Cellular mechanisms associated with spontaneous and ciliary neurotrophic factor-cAMP-induced survival and axonal regeneration of adult retinal ganglion cells.
We have shown previously that intraocular elevation of cAMP using the cAMP analog 8-(4-chlorophenylthio)-cAMP (CPT-cAMP) failed to promote axonal regeneration of axotomized adult retinal ganglion cells (RGCs) into peripheral nerve (PN) grafts but significantly potentiated ciliary neurotrophic factor (CNTF)-induced axonal regeneration. Using the PN graft model, we now examine the mechanisms underlying spontaneous and CNTF/CPT-cAMP-induced neuronal survival and axonal regrowth. We found that blockade of the cAMP pathway executor protein kinase A (PKA) using the cell-permeable inhibitor KT5720 did not affect spontaneous survival and axonal regeneration but essentially abolished the CNTF/CPT-cAMP-induced RGC survival and axonal regeneration. Blockade of CNTF signaling pathways such as phosphotidylinositol 3-kinase (PI3K)/akt by 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002), mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) by 2-(2-diamino-3-methoxyphenyl-4H-1-benzopyran-4-one (PD98059), or Janus kinase (JAK)/signal transducer and activators of transcription (STAT3) by tyrphostin AG490 also blocked the CNTF/CPT-cAMP-dependent survival and regeneration effects. PKA activity assay and Western blots showed that KT5720, LY294002, and PD98059 almost completely inhibited PKA, PI3K/akt, and MAPK/ERK signal transduction, respectively, whereas AG490 substantially decreased JAK/STAT3 signal transduction. Intraocular injection of CPT-cAMP resulted in a small PKA-dependent increase in CNTF receptor alpha mRNA expression in the retinas, an effect that may facilitate CNTF action on survival and axonal regeneration. Surprisingly, in the absence of CNTF/CPT-cAMP, LY294002, PD98059, and AG490, but not KT5720, significantly enhanced spontaneous RGC survival, suggesting differential roles of these pathways in RGC survival under different conditions. Our data suggest that CNTF/CPT-cAMP-induced RGC survival and axonal regeneration are a result of multiple pathway actions, with PKA as an essential component, but that these pathways can function in an antagonistic manner under different conditions.
Park
Kevin
K
School of Anatomy and Human Biology and Western Australian Institute for Medical Research, The University of Western Australia, Crawley, Perth WA 6009, Australia.
Luo
Jian-Min
JM
Hisheh
Susan
S
Harvey
Alan R
AR
Cui
Qi
Q
D000818:Animals; D001369:Axons / Q000187:drug effects / Q000502:physiology; D002227:Carbazoles / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D002867:Chromones / Q000494:pharmacology; D020934:Ciliary Neurotrophic Factor / Q000037:antagonists & inhibitors / Q000494:pharmacology / Q000502:physiology*; D000242:Cyclic AMP / Q000031:analogs & derivatives* / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors; D005419:Flavonoids / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D020935:MAP Kinase Signaling System / Q000187:drug effects; D009025:Morpholines / Q000494:pharmacology; D009416:Nerve Regeneration / Q000187:drug effects; D020221:Optic Nerve Injuries / Q000201:enzymology / Q000503:physiopathology; D010543:Peroneal Nerve / Q000637:transplantation; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors; D051057:Proto-Oncogene Proteins c-akt; D011758:Pyrroles / Q000494:pharmacology; D051381:Rats; D011916:Rats, Inbred F344; D020801:Receptor, Ciliary Neurotrophic Factor / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics; D012165:Retinal Ganglion Cells / Q000187:drug effects / Q000502:physiology*; D050796:STAT3 Transcription Factor; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013873:Thionucleotides / Q000494:pharmacology; D015534:Trans-Activators / Q000037:antagonists & inhibitors; D020032:Tyrphostins / Q000494:pharmacology; D015854:Up-Regulation
D002227:Carbazoles; D002867:Chromones; D020934:Ciliary Neurotrophic Factor; D004268:DNA-Binding Proteins; D005419:Flavonoids; D007211:Indoles; D009025:Morpholines; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011518:Proto-Oncogene Proteins; D011758:Pyrroles; D020801:Receptor, Ciliary Neurotrophic Factor; D050796:STAT3 Transcription Factor; C494088:Stat3 protein, rat; D013873:Thionucleotides; D015534:Trans-Activators; D020032:Tyrphostins; C095512:alpha-cyano-(3,4-dihydroxy)-N-benzylcinnamide; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; C057416:KT 5720; D000242:Cyclic AMP; D011505:Protein-Tyrosine Kinases; C494920:Akt1 protein, rat; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D017868:Cyclic AMP-Dependent Protein Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one