title: Functional, endogenously expressed corticotropin-releasing factor receptor type 1 (CRF1) and CRF1 receptor mRNA expression in human neuroblastoma SH-SY5Y cells.
abstract: Corticotropin-releasing factor (CRF) is a hypothalamic 41-amino acid peptide which stimulates corticotropin (ACTH) release from the anterior pituitary and is also involved in the body response to stress. CRF1 receptors represent a potential target for novel antidepressant/anxiolytic drugs. The aim of the present study was to search for a human cell line expressing native, functional CRF1 receptors as a starting material for screening purposes. We identified CRF1 receptors functionally coupled to cAMP formation in human neuroblastoma SH-SY5Y cells. CRF induced concentration-dependent increases in cAMP accumulation in SH-SY5Y cells (maximal increase 6.9 +/- 0.9 fold over basal values, n = 14). This effect was mimicked by related peptides with similar potencies: (mean pEC50 value) human/rat CRF (8.63), rat urocortin (9.32), sauvagine (8.97), urotensin I (8.93), ovine CRF (8.81). The efficacies of these agonists were nearly the same, with the exception of ovine CRF which was slightly less efficacious (75% the Emax of CRF). The responses to CRF were competitively antagonised by the following peptide fragments (mean pKB value): alpha-helical-CRF (9-41) (7.54), [D-Phe12,Nle21,38,C alpha MeLeu37]CRF (12-41) (8.36) and [D-Tyr12]astressin (9.49) and by the selective, non-peptidic CRF1 receptor antagonists, CP-154,526 (7.76) and antalarmin (9.19). Estimation of receptor density by [125I]Tyr0-ovine CRF saturation binding yielded a modest number of binding sites (Bmax 12 fmol/mg protein, KD 0.2 nM). Analysis of mRNA by reverse transcription-polymerase chain reaction clearly revealed the presence of mRNA for CRF1 receptors in SH-SY5Y cells. A slight signal for CRF2 receptor mRNA was also observed. We conclude that neuroblastoma SH-SY5Y cells are endowed with native CRF1 receptors positively coupled to cAMP formation. They therefore constitute a useful functional model for the search of CRF1 selective compounds with potential anxiolytic/antidepressant activity.
authors.affiliation: Nervous System Research, Mental Health Unit, Novartis Pharma AG, CH-4002 Basel, Switzerland.
authors.forename: P
authors.initials: P
authors.lastname: Schoeffter
authors.forename: D
authors.initials: D
authors.lastname: Feuerbach
authors.forename: I
authors.initials: I
authors.lastname: Bobirnac
authors.forename: L
authors.initials: L
authors.lastname: Gazi
authors.forename: R
authors.initials: R
authors.lastname: Longato
mesh_terms: D000818:Animals; D000242:Cyclic AMP / Q000096:biosynthesis; D006801:Humans; D009447:Neuroblastoma; D012333:RNA, Messenger / Q000378:metabolism*; D011869:Radioligand Assay; D051381:Rats; D018019:Receptors, Corticotropin-Releasing Hormone / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012756:Sheep; D014407:Tumor Cells, Cultured; D000099052:CRF Receptor, Type 1
chemical_list: D012333:RNA, Messenger; D018019:Receptors, Corticotropin-Releasing Hormone; D000099052:CRF Receptor, Type 1; D000242:Cyclic AMP