Demonstration of the neurotransmitter role of calcitonin gene-related peptides (CGRP) by immunoblockade with anti-CGRP monoclonal antibodies.
1. Monoclonal antibodies (MAbs) against rat alpha-calcitonin gene-related peptide (alpha CGRP) were produced. Those which bound CGRP in a radioimmunoassay and inhibited the binding of 2-[125I]-iodohistidyl10-CGRP in a receptor binding assay were selected for immunoblockade experiments. 2. The effect of MAbs on CGRP inhibition of electrically stimulated contractions of the rat isolated vas deferens was characterized. Four out of 11 MAbs tested shifted the concentration-response curve of CGRP to the right compared with vehicle or irrelevant MAb control. MAb C4.19 produced equipotent blockade of rat alpha CGRP and rat beta CGRP and was chosen for further studies. MAb C4.19 had no pharmacologically significant effect on the concentration-response relationship of isoprenaline, rat beta-endorphin or somatostatin. 3. We demonstrated that the pharmacological response to CGRP in the presence of MAb C4.19 could be predicted when the dissociation constant and concentration of binding sites of the antibody were known. Comparison of experimental and computer simulated data showed good agreement for EC50 and maximum effect of CGRP in the presence of MAb C4.19. 4. Capsaicin at 1 microM inhibited the electrically stimulated contractions by 60.8% (95% confidence interval 51.8% to 69.9%). This effect was significantly attenuated by MAb C4.19 to 26.0% (95% confidence interval 15.2% to 36.8%; P < 0.003). 5. The immunoblockade of exogenous and endogenous CGRP described here, together with complementary evidence from other studies, strongly suggest that CGRP has a major neurotransmitter role at the neuroeffector junction of the rat vas deferens.
Tan
K K
KK
Clinical Pharmacology Unit, Addenbrooke's Hospital, Cambridge.
Brown
M J
MJ
Longmore
J
J
Plumpton
C
C
Hill
R G
RG
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology*; D000918:Antibody Specificity; D001667:Binding, Competitive; D015740:Calcitonin Gene-Related Peptide / Q000276:immunology* / Q000494:pharmacology / Q000502:physiology*; D002211:Capsaicin / Q000494:pharmacology; D004558:Electric Stimulation; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008954:Models, Biological; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology; D009469:Neuromuscular Junction / Q000187:drug effects / Q000502:physiology; D018377:Neurotransmitter Agents / Q000037:antagonists & inhibitors* / Q000494:pharmacology / Q000502:physiology*; D011863:Radioimmunoassay; D051381:Rats; D017207:Rats, Sprague-Dawley; D017208:Rats, Wistar; D009435:Synaptic Transmission / Q000187:drug effects / Q000502:physiology; D014649:Vas Deferens / Q000294:innervation
D000911:Antibodies, Monoclonal; D018377:Neurotransmitter Agents; D015740:Calcitonin Gene-Related Peptide; D002211:Capsaicin