title: Methylmercury decreases NGF-induced TrkA autophosphorylation and neurite outgrowth in PC12 cells.
abstract: Neurotrophin signaling through Trk receptors is important for differentiation and survival in the developing nervous system. The present study examined the effects of CH(3)Hg on (125)I-nerve growth factor (NGF) binding to the TrkA receptor, NGF-induced activation of the TrkA receptor, and neurite outgrowth in an in vitro model of differentiation using PC12 cells. Whole-cell binding assays using (125)I-NGF revealed a single binding site with a K(d) of approximately 1 nM. Methylmercury (CH(3)Hg) at 30 nM (EC(50) for neurite outgrowth inhibition) did not affect NGF binding to TrkA. TrkA autophosphorylation was measured by immunoblotting with a phospho-specific antibody. TrkA autophosphorylation peaked between 2.5 and 5 min of exposure and then decreased but was still detectable at 60 min. Concurrent exposure to CH(3)Hg and NGF for 2.5 min resulted in a concentration-dependent decrease in TrkA autophosphorylation, which was significant at 100 nM CH(3)Hg. To determine whether the observed inhibition of TrkA was sufficient to alter cell differentiation, NGF-stimulated neurite outgrowth was examined in PC12 cells after exposure to 30 nM CH(3)Hg, a concentration that inhibited TrkA autophosphorylation by approximately 50%. For comparison, a separate group of PC12 cells were exposed to a concentration of the selective Trk inhibitor K252a (30 nM), which had been shown to produce significant inhibition of TrkA autophosphorylation. Twenty-four hour exposure to either CH(3)Hg or K252a reduced neurite outgrowth to a similar degree. Our results suggest that CH(3)Hg may inhibit differentiation of PC12 cells by interfering with NGF-stimulated TrkA autophosphorylation.
authors.affiliation: Curriculum in Toxicology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Damani K
authors.initials: DK
authors.lastname: Parran
authors.forename: Stanley
authors.initials: S
authors.lastname: Barone
authors.forename: William R
authors.initials: WR
authors.lastname: Mundy
mesh_terms: D000818:Animals; D001665:Binding Sites / Q000187:drug effects / Q000502:physiology; D001667:Binding, Competitive / Q000187:drug effects / Q000502:physiology; D002352:Carrier Proteins / Q000187:drug effects* / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects* / Q000502:physiology; D002490:Central Nervous System / Q000002:abnormalities / Q000187:drug effects* / Q000473:pathology; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D008565:Membrane Proteins / Q000187:drug effects* / Q000378:metabolism; D020262:Mercury Poisoning, Nervous System / Q000201:enzymology* / Q000473:pathology / Q000503:physiopathology; D008767:Methylmercury Compounds / Q000633:toxicity*; D020932:Nerve Growth Factor / Q000037:antagonists & inhibitors* / Q000378:metabolism; D016501:Neurites / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D016716:PC12 Cells; D010766:Phosphorylation / Q000187:drug effects; D011247:Pregnancy; D011297:Prenatal Exposure Delayed Effects; D051381:Rats; D020917:Receptor, trkA
chemical_list: D002352:Carrier Proteins; D004791:Enzyme Inhibitors; D008565:Membrane Proteins; D008767:Methylmercury Compounds; D020932:Nerve Growth Factor; D020917:Receptor, trkA