Epac1-mediated Rap1 activation is not required for the production of nitric oxide in BV2, murine microglial cells.
This study demonstrates that cyclic AMP (cAMP) production is induced by lipopolysaccharide (LPS) stimulation and activates two different pathways in murine BV2 microglial cells. Two principal effector proteins for cAMP are protein kinase A (PKA) and cAMP-responsive guanine nucleotide exchange factor (Epac), a Rap GDP exchange factor. When cells were treated with various cAMP level modulators, nitric oxide (NO) production increased as the result of posttreatment with Type IV phosphodiesterase (PDE4) inhibitor, rolipram or dibutyryl-cAMP (dbcAMP), at 2 hr after LPS stimulation. Intracellular cAMP increased due to LPS stimulation and the cAMP modulators phosphorylate transcription factor CREB, which is enhanced in turn by posttreatment with dbcAMP. In contrast, the Epac-specific cAMP analog 8-(4-chloro-phenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate (8CPT-2Me-cAMP) activates Rap1 in the BV2 cells, but does not induce PKA activation, as judged by CREB phosphorylation. NO production was enhanced by posttreatment with dbcAMP but not by treatment with 8CPT-2Me-cAMP. This suggests that LPS-stimulated NO production is mainly PKA-dependent and also that Epac1-mediated Rap1 activation is not required for the induction of NO production.
Moon
Eun-Yi
EY
Laboratory of Human Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Taejeon, Korea. eunyi@kribb.re.kr
Oh
Su-Young
SY
Han
Gyoon-Hee
GH
Lee
Chul-Sang
CS
Park
Song-Kyu
SK
D000704:Analysis of Variance; D000818:Animals; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D020662:Guanine Nucleotide Exchange Factors / Q000502:physiology*; D007424:Intracellular Fluid / Q000378:metabolism; D008070:Lipopolysaccharides / Q000008:administration & dosage / Q000276:immunology; D051379:Mice; D017628:Microglia / Q000276:immunology / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis*; D015398:Signal Transduction / Q000502:physiology; D018709:Statistics, Nonparametric; D020827:rap1 GTP-Binding Proteins / Q000378:metabolism*
C482601:Epac protein, mouse; D020662:Guanine Nucleotide Exchange Factors; D008070:Lipopolysaccharides; D009569:Nitric Oxide; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D020827:rap1 GTP-Binding Proteins