inhibitor of protein kinase G) had no effect on (S)-3,5-DHPG-mediated hyperalgesia. We also show for the first time that i.t. injection of pSM2 (pShag Magic version 2)-grin2b (coding for an short-hairpin RNA to the NR2B subunit of the NMDAR) resulted in a dose-dependent decrease in the NR2B protein and blockade of hyperalgesia induced by activation of the mGluR1/5 in (S)-3,5-DHPG-treated mice. Taken together, our results suggest the hypothesis that mGluRs are coupled to the NMDAR channels through the NR2B subunit in the spinal cord and that this coupling involves the activation of protein kinase C and protein kinase A.
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PMID:Decrease in N-methyl-D-aspartic acid receptor-NR2B subunit levels by intrathecal short-hairpin RNA blocks group I metabotropic glutamate receptor-mediated hyperalgesia. 1740 69

The contribution of metabotropic glutamate receptors to brain injury after in vivo cerebral ischemia remains to be determined. We investigated the effects of the metabotropic glutamate receptor 1 (mGluR1) antagonist LY367385 on brain injury after transient (90 min) middle cerebral artery occlusion in the rat and sought to explore their mechanisms. The intravenous administration of LY367385 (10 mg/kg) reduced the infarct volume at 24 h after the start of reperfusion. As the Gq-coupled mGluR1 receptor is known to activate the PKC/Src family kinase cascade, we focused on changes in the activation and amount of these kinases. Transient focal ischemia increased the amount of activated tyrosine kinase Src and PKC in the post-synaptic density (PSD) at 4 h of reperfusion. The administration of LY367385 attenuated the increases in the amounts of PSD-associated PKCgamma and Src after transient focal ischemia. We further investigated phosphorylation of the NMDA receptor, which is a major target of Src family kinases to modulate the function of the receptor. Transient focal ischemia increased the tyrosine phosphorylation of NMDA receptor subunits NR2A and NR2B. Tyrosine phosphorylation of NR2A, but not that of NR2B, in the PSD at 4 h of reperfusion was inhibited by LY367385. These results suggest that the mGluR1 after transient focal ischemia is involved in the activation of Src, which may be linked to the modification of properties of the NMDA receptor and the development of cerebral infarction.
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PMID:mGluR1 antagonist decreases tyrosine phosphorylation of NMDA receptor and attenuates infarct size after transient focal cerebral ischemia. 1824 25

Src family protein kinases (SFKs) -mediated tyrosine-phosphorylation regulates N-methyl-D-aspartate (NMDA) receptor synaptic function. Some members of the membrane-associated guanylate kinase (MAGUK) family of proteins bind to both SFKs and NMDA receptors, but it is unclear whether the MAGUK family of proteins is required for SFKs-mediated tyrosine-phosphorylation of the NMDA receptors. Here, we showed by co-immunoprecipitation that post-synaptic density (PSD) -93, a member of the MAGUK family of proteins, interacts with the NMDA receptor subunits NR2A and NR2B as well as with Fyn, a member of the SFKs, in mouse cerebral cortex. Using a biochemical fractionation approach to isolate subcellular compartments revealed that the expression of Fyn, but not of other members of the SFKs (Lyn, Src, and Yes), was significantly decreased in synaptosomal membrane fractions derived from the cerebral cortex of PSD-93 knockout mice. Interestingly, we found that PSD-93 disruption causes reduction of tyrosine-phosphorylated NR2A and NR2B in the same fraction. Moreover, PSD-93 deletion markedly blocked the SFKs-mediated increase in tyrosine-phosphorylated NR2A and NR2B through the protein kinase C pathway after induction with 4-phorbol 12-myristate 13-acetate in cultured cortical neurons. Our findings indicate that PSD-93 appears to mediate tyrosine-phosphorylation of the NMDA receptors and synaptic localization of Fyn.
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PMID:Post-synaptic density-93 mediates tyrosine-phosphorylation of the N-methyl-D-aspartate receptors. 1842 99

The purpose of this study was to determine the effect of protein kinase C (PKC) activation by 100 nM phorbol 12-myristate 13-acetate (PMA) on N-methyl-d-aspartate (NMDA) receptor function with the whole-cell patch-clamp technique. Receptors expressed in primary cultured cerebellar granule cells at days in vitro that result in different NMDA NR2A and NR2B subunit composition were assessed. The effect of temperature during PMA exposure on NMDA-induced current amplitudes as well as PMA-induced translocation of PKC isoform-specific immunoreactivity was also assessed. We observed that PMA augmented NMDA-induced peak current amplitude regardless of NR2 subunit composition and augmentation of NMDA-induced steady-state current amplitudes was only observed in 13 and older days in vitro cerebellar granule cells. PMA treatment did not affect the desensitized state (steady-state to peak current ratios) of the receptor. Augmentation of NMDA-induced current amplitude was seen by 12.5 min PMA exposure, a time that corresponded with translocation of all PMA-sensitive PKC isoform immunoreactivity. PMA exposure at 37 degrees C resulted in a significant enhancement of NMDA-induced current amplitude compared to augmentation of receptor function following a PMA exposure at 23 degrees C. Translocation of PKC immunoreactivity was also greatly attenuated at 23 degrees C compared to treatment at 37 degrees C. While our data support previous observations that activation of PKC by PMA enhances NMDA receptor function, this augmentation does not appear to be dependent upon NR2 subunit composition. Furthermore our data emphasize the importance of conducting experiments at physiological temperatures when assessing PKC effects on native NMDA receptors.
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PMID:Activation of protein kinase C enhances NMDA-induced currents in primary cultured cerebellar granule cells: Effect of temperature and NMDA NR2 subunit composition. 1876 83

Platelet-derived growth factor (PDGF) beta receptor activation inhibits N-methyl-d-aspartate (NMDA)-evoked currents in hippocampal and cortical neurons via the activation of phospholipase Cgamma, PKC, the release of intracellular calcium, and a rearrangement of the actin cytoskeleton. In the hippocampus, the majority of NMDA receptors are heteromeric; most are composed of 2 NR1 subunits and 2 NR2A or 2 NR2B subunits. Using NR2B- and NR2A-specific antagonists, we demonstrate that PDGF-BB treatment preferentially inhibits NR2B-containing NMDA receptor currents in CA1 hippocampal neurons and enhances long-term depression in an NR2B subunit-dependent manner. Furthermore, treatment of hippocampal slices or cultures with PDGF-BB decreases the surface localization of NR2B but not of NR2A subunits. PDGFbeta receptors colocalize to a higher degree with NR2B subunits than with NR2A subunits. After neuronal injury, PDGFbeta receptors and PDGF-BB are up-regulated and PDGFbeta receptor activation is neuroprotective against glutamate-induced neuronal damage in cultured neurons. We demonstrate that the neuroprotective effects of PDGF-BB are occluded by the NR2B antagonist, Ro25-6981, and that PDGF-BB promotes NMDA signaling to CREB and ERK1/2. We conclude that PDGFbetaR signaling, by preferentially targeting NR2B receptors, provides an important mechanism for neuroprotection by growth factors in the central nervous system.
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PMID:Platelet-derived growth factor selectively inhibits NR2B-containing N-methyl-D-aspartate receptors in CA1 hippocampal neurons. 1910 10

Pregnenolone sulfate (PS), an endogenously occurring neurosteroid, has been shown to modulate the activity of several neurotransmitter-gated channels, including the N-methyl-D-aspartate receptor (NMDAR). NMDARs are glutamate-gated ion channels involved in excitatory synaptic transmission, synaptic plasticity, and excitotoxicity. To determine the mechanism that controls PS sensitivity of NMDARs, we measured NMDAR responses induced by exogenous agonist application in voltage-clamped HEK293 cells expressing NR1/NR2B NMDARs and cultured rat hippocampal neurons. We report that PS potentiates the amplitude of whole-cell recorded NMDAR responses in cultured hippocampal neurons and HEK293 cells; however, the potentiating effect of PS on NMDAR in outside-out patches isolated from cultured hippocampal neurons and HEK293 cells was lost within 2 min after patch isolation in a neurosteroid-specific manner. The rate of diminution of the PS potentiating effect was slowed by protein phosphatase inhibitors. Treatment of cultured hippocampal neurons with a nonspecific protein kinase inhibitor and a specific protein kinase A (PKA) inhibitor diminished PS-induced potentiation, which was recovered by adding a PKA, but not a protein kinase C (PKC), activator. These results suggest that the effect of PS on NMDARs is controlled by cellular mechanisms that are mediated by dephosphorylation/phosphorylation pathways.
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PMID:Pregnenolone sulfate modulation of N-methyl-D-aspartate receptors is phosphorylation dependent. 1927 23

Long-term potentiation (LTP) at hippocampal CA3-CA1 synapses is thought to be mediated, at least in part, by an increase in the postsynaptic surface expression of alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA) receptors induced by N-methyl-d-aspartate (NMDA) receptor activation. While this process was originally attributed to the regulated synaptic insertion of GluA1 (GluR-A) subunit-containing AMPA receptors, recent evidence suggests that regulated synaptic trafficking of GluA2 subunits might also contribute to one or several phases of potentiation. However, it has so far been difficult to separate these two mechanisms experimentally. Here we used genetically modified mice lacking the GluA1 subunit (Gria1(-/-) mice) to investigate GluA1-independent mechanisms of LTP at CA3-CA1 synapses in transverse hippocampal slices. An extracellular, paired theta-burst stimulation paradigm induced a robust GluA1-independent form of LTP lacking the early, rapidly decaying component characteristic of LTP in wild-type mice. This GluA1-independent form of LTP was attenuated by inhibitors of neuronal nitric oxide synthase and protein kinase C (PKC), two enzymes known to regulate GluA2 surface expression. Furthermore, the induction of GluA1-independent potentiation required the activation of GluN2B (NR2B) subunit-containing NMDA receptors. Our findings support and extend the evidence that LTP at hippocampal CA3-CA1 synapses comprises a rapidly decaying, GluA1-dependent component and a more sustained, GluA1-independent component, induced and expressed via a separate mechanism involving GluN2B-containing NMDA receptors, neuronal nitric oxide synthase and PKC.
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PMID:Induction and expression of GluA1 (GluR-A)-independent LTP in the hippocampus. 1930 50

We have previously reported that activation of protein kinase C (PKC) by phorbol 12-myristate 13-acetate (PMA) results in potentiation of N-methyl-D-aspartate-induced currents (I(NMDA))of receptors contained in primary cultured cerebellar granule cells (CGCs). The purpose of this study was to identify which PKC isoform(s) was responsible for this effect by using the whole-cell patch-clamp technique. Experiments were conducted on CGCs that expressed both the NR2A and NR2B NMDA receptor subunits as well as the PMA-sensitive PKC isoforms alpha, betaI, betaII, delta, epsilon, gamma, and . As observed previously, N-methyl-D-aspartate-induced peak currents (I(Pk)) were enhanced by a 12.5-min, 100 nM PMA exposure at 37 degrees C under normal recording conditions. Potentiation of receptor function was not observed when extracellular Ca(2+) was removed and 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid was present inside the cell. PMA-induced potentiation of I(Pk) did not occur when PKCalpha-specific antibody was introduced into the cell via the recording electrode. However, in similar experiments with antibodies specific for PKCbetaII, delta, epsilon, gamma, and , PMA potentiation of I(Pk) was observed. Down-regulation of PMA-sensitive PKC isoforms by an overnight exposure of 100 nM PMA resulted in lack of potentiation by PMA that was rescued when catalytically active PKCalpha was introduced into the cell via the patch electrode. PMA potentiation of I(Pk) was not recovered when catalytically active PKCbetaI, PKCbetaII, or PKCgamma was introduced into the cell via the patch electrode. Collectively, our data provide strong evidence that PMA-enhanced function of native NMDA receptors expressed in primary cultured CGCs is mediated by activation of PKCalpha.
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PMID:Phorbol 12-myristate 13-acetate potentiation of N-methyl-D-aspartate-induced currents in primary cultured cerebellar granule cells is mediated by protein kinase C alpha. 1942 93


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