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hCGRP8–37 antagonizes vasodilations and cAMP responses to rat calcitonin gene-related peptide (rCGRP) in rat caudal artery
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Session IV: Cardiovascular and regional hemodynamic actions of CGRP 47 CHARACTERISTICS
OF
CGRP-INDUCED
RELAXATION
IN R A B B I T
OPHTHALMIC
ARTERY
A. Z S C H A U E R I, H. U U S I T A L O 1 and J.E. B R A Y D E N 2, iDepartment of Anatomy, U n i v e r s i t y of Helsinki, 00170 Helsinki, Finland, and 2 D e p a r t m e n t of Pharmacology, College of Medicine, The U n i v e r s i t y of Vermont, Burlington, V e r m o n t 05405, U.S.A. The e l e c t r o m e c h a n i c a l e f f e c t s of c a l c i t o n i n gene r e l a t e d p e p t i d e (CGRP) on intact and e n d o t h e l i u m d e n u d e d rabbit o p h t h a l m i c artery w e r e studied. This study was p e r f o r m e d u s i n g m i c r o e l e c t r o d e and isometric t e n s i o n r e c o r d i n g t e c h n i q u e s for m e a s u r i n g c h a n g e s in m e m b r a n e p o t e n t i a l and in c o n t r a c t i l e force. CGRP induced a d o s e d e p e n d e n t r e l a x a t i o n both on intact and e n d o t h e l i u m denuded arteries s t i m u l a t e d by 5 uM n o r e p i n e p h r i n e (NE). CGRP (i0 nM) h y p e r p o l a r i z e d the smooth m u s c l e m e m b r a n e by 20.2 ~ 9.0 mV (n=5) a n d 8 . 0 Z 0.9 m V (n=5) in e n d o t h e l i u m d e n u d e d and intact arteries, respectively. Both the r e l a x a t i o n and hyperp o l a r i z a t i o n induced b y C G R P was p a r t l y inhibited b y g l y b e n c l a m i d e , a b l o c k e r cf the A T P - s e n s i t i v e K channels. This result indicates that C G R P - i n d u c e d relaxation is p a r t l y due to a h y p e r p o l a r i z a t i o n t h r o u g h a c t i v a t i o n of A T P - s e n s i t i v e K c h a n n e l s as has b e e n shown by Nelson et al (1990). In intact arteries after w a s h o u t of CGRP the c o n t r a c t i l e r e a c t i v i t y to NE was increased whereas in e n d o t h e l i u m d e n u d e d a r t e r i e s the CGRP induced r e l a x a t i o n p r o l o n g e d and after w a s h o u t of CGRP the c o n t r a c t i l e s e n s i t i v i t y to NE was diminished. In intact a r t e r i e s NE c o n t r a c t i o n s w e r e e n h a n c e d by L - N M M A (L-N-monomethyl arginine), an i n h i b i t o r of EDRF (endothelium d e r i v e d r e l a x i n g factor) synthesis, and in the p r e s e n c e of L-NMMA, C G R P - i n d u c e d r e l a x a t i o n s r e s e m b l e d those seen in e n d o t h e l i u m d e n u d e d arteries. This result suggests that there is an i n c r e a s e d EDRF synthesis in intact a r t e r i e s d u r i n g N E - s t i m u l a t i o n and that CGRP may inhibit either the s y n t h e s i s or the a c t i v i t y of EDRF. I n c r e a s e d c o n t r a c t i l e s e n s i t i v i t y to NE after of CGRP c o u l d be e x p l a i n e d if r e c o v e r y of EDRF synthetic a c t i v i t y is slow.
48 hCGRPa_37 ANTAGONIZES VASODILATIONS AND cAMP RESPONSES TO RAT CALCITONIN GENE-RELATED PEP'ITDE (rCGRP) IN RAT CAUDAL ARTERY R.R. FISCUS, X. WANG and H. HAO. Sanders-Brown Res. Center on Aging & Dept. Physiol. & Biophysics, Univ. Kentucky College of Medicine, Lexington, KY 40536-0230, USA. CGRP is an endogenous and potent hypotensive neuropeptide in rat and human. In rat, rCGRP relaxes isolated aorta and caudal artery rings in endothelium-dependent and endothelium-independent manners, respectively. In caudal artery, rCGRP also causes endothelium-independent cAMP elevations, suggesting that cAMP serves as a second messenger in rCGRP-induced relaxations of rat caudal artery. Recently, a fragment of human CGRP (hCGRPa_a7) has been proposed as a CGRP receptor antagonist. In the present study, we tested whether hCGRPs.37 can antagonize rCGRP-induced relaxations and cAMP elevations in rat caudal artery, rCGRP (0.3 to 300 nM) relaxed caudal arterial rings precontracted with norepinephrine (ECs0 for each ring, approx. 10 nM). hCGRPa.37 (3 pM) shifted to the right 14-fold the dose-response curve for rCGRP-induced vasodilation (ECso for rCGRP = 3 nM & 42 nM without and with hCGRPa.37). hCGRPa.37 completely inhibited relaxations at low concentrations (1 and 3 nM), but not high concentrations (100 and 300 nM), of rCGRP. Therefore, hCGRPs.37 inhibited rCGRP-induced relaxations of rat caudal artery in a competitive manner, hCGRPa.37 (3 pM) also significantly inhibited cAMP elevations in rat caudal artery induced by rCGRP (10 - 300 nM). We conclude that hCGRPs.37 is a competitive CGRP receptor antagonist in rat caudal artery and should be a useful pharmacological tool for determining the role of endogenous CGRP in rat models. (Supported by Stroke Program, Sanders-Brown Center on Aging, University of Kentucky)
Session IV: Cardiovascular and regional hemodynamic actions of CGRP 47 CHARACTERISTICS
OF
CGRP-INDUCED
RELAXATION
IN R A B B I T
OPHTHALMIC
ARTERY
A. Z S C H A U E R I, H. U U S I T A L O 1 and J.E. B R A Y D E N 2, iDepartment of Anatomy, U n i v e r s i t y of Helsinki, 00170 Helsinki, Finland, and 2 D e p a r t m e n t of Pharmacology, College of Medicine, The U n i v e r s i t y of Vermont, Burlington, V e r m o n t 05405, U.S.A. The e l e c t r o m e c h a n i c a l e f f e c t s of c a l c i t o n i n gene r e l a t e d p e p t i d e (CGRP) on intact and e n d o t h e l i u m d e n u d e d rabbit o p h t h a l m i c artery w e r e studied. This study was p e r f o r m e d u s i n g m i c r o e l e c t r o d e and isometric t e n s i o n r e c o r d i n g t e c h n i q u e s for m e a s u r i n g c h a n g e s in m e m b r a n e p o t e n t i a l and in c o n t r a c t i l e force. CGRP induced a d o s e d e p e n d e n t r e l a x a t i o n both on intact and e n d o t h e l i u m denuded arteries s t i m u l a t e d by 5 uM n o r e p i n e p h r i n e (NE). CGRP (i0 nM) h y p e r p o l a r i z e d the smooth m u s c l e m e m b r a n e by 20.2 ~ 9.0 mV (n=5) a n d 8 . 0 Z 0.9 m V (n=5) in e n d o t h e l i u m d e n u d e d and intact arteries, respectively. Both the r e l a x a t i o n and hyperp o l a r i z a t i o n induced b y C G R P was p a r t l y inhibited b y g l y b e n c l a m i d e , a b l o c k e r cf the A T P - s e n s i t i v e K channels. This result indicates that C G R P - i n d u c e d relaxation is p a r t l y due to a h y p e r p o l a r i z a t i o n t h r o u g h a c t i v a t i o n of A T P - s e n s i t i v e K c h a n n e l s as has b e e n shown by Nelson et al (1990). In intact arteries after w a s h o u t of CGRP the c o n t r a c t i l e r e a c t i v i t y to NE was increased whereas in e n d o t h e l i u m d e n u d e d a r t e r i e s the CGRP induced r e l a x a t i o n p r o l o n g e d and after w a s h o u t of CGRP the c o n t r a c t i l e s e n s i t i v i t y to NE was diminished. In intact a r t e r i e s NE c o n t r a c t i o n s w e r e e n h a n c e d by L - N M M A (L-N-monomethyl arginine), an i n h i b i t o r of EDRF (endothelium d e r i v e d r e l a x i n g factor) synthesis, and in the p r e s e n c e of L-NMMA, C G R P - i n d u c e d r e l a x a t i o n s r e s e m b l e d those seen in e n d o t h e l i u m d e n u d e d arteries. This result suggests that there is an i n c r e a s e d EDRF synthesis in intact a r t e r i e s d u r i n g N E - s t i m u l a t i o n and that CGRP may inhibit either the s y n t h e s i s or the a c t i v i t y of EDRF. I n c r e a s e d c o n t r a c t i l e s e n s i t i v i t y to NE after of CGRP c o u l d be e x p l a i n e d if r e c o v e r y of EDRF synthetic a c t i v i t y is slow.
48 hCGRPa_37 ANTAGONIZES VASODILATIONS AND cAMP RESPONSES TO RAT CALCITONIN GENE-RELATED PEP'ITDE (rCGRP) IN RAT CAUDAL ARTERY R.R. FISCUS, X. WANG and H. HAO. Sanders-Brown Res. Center on Aging & Dept. Physiol. & Biophysics, Univ. Kentucky College of Medicine, Lexington, KY 40536-0230, USA. CGRP is an endogenous and potent hypotensive neuropeptide in rat and human. In rat, rCGRP relaxes isolated aorta and caudal artery rings in endothelium-dependent and endothelium-independent manners, respectively. In caudal artery, rCGRP also causes endothelium-independent cAMP elevations, suggesting that cAMP serves as a second messenger in rCGRP-induced relaxations of rat caudal artery. Recently, a fragment of human CGRP (hCGRPa_a7) has been proposed as a CGRP receptor antagonist. In the present study, we tested whether hCGRPs.37 can antagonize rCGRP-induced relaxations and cAMP elevations in rat caudal artery, rCGRP (0.3 to 300 nM) relaxed caudal arterial rings precontracted with norepinephrine (ECs0 for each ring, approx. 10 nM). hCGRPa.37 (3 pM) shifted to the right 14-fold the dose-response curve for rCGRP-induced vasodilation (ECso for rCGRP = 3 nM & 42 nM without and with hCGRPa.37). hCGRPa.37 completely inhibited relaxations at low concentrations (1 and 3 nM), but not high concentrations (100 and 300 nM), of rCGRP. Therefore, hCGRPs.37 inhibited rCGRP-induced relaxations of rat caudal artery in a competitive manner, hCGRPa.37 (3 pM) also significantly inhibited cAMP elevations in rat caudal artery induced by rCGRP (10 - 300 nM). We conclude that hCGRPs.37 is a competitive CGRP receptor antagonist in rat caudal artery and should be a useful pharmacological tool for determining the role of endogenous CGRP in rat models. (Supported by Stroke Program, Sanders-Brown Center on Aging, University of Kentucky)