Characterization and effects on cAMP accumulation of adrenomedullin and calcitonin gene‐related peptide (CGRP) receptors in dissociated rat spinal cord cell culture

• Published in: British journal of pharmacology 2006-06, Vol.148 (4), p.459-468
• Main Authors: Takhshid, Mohammad A, Poyner, David R, Chabot, Jean‐Guy, Fournier, Alain, Ma, Weiya, Zheng, Wen‐Hua, Owji, Ali A, Quirion, Remi
• Format: Article
• Language: English
• Subjects: Adrenomedullin; Animals; BIBN4096BS; Binding Sites; Binding, Competitive; Biological and medical sciences; Calcitonin Gene-Related Peptide; Calcitonin Gene-Related Peptide - pharmacology; calcitonin receptor‐like receptor; cAMP; Cells, Cultured; CGRP; Cyclic AMP; Cyclic AMP - physiology; Female; Immunohistochemistry; Life Sciences; Medical sciences; Peptide Fragments; Peptide Fragments - pharmacology; Pharmacology. Drug treatments; Piperazines; Piperazines - pharmacology; Quinazolines; Quinazolines - pharmacology; Rats; Rats, Sprague-Dawley; receptor binding assays; Receptors, Adrenomedullin; Receptors, Calcitonin Gene-Related Peptide; Receptors, Calcitonin Gene-Related Peptide - metabolism; Receptors, G-Protein-Coupled; Receptors, G-Protein-Coupled - metabolism; receptor‐activity‐modifying protein; Spinal Cord; Spinal Cord - cytology; Spinal Cord - embryology; Spinal Cord - metabolism; Toxicology
• ISSN: 0007-1188; 1476-5381
• DOI: 10.1038/sj.bjp.0706750

1 Adrenomedullin (AM) and calcitonin gene‐related peptide (CGRP) have structural similarities, interact with each others receptors (calcitonin receptor‐like receptor (CLR)/receptor‐activity‐modifying proteins (RAMPs)) and show overlapping biological activities. AM and CGRP receptors are chiefly coupled to cAMP production. In this study, a method of primary dissociated cell culture was used to investigate the presence of AM and CGRP receptors and their effects on cAMP production in embryonic spinal cord cells. 2 Both neuronal and non‐neuronal CLR immunopositive cells were present in our model. 3 High affinity, specific [125I]‐AM binding sites (Kd 79±9 pM and Bmax 571±34 fmol mg−1 protein) were more abundant than specific [125I]‐CGRP binding sites (Kd 12±0.7 pM and Bmax 32±2 fmol mg−1 protein) in embryonic spinal cord cells. 4 Specific [125I]‐AM binding was competed by related molecules with a ligand selectivity profile of rAM>hAM22–52>rCGRPα>CGRP8–37≫[r‐(r*,s*)]‐N‐[2‐[[5‐amino‐1‐[[4‐(4‐pyridinyl)‐1‐piperazinyl]carbonyl]pentyl]amino]‐1‐[(3,5‐dibromo‐4‐hydroxyphenyl)methyl]‐2‐oxoethyl]‐4‐(1,4‐dihydro‐2‐oxo‐3(2H)‐quinazolinyl)‐,1‐piperidinecarboxamide (BIBN4096BS). 5 Specific [125I]‐CGRP binding was competed by rCGRPα>rAMCGRP8–37BIBN4096BS>hAM22–52. 6 Cellular levels of cAMP were increased by AM (pEC50 10.2±0.2) and less potently by rCGRPα (pEC50 8.9±0.4). rCGRPα‐induced cAMP accumulation was effectively inhibited by CGRP8–37 (pA2 7.63±0.44) and hAM22–52 (pA2 6.18±0.21) while AM‐stimulation of cAMP levels was inhibited by CGRP8–37 (pA2 7.41±0.15) and AM22–52 (pA2 7.26±0.18). BIBN4096BS only antagonized the effects of CGRP (pA2 8.40±0.30) on cAMP accumulation. 7 These pharmacological profiles suggest that effects of CGRP are mediated by the CGRP1 (CLR/RAMP1) receptor in our model while those of AM are related to the activation of the AM1 (CLR/RAMP2) receptor subtype. British Journal of Pharmacology (2006) 148, 459–468. doi:10.1038/sj.bjp.0706750