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Antagonism by Ca2+ of the inhibitory action of PGE1 on the electrically induced vasoconstriction in the rabbit ear artery
393
European Journal o f Pharmacology, 34 ( 1 9 7 5 ) 3 9 3 - - 3 9 5 © N o r t h - H o l l a n d Publishing C o m p a n y , A m s t e r d a m -- P r i n t e d in T h e N e t h e r l a n d s
Short communication ANTAGONISM BY Ca 2÷ OF THE INHIBITORY ACTION OF PGEI ON THE ELECTRICALLY INDUCED VASOCONSTRICTION IN THE RABBIT EAR ARTERY P. H A D H A Z Y , S. T O D O R O V a n d T. N/~DOR
Department o f Pharmacology, Semmelweis University o f Medicine, 1085 Budapest VIH, Hungary Received 16 S e p t e m b e r 1975, a c c e p t e d 9 O c t o b e r 1 9 7 5
P. HADH/~ZY, S. T O D O R O V a n d T. N/kDOR, Antagonism by Ca :+ o f the inhibitory action o f PGE1 on the electrically induced vasoconstriction in the rabbit ear artery, E u r o p e a n J. P h a r m a c o l . 34 ( 1 9 7 5 ) 3 9 3 - - 3 9 5 . T h e i n h i b i t o r y a c t i o n o f PGE1 o n v a s o c o n s t r i c t o r responses to nerve s t i m u l a t i o n of isolated ear arteries of r a b b i t s was s t u d i e d in t h e p r e s e n c e of 1.0 to 7.5 m M e x t e r n a l calcium c o n c e n t r a t i o n . Raising t h e [ C a ] 0 in t h e m e d i u m increased t h e electrically i n d u c e d v a s o c o n s t r i c t i o n a n d a n t a g o n i z e d t h e i n h i b i t o r y e f f e c t of PGEI o n c o n s t r i c t o r responses. This a n t a g o n i s m is p r o b a b l y due to an i n t e r a c t i o n b e t w e e n Ca 2+ a n d PGE1 in the sympat h e t i c nerves to t h e vessel. R a b b i t ear a r t e r y
Nerve s t i m u l a t i o n
Calcium
1. Introduction The prostaglandins E (PGEs) have been shown to modulate constrictor responses to nerve activity in different blood vessels (Hedqvist, 1972; Kadowitz et al., 1972). The inhibitory action of PGEs on adrenergic neurotransmission may be due to the decrease of the available calcium in the nerve endings (Hedqvist, 1970; Johnson et al., 1971; Stj~irne, 1973). Recently, it was found that PGE2 (De la Lande, 1975) and PGEI (Hadhdzy et al., in press) inhibited the nerve-evoked constriction of the rabbit ear artery as well as the stimulation-induced outflow of tritiated noradrenaline from the vessel (Hadhdzy et al., in press). The present work was done to study the calcium dependence of the inhibitory effect of PGEI on sympathetic neurotransmission in the isolated central ear artery of the rabbit.
2. Materials and methods A 3--4 cm portion of the rabbit ear artery was dissected free, cannulated at both ends,
PGEI
Vasoconstriction
m o u n t e d in an organ bath containing 4 ml Krebs solution and perfused with Krebs solution at a constant rate of 2.7 ml/min. The nutritive fluid was oxygenated with 5% CO2 in 02 and kept at 37°C. A DISA square wave generator was used for field stimulation. Trains of 10 pulses (0.5 msec in duration) were applied at 5 Hz every 2 min; the voltage was supramaximal (about 60 V). The resulting increase in pressure, which reflected vasoconstriction, was measured and recorded. PGE1 (Upjohn) was added to the bath in a cumulative manner. The external calcium concentration was varied from 1.0 to 7.5 mM. 3. Results Raising the [Ca] 0 in the bathing and perfusing fluid was accompanied by an increase in vasoconstrictor responses to sympathetic nerve stimulation. The magnitude of vasoconstriction was 11 + 1.7 (mean -+ S.E.), 20-+ 2.5, 31 +3.2, 34 -+ 5.7, and 45 +- 4.9 mm Hg in the presence of 1.0, 1.75, 2.5, 4.0 and 7.5 mM Ca 2÷ respectively.
4
394
P. H A D H A Z Y E T AL.
A mml-.Ig
30
10 rain : .
: .
.
-'-.
.
" ~
.
.
.
.
.
.
10 rain .................
~ .
.
.
.
.
.
.
.
.
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iI1441~I
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.
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I-'-'I:-'-';.'.'I-'-'~;i • i i i l l l l l l l l l l l l l l l
,IIlIJlIIHIIIIII -
-
i . . . . . . . . . . . . . . . .
HHH[H[~L-: .~..H~... !!~!!!!!':!!!!'!!!!!!!i- - ~{{~H~H[~H}I
0
ii-i'.'.-ii'.ili'.-iiiiii~..~ iiiiiiiiiiiiiiiii ii~iiiiiii~iiiiiiiiiiiii!iiiiiiiiii~!!!
~
............ :"....... " ' !
"'.:----'-::
2
/,
8
2
•
48
32
16
m H Co + +
I
PGE 1
¢~
w 2,5
nglmt
7,5 rnM Co + + !
4. Discussion
Fig. 1. A n t a g o n i s m b y Ca 2+ of t h e i n h i b i t o r y a c t i o n o f PGE] o n v a s o c o n s t r i c t o r r e s p o n s e s to nerve stimul a t i o n (5 Hz, 10 pulses) o f the p e r f u s e d ear a r t e r y of £he r a b b i t .
PGE] reduced the electrically induced vasoconstriction; the degree of this reduction was inversely related to [Ca] 0. Fig. 1 demonstrates the technique for establishing the interaction between PGE~ and calcium. At a higher calcium concentration, a higher a m o u n t of PGE~ was needed to produce an inhibition comparable to that obtained at a lower [Ca] 0. The quantitative relationship of the inhibitory effect of PGE1 and [Ca] 0 is presented in Fig. 2. The inhibition curve for PGE~ was shifted to
100 90
l
T
80
//' /./'
)ZI/'
/
"tO 60 50 40
~o/
/:/~,/
30 20 l
10
03
1,4
the right when the external calcium concentration was increased. The magnitude of this shift was not entirely proportional to the elevation of [Ca] 0 : it was more pronounced at the lower than at the higher range of [Ca] 0. The concentrations of PGE~ producing a 50% inhibition (ICs 0) were calculated from the curves. The ICs 0 values were: 2.1, 5.0, 15.4, 21 and 39 × 10 -9 M in the presence of 1.0, 1.75, 2.5, 4.0 and 7.5 mM Ca 2÷respectively.
2,8
5,5 X
10 -9
I1 1,1
22
45
gO
180
PGE 1
Fig. 2. I n h i b i t o r y e f f e c t o f POE1 o n v a s o c o n s t r i c t o r r e s p o n s e s t o s y m p a t h e t i c n e r v e s t i m u l a t i o n (5 Hz, 10 pulses), as a f u n c t i o n o f e x t e r n a l calcium c o n c e n t r a t i o n in p e r f u s e d ear arteries o f rabbits. T h e i n h i b i t i o n curves were o b t a i n e d in t h e p r e s e n c e o f 1.0 (e), 1.75 (©), 2.5 (A), 4.0 (~) a n d 7.5 (•) m M Ca 2+. Vertical bars i n d i c a t e s t a n d a r d errors.
The results described in this paper provide evidence that Ca 2÷ reduces the inhibitory action of PGE~ on vasoconstrictor responses to nerve stimulation of the rabbit ear artery. Raising the [Ca]0 has also been reported to antagonize the inhibitory effect of PGE2 on the release of noradrenaline by nerve stimulation from the cat spleen (Hedqvist, 1970) and from the vas deferens of the guinea pig (Stj~irne, 1973). Ca 2÷ may also have reduced the inhibitory action of PGE~ in our experiments by interfering with the secretion process of neurotransmission. This assumption is supported by the finding of Al Tai and Graham (1972) who reported that elevation of Ca 2÷failed to reduce significantly the dilator effect of PGE~ on the smooth muscle cells of the rabbit ear artery. We observed that the potency of PGE~ was inversely related to the calcium concentration (fig. 2); however the inhibitory action of PGE~ was more effectively antagonized by Ca 2÷ within the lower than the higher range of [Ca] 0. This is in accordance with the finding of Stj~irne (1973) who found that the inhibitory effect of PGE~ on stimulation-induced release of noradrenaline was a distinct function of the external Ca 2÷ when increasing the level of this ion up to 3--4 mM.
References AI Tai, S.A. a n d J.D.P. G r a h a m , 1 9 7 2 , T h e a c t i o n of p r o s t a g l a n d i n s E1 a n d F2~ o n t h e p e r f u s e d vessels
Ca--PGEI INTERACTION IN BLOOD VESSEL of the isolated rabbit ear, Brit. J. Pharmacol. 44, 699. De la Lande, I.S., R.C. Hall, J.A. Kennedy and G.D. Higgins, 1975, Prostaglandins, antipyretic analgesics and adrenergic stimuli on the isolated artery, European J. Pharmacol. 30,319. Hedqvist, P., 1970, Antagonism by calcium of the inhibitory action of prostaglandin E 2 on sympathetic neurotransmission in the cat spleen, Acta Physiol. Scand. 80, 269. Hedqvist, P., 1972, Prostaglandin-induced inhibition of vascular tone and reactivity in the cat's hindleg in vivo, European d. Pharmacol. 17, 157.
395 Johnson, D.G., N.B. Thoa, R. Weinshilboum, J. Axelrod and I.J. Kopin, 1971, Enhanced release of dopamine /3-hydroxylase from sympathetic nerves by calcium and phenoxybenzamine and its reversal by prostaglandins, Proc. Nat. Acad. Sci. U.S. 68, 2227. Kadowitz, P.J., C.S. Sweet and M.J. Brody, 1972, Influence of prostaglandins on adrenergic transmission to vascular smooth muscle, Circulation Res. 31, Suppl. II, 36. Stj/irne, L., 1973, Inhibitory effect of prostaglandin E 2 on noradrenaline secretion from sympathetic nerves as a function of external calcium, Prostaglandins 3, 105.
European Journal o f Pharmacology, 34 ( 1 9 7 5 ) 3 9 3 - - 3 9 5 © N o r t h - H o l l a n d Publishing C o m p a n y , A m s t e r d a m -- P r i n t e d in T h e N e t h e r l a n d s
Short communication ANTAGONISM BY Ca 2÷ OF THE INHIBITORY ACTION OF PGEI ON THE ELECTRICALLY INDUCED VASOCONSTRICTION IN THE RABBIT EAR ARTERY P. H A D H A Z Y , S. T O D O R O V a n d T. N/~DOR
Department o f Pharmacology, Semmelweis University o f Medicine, 1085 Budapest VIH, Hungary Received 16 S e p t e m b e r 1975, a c c e p t e d 9 O c t o b e r 1 9 7 5
P. HADH/~ZY, S. T O D O R O V a n d T. N/kDOR, Antagonism by Ca :+ o f the inhibitory action o f PGE1 on the electrically induced vasoconstriction in the rabbit ear artery, E u r o p e a n J. P h a r m a c o l . 34 ( 1 9 7 5 ) 3 9 3 - - 3 9 5 . T h e i n h i b i t o r y a c t i o n o f PGE1 o n v a s o c o n s t r i c t o r responses to nerve s t i m u l a t i o n of isolated ear arteries of r a b b i t s was s t u d i e d in t h e p r e s e n c e of 1.0 to 7.5 m M e x t e r n a l calcium c o n c e n t r a t i o n . Raising t h e [ C a ] 0 in t h e m e d i u m increased t h e electrically i n d u c e d v a s o c o n s t r i c t i o n a n d a n t a g o n i z e d t h e i n h i b i t o r y e f f e c t of PGEI o n c o n s t r i c t o r responses. This a n t a g o n i s m is p r o b a b l y due to an i n t e r a c t i o n b e t w e e n Ca 2+ a n d PGE1 in the sympat h e t i c nerves to t h e vessel. R a b b i t ear a r t e r y
Nerve s t i m u l a t i o n
Calcium
1. Introduction The prostaglandins E (PGEs) have been shown to modulate constrictor responses to nerve activity in different blood vessels (Hedqvist, 1972; Kadowitz et al., 1972). The inhibitory action of PGEs on adrenergic neurotransmission may be due to the decrease of the available calcium in the nerve endings (Hedqvist, 1970; Johnson et al., 1971; Stj~irne, 1973). Recently, it was found that PGE2 (De la Lande, 1975) and PGEI (Hadhdzy et al., in press) inhibited the nerve-evoked constriction of the rabbit ear artery as well as the stimulation-induced outflow of tritiated noradrenaline from the vessel (Hadhdzy et al., in press). The present work was done to study the calcium dependence of the inhibitory effect of PGEI on sympathetic neurotransmission in the isolated central ear artery of the rabbit.
2. Materials and methods A 3--4 cm portion of the rabbit ear artery was dissected free, cannulated at both ends,
PGEI
Vasoconstriction
m o u n t e d in an organ bath containing 4 ml Krebs solution and perfused with Krebs solution at a constant rate of 2.7 ml/min. The nutritive fluid was oxygenated with 5% CO2 in 02 and kept at 37°C. A DISA square wave generator was used for field stimulation. Trains of 10 pulses (0.5 msec in duration) were applied at 5 Hz every 2 min; the voltage was supramaximal (about 60 V). The resulting increase in pressure, which reflected vasoconstriction, was measured and recorded. PGE1 (Upjohn) was added to the bath in a cumulative manner. The external calcium concentration was varied from 1.0 to 7.5 mM. 3. Results Raising the [Ca] 0 in the bathing and perfusing fluid was accompanied by an increase in vasoconstrictor responses to sympathetic nerve stimulation. The magnitude of vasoconstriction was 11 + 1.7 (mean -+ S.E.), 20-+ 2.5, 31 +3.2, 34 -+ 5.7, and 45 +- 4.9 mm Hg in the presence of 1.0, 1.75, 2.5, 4.0 and 7.5 mM Ca 2÷ respectively.
4
394
P. H A D H A Z Y E T AL.
A mml-.Ig
30
10 rain : .
: .
.
-'-.
.
" ~
.
.
.
.
.
.
10 rain .................
~ .
.
.
.
.
.
.
.
.
.~i~..L~ Ji~iiii:.i:. .~.~--:::
iI1441~I
"..,,II,|
.
:JiJ'JJiiiiiJJJJ
I-'-'I:-'-';.'.'I-'-'~;i • i i i l l l l l l l l l l l l l l l
,IIlIJlIIHIIIIII -
-
i . . . . . . . . . . . . . . . .
HHH[H[~L-: .~..H~... !!~!!!!!':!!!!'!!!!!!!i- - ~{{~H~H[~H}I
0
ii-i'.'.-ii'.ili'.-iiiiii~..~ iiiiiiiiiiiiiiiii ii~iiiiiii~iiiiiiiiiiiii!iiiiiiiiii~!!!
~
............ :"....... " ' !
"'.:----'-::
2
/,
8
2
•
48
32
16
m H Co + +
I
PGE 1
¢~
w 2,5
nglmt
7,5 rnM Co + + !
4. Discussion
Fig. 1. A n t a g o n i s m b y Ca 2+ of t h e i n h i b i t o r y a c t i o n o f PGE] o n v a s o c o n s t r i c t o r r e s p o n s e s to nerve stimul a t i o n (5 Hz, 10 pulses) o f the p e r f u s e d ear a r t e r y of £he r a b b i t .
PGE] reduced the electrically induced vasoconstriction; the degree of this reduction was inversely related to [Ca] 0. Fig. 1 demonstrates the technique for establishing the interaction between PGE~ and calcium. At a higher calcium concentration, a higher a m o u n t of PGE~ was needed to produce an inhibition comparable to that obtained at a lower [Ca] 0. The quantitative relationship of the inhibitory effect of PGE1 and [Ca] 0 is presented in Fig. 2. The inhibition curve for PGE~ was shifted to
100 90
l
T
80
//' /./'
)ZI/'
/
"tO 60 50 40
~o/
/:/~,/
30 20 l
10
03
1,4
the right when the external calcium concentration was increased. The magnitude of this shift was not entirely proportional to the elevation of [Ca] 0 : it was more pronounced at the lower than at the higher range of [Ca] 0. The concentrations of PGE~ producing a 50% inhibition (ICs 0) were calculated from the curves. The ICs 0 values were: 2.1, 5.0, 15.4, 21 and 39 × 10 -9 M in the presence of 1.0, 1.75, 2.5, 4.0 and 7.5 mM Ca 2÷respectively.
2,8
5,5 X
10 -9
I1 1,1
22
45
gO
180
PGE 1
Fig. 2. I n h i b i t o r y e f f e c t o f POE1 o n v a s o c o n s t r i c t o r r e s p o n s e s t o s y m p a t h e t i c n e r v e s t i m u l a t i o n (5 Hz, 10 pulses), as a f u n c t i o n o f e x t e r n a l calcium c o n c e n t r a t i o n in p e r f u s e d ear arteries o f rabbits. T h e i n h i b i t i o n curves were o b t a i n e d in t h e p r e s e n c e o f 1.0 (e), 1.75 (©), 2.5 (A), 4.0 (~) a n d 7.5 (•) m M Ca 2+. Vertical bars i n d i c a t e s t a n d a r d errors.
The results described in this paper provide evidence that Ca 2÷ reduces the inhibitory action of PGE~ on vasoconstrictor responses to nerve stimulation of the rabbit ear artery. Raising the [Ca]0 has also been reported to antagonize the inhibitory effect of PGE2 on the release of noradrenaline by nerve stimulation from the cat spleen (Hedqvist, 1970) and from the vas deferens of the guinea pig (Stj~irne, 1973). Ca 2÷ may also have reduced the inhibitory action of PGE~ in our experiments by interfering with the secretion process of neurotransmission. This assumption is supported by the finding of Al Tai and Graham (1972) who reported that elevation of Ca 2÷failed to reduce significantly the dilator effect of PGE~ on the smooth muscle cells of the rabbit ear artery. We observed that the potency of PGE~ was inversely related to the calcium concentration (fig. 2); however the inhibitory action of PGE~ was more effectively antagonized by Ca 2÷ within the lower than the higher range of [Ca] 0. This is in accordance with the finding of Stj~irne (1973) who found that the inhibitory effect of PGE~ on stimulation-induced release of noradrenaline was a distinct function of the external Ca 2÷ when increasing the level of this ion up to 3--4 mM.
References AI Tai, S.A. a n d J.D.P. G r a h a m , 1 9 7 2 , T h e a c t i o n of p r o s t a g l a n d i n s E1 a n d F2~ o n t h e p e r f u s e d vessels
Ca--PGEI INTERACTION IN BLOOD VESSEL of the isolated rabbit ear, Brit. J. Pharmacol. 44, 699. De la Lande, I.S., R.C. Hall, J.A. Kennedy and G.D. Higgins, 1975, Prostaglandins, antipyretic analgesics and adrenergic stimuli on the isolated artery, European J. Pharmacol. 30,319. Hedqvist, P., 1970, Antagonism by calcium of the inhibitory action of prostaglandin E 2 on sympathetic neurotransmission in the cat spleen, Acta Physiol. Scand. 80, 269. Hedqvist, P., 1972, Prostaglandin-induced inhibition of vascular tone and reactivity in the cat's hindleg in vivo, European d. Pharmacol. 17, 157.
395 Johnson, D.G., N.B. Thoa, R. Weinshilboum, J. Axelrod and I.J. Kopin, 1971, Enhanced release of dopamine /3-hydroxylase from sympathetic nerves by calcium and phenoxybenzamine and its reversal by prostaglandins, Proc. Nat. Acad. Sci. U.S. 68, 2227. Kadowitz, P.J., C.S. Sweet and M.J. Brody, 1972, Influence of prostaglandins on adrenergic transmission to vascular smooth muscle, Circulation Res. 31, Suppl. II, 36. Stj/irne, L., 1973, Inhibitory effect of prostaglandin E 2 on noradrenaline secretion from sympathetic nerves as a function of external calcium, Prostaglandins 3, 105.