Nerve Growth Factor Promotes the Survival of Sympathetic Neurons through the Cooperative Function of the Protein Kinase C and Phosphatidylinositol 3-Kinase Pathways

Abstract

The signaling pathways activated by nerve growth factor (NGF) that account for its ability to promote the survival of neurons are not completely understood. Phosphatidylinositol 3-kinase (PI3K) is critical for the survival of several cell types, including neurons. To determine whether additional signaling pathways cooperate with PI3K to promote survival, we examined other pathways known to be activated by NGF. NGF activated protein kinases C (PKCs) in sympathetic neurons, and pharmacologic PKC activation rescued neurons from apoptosis induced by the withdrawal of NGF. Inhibition of PKCs did not inhibit the survival of NGF-maintained neurons. Similarly, inhibition of PI3K caused only a modest attrition of neurons in the presence of NGF. In contrast, the simultaneous inhibition of both PKCs and PI3K induced the apoptotic death of NGF-maintained sympathetic neurons. Inhibition of both PI3K and PKCs promoted the expression and phosphorylation of the proapoptotic transcription factor c-Jun, indicating that these pathways inhibit programmed cell death at the stage of proapoptotic gene expression. In culture conditions under which PI3K inhibition alone kills NGF-maintained neurons, PKC inhibition also led to a significant loss of viability, indicating that both pathways are required. Therefore, PKC and PI3K, regardless of the culture conditions, cooperate to promote the NGF-dependent survival of sympathetic neurons. The signaling pathways activated by nerve growth factor (NGF) that account for its ability to promote the survival of neurons are not completely understood. Phosphatidylinositol 3-kinase (PI3K) is critical for the survival of several cell types, including neurons. To determine whether additional signaling pathways cooperate with PI3K to promote survival, we examined other pathways known to be activated by NGF. NGF activated protein kinases C (PKCs) in sympathetic neurons, and pharmacologic PKC activation rescued neurons from apoptosis induced by the withdrawal of NGF. Inhibition of PKCs did not inhibit the survival of NGF-maintained neurons. Similarly, inhibition of PI3K caused only a modest attrition of neurons in the presence of NGF. In contrast, the simultaneous inhibition of both PKCs and PI3K induced the apoptotic death of NGF-maintained sympathetic neurons. Inhibition of both PI3K and PKCs promoted the expression and phosphorylation of the proapoptotic transcription factor c-Jun, indicating that these pathways inhibit programmed cell death at the stage of proapoptotic gene expression. In culture conditions under which PI3K inhibition alone kills NGF-maintained neurons, PKC inhibition also led to a significant loss of viability, indicating that both pathways are required. Therefore, PKC and PI3K, regardless of the culture conditions, cooperate to promote the NGF-dependent survival of sympathetic neurons. Nerve growth factor (NGF) 1The abbreviations used are: NGF, nerve growth factor; PI3K, phosphatidylinositol 3-kinase; PKC, protein kinase C; PMA, phorbol 12-myristate 13-acetate; BAF, boc-aspartyl fluoromethyl ketone; MAPK, mitogen-activated protein kinase; BSA, bovine serum albumin; TBS, Tris-buffered saline; P-PKC, phosphorylated PKC; PKD, protein kinase D. can maintain the survival of sympathetic neurons in vitro and is required for their target-dependent survival in vivo (1Levi-Montalcini R. Science. 1987; 237: 1154-1162Crossref PubMed Scopus (2701) Google Scholar, 2Oppenheim R.W. Annu. Rev. Neurosci. 1991; 14: 453-501Crossref PubMed Scopus (2766) Google Scholar). NGF promotes survival via activation of its receptor tyrosine kinase, TrkA (3Segal R.A. Greenberg M.E. Annu. Rev. Neurosci. 1996; 19: 463-489Crossref PubMed Scopus (908) Google Scholar, 4Bothwell M. Annu. Rev. Neurosci. 1995; 18: 223-253Crossref PubMed Scopus (767) Google Scholar). However, the signaling pathways that account for the survival promoting activities of NGF are not completely understood. Although phosphatidylinositol 3-kinase (PI3K) is important for the NGF-dependent survival of sympathetic neurons (5Kaplan D.R. Miller F.D. Curr. Opin. Neurobiol. 2000; 10: 381-391Crossref PubMed Scopus (1670) Google Scholar), whether PI3K is the only required signaling pathway is unresolved. Under certain culture conditions, PI3K appears to be necessary and sufficient for NGF-dependent survival (6Crowder R.J. Freeman R.S. J. Neurosci. 1998; 18: 2933-2943Crossref PubMed Google Scholar, 7Mazzoni I.E. Said F.A. Aloyz R. Miller F.D. Kaplan D. J. Neurosci. 1999; 19: 9716-9727Crossref PubMed Google Scholar, 8Vaillant A.R. Mazzoni I. Tudan C. Boudreau M. Kaplan D.R. Miller F.D. J. Cell Biol. 1999; 146: 955-966Crossref PubMed Scopus (211) Google Scholar), whereas under different circumstances PI3K is dispensable (9Virdee K. Xue L. Hemmings B.A. Goemans C. Heumann R. Tolkovsky A.M. Brain Res. 1999; PubMed Google Scholar, 10Philpott K.L. McCarthy M.J. Klippel A. Rubin L.L. J. Cell Biol. 1997; 139: 809-815Crossref PubMed Scopus (220) Google Scholar, 11Tsui-Pierchala B.A. Putcha G.V. Johnson Jr., E.M. J. Neurosci. 2000; 20: 7228-7237Crossref PubMed Google Scholar). Therefore, additional signaling pathways probably function in the NGF-dependent survival of sympathetic neurons. The protein kinase C (PKC) family of serine/threonine kinases encompasses at least 12 members that participate in cellular functions ranging from proliferation to apoptosis (12Dempsey E.C. Newton A.C. Mochly-Rosen D. Fields A.P. Reyland M.E. Insel P.A. Messing R.O. Am. J. Physiol. 2000; 279: L429-L438Crossref PubMed Google Scholar, 13Battaini F. Pharm. Res. (N. Y.). 2001; 44: 353-361Crossref Scopus (96) Google Scholar). The PKC family can be separated into three groups based on their regulation. The conventional PKCs (α, βI, βII, and γ) are activated by DAG and phorbol esters and also require calcium, whereas the novel PKCs (δ, ϵ, θ, η, μ, and ν) require DAG or phorbol esters for activation but are insensitive to calcium. The atypical PKCs (ξ and ι/λ) are not sensitive to DAG, phorbol esters, or calcium. PKCμ and PKCν, although they are activated by DAG and phorbol esters, lack the autoinhibitory pseudosubstrate domain and are therefore probably regulated quite differently than other novel PKCs and are frequently termed PKDs to distinguish them. Growth factor receptors regulate PKCs via the activation of phospholipase C-γ. Phospholipase C-γ, upon binding to phosphorylated Tyr785 in TrkA, is itself phosphorylated and activated (14Loeb D.M. Stephens R.M. Copeland T. Kaplan D.R. Greene L.A. J. Biol. Chem. 1994; 269: 8901-8910Abstract Full Text PDF PubMed Google Scholar), hydrolyzing phosphatidylinositol 4,5-bisphosphate to produce DAG and inositol trisphosphate and thus activating PKCs. Growth factor receptors also influence PKC activity through the phosphorylation of the activation loop of PKC by phosphoinositide-dependent kinase-1 (15Toker A. Mol. Pharmacol. 2000; 57: 652-658Crossref PubMed Scopus (284) Google Scholar, 16Parekh D.B. Ziegler W. Parker P.J. EMBO J. 2000; : 496-503Crossref PubMed Scopus (513) Google Scholar). Although PKCs have been implicated in the regulation of survival, their function varies depending upon the cell type and apoptotic signal (17Cross T.G. Scheel-Toellner D. Henriquez N.V. Deacon E. Salmon M. Lord J.M. Exp. Cell Res. 2000; 256: 34-41Crossref PubMed Scopus (620) Google Scholar, 18Musashi M. Ota S. Shiroshita N. Int. J. Hematol. 1999; 72: 12-19Google Scholar). PKC-α, -β, -ϵ, and -ξ can function as suppressors of apoptosis, whereas PKC-δ and -θ appear to be proapoptotic in function. PKC-δ appears to be involved in apoptosis induced by several stimuli and is cleaved by caspase-3 to generate a catalytically active fragment (19Emoto Y. Manome Y. Meinhardt G. Kisaki H. Kharbanda S. Robertson M. Ghayur T. Wong W.W. Kamen R. Weichselbaum R. Kufe D. EMBO J. 1995; 14: 6148-6156Crossref PubMed Scopus (652) Google Scholar). Whether PKCs function in the trophic factor-dependent survival of neurons is unclear. Tyr785 of TrkA, and thus phospholipase C-γ, is required for neurite outgrowth stimulated by NGF in the sympathetic neuron-like cell line PC12 but is not required for survival (20Obermeier A. Bradshaw R.A. Seedorf K. Choidas A. Schlessinger J. Ullrich A. EMBO J. 1994; 13: 1585-1590Crossref PubMed Scopus (268) Google Scholar, 21Stephens R.M. Loeb D.M. Copeland T.D. Pawson T. Greene L.A. Kaplan D.R. Neuron. 1994; 12: 691-705Abstract Full Text PDF PubMed Scopus (471) Google Scholar). Atypical PKCs, however, may function in the survival of PC12 cells via activation of NF-κB (22Wooten M.W. J. Neurosci. Res. 1999; 58: 607-611Crossref PubMed Scopus (77) Google Scholar). We examined to what extent PKC signaling is involved in the survival of sympathetic neurons and here provide evidence that PKCs have a cooperative function in NGF-dependent survival along with PI3K. Materials—LY294002 and GF109203X were obtained from Biomol (Plymouth Meeting, PA); Bryostatin 1 and PMA were from Calbiochem; and boc-aspartyl fluoromethyl ketone (BAF) was obtained from Enzyme Systems Products (Livermore, CA). These compounds were dissolved in Me2SO and used at the concentrations under and were obtained from Cell was from and were obtained from was from were used for at a were from and from 1 were and culture or as Johnson Jr., E.M. J. Cell Biol. PubMed Scopus Google Scholar). were for in vitro in bovine and NGF were as under the NGF in the survival and the neurons were with not NGF and was for the of In were to the of as the other conditions, and the type of the alone was the NGF of neurons to PKC sympathetic neurons were with and in for was used in these of the from was NGF was to and was The of NGF in these was conditions, the neurons were and in the for in The neurons were to 12 and 1 and NGF and the were 1 in the presence of the NGF the neurons were of NGF for or The sympathetic neurons were with and with or NGF for The neurons were with and to NGF The the was with NGF and for additional was as B.A. Putcha G.V. Johnson Jr., E.M. J. Neurosci. 2000; 20: 7228-7237Crossref PubMed Google Scholar). the were with and for with at were for in and the was by the for additional in The cells were by that concentrations of to by three were with cell that were for were and were survival were obtained in a were on on three or The are as the and significant groups to with a of the sympathetic neurons in culture were with and with and The were of the with a cell to and for The were and to and was on these by CA). were to and with were with a of for at and for in the and were three with BSA, with the in for with BSA, and with were by the with and was used as a of were by the from the by in for at by three with of were as B.A. J. Johnson Jr., E.M. Neuron. Full Text Full Text PDF PubMed Scopus Google Scholar). the the neurons were with and for in 1 and at with The were by and were to the along with protein of a and the were for at The were three with and by the of of by in a NGF PKC in determine whether NGF induced PKC activity in sympathetic neurons, in vitro neurons that been of NGF were stimulated with NGF for The neurons were to cellular and the were to with that phosphorylated on PKC NGF induced the phosphorylation of a of on PKC and their phosphorylation was for These were phosphorylated sympathetic neurons were with PMA, a phorbol that conventional and novel PKCs indicating that the PKC than of To determine the extent to which of PKCs can promote the phosphorylation of these sympathetic neurons that were of NGF were with PMA or PKC for The phosphorylation of PKC was by of cell from these neurons. PMA and 1 promoted but PKC activity in sympathetic neurons In PMA and 1 in the of the phosphorylation of PKC These that the that are phosphorylated by NGF of phorbol PKCs. PKCs are phosphorylated on several that regulate their activity D.B. Ziegler W. Parker P.J. EMBO J. 2000; : 496-503Crossref PubMed Scopus (513) Google Scholar). PKC are phosphorylated NGF we whether NGF promoted the phosphorylation of PKCs by that conventional PKCs phosphorylated on that the kinase of the or phosphorylated on the phosphorylation The is also and appears to the A.C. J. PubMed Scopus Google Scholar). of both and was by NGF in sympathetic neurons although the phosphorylation of was with However, in the of NGF a significant of PKC was phosphorylated at these that other NGF account for of the PKC phosphorylation in sympathetic neurons. to the of phorbol on PKC PKC phosphorylation on and was in the presence of PMA and 1 the of conventional PKCs were with PMA and 1 to a N. A. J. Biol. Chem. Full Text PDF PubMed Google Scholar). of PKCs is of the of PKCs their and phorbol is used as a of PKC activity in In to phorbol esters, PKC are regulated by PI3K via phosphorylation of in the PKC activation loop by phosphoinositide-dependent kinase-1 W. Johnson J. Newton A.C. A. Curr. Biol. 1998; Full Text Full Text PDF PubMed Google Scholar, Ziegler D.B. D.R. Parker P.J. Science. 1998; PubMed Scopus Google Scholar). To determine whether PI3K regulated NGF-dependent PKC activity in sympathetic neurons, neurons were with the PI3K and PKC activity was by of cell from these neurons. PI3K inhibition the phosphorylation of PKC that was whereas the other PKC were In contrast, of NGF-maintained sympathetic neurons with a PKC that is for conventional PKCs, the phosphorylation of the of PKC in sympathetic neurons, although were to PMA promoted a phosphorylation of PKC than NGF NGF for led to the of only the and other PKC were at To determine whether PKC inhibition led to the inhibition of the PI3K or the phosphorylation of and kinases was examined by these with to the phosphorylated of and phosphorylation but did not inhibit kinase phosphorylation Inhibition of PKCs with GF109203X did not phosphorylation and kinase phosphorylation only these that PI3K regulated only a of PKC activity in neurons only in the and that the PKC GF109203X PKC phosphorylation in sympathetic neurons. PKC inhibition in sympathetic neurons did not the or PI3K PKCs are to regulate the activity of as the growth factor receptor J. Parker P.J. J. Biol. Chem. Full Text PDF PubMed Google Scholar, Mol. Biol. PubMed Scopus Google Scholar), we whether the inhibition of PKCs in sympathetic neurons TrkA NGF-maintained sympathetic neurons were with GF109203X or alone and were and TrkA phosphorylation was by TrkA from these GF109203X did not TrkA or the of TrkA as as of PKC inhibition Therefore, conventional PKCs did not regulate the or of TrkA in sympathetic neurons. PKC from by NGF NGF induced PKC activity in sympathetic neurons, we examined whether PKCs promote their sympathetic neurons were of NGF for or of NGF in the presence of the PKC PMA or The of neurons were and NGF led to the apoptosis of of neurons and both PMA and 1 the death of these neurons The survival of PMA and 1 however, modest and additional of NGF PMA 1 caused of survival not to a S. T. Neurosci. 2001; PubMed Scopus Google Scholar). of PKC activation on survival is probably caused by the of the of these compounds on PKC activity To the ability of PKC activation with NGF, were to the of PMA and NGF at apoptosis of trophic neurons with thus the of PMA on PKC neurons were of NGF for or and to NGF, PMA, or alone for to neurons that have to in that neurons were with NGF for the and the of neurons was and with neurons that were not of NGF the of of cells with PMA, thus of the of phorbol PKCs. of NGF PMA the death of the of neurons from the that have to in and PMA was as as NGF was in the cell death of these neurons Similarly, of NGF PMA was as as NGF at the of neurons that have to additional and of NGF that PKCs in the cell death pathway to sympathetic neurons from Therefore, activation of phorbol PKCs the death of sympathetic neurons that the cell death NGF PKCs with PI3K to that activation of PKCs the death of neurons we whether PKCs were required for the NGF-dependent survival of sympathetic neurons. sympathetic neurons were of NGF, of neurons In to the of NGF withdrawal on survival, inhibition of conventional PKCs with GF109203X in the presence of NGF on their survival that conventional PKCs were not required for the survival of sympathetic neurons. PI3K inhibition of NGF-maintained sympathetic neurons on their We have that PI3K inhibition of sympathetic neurons a modest and apoptotic of neurons of with at a that completely activation of B.A. Putcha G.V. Johnson Jr., E.M. J. Neurosci. 2000; 20: 7228-7237Crossref PubMed Google Scholar). We these of death NGF-maintained sympathetic neurons were with Therefore, under culture conditions, PI3K was not required for the of NGF. To determine whether PKCs and PI3K cooperate to promote the survival of sympathetic neurons, NGF-maintained neurons were with both and and the of neurons was The inhibition of both PKCs and PI3K caused the death of sympathetic neurons to a extent and with as the withdrawal of NGF with NGF M. J. P.A. Johnson Jr., E.M. J. Cell Biol. 1996; PubMed Scopus Google Scholar), the cell death caused by PKC and PI3K inhibition was completely with the indicating that the inhibition of these signaling pathways induced a apoptotic death to that caused by NGF These are with the that PKCs and PI3K to promote the survival of sympathetic neurons. To determine whether other of PKC activity also induced programmed cell death in with PI3K PMA of neurons was to the of NGF-maintained sympathetic neurons with PMA alone on their survival In contrast, with both PMA and neurons cell death as with alone PMA and induced apoptosis with as with NGF withdrawal which was to a extent apoptosis was induced with and GF109203X The of apoptosis induced by PMA and may be by the that PMA is a PKC than is GF109203X and and that PMA not phorbol PKCs N. A. J. Biol. Chem. Full Text PDF PubMed Google Scholar, M. R. L.L. J. Biol. Chem. 1994; Full Text PDF Google Scholar, A.R. Parker P.J. J. Cell Physiol. PubMed Scopus (77) Google Scholar). In the PI3K and PKC pathways function in a cooperative to promote the survival of sympathetic neurons in NGF. PKC and PI3K for NGF-dependent of under that PI3K is not required for the survival of sympathetic neurons B.A. Putcha G.V. Johnson Jr., E.M. J. Neurosci. 2000; 20: 7228-7237Crossref PubMed Google Scholar), have that PI3K for the of the of NGF (6Crowder R.J. Freeman R.S. J. Neurosci. 1998; 18: 2933-2943Crossref PubMed Google Scholar, 7Mazzoni I.E. Said F.A. Aloyz R. Miller F.D. Kaplan D. J. Neurosci. 1999; 19: 9716-9727Crossref PubMed Google Scholar, 8Vaillant A.R. Mazzoni I. Tudan C. Boudreau M. Kaplan D.R. Miller F.D. J. Cell Biol. 1999; 146: 955-966Crossref PubMed Scopus (211) Google Scholar). In of these sympathetic neurons were in that than PI3K may be at apoptosis in neurons in NGF is in activating PKCs in the of To determine to what extent PKCs were activated by NGF in conditions, sympathetic neurons that were in or were stimulated with NGF, and the extent of PKC phosphorylation was of sympathetic neurons with NGF induced PKC activity as by phosphorylation of several on PKC NGF induced the phosphorylation of PKC in neurons under conditions as the of phosphorylation are different neurons in and that were phosphorylated NGF under conditions were not under conditions with and The of PKC phosphorylation was in neurons in serum as These that the ability of NGF to PKC and which are upon the culture conditions in which the neurons are these we to determine the of both the PI3K pathway and the PKC pathway in the NGF-dependent survival of sympathetic neurons in neurons were and to the in were to a the of NGF for additional to NGF-maintained neurons in NGF of neurons in caused the apoptotic cell death of the neurons In to neurons in PI3K inhibition induced a apoptotic death of NGF-maintained sympathetic neurons in that PI3K was required for survival under these NGF-dependent neurons in were to PKC inhibition with a significant than by cell death in to neurons by NGF in The of neurons with both GF109203X and under conditions did not the of neurons that did the of their as with alone These that both PKCs and PI3K were required for the survival of NGF-maintained sympathetic neurons under conditions, whereas of these signaling pathways were in the presence of PKCs to in the Cell at which activated PKCs the apoptotic we examined in the apoptotic pathway that were by PKC To determine whether PKC and PI3K inhibition promoted the of apoptotic gene NGF-maintained sympathetic neurons under conditions were with or PMA alone or in for and NGF-maintained neurons were with and were of NGF, these the neurons were to and by that is required for the expression of apoptotic necessary for programmed cell death in sympathetic neurons is the activation of the transcription factor M. S. C. Johnson R.S. R. J. Cell Biol. PubMed Scopus Google Scholar, S. Freeman R.S. M. R. Johnson Jr., E.M. J. Cell Biol. 1994; PubMed Scopus Google Scholar, J. C. J. D. M. Rubin L.L. Neuron. 1995; 14: Full Text PDF PubMed Scopus Google Scholar). to NGF of sympathetic neurons induced the phosphorylation of on a transcription Inhibition of PI3K also induced the phosphorylation of with B.A. Putcha G.V. Johnson Jr., E.M. J. Neurosci. 2000; 20: 7228-7237Crossref PubMed Google Scholar), but phosphorylation was than that induced by NGF Inhibition of PKCs with GF109203X or PMA also induced phosphorylation on but promoted phosphorylation than NGF induced In contrast, the simultaneous inhibition of both PKCs and PI3K induced phosphorylation to the extent as NGF withdrawal In a its expression by its phosphorylation and to inhibition of both PKC and PI3K activity induced expression to the extent as NGF whereas inhibition of PKCs or PI3K alone was To determine whether proapoptotic gene expression was we the expression of the family that is important for NGF cell death G.V. K.L. A. Johnson E.M. Neuron. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar, G.V. S. S. K. S. R.J. A. A.C. Johnson E.M. Neuron. Full Text Full Text PDF PubMed Scopus Google Scholar). Inhibition of PKCs or PI3K alone induced or in to NGF which promoted the expression of Inhibition of both PKC and PI3K activity promoted expression to a extent than did inhibition of signaling pathway but simultaneous PKC and PI3K caused expression than did NGF The expression of proapoptotic family was also examined by and a as did simultaneous PKC and PI3K inhibition promoted expression of than did NGF not Therefore, PKC with PI3K to inhibit the cell death pathway to the of activation and proapoptotic gene expression. The modest of simultaneous PKC and PI3K inhibition on the expression of proapoptotic as as with NGF may the of apoptosis induced by PKC and PI3K inhibition The PKC family a of signaling that participate in functions as as and we that NGF promoted the phosphorylation activation of PKCs, as as the phosphorylation of PKC in sympathetic neurons. PKC activation with phorbol esters rescued trophic sympathetic neurons as as NGF PKCs with PI3K to promote the survival of NGF-dependent sympathetic neurons, and these pathways for the of the of NGF. The simultaneous inhibition of the PI3K and PKC pathways caused the phosphorylation and of the proapoptotic transcription factor c-Jun, that these signaling pathways to inhibit programmed cell death at the of proapoptotic gene expression. In the PKC pathway a critical function in the survival of sympathetic neurons by NGF, regardless of the culture conditions the and of the PKC important is which PKC family functions in trophic factor-dependent The ability of PMA and GF109203X to inhibit the survival of sympathetic neurons that the PKC was a conventional or novel PKC, which different a which PKC was we examined sympathetic neurons from and with neurons from were significant in the of death NGF withdrawal or in their to PI3K inhibition not however, to that of the PKC by or may be the of of PKCs that be involved and the with the expression in sympathetic neurons. that was to determine which PKC is involved in survival was a pharmacologic several to inhibit of PKCs. of the compounds as and were to sympathetic neurons, at concentrations in the to inhibit PKC thus their in cell death the by these compounds did not have the or of apoptosis, that they inhibit other Although these to determine which PKC to trophic factor-dependent survival signaling were important The here a of in the in vitro conditions in which the neurons are their signaling of whether or a is both the PKC and PI3K pathways in survival by NGF. The that these is which culture the in vivo for sympathetic neurons. neurons in vivo not be to serum they be to the of of the of as and thus to the for the of both the PI3K and the PKC pathways in survival be with or However, the to the PI3K and PKC may a PKC inhibit Although PKCs appear to to which are required for function are unclear. can with kinase and on from and thus activity C. E. PubMed Scopus Google Scholar), which is with However, NGF not appear to PKDs in sympathetic neurons, in to PMA of these neurons, based on that PKDs phosphorylated on activation loop necessary for activity not on which its activity T. J. Biol. Chem. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar, J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). Although a the function of PKCs to of the regulated by In to PKCs promote survival in neurons, we several to of the PKC phosphorylated NGF in a but were the protein that is regulated by NGF was not or of which have and not These were obtained by with to and and by neurons from the by which PKCs function to inhibit apoptosis in neurons unclear. The that PI3K and PKC inhibition did not the proapoptotic as that are necessary for programmed cell death that additional signaling pathways the apoptotic pathway in sympathetic neurons, at a or is the phosphorylation and of Greene L.A. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar), which the of PI3K and PKC inhibition to survival signaling as that regulate the transcription protein and NF-κB Freeman R.S. Mol. Neurosci. 2001; 18: PubMed Scopus Google Scholar, A. S. Science. 1999; PubMed Scopus Google Scholar, S. Freeman R.S. J. Neurosci. 1998; 18: PubMed Google and that proapoptotic Greene L.A. Neuron. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar), may also account for the of cell death induced by PI3K and PKC The of signaling used by neurons to maintain survival the of of cells from or The PKC pathway is pathway in with PI3K, the expression of for programmed cell We for We A. for and critical of the We also and for and critical of and for