共查询到18条相似文献,搜索用时 0 毫秒
1.
The Metabotropic Glutamate Receptor mGluR5 Induces Calcium Oscillations in Cultured Astrocytes via Protein Kinase C Phosphorylation 总被引:6,自引:3,他引:3
Kiyoshi Nakahara Masamichi Okada Shigetada Nakanishi 《Journal of neurochemistry》1997,69(4):1467-1475
Abstract: The metabotropic glutamate receptor mGluR5, but not the closely related mGluR1, is expressed in cultured astrocytes, and this expression is up-regulated by specific growth factors. We investigated the capability and underlying mechanisms of mGluR5 to induce oscillatory responses of intracellular calcium concentration ([Ca2+ ]i ) in cultured rat astrocytes. Single-cell [Ca2+ ]i recordings indicated that an mGluR-selective agonist, (1 S ,3 R )-1-aminocyclopentane-1,3-dicarboxylate (1 S ,3 R -ACPD), elicits [Ca2+ ]i oscillations in good agreement with the growth factor-induced up-regulation of mGluR5 in cultured astrocytes. A protein kinase C (PKC) inhibitor, bisindolylmaleimide I, converted a 1 S ,3 R -ACPD-mediated oscillatory response into a nonoscillatory response. In addition, the PKC activator phorbol 12-myristate 13-acetate completely abolished the [Ca2+ ]i increase. These and other pharmacological properties of 1 S ,3 R -ACPD-induced [Ca2+ ]i oscillations correlate well with those of the cloned mGluR5 characterized in heterologous expression systems. Furthermore, the potential involvement of protein phosphatases in [Ca2+ ]i oscillations is suggested. The present study demonstrates that mGluR5 is capable of inducing [Ca2+ ]i oscillations in cultured astrocytes and that phosphorylation/dephosphorylation of mGluR5 is critical in [Ca2+ ]i oscillations, analogous to the cloned mGluR5 expressed in heterologous cell lines. 相似文献
2.
Eleanor T. Coffey Inmaculada Herrero Talvinder S. Sihra José Sánchez-Prieto David G. Nicholls 《Journal of neurochemistry》1994,63(4):1303-1310
Abstract: 4-Aminopyridine evokes repetitive firing of synaptosomes and exocytosis of glutamate by inhibiting a dendrotoxin-sensitive K+ channel responsible for stabilizing the membrane potential. We have shown previously that activation of protein kinase C (PKC) by high concentrations of phorbol ester (4β-phorbol dibutyrate) can increase release by inhibiting a dendrotoxin-insensitive ion channel, whereas the metabotropic glutamate receptor (mGluR) agonist (1 S ,3 R )-1-aminocyclopentane-1,3-dicarboxylate [(1 S ,3 R )-ACPD] mimics the action of 4β-phorbol dibutyrate, but only in the presence of 2 µ M arachidonic acid (AA). In this article, we investigate the role of AA. AA plus (1 S ,3 R )-ACPD is without effect on KCl-induced glutamate exocytosis, indicating that the regulatory pathway acts upstream of the release-coupled Ca2+ channel or Ca2+ -secretion coupling. Diacylglycerol concentrations are greatly enhanced by (1 S ,3 R )-ACPD alone, independently of AA, indicating that AA acts downstream of phospholipase C. Myristoylated alanine-rich C kinase substrate (MARCKS) is the major presynaptic substrate for PKC. mGluR activation by (1 S ,3 R )-ACPD enhances phosphorylation of MARCKS, but only in the presence of AA. These results strongly suggest that AA acts on presynaptic PKC synergistically with diacylglycerol generated by the phospholipase-coupled mGluR, consistent with the known behaviour of certain purified PKC isoforms. The magnitude of the effects observed in a population of rat cerebrocortical synaptosomes suggests that this is a major mechanism regulating the release of the brain's dominant excitatory neurotransmitter and supports the concept that AA, or a related compound with a similar locus of action, may in certain circumstances play a role in synaptic plasticity. 相似文献
3.
Cellular Mechanisms of Protection by Metabotropic Glutamate Receptors During Anoxia and Nitric Oxide Toxicity 总被引:3,自引:2,他引:1
Abstract: Metabotropic glutamate receptors, nitric oxide (NO), and the signal transduction pathways of protein kinase C (PKC) and protein kinase A (PKA) can independently alter ischemic-induced neuronal cell death. We therefore examined whether the protective effects of metabotropic glutamate receptors during anoxia and NO toxicity were mediated through the cellular pathways of PKC or PKA in primary hippocampal neurons. Pretreatment with the metabotropic glutamate receptor agonists (±)-1-aminocyclopentane- trans -1,3-dicarboxylic acid, (1 S ,3 R )-1-aminocyclopentane-1,3-dicarboxylic acid (1 S ,3 R -ACPD), and l (+)-2-amino-4-phosphonobutyric acid ( l -AP4) 1 h before anoxia or NO exposure increased hippocampal neuronal cell survival from ∼30 to 70%. In addition, posttreatment with 1 S ,3 R -ACPD or l -AP4 up to 6 h following an insult attenuated anoxic- or NO-induced neurodegeneration. In contrast, treatment with l -(+)-2-amino-3-phosphonopropionic acid, an antagonist of the metabotropic glutamate receptor, did not significantly alter neuronal survival during anoxia or NO exposure. Protection by the ACPD-sensitive metabotropic receptors, such as the subtypes mGluR1α, mGluR2, and mGluR5, appears to be dependent on the modulation of PKC activity. In contrast, l -AP4-sensitive metabotropic glutamate receptors, such as the subtype mGluR4, may increase neuronal survival through PKA rather than PKC. Thus, activation of specific metabotropic glutamate receptors is protective during anoxia and NO toxicity, but the signal transduction pathways mediating protection differ among the metabotropic glutamate receptor subtypes. 相似文献
4.
Abstract: KCl-evoked glutamate exocytosis from cerebrocortical synaptosomes can be inhibited by the adenosine A1 receptor agonist cyclohexyladenosine (CHA). Inhibition is associated with a decreased KCl-evoked Ca2+ level elevation, and the effect of the agonist is occluded by prior incubation with the Agelenopsis aperta neurotoxin ω-agatoxin-IVA at 250 n M . The inhibition is suppressed in the presence of 3 n M phorbol dibutyrate (PDBu) or by activation of the protein kinase C (PKC)-coupled metabotropic glutamate receptor by 100 µ M (1 S ,3 R )-1-aminocyclopentane-1,3-dicarboxylate [(1 S ,3 R )ACPD]. A tonic inhibition of release by leaked exogenous adenosine can be reversed by adenosine deaminase or by PDBu addition. The CHA-induced inhibition can be enhanced by the PKC inhibitor Ro 31-8220. The mechanism for the suppression of the adenosine A1 receptor-mediated inhibition is distinct from that previously described for the (1 S ,3 R )ACPD-evoked, PKC-mediated, facilitatory pathway, which enhances phosphorylation of the MARCKS protein, 4-aminopyridine-induced action potentials, and release of glutamate because the latter requires at least 100 n M PDBu [or the combination of (1 S ,3 R )ACPD and arachidonic acid] and is not seen following KCl depolarization. Both PKC-mediated pathways may be involved in the presynaptic events associated with the establishment of synaptic plasticity. 相似文献
5.
Abstract: The ability of various stimuli to convert Ca2+/calmodulin-dependent protein kinase II (CaMKII) into a Ca2+-independent (autonomous) form was examined in cultured embryonic rat hippocampal pyramidal neurons. The most effective stimulation by far was observed when cells were equilibrated in buffer containing low extracellular [Ca2+] ([Ca2+]o) (~50 nM) and then shifted to normal [Ca2+]o (~1.26 mM) by addition of CaCl2 (referred to as “Ca2+ stimulation”). Virtually complete (>90%) conversion of the kinase to the autonomous form occurred within 30–50 s, with a return to baseline within 5 min. By contrast, depolarization of cells with high [K+] or treatment with glutamate or a Ca2+ ionophore caused insignificant increases (<10%) in levels of the autonomous form. The Ca2+-stimulated increase in CaMKII autonomy coincided with a two- to threefold increase in kinase subunit phosphorylation. In the first 40 s of Ca2+ stimulation, 32P incorporation into the immunoprecipitated subunits of CaMKII occurred exclusively on threonine residues, including Thr286Thr287 of the α/β subunits. Longer incubation of cells resulted in a decline of phosphothreonine content, whereas levels of phosphoserine-containing peptides showed a significant increase. The activation of CaMKII by Ca2+ stimulation was accompanied by only a small rise in intracellular [Ca2+]. Inhibitor studies showed that Na+-dependent action potentials and Ca2+ influx through glutamate receptors or voltage-sensitive Ca2+ channels did not contribute to the activation. Moreover, CaMKII was not activated by extracellular addition of other cations, including Mn2+, Mg2+, Co2+, or Gd3+. Although the mechanism of Ca2+ stimulation is presently unclear, it may involve either activation of extracellular calcium receptors or capacitative calcium entry. The dramatic rise in CaMKII autonomy and the Ca2+ selectivity of the response suggest a direct and specific relationship between [Ca2+]o and the state of activation of the kinase in intact neurons. 相似文献
6.
Activation of Mitogen-Activated Protein Kinase in Cultured Rat Hippocampal Neurons by Stimulation of Glutamate Receptors 总被引:8,自引:4,他引:4
†Masahito Kurino Kohji Fukunaga †Yukitaka Ushio Eishichi Miyamoto 《Journal of neurochemistry》1995,65(3):1282-1289
Abstract: Mitogen-activated protein kinase (MAP kinase) was activated by stimulation of glutamate receptors in cultured rat hippocampal neurons. Ten micromolar glutamate maximally stimulated MAP kinase activity, which peaked during 10 min and decreased to the basal level within 30 min. Experiments using glutamate receptor agonists and antagonists revealed that glutamate stimulated MAP kinase through NMDA and metabotropic glutamate receptors but not through non-NMDA receptors. Glutamate and its receptor agonists had no apparent effect on MAP kinase activation in cultured cortical astrocytes. Addition of calphostin C, a protein kinase C (PKC) inhibitor, or down-regulation of PKC activity partly abolished the stimulatory effect by glutamate, but the MAP kinase activation by treatment with ionomycin, a Ca2+ ionophore, remained intact. Lavendustin A, a tyrosine kinase inhibitor, was without effect. In experiments with 32 P-labeled hippocampal neurons, MAP kinase activation by glutamate was associated with phosphorylation of the tyrosine residue located on MAP kinase. However, phosphorylation of Raf-1, the c- raf protooncogene product, was not stimulated by treatment with glutamate. Our observations suggest that MAP kinase activation through glutamate receptors in hippocampal neurons is mediated by both the PKC-dependent and the Ca2+ -dependent pathways and that the activation of Raf-1 is not involved. 相似文献
7.
Phosphorylation of Mitogen-Activated Protein Kinase in Cultured Rat Cortical Glia by Stimulation of Metabotropic Glutamate Receptors 总被引:6,自引:3,他引:3
Abstract: Activation of metabotropic glutamate receptors (mGluRs) in glia results in significant physiological effects for both the glia and the neighboring neurons; but in many cases, the mGluR subtypes and signal transduction mechanisms mediating these effects have not been determined. In this study, we report that mGluR activation in primary cultures of rat cortical glia results in tyrosine phosphorylation of several proteins, including p44/p42 mitogen-activated protein kinases, also referred to as extracellular signal-regulated kinases (ERK1/2). Incubation of glial cultures with the general mGluR agonist 1-aminocyclopentane-1 S ,3 R -dicarboxylate and the mGluR group I-selective agonists ( RS )-3,5-dihydroxyphenylglycine (DHPG) and l -quisqualate resulted in increased tyrosine phosphorylation of ERK1/2. The group II-selective agonist (2 S ,2' R ,3' R )-2-(2',3'-dicarboxycyclopropyl)glycine and group III-selective agonist l (+)-2-amino-4-phosphonobutyric acid had no effect on tyrosine phosphorylation. DHPG-induced ERK1/2 phosphorylation could be inhibited by an antagonist that acts at group I or group II mGluRs but not by antagonists for group II and group III mGluRs. Protein kinase C (PKC) activators also induced ERK1/2 phosphorylation, but the PKC inhibitor bisindolylmaleimide I did not inhibit DHPG-induced ERK1/2 phosphorylation at a concentration that inhibited the response to phorbol 12,13-dibutyrate. These data suggest that mGluR activation of ERK1/2 in cultured glia is mediated by group I mGluRs and that this effect is independent of PKC activation. Furthermore, immunoblots with antibodies against various mGluR subtypes show expression of mGluR5, but no other mGluRs in our cultures. Taken together, these results suggest that mGluR5 stimulation results in tyrosine phosphorylation of ERK1/2 and other glial proteins. 相似文献
8.
Chronic Lithium-Induced Down-Regulation of MARCKS in Immortalized Hippocampal Cells: Potentiation by Muscarinic Receptor Activation 总被引:2,自引:0,他引:2
Abstract: Previous studies in our laboratory have demonstrated that exposure of rats to chronic lithium results in a significant reduction in the hippocampus of levels of the protein kinase C (PKC) phosphoprotein substrate MARCKS (myristoylated alanine-rich C kinase substrate), which persists after withdrawal and is not observed following acute administration. In an immortalized hippocampal cell line (HN33), we have determined that phorbol esters rapidly down-regulate PKC activity and lead to a subsequent PKC-dependent reduction in content of MARCKS protein. We now report that chronic exposure of HN33 cells to LiCl (1–10 m M ) produces a dose- and time-dependent down-regulation of MARCKS protein. The lithium-induced reduction in MARCKS is dependent on the concentration of inositol present in the medium and is reversed and prevented in the presence of elevated inositol concentrations. When HN33 cells were exposed to lithium at clinically relevant concentrations (1 m M ) under limiting inositol conditions, activation of muscarinic receptor-coupled phosphoinositide signaling significantly potentiated the lithium-induced down-regulation of MARCKS protein. It has been suggested that a major action of lithium in the brain is linked to its inositol monophosphatase inhibitory activity in receptor-mediated signaling through the inositol trisphosphate/diacylglycerol pathway, resulting in a relative inositol depletion. Our data provide evidence that this initial action of lithium may translate into a PKC-dependent long-term down-regulation of MARCKS protein expression in the hippocampus. 相似文献
9.
Abstract: Neurotransmitter receptors that increase phosphatidylinositol hydrolysis generate second messengers that activate protein kinase C. Here, we used metabotropic glutamate receptor agonists to increase both phosphatidylinositol hydrolysis and secretion of the soluble extracellular fragment of amyloid precursor protein (APPs) from cortical astrocyte cultures. The increase in APPs secretion was mimicked by direct activation of protein kinase C with phorbol ester and was suppressed by the metabotropic glutamate receptor antagonist l -(+)-2-amino-3-phosphonopropionic acid or by the protein kinase C inhibitor GF109203X. Ionotropic glutamate agonists did not increase APPs secretion. Forskolin or dibutyryl cyclic AMP inhibited the increase in APPs secretion caused by metabotropic glutamate receptor agonists or by phorbol ester treatment but did not affect basal APPs levels. Therefore, glutamatergic agonists that increase protein kinase C activation or decrease cyclic AMP formation may enhance the conversion of full-length APP to nonamyloidogenic APPs in Alzheimer's disease. 相似文献
10.
11.
J. L. Weiner C. F. Valenzuela P. L. Watson †C. J. Frazier †‡T. V. Dunwiddie 《Journal of neurochemistry》1997,68(5):1949-1959
Abstract: The ability of ethanol to enhance GABAA receptor function remains controversial; conflicting observations have been made even in the same brain region, and when using apparently similar methodologies. In this study we characterized a single protocol variable, the initial incubation temperature of brain slices, that had dramatic effects on the ethanol sensitivity of GABAA inhibitory postsynaptic currents (IPSCs) recorded from rat hippocampal CA1 pyramidal neurons. Incubation of hippocampal slices at relatively low temperatures (11–15°C) immediately after slice preparation significantly affected a number of physiological and biochemical parameters. Such slices showed a decrease in extracellular inhibitory postsynaptic potential amplitude, a significant increase in the ethanol sensitivity of GABAA IPSCs in CA1 pyramidal neurons, no change in pentobarbital or flunitrazepam potentiation of IPSCs, and an increase in basal protein kinase C (PKC) activity relative to slices incubated at 31–33°C. In addition, the increase in ethanol sensitivity of GABAA IPSCs was blocked by chelerythrine, a selective inhibitor of PKC. These results suggest that differences in hippocampal slice incubation protocols may have contributed to the disparate results of previous investigations of ethanol modulation of GABAA receptor-mediated synaptic transmission in the rat hippocampus. In addition, these findings provide further evidence that PKC activity positively modulates the interaction between ethanol and GABAA receptors in the mammalian brain. 相似文献
12.
蛋白激酶C对大鼠缺血海马突触体谷氨酸摄取的调控作用 总被引:1,自引:0,他引:1
采用大鼠海马脑片体外缺血模型,观察海马突触体内蛋白激酶C(PKC)活性的变化,以及这种变化对突触体谷氨酸(GLU)摄取的影响。结果显示:海马脑片体外“缺血”10min,其突触体内PKC活性基本不变,而缺血30min,突触体内PKC活性显著上升(P<0.01,n=6);非N-甲基-D-天门冬氨酸(NMDA)受体拮抗剂DNQX有效地抑制PKC活性的同时,可降低胞外GLU的堆积,而NMDA受体阻断剂AP_5无作用。进一步实验证明,PKC激动剂PDB浓度依赖性地抑制突触体对3H-GLU的摄取(IC50=131±10μmol/L),此抑制作用可由PKC抑制剂H-7(100μmol/L)抵消。提示脑缺血诱发GLU堆积的作用机理可能是:脑缺血引发钙内流导致GLU过量释放,GLU又通过突触前非NMDA受体激活PKC,抑制其自身摄取,正反馈性加重胞外GLU的堆积。 相似文献
13.
Alberto Alcázar Cristina Martín de la Vega Eulalia Bazán Juan L. Fando Matilde Salinas 《Journal of neurochemistry》1997,69(4):1703-1708
Abstract: Protein synthesis plays an important role in the viability and function of the cell. There is evidence indicating that Ca2+ may be a physiological regulator of the translational process. In the present study, the effect of agents that increase intracellular calcium levels by different mechanisms, as well as repercussion on the rate of protein synthesis, including phosphorylation of initiation factor 2α subunit, and double-stranded RNA-dependent eIF-2α kinase (PKR) activity were analyzed. Glutamate (100 µ M ) and K+ (60 m M ), which increase intracellular calcium levels (the former mostly by the influx of extracellular calcium via voltage-sensitive calcium channels, and the latter by receptor-operated calcium channels), and carbachol (1 m M ), as well as glutamate, which mobilizes intracellular calcium from the endoplasmic reticulum via activation of inositol 1,4,5-trisphosphate receptor, did not modify any of the analyzed parameters. Nevertheless, 100 n M ryanodine, which increases intracellular calcium concentration by activating the ryanodine receptor, promoted a significant decrease in the rate of protein synthesis and increased both initiation factor 2α subunit phosphorylation and PKR activity. From our results, we can conclude that inhibition of protein synthesis is dependent on the mobilization of intracellular calcium from internal stores. Moreover, they strongly suggest that this inhibition is only promoted when calcium is increased via ryanodine receptor, and possibly by activation of PKR activity. 相似文献
14.
Presynaptic GABAB Receptor Modulation of Glutamate Exocytosis from Rat Cerebrocortical Nerve Terminals: Receptor Decoupling by Protein Kinase C 总被引:1,自引:1,他引:0
Abstract: GABA and the GABAB receptor agonist (−)-baclofen inhibited 4-aminopyridine (4AP)- and KCl-evoked, Ca2+ -dependent glutamate release from rat cerebrocortical synaptosomes. The GABAB receptor antagonist CGP 35348, prevented this inhibition of glutamate release, but phaclofen had no effect. (−)-Baclofen-mediated inhibition of glutamate release was insensitive to 2 µg/ml pertussis toxin. As determined by examining the mechanism of GABAB receptor modulation of glutamate release, (−)-baclofen caused a significant reduction in 4AP-evoked Ca2+ influx into synaptosomes. The agonist did not alter the resting synaptosomal membrane potential or 4AP-mediated depolarization; thus, the inhibition of Ca2+ influx could not be attributed to GABAB receptor activation causing a decrease in synaptosomal excitability. Ionomycin-mediated glutamate release was not affected by (−)-baclofen, indicating that GABAB receptors in this preparation are not coupled directly to the exocytotic machinery. Instead, the data invoke a direct coupling of GABAB receptors to voltage-dependent Ca2+ channels linked to glutamate release. This coupling was subject to regulation by protein kinase C (PKC), because (−)-baclofen-mediated inhibition of 4AP-evoked glutamate release was reversed when PKC was stimulated with phorbol ester. This may therefore represent a mechanism by which inhibitory and facilitatory presynaptic receptor inputs interplay to fine-tune transmitter release. 相似文献
15.
Jean Christophe Deloulme Thierry Janet Brigitte Pettmann Pascal Laeng Marie-France Knoetgen Monique Sensenbrenner Jacques Baudier 《Journal of neurochemistry》1992,58(2):567-578
Basic fibroblast growth factor (bFGF) is a well-characterized peptide hormone that has mitogenic activity for various cell types and elicits a characteristic set of responses on the cell types investigated. In this report we confirmed that bFGF is a potent mitogen for rat brain-derived oligodendrocyte (OL) precursor cells as well as for differentiated OL in secondary culture. bFGF was shown to induce expression of the protooncogene c-fos in OL. The role of protein kinase C (PKC) in mediating bFGF-stimulated proliferation as well as c-fos expression in OL was investigated. The PKC activator phorbol 12-myristate 13-acetate (PMA) stimulated c-fos expression but did not trigger cell proliferation. When PKC was down-regulated by pretreatment of OL with PMA for 20 h, the bFGF-mediated stimulations of OL proliferation and c-fos mRNA expression were still observed, whereas the induction of c-fos mRNA by PMA was totally inhibited. These data demonstrate that the bFGF mitogenic signaling pathway in OLs does not require PKC. On the other hand, bFGF was found to stimulate specifically the phosphorylation of a limited number of PKC substrates in oligodendroglial cells, including the MARCKS protein. The bFGF-dependent phosphorylation of MARCKS protein was totally inhibited when PKC was first down-regulated, indicating that the phosphorylation of this protein is PKC dependent. Tryptic digestion of the phosphorylated MARCKS protein revealed that bFGF stimulated specifically the phosphorylation of the MARCKS protein on a single phosphopeptide. We provide evidence that bFGF also stimulated fatty acylation of the MARCKS protein, which might explain the observed specific bFGF-dependent phosphorylation of this protein in OL. We propose that bFGF-dependent fatty acylation and phosphorylation of the MARCKS protein are not essential for the transduction of the bFGF mitogenic signal but are probably linked to differentiation processes elicited by bFGF on OL. 相似文献
16.
Khoa T. D. Nguyen Jae-Won Shin Caroline Rae Ellas K. Nanitsos Gabriela B. Acosta David V. Pow Vlado Buljan Maxwell R. Bennett Paul L. Else Vladimir J. Balcar 《Neurochemical research》2009,34(10):1767-1774
The naturally occurring toxin rottlerin has been used by other laboratories as a specific inhibitor of protein kinase C-delta
(PKC-δ) to obtain evidence that the activity-dependent distribution of glutamate transporter GLAST is regulated by PKC-δ mediated
phosphorylation. Using immunofluorescence labelling for GLAST and deconvolution microscopy we have observed that d-aspartate-induced redistribution of GLAST towards the plasma membranes of cultured astrocytes was abolished by rottlerin.
In brain tissue in vitro, rottlerin reduced apparent activity of (Na+, K+)-dependent ATPase (Na+, K+-ATPase) and increased oxygen consumption in accordance with its known activity as an uncoupler of oxidative phosphorylation
(“metabolic poison”). Rottlerin also inhibited Na+, K+-ATPase in cultured astrocytes. As the glutamate transport critically depends on energy metabolism and on the activity of
Na+, K+-ATPase in particular, we suggest that the metabolic toxicity of rottlerin and/or the decreased activity of the Na+, K+-ATPase could explain both the glutamate transport inhibition and altered GLAST distribution caused by rottlerin even without
any involvement of PKC-δ-catalysed phosphorylation in the process. 相似文献
17.
18.