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Titolo:
Simultaneous measurement of tyrosine hydroxylase activity and phosphorylation in bovine adrenal chromaffin cells
Autore:
Cheah, TB; Bobrovskaya, L; Goncalves, CA; Hall, A; Elliot, R; Lengyel, I; Bunn, SJ; Marley, PD; Dunkley, PR;
Indirizzi:
Univallaghan,e, Fac Med & Hlth Sci, Neurosci Grp, Discipline Med Biochem, C Univ Newcastle Callaghan NSW Australia 2308 rp, Discipline Med Biochem, C Univ Melbourne, Dept Pharmacol, Parkville, Vic 3052, Australia Univ Melbourne Parkville Vic Australia 3052 arkville, Vic 3052, Australia UFRGS, Dept Bioquim, Porto Alegre, RS, Brazil UFRGS Porto Alegre RS Brazil RGS, Dept Bioquim, Porto Alegre, RS, Brazil Univ Otago, Dept Anat & Struct Biol, Sch Med Sci, Dunedin, New Zealand Univ Otago Dunedin New Zealand Biol, Sch Med Sci, Dunedin, New Zealand
Titolo Testata:
JOURNAL OF NEUROSCIENCE METHODS
fascicolo: 2, volume: 87, anno: 1999,
pagine: 167 - 174
SICI:
0165-0270(19990301)87:2<167:SMOTHA>2.0.ZU;2-1
Fonte:
ISI
Lingua:
ENG
Soggetto:
DEPENDENT PROTEIN-KINASE; STIMULATED PHOSPHORYLATION; II INHIBITOR; ACTIVATION; SER(19); SER(40); BIOSYNTHESIS; SER(31); MEDIATE; SITES;
Keywords:
chromaffin cells; digitonin permeabilisation; tyrosine hydroxylase activity; protein phosphorylation; catecholamines;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
25
Recensione:
Indirizzi per estratti:
Indirizzo: Dunkley, PR Univallaghan,e, Fac Med & Hlth Sci, Neurosci Grp, Discipline Med Biochem, C Univ Newcastle Callaghan NSW Australia 2308 ne Med Biochem, C
Citazione:
T.B. Cheah et al., "Simultaneous measurement of tyrosine hydroxylase activity and phosphorylation in bovine adrenal chromaffin cells", J NEUROSC M, 87(2), 1999, pp. 167-174

Abstract

A method for simultaneous measurement of tyrosine hydroxylase (TH) activation and phosphorylation in permeabilised and intact bovine adrenal chromaffin cells (BACCs) was established. Permeabilised cells were stimulated with cyclic AMP (1-10 mu M) in the presence of [P-32]ATP and L-[carboxyl-C-14]tyrosine. Intact BACCs were preincubated with P-32(i) for 3 h and stimulated with forskolin (1-5 mu M) in the presence of L-[carboxyl-C-14]tyrosine. On stimulation each well was covered with a sealed 'chimney' fitted with a small plastic cup containing 300 mu l of 1.0 M NaOH that trapped the (CO2)-C-14 released. TH activity was determined by measuring C-14 radioactivity. TH phosphorylation was measured in the same cells by separating the solubilized proteins on SDS PAGE followed by autoradiography and/or HPLC analysis. Itwas found that H89, a protein kinase A inhibitor, significantly blocked both TH phosphorylation and activation in response to cyclic AMP in permeabilised cells. However, in intact cells, H89 was effective only in respect to forskolin-stimulated TH activity and did not block the forskolin-stimulatedTH phosphorylation of Ser-40. The reason(s) for this lack of correlation between TH activation and phosphorylation is presently not understood. (C) 1999 Elsevier Science B.V. All rights reserved.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 01/04/20 alle ore 11:39:05