Abstract
5-Hydroxytryptamine (5-HT)2A receptors have been implicated to play a role in both the treatment and pathophysiology of a number of psychiatric disorders. Therefore, the coupling of this receptor to signals, such as extracellular signal-regulated kinase (ERK), that elicit long-term neuronal changes may be relevant. In the present study we examined the coupling of the Gq-coupled receptor to ERK in PC12 cells, a cell line commonly used as a neuronal model system. Activation of ERK occurred through a pathway different than the protein kinase C-dependent pathways described previously in studies of non-neuronal cells. Activation of ERK, in PC12 cells, was inhibited by both chelation of extracellular Ca2+ and by depletion of intracellular Ca2+ stores. Surprisingly, activation was not inhibited, but actually potentiated, by a variety of protein kinase C inhibitors covering all known protein kinase C isoforms. In contrast, the coupling of receptor to activation of ERK was found to be sensitive to N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (W7) andN-(4-aminobutyl)-5-chloro-1-naphthalenesulfonamide (W13), inhibitors of calmodulin, but not to 1-(N,O-bis[5-isoquinolinesulfonyl]-N-methyl-l-tyrosyl)-4-phenylpiperazine (KN62) and 2-[N-(2-hydroxyethyl)]-N-4-methoxybenzenesulfonyl)]amino-N-(4-chlorocinnamyl)-N-methylbenzylamine) (KN93), inhibitors of calmodulin-dependent protein kinase. Additionally, the general tyrosine kinase inhibitor genistein, as well as the Src inhibitor PP1 and the epidermal growth factor receptor kinase inhibitor 4-(3-chloroanilino)-6,7-dimethoxyquinazoline (AG 1478), inhibited receptor-mediated activation of ERK, suggesting a role for tyrosine kinases. In fact, 5-HT was found to stimulate tyrosine phosphorylation of a number of proteins, and this phosphorylation was inhibited by W7. 5-HT2A receptor-activation of ERK through a protein kinase C-independent pathway requiring Ca2+/calmodulin/tyrosine kinases represents a pathway distinct from those described in studies of non-neuronal cells.
Footnotes
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This study was supported by National Institute of Mental Health Grant MH60100 (to D.S.C).
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DOI: 10.1124/jpet.102.038083
- Abbreviations:
- 5-HT
- 5-hydroxytryptamine
- ERK
- extracellular signal-regulated kinase
- PKC
- protein kinase C
- H89
- N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide
- PI
- phosphoinositide
- phorbol 12-myristate-13-acetate
- CaM, calmodulin
- PKA
- protein kinase A
- EGF
- epidermal growth factor
- AG 17
- tyrphostin A9
- W7
- N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride
- W13
- N-(4-aminobutyl)-5-chloro-1-naphthalenesulfonamide
- AG 1478
- 4-(3-chloroanilino)-6,7-dimethoxyquinazoline
- KN62
- 1-(N,O-bis[5-isoquinolinesulfonyl]-N-methyl-l-tyrosyl)-4-phenylpiperazine
- KN93
- 2-[N-(2-hydroxyethyl)]-N-(4-methyoxybenzenesulfonyl)]amino-N-(4-chlorocinnamyl)-N-methylbenzylamine)
- MDL 100907
- R-(+)-α-(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenyl)ethyl]-4-piperidinemethanol
- Go6983
- 2-[1-(3-dimethylaminopropyl)-5-methoxyindol-3-yl]-3-(1H-indol-3-yl) maleimide
- Go6976
- 12-(2-cyanoethyl)-6,7,12,13-tetrahydro-13-methyl-5-oxo-5H-indolo(2,3-a)pyrrolo(3,4-c)-carbazole
- Ro-31-8220
- 3-[1-[3-amidinothio)propyl-1H-indol-3-yl]-3-(1-methyl-1H-indol-3-yl)maleimide
- PP1
- {4-amino-1-tert-butyl-3-(1′-naphthyl)pyrazolo[3,4-d]pyrimidine}
- SB206553
- N-3-pyridinyl-3,5-dihydro-5-methylbenzo(1,2-b:4,5-b′)dipyrrole-1(2H)carboxamide
- LY-278,584
- 1-methyl-N-(8-methyl-8-azabicyclo[3.2.1]-oct-3-yl)-1H-indazole-3-carboxamide
- Received April 26, 2002.
- Accepted August 8, 2002.
- The American Society for Pharmacology and Experimental Therapeutics
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