Phosphorylation of p38 MAPK Induced by Oxidative Stress Is Linked to Activation of Both Caspase-8- and -9-mediated Apoptotic Pathways in Dopaminergic Neurons

Abstract

We evaluated the contribution of p38 mitogen-activated protein kinase and the events upstream/downstream of p38 leading to dopaminergic neuronal death. We utilized MN9D cells and primary cultures of mesencephalic neurons treated with 6-hydroxydopamine. Phosphorylation of p38 preceded apoptosis and was sustained in 6-hydroxydopamine-treated MN9D cells. Co-treatment with PD169316 (an inhibitor of p38) or expression of a dominant negative p38 was neuroprotective in death induced by 6-hydroxydopamine. The superoxide dismutase mimetic and the nitric oxide chelator blocked 6-hydroxydopamine-induced phosphorylation of p38, suggesting a role for superoxide anion and nitric oxide in eliciting a neurotoxic signal by activating p38. Following 6-hydroxydopamine treatment, inhibition of p38 prevented both caspase-8- and -9-mediated apoptotic pathways as well as generation of truncated Bid. Consequently, 6-hydroxydopamine-induced cell death was rescued by blockading activation of caspase-8 and -9. In primary cultures of mesencephalic neurons, the phosphorylation of p38 similarly appeared in tyrosine hydroxylase-positive, dopaminergic neurons after 6-hydroxydopamine treatment. This neurotoxin-induced phosphorylation of p38 was inhibited in the presence of superoxide dismutase mimetic or nitric oxide chelator. Co-treatment with PD169316 deterred 6-hydroxydopamine-induced loss of dopaminergic neurons and activation of caspase-3 in these neurons. Furthermore, inhibition of caspase-8 and -9 significantly rescued 6-hydroxydopamine-induced loss of dopaminergic neurons. Taken together, our data suggest that superoxide anion and nitric oxide induced by 6-hydroxydopamine initiate the p38 signal pathway leading to activation of both mitochondrial and extramitochondrial apoptotic pathways in our culture models of Parkinson's disease. We evaluated the contribution of p38 mitogen-activated protein kinase and the events upstream/downstream of p38 leading to dopaminergic neuronal death. We utilized MN9D cells and primary cultures of mesencephalic neurons treated with 6-hydroxydopamine. Phosphorylation of p38 preceded apoptosis and was sustained in 6-hydroxydopamine-treated MN9D cells. Co-treatment with PD169316 (an inhibitor of p38) or expression of a dominant negative p38 was neuroprotective in death induced by 6-hydroxydopamine. The superoxide dismutase mimetic and the nitric oxide chelator blocked 6-hydroxydopamine-induced phosphorylation of p38, suggesting a role for superoxide anion and nitric oxide in eliciting a neurotoxic signal by activating p38. Following 6-hydroxydopamine treatment, inhibition of p38 prevented both caspase-8- and -9-mediated apoptotic pathways as well as generation of truncated Bid. Consequently, 6-hydroxydopamine-induced cell death was rescued by blockading activation of caspase-8 and -9. In primary cultures of mesencephalic neurons, the phosphorylation of p38 similarly appeared in tyrosine hydroxylase-positive, dopaminergic neurons after 6-hydroxydopamine treatment. This neurotoxin-induced phosphorylation of p38 was inhibited in the presence of superoxide dismutase mimetic or nitric oxide chelator. Co-treatment with PD169316 deterred 6-hydroxydopamine-induced loss of dopaminergic neurons and activation of caspase-3 in these neurons. Furthermore, inhibition of caspase-8 and -9 significantly rescued 6-hydroxydopamine-induced loss of dopaminergic neurons. Taken together, our data suggest that superoxide anion and nitric oxide induced by 6-hydroxydopamine initiate the p38 signal pathway leading to activation of both mitochondrial and extramitochondrial apoptotic pathways in our culture models of Parkinson's disease. Parkinson's disease (PD) 1The abbreviations used are: PD, Parkinson's disease; JNK, c-Jun N-terminal kinases; MPTP, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine; MPP+, 1-methyl-4-phenylpyridinium; NO, nitric oxide; 6-OHDA, 6-hydroxydopamine; BAF, t-butoxycarbonyl-(Asp)-fluoromethyl ketone; Z, benzyloxycarbonyl; fmk, fluoromethyl ketone; AMC, 7-amino-4-methylcoumarin; AFC, 7-amino-4-trifluoromethylcoumarin; pNA, p-nitroanaline; DAF-FM diacetate, 4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate; Mn-TBAP, Mn(III)tetrakis(4-benzoic acid)porphyrin chloride; carboxy-PTIO, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimida-zoline-1-oxyl-3-oxide; MTT, 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide; LDH, lactate dehydrogenase; ROS, reactive oxygen species; TH, tyrosine hydroxylase; phospho-p38, phosphorylation of p38; ANOVA, analysis of variance; CHAPS, 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonic acid. 1The abbreviations used are: PD, Parkinson's disease; JNK, c-Jun N-terminal kinases; MPTP, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine; MPP+, 1-methyl-4-phenylpyridinium; NO, nitric oxide; 6-OHDA, 6-hydroxydopamine; BAF, t-butoxycarbonyl-(Asp)-fluoromethyl ketone; Z, benzyloxycarbonyl; fmk, fluoromethyl ketone; AMC, 7-amino-4-methylcoumarin; AFC, 7-amino-4-trifluoromethylcoumarin; pNA, p-nitroanaline; DAF-FM diacetate, 4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate; Mn-TBAP, Mn(III)tetrakis(4-benzoic acid)porphyrin chloride; carboxy-PTIO, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimida-zoline-1-oxyl-3-oxide; MTT, 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide; LDH, lactate dehydrogenase; ROS, reactive oxygen species; TH, tyrosine hydroxylase; phospho-p38, phosphorylation of p38; ANOVA, analysis of variance; CHAPS, 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonic acid. is a common neurodegenerative disorder characterized by a progressive loss of dopaminergic neurons in the substantia nigra. Although its precise etiology is unknown, such factors as oxidative stress, impairment of mitochondrial respiration, and abnormal protein aggregation may play roles in its pathogenesis (1Dauer W. Przedborski S. Neuron. 2003; 39: 889-909Abstract Full Text Full Text PDF PubMed Scopus (4057) Google Scholar, 2Jenner P. Ann. Neurol. 2003; 53: S26-S38Crossref PubMed Scopus (1701) Google Scholar). Although the role of apoptosis in the process of dopaminergic neuronal death has been highlighted in studies using postmortem brains and experimental models of PD, other evidence implicates both apoptosis and non-apoptotic death in PD (3Hartmann A. Hirsch E.C. Adv. Neurol. 2001; 86: 143-153PubMed Google Scholar). Regardless, it is not clearly understood which signals may be involved in determining the mode of dopaminergic neuronal death. Among the upstream signals leading to neuronal degeneration, one may include the stress-activated protein kinases. These are c-Jun N-terminal kinases (JNK) and p38 kinases, which are activated in response to various stimuli and are potent effectors of neuronal apoptosis (4Mielke K. Herdegen T. Prog. Neurobiol. 2000; 61: 45-60Crossref PubMed Scopus (437) Google Scholar). Most interesting, the phosphorylated forms of JNK and p38 are expressed in postmortem brains of PD (5Ferrer I. Blanco R. Carmona M. Puig B. Barrachina M. Gomez C. Ambrosio S. J. Neural. Transm. 2001; 108: 1383-1396Crossref PubMed Scopus (142) Google Scholar). Moreover, evidence from experimental models of PD has implicated JNK in the process of dopaminergic neuronal death. For example, activation of JNK has been demonstrated in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. Blockade of JNK by JNK-interacting protein-1 or by a synthetic inhibitor of the mixed lineage kinase family is neuroprotective (6Saporito M.S. Brown E.M. Miller M.S. Carswell S. J. Pharmacol. Exp. 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Blockade of JNK in cultures has also been shown to be neuroprotective (12Hashimoto M. Hsu L.J. Rockenstein E. Takenouchi T. Mallory M. Masliah E. J. Biol. Chem. 2002; 277: 11465-11472Abstract Full Text Full Text PDF PubMed Scopus (171) Google Scholar, 13Sawada H. Ibi M. Kihara T. Honda K. Nakamizo T. Kanki R. Nakanishi M. Sakka N. Akaike A. Shimohama S. Neuropharmacology. 2002; 42: 1056-1064Crossref PubMed Scopus (97) Google Scholar). Although its role is less defined, p38 activation has been investigated following treatment with dopamine, MPTP, or MPP+ (14De Girolamo L.A. Hargreaves A.J. Billett E.E. J. Neurochem. 2001; 76: 650-660Crossref PubMed Scopus (45) Google Scholar, 15Du Y. Ma Z. Lin S. Dodel R.C. Gao F. Bales K.R. Triarhou L.C. Chernet E. Perry K.W. Nelson D.L. Luecke S. Phebus L.A. Bymaster F.P. Paul S.M. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 14669-14674Crossref PubMed Scopus (690) Google Scholar, 16Junn E. Mouradian M.M. J. Neurochem. 2001; 78: 374-383Crossref PubMed Scopus (191) Google Scholar). As typified in the MPTP mouse model and in MPP+-treated culture models, protection of neuronal degeneration by minocycline is mediated by inhibiting drug-induced glial inducible NOS expression and nitric oxide (NO)-induced phosphorylation of p38 (15Du Y. Ma Z. Lin S. Dodel R.C. Gao F. Bales K.R. Triarhou L.C. Chernet E. Perry K.W. Nelson D.L. Luecke S. Phebus L.A. Bymaster F.P. Paul S.M. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 14669-14674Crossref PubMed Scopus (690) Google Scholar). However, a role for the p38 signal in dopaminergic neuronal degeneration is controversial (17Gomez-Santos C. Ferrer I. Reiriz J. Vinals F. Barrachina M. Ambrosio S. Brain Res. 2002; 935: 32-39Crossref PubMed Scopus (134) Google Scholar, 18Halvorsen E.M. Dennis J. Keeney P. Sturgill T.W. Tuttle J.B. Benett Jr., J.B. Brain Res. 2002; 952: 98-110Crossref PubMed Scopus (54) Google Scholar, 19Shimoke K. Kudo M. Brain Res. Dev. Brain Res. 2002; 133: 105-114Crossref PubMed Scopus (11) Google Scholar). Furthermore, cellular events upstream and downstream of p38 activation leading to dopaminergic neuronal death are not yet understood. Here, we attempted to evaluate the following: (i) to what extent p38 contributes to 6-hydroxydopamine (6-OHDA)-induced death; (ii) whether p38 is linked to activation of the mitochondrial and extramitochondrial apoptotic pathways; and (iii) the temporal sequence of cell death with regard to oxidative stress, and the activation of p38 and caspases using MN9D dopaminergic neuronal cells and primary cultures of mesencephalic neurons. Our data show that superoxide anion and NO induced by 6-OHDA activate p38 that, in turn, activates both the caspase-8 and -9-dependent apoptotic pathways in culture models of PD. Materials—t-Butoxycarbonyl-(Asp)-fluoromethyl ketone (BAF), Z-LEHD-fmk, and Z-IETD-fmk from and from The for using was from The reactive oxygen used superoxide anion and 4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate; a nitric oxide and from of p38 was from Mn(III)tetrakis(4-benzoic acid)porphyrin a superoxide dismutase was from and a nitric oxide was from and from other 6-OHDA and from and dopaminergic neuronal cells by a mesencephalic neurons and cells L.A. A. Brain Res. PubMed Scopus Google Scholar). MN9D cells with a expression dominant negative p38 and B. J. T. J. PubMed Scopus Google and in the presence of as J. Neurochem. 2001; PubMed Scopus Google Scholar). MN9D and cells culture or in with in of for and to 6-OHDA in the presence or the of various experimental Following treatment, the of cell was by the as J. Neurochem. 2001; PubMed Scopus Google Scholar). The of cell death was by the of lactate by cells the J. PubMed Scopus Google Scholar). or primary cultures of dopaminergic neurons treated with 6-OHDA in the presence or the of various Following treatment, cells with in a for and with a a of from was a and for used that p38, phosphorylated of p38, forms of and -9 used of caspase-8 and of a from S. J. the For the mitochondrial of the cells treated with 6-OHDA in the presence or the of the and as with A. J. J. PubMed Scopus Google Scholar). used was a mouse mouse with was used to the expression of dominant negative of p38. For a mouse and a by of cells and cells with 6-hydroxydopamine or in with with for and with a as J. Res. 1999; PubMed Scopus Google Scholar). also with DAF-FM for with and in for for or DAF-FM using a was by a or following treatment with 6-OHDA in the presence or the of MN9D cells in a with for for or for for The in a for or or the by the for was for and for or with a for using a of and of primary cultures of mesencephalic neurons, mesencephalic from the brains of cells and as in our H.S. J. 2003; PubMed Google Scholar). or in cultures with to and treated with 6-OHDA or in with various experimental the of dopaminergic neurons following treatment, primary cultures to of as H.S. J. 2003; PubMed Google Scholar). by to the treatment of the neurons the of the of the phosphorylated of p38, MN9D cells with for in for and with a of p38 with cells with for For a of and phosphorylated of p38 or primary cultures of mesencephalic neurons treated with 6-OHDA or in with PD169316 as H.S. J. 2003; PubMed Google Scholar). The primary used a mouse a of p38 and a that caspase-3 Following with cultures with and for cells with and a by to the treatment of the phosphorylated forms of or neurons neurons. from the of of 6-OHDA treated and with various was by and of as of p38 Following 6-OHDA demonstrated J. Res. 1999; PubMed Scopus Google Scholar, H.S. J. 2003; PubMed Google Scholar, M. Neurobiol. PubMed Scopus (54) Google Scholar, 2003; PubMed Scopus Google that 6-OHDA of of MN9D cells found to be following treatment with 6-OHDA for As of was to whether and to what extent p38 to neurotoxin-induced cell death. As shown in phosphorylation of p38 in MN9D cells was as as following 6-OHDA treatment and to in a to after 6-OHDA treatment, of to The of p38 the As shown in also that MN9D cells for following 6-OHDA treatment. Among the of PD169316 (an inhibitor of p38; we found the of p38 inhibitor that blocked generation of The of was significantly blocked in the presence of PD169316 or in MN9D cells a dominant negative p38 and Co-treatment with PD169316 or expression of a dominant negative p38 significantly blocked cell death as by and and of p38 of the and of Following 6-OHDA and J. Res. 1999; PubMed Scopus Google Scholar, H.S. J. 2003; PubMed Google Scholar, M. Neurobiol. PubMed Scopus (54) Google Scholar, 2003; PubMed Scopus Google Scholar, J. J. 1999; PubMed Google the that generation of and signal pathways play role in cell death in MN9D cells and primary cultures of and mesencephalic neurons. We that induced by 6-OHDA contributes to the of in MN9D cells. we of by using superoxide anion J. Miller J. PubMed Google and DAF-FM (an NO Y. Ma H. M. K. Y. H. T. N. 2000; PubMed Scopus Google following 6-OHDA treatment in the presence or the of PD169316 or in cells. As shown in of superoxide anion and NO induced of studies demonstrated that with PD169316 or expression of a dominant negative p38 not the of superoxide anion and NO generation following 6-OHDA treatment for to We whether with the of As by analysis of following 6-OHDA treatment was inhibited in the presence of a superoxide dismutase mimetic or by NO that one of the of the p38 activation is involved in MN9D cell we attempted to the temporal cell death in to p38 and As shown in with a not the of with inhibitor was to and -9-mediated Following 6-OHDA has been demonstrated that both caspase-8 and -9 are the that to activation of of caspases and 2000; PubMed Scopus Google Scholar). we the of as upstream of activation following 6-OHDA treatment, we attempted to whether may be linked to the activation of the caspase-8 and -9-mediated apoptotic in MN9D cells treated with 6-OHDA for various using and or in the presence or the of As shown in following 6-OHDA treatment, and the in a to Co-treatment with PD169316 significantly blocked activation of these p38 signal the activation of caspases following 6-OHDA treatment, cellular to analysis using the forms of As shown in of the of caspase-8 appeared as as following 6-OHDA treatment and to Although of the forms of appeared as as following 6-OHDA treatment, a in the forms of by In analysis that the of forms of caspase-3 and following 6-OHDA treatment. of p38 signal by PD169316 or by expression of a dominant negative p38 generation of forms of caspases with a and by and Following 6-OHDA response to and various the and the activation of the mitochondrial apoptotic In of mitochondrial of the and to the activation of 2000; PubMed Scopus Google Scholar). MN9D cells treated with 6-OHDA in the presence or the of The of was by following 6-OHDA treatment and to Co-treatment with PD169316 inhibited generation of of was also inhibited in the presence of Z-IETD-fmk caspase-8 not by that activation of caspase-8 to generation of following 6-OHDA treatment. As shown in and of the was inhibited by PD169316 not by Although it to be that following 6-OHDA treatment, the mitochondrial of the may be induced by and Co-treatment of MN9D cells with Z-IETD-fmk or with cells from death as by the and in Although it was not the extent of the protection was Z-IETD-fmk and and with that in the This that the of the p38 signal induced by 6-OHDA is linked to caspase-8 and -9-mediated apoptotic pathways in MN9D with caspase-8 inhibitor inhibitor is neuroprotective in MN9D cell death. MN9D cells treated with 6-OHDA for in the presence or the of Z-IETD-fmk the of and or was by the from treatment expressed as a of the the from in with p38 in of Following 6-OHDA we the temporal sequence of cell death in MN9D cells with regard to ROS, and activation of p38 and we to these using primary cultures of mesencephalic neurons. As we demonstrated in our H.S. J. 2003; PubMed Google treatment with 6-OHDA for in a loss of of neurons. of was for As shown in of was in a In a of and was in primary cultures of mesencephalic neurons treated with of or Following 6-OHDA treatment, neurons for The neurons neurons by following 6-OHDA treatment of neurons the cultures for of cells for in of glial cells for of p38 in of Following 6-OHDA our H.S. J. 2003; PubMed Google we demonstrated that with the loss of neurons induced by we whether the of and inhibition of p38 loss of neurons. As shown in the of neurons induced by 6-OHDA treatment was in the presence of such as Mn-TBAP, and and for 6-OHDA Furthermore, loss of neurons was significantly blocked in the presence of PD169316 Among the of PD169316 we found PD169316 the protection PD169316 significantly inhibited the loss of neurons Although was in neurons, PD169316 significantly rescued neurons. This may that a of neurons is p38 activation of p38 in of Following 6-OHDA whether inhibition of the p38 signal primary cultures of mesencephalic neurons treated with 6-OHDA in the presence or the of of the and cultures for by a of and of and the of caspase-3 prevented in the presence of As demonstrated in our H.S. J. 2003; PubMed Google of the neurons neurons following 6-OHDA treatment for of neurons for We investigated whether the caspase-8 and -9 pathways are involved in apoptosis in primary cultures of mesencephalic neurons. Among the of and -9 inhibitor we found of inhibitor the This is that used by in primary cultures of mesencephalic neurons A. S. K. B.A. A. M. Y. Hirsch E.C. J. 2001; PubMed Google Scholar). Z-IETD-fmk or significantly blocked loss of neurons and neurons in 6-OHDA Although not of caspase-8 and -9 protection This protection was to that by PD169316 by H.S. J. 2003; PubMed Google of caspase-8 -9 is neuroprotective following 6-OHDA treatment. cultures of mesencephalic neurons treated with 6-OHDA for in the presence or of Z-IETD-fmk or or in for TH, and the of neurons was from treatment expressed as a of the the from with We demonstrated that 6-OHDA apoptosis in the MN9D dopaminergic neuronal cell J. Res. 1999; PubMed Scopus Google Scholar, M. Neurobiol. PubMed Scopus (54) Google in primary cultures of dopaminergic neurons H.S. J. 2003; PubMed Google and in neurons 2003; PubMed Scopus Google Scholar). our we that the death signal by superoxide anion and NO a role in apoptosis in dopaminergic neurons. Furthermore, we demonstrated that the p38 signal in response to 6-OHDA is linked to activation of both and apoptotic Most interesting, in the of forms of and preceded the of forms of by This as both caspase-8- and apoptotic pathways to activate the of and 2000; PubMed Scopus Google Scholar). In generation of was blocked in the presence of caspase-8 We are to that the may play role in neuronal cell death. inhibition of the p38 signal significantly of activation and cell death. We the presence of death sequence with regard to activation of p38 and activation of caspase-8- and -9-mediated pathways in primary cultures of mesencephalic neurons treated with The in the of cell death protection and and inhibition of activation and MN9D cells with PD169316 and cells may the presence of of the p38 may be that a dominant negative of p38 not p38 activation in MN9D cells is that oxidative contributes to the leading to degeneration of dopaminergic neurons in PD. is evidence that oxidative in PD may from the of NO P. Ann. Neurol. 2003; 53: S26-S38Crossref PubMed Scopus (1701) Google Scholar). Among the of of NO, (15Du Y. Ma Z. Lin S. Dodel R.C. Gao F. Bales K.R. Triarhou L.C. Chernet E. Perry K.W. Nelson D.L. Luecke S. Phebus L.A. Bymaster F.P. Paul S.M. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 14669-14674Crossref PubMed Scopus (690) Google that NO phosphorylation of p38 and that the p38 inhibitor NO in neurons. inhibition of p38 apoptotic death in in primary cultures of neurons, and in cells S. S. Y. M. Z. J. Biol. 2000; PubMed Scopus Google Scholar, A. S. J. Biol. Chem. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). Although a a role of NO in cell death in cells W.D. J. 2003; PubMed Scopus Google our data that with NO and cell death a role of NO p38 pathway in dopaminergic neurons. Furthermore, we the that not NO also superoxide anion is for the activation of the p38 Although is evidence for superoxide anion and for its role in PD, of in and dopaminergic cells as well as cell death S. S. S. E. J. PubMed Google Scholar, M. H. PubMed Scopus Google Scholar, M. S. A. J. J. Neurochem. 2002; PubMed Scopus Google Scholar). in and is following of the pathway P. Brain Res. 2001; PubMed Scopus Google Scholar). In MN9D we J. Res. 1999; PubMed Scopus Google a of following In the with the superoxide dismutase mimetic significantly and cell death in MN9D cells and primary cultures of dopaminergic neurons. Most interesting, inhibition of superoxide anion or NO generation cells from death Although one that following 6-OHDA treatment, superoxide anion are also for the activation of the p38 studies that such as and stress-activated protein kinase and p38) is implicated in the in such as and disease M. J. 2002; PubMed Scopus Google Scholar, Perry M.A. 2002; PubMed Scopus Google Scholar). inhibition of p38 as a in and in various S. S. A.J. K. Res. 2001; PubMed Scopus Google Scholar, J. 2002; PubMed Scopus Google Scholar). However, is evidence to that the p38 signal is involved in the pathogenesis of PD is evidence for its role in models of dopaminergic neuronal For example, Ferrer (5Ferrer I. Blanco R. Carmona M. Puig B. Barrachina M. Gomez C. Ambrosio S. J. Neural. Transm. 2001; 108: 1383-1396Crossref PubMed Scopus (142) Google demonstrated that is found in in the of or in with or in a of neurons of the in PD. studies using culture models neurons, and mesencephalic dopaminergic the of a role for p38 in neuronal apoptosis (14De Girolamo L.A. Hargreaves A.J. Billett E.E. J. Neurochem. 2001; 76: 650-660Crossref PubMed Scopus (45) Google Scholar, 15Du Y. Ma Z. Lin S. Dodel R.C. Gao F. Bales K.R. Triarhou L.C. Chernet E. Perry K.W. Nelson D.L. Luecke S. Phebus L.A. Bymaster F.P. Paul S.M. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 14669-14674Crossref PubMed Scopus (690) Google Scholar, 16Junn E. Mouradian M.M. J. Neurochem. 2001; 78: 374-383Crossref PubMed Scopus (191) Google Scholar, H. T. Shimohama S. J. T. Y. E. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, K. W. H. T. M. K. Res. 1999; PubMed Scopus Google Scholar, P. J. Brain Res. 2001; PubMed Scopus Google Scholar, B. R.C. Ann. N. Y. Acad. Sci. 2002; PubMed Scopus Google Scholar). In these inhibition of the p38 signal activated by MPP+, dopamine, or is are whether activation of p38 and inhibition of the p38 signal a neuroprotective (17Gomez-Santos C. Ferrer I. Reiriz J. Vinals F. Barrachina M. Ambrosio S. Brain Res. 2002; 935: 32-39Crossref PubMed Scopus (134) Google demonstrated that, in with the in and apoptotic death in cells treated with MPP+, activation of not JNK and p38 is inhibition of the by Furthermore, inhibition of p38 signal in and cells cell death in response to treatment E.M. Dennis J. Keeney P. Sturgill T.W. Tuttle J.B. Benett Jr., J.B. Brain Res. 2002; 952: 98-110Crossref PubMed Scopus (54) Google Scholar, 19Shimoke K. Kudo M. Brain Res. Dev. Brain Res. 2002; 133: 105-114Crossref PubMed Scopus (11) Google Scholar). Although is not our data from MN9D cells and primary cultures of mesencephalic neurons of p38 activation and its role dopaminergic neuronal cell death. postmortem studies demonstrated that forms of caspase-8 are in dopaminergic neurons A. S. K. B.A. A. M. Y. Hirsch E.C. J. 2001; PubMed Google Scholar, Y. R. S. F. J. 2001; PubMed Google Scholar). Although it is not clearly understood whether activation of caspase-8 in PD is mediated by the cell death is evidence suggesting that caspase-8 be activated by of the death pathway in both in and in J. 2000; PubMed Scopus Google Scholar, N. S. S. A. Biol. 2001; PubMed Scopus Google Scholar, C. T. F. K. B. 2003; PubMed Scopus Google Scholar). and J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google demonstrated that death and and are in MN9D cells as by In our treatment with or with not MN9D cells As demonstrated N. S. S. A. Biol. 2001; PubMed Scopus Google activation of caspase-8 by p38 in our culture models of PD to be death Taken together, our role of p38 pathway leading to a of activation dopaminergic neuronal death. As a of the p38 pathway has been as one of that the of PD (15Du Y. Ma Z. Lin S. Dodel R.C. Gao F. Bales K.R. Triarhou L.C. Chernet E. Perry K.W. Nelson D.L. Luecke S. Phebus L.A. Bymaster F.P. Paul S.M. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 14669-14674Crossref PubMed Scopus (690) Google Scholar, H. P. J. R. J. 2003; PubMed Google Scholar). studies the to p38 activation to activation may our to for PD. As inhibition of p38 signal protection in our culture models of PD, of death is also We A. for with the MN9D cell