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Inhibitory effects of piribedil on adrenergic neurotransmission
European Journal of Pharmacology, 52 (1978) 99--107
99
© Elsevier/North-Holland Biomedical Press
INHIBITORY EFFECTS OF PIRIBEDIL ON ADRENERGIC NEUROTRANSMISSION MICHEL LAUBIE a:ad HENRI SCHMITT
Department of Cardiovascular Pharmacology, I.R.S., 14 Rue du Val d'Or, 92150 Suresnes, France Received 17 March 1978, revised MS received 4 July 1978, accepted 6 July 1978
M. LAUBIE and H. SCHMITT, Inhibitory effects of piribedil on adrenergic neurotmnsmission, European J. Pharmacol. 52 (1978) 99--107. The effects of piribedil on responses to sympathetic stimulation were investigated in anaesthetized dogs. Piribedil (1 mg/kg i.v.) impaired the vasoconstrictor responses to lumbar sympathetic chain stimulation of the perfused hindlimb without changing the effects of noradrenaline. Piribedil (2 mg/kg i.v.) depressed the chronotropic responses to stimulation of the right anterior ansa and the inotropic response to stimulation of the left anterior ansa. Stimulation of the splanchnic nerve induced frequency dependent increases in systemic blood pressure. Piribedil antagonized this effect. Piribedil (1 mg/kg i.v.) attenuated the constrictor responses of the perfused mesenteric artery to postganglionic sympathetic stimulation and reduced the decreases in renal blood flow caused by stimulation of sympathetic renal nerves. The inhibitory effects of piribedil were preferential on responses induced by low frequency stimulation of nerves. The hypertensive, vasoconstrictor and tachycardic effects of noradrenaline and tyramine were not affected. The effects of piribedil were reversed by haloperidol (0.5 mg/kg i.v.) or pimozide (0.2 mg]kg i.v.). Adrenal medulla
Piribedil
Presynaptic dopamine receptors
1. Introduction Piribedil was shown to increase femoral blood flow when injected in small doses into the femoral artery, likely by an action on dopamine presynpatic receptors (Laubrie et al., 1977). This effect of piribedil was abolished by inhibition of the sympathetic tone (Laubie et al., 1969, 1977) and in addition it was antagonized by previous administration of the dopamine receptor blocking agents, haloperidol and pimozide (Buylaert, 1977; Laubie et al., 1977). It was therefore suggested that piribedil induced femoral vasodilatation by inhibiting the release of noradrenaline probably by an action on dopamine presynaptic receptors. However, it is not yet known whether piribedil could inhibit sympathetic transmission in other cardiovascular areas. Dopamine is known to impair peripheral sympathetic transmission by reducing the
Sympathetic nerve stimulation
release of noradrenaline from postganglionic sympathetic nerves and by an action on sympathetic ganglia. Due to the experimental conditions it was not possible to distinguish the part played by each mechanism in all the experiments reported here. However it is clearly shown that piribedil could reduce the effect of sympathetic nerve stimulation in all the cardiovascular areas investigated. Dopamine and the dopaminergic agonistic agent, apomorphine have been shown to reduce the release of noradrenaline induced by stimulation of sympathetic nerves in many in vitro preparations e.g., isolated rabbit ear artery (Mc Culloch et al., 1973; Hope et al., 1975), isolated nictating membrane of the cat (Enero and Langer, 1975). In addition, haloperidol has been shown to increase the response of the guinea pig atria to transmural stimulation (Ilhan et al., 1976a,b) suggesting a negative feedback mechanism. This mechanism has been shown to operate
100 in vivo. Do p a m i ne and t he dopaminergic agonistic agents, a p o m o r p h i n e and 5,6-dihyd r o x y - 2 - m e t h y l a m i n o t e t r a l i n e reduced the cardioacceleration elicited by stimulation of the cardiac nerves in cats and dogs (Long et al., 1975), the c o n t r a c t i o n o f t he spleen induced by stimulation of t he splenic nerves in dogs (Sharabi et al., 1977) and the decrease in renal blood flow induced by stimulation of the renal nerves (Lokhandwala and Buckley, 1977). These experiments suggested t hat the inhibition o f noradrenaline release by dopaminergic agonists m a y be a general phenomenon. F u r t h e r evidence for this view is provided by th e present experiments on the effects o f a dopaminergic agonist, piribedil on the response o f different cardiovascular regions to s y m p a t h e t i c stimulation.
2. Materials and m e t h o d s
2.1. General procedures Mongrel dogs of either sex weighing 15--21 kg were anaesthetized with pentobarbital (30 mg/kg i.e.). The trachea was i nt ubat ed and the dogs were ventilated with a Mark VII Bird respirator. Blood pressure was recorded by means o f a c a t he t e r i n t r o d u c e d into the abdominal aorta via the right femoral artery and c o n n e c t e d to a Statham P23 Db pressure transducer on one channel o f a Brush 440 recorder. Mean blood pressure was obtained by electrical integration of t he blood pressure.
2.2. Perfusion o f the hindlimb After transection of the spinal cord (C~-C2) a p o l y e t h y l e n e c a t he t e r was i n t r o d u c e d into the abdominal aorta via t he left femoral artery; the caudal end o f the cat het er was inserted into the distal end o f the femoral artery. Blood was infused f r om the aorta to the femoral artery by means of a Sigmamotor pump. The perfusion pressure was measured by means o f a Statham P23 Db transducer and was adjusted to 100 m m Hg at t he
M. LAUBIE, H. SCHMITT beginning of the experiment. The left lumbar sym pat het i c t runk was exposed at the level of L3 and placed on a bipolar platinum electrode. The nerve was stimulated at supramaximal voltage (7 V) for 20 sec at frequencies ranging from 0.5 to 16 Hz and a pulse duration of 0.5 msec.
2.3. Stimulation of the sympathetic cardiac nerves In these experiments, the spinal cord was transected at the level of C1-C: and bot h vagus nerves cut. The chest was opened on the right or the left side at the level of the second intercostal space. Either the left or the right anterior ansa was isolated, placed on a pair of platinum electrodes and stimulated at supramaximal voltage (7 V), at frequencies from 0.3 to 10 Hz with a pulse width of 0.5 msec for 20 sec. Stimulation of t he right anterior ansa induced mainly an increase in heart rate as c o u n t e d on the pulse record. In the group o f dogs in which the left anterior ansa was stimulated, a cat het er was i nt roduced into the left ventricle via the left carotid artery. The maximal rate of rise of the left ventricular pressure ( d p / d t max) was recorded by differentiating the signal of the left ventricular pressure. Electrical stimulation of the left anterior ansa induced a m arked inotropic response with a minimal increase in heart rate (Norris and Randall, 1977).
2.4. Perfusion o f the mesenteric artery The mesenteric artery was isolated, its distal end cannulated and infused with bl ood taken from the abdominal aorta. A Si gm am ot or pum p was used to infuse the blood at a constant flow rate and the perfusion pressure was measured by means o f a Statham P23 Db transducer. The perfusion pressure was adjusted at 100 m m Hg at the beginning of the experiment. The mesenteric sympathetic postganglionic nerves were ligated and cut distally to the coeliac ganglion. The nerves were stimulated at supramaximal voltage (7 V)
DOPAMINE RECEPTORS AND ADRENERGIC MECHANISMS
at frequencies from 4 to 16 Hz, a pulse width of 1 msec for 30 sec.
2.5. Stimulation of the splanchnic nerve The spinal cord was transected at the level of CI-C2, the splanchnic nerve was isolated at its exit from the diaphragm, cut and its distal end placed on a pair of platinum electrodes. The nerve was stimulated at supramaximal voltage (7 V) at frequencies ranging from 1 to 8 Hz, a pulse width of 0.5 msec for 30 sec.
2.6. Stimulation nerves
of
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3. Results
3.1. Effects of piribedil, haloperidol and pimozide on blood pressure and heart rate Piribedil (1--2 mg/kg i.e.) was administered slowly; under these conditions, blood pressure and heart rate did not change significantly. Haloperidol (0.5 mg/kg i.e.) and pimozide (0.2 mg/kg i.e.) induced a transient decrease in blood pressure; stimulation of the sympathetic nerves was always performed after the blood pressure had recovered.
the renal sympathetic
The left kidney was exposed b y a flank incision, the renal artery was dissected and an electromagnetic flow probe (Statham, internal diameter 3 mm) was placed on the renal artery near the aorta. The renal nerves were isolated and a ligature placed at their central end. The renal plexus was placed on a pair of platinum electrodes and stimulated at supramaximal voltage (7 V) at frequencies from 1 to 4 Hz, a pulse width of 1 msec for 20 sec. The renal blood flow was reduced at frequencies of stimulation greater tha 1 Hz.
2.7. Drugs used The following drugs were used: piperoxan hydrochloride (synthetized b y Dr. R6gnier, Servier), piribedil hydrochloride (Servier), haloperidol hydrochloride (Janssen Pharm.), pimozide hydrochloride (Janssen Pharm.), (--)noradrenaline bitartrate (RhSne Poulenc) and tyramine hydrochloride (RhSne Poulenc). All solutions were made in saline except for pimozide which was dissolved in a solution of tartaric acid (10% W/V) and diluted to 5 ml with 5% glucose solution. The doses of the drugs are given as the base.
2.8. Statistical analyses Statistical analyses were performed using Student's t-test for paired comparisons.
3.2. Effect o f piribedil on the pressor response and the tachycardia induced by noradrenaline and tyramine In dogs with both vagus nerves cut, piribedil (2 mg/kg i.e.) did n o t change the pressor responses to noradrenaline (0.5 to 2 pg/kg i.e.) and tyramine (100/~g/kg i.e.) (5 experiments). In dogs with spinal cord transection and both vagus nerves cut, piribedil (2 mg/kg i.e.) did not change the pressor response and the tachycardia after noradrenaline and tyramine (5 experiments; table 1).
3.3. Effects of piribedil on the responses o f the perfused hindlimb to stimulation of the lumbar sympathetic chain In dogs with transection of the spinal cord, electrical stimulation of the lumbar sympathetic chain induced a frequency dependent increase in the perfusion pressure of the hindlimb. The average increase in perfusion pressure ranged from 25 + 3 mm Hg at 0.5 Hz to 120 + 11 mm Hg at 16 Hz. Piribedil (1 mg/ kg i.e.) induced a transient increase in blood pressure from 90 + 4 to 120 + 10 mm Hg and in the perfusion pressure of the hindlimb from 1 0 4 + 5 to 1 1 5 + 7 m m H g . After recovery, the increases in perfusion pressure induced b y stimulation of the lumbar sympathetic nerve were reduced (fig. 1--7 experiments). This reduction was greater at low frequencies of stimulation (0.5--4 Hz) that at
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M. L A U B I E , H. S C H M I T T
TABLE 1 Effects of piribedil (2 m g / k g i.v.) on the tachycardia and hypertensive responses to noradrenaline and t y r a m i n e in dogs with spinal cord transection and b o t h vagus nerves cut (5 experiments). Each value is the mean ± S.E.M. Increase in b l o o d pressure (A m m Hg * S.E.M.)
Increase in heart rate (A b e a t s / m i n ± S.E.M.)
Before piribedil
After piribedil
Before piribedil
A f t e r piribedil
Noradrenaline (p g/kg i.v.) 0.1 0.3 1.0
23+_ 5 35+_ 8 66 _+ 16
26_+ 6 34_+ 7 65 _+ 16
10+_4 20+_5 40 +_ 9
8+_ 1 20_+ 6 44 + 13
Tyramine (pg/kg i.v.) 50 100
32+_ 9 74+_15
34_+ 7 69+10
19+_6 38_+6
21+_ 39+_
high frequencies. In contrast, piribedil did not change the increase in perfusion pressure (60 + 4 mm Hg) induced by injection of noradrenaline (3 pg) into the femoral artery. Haloperidol (0.5 mg/kg i.v.) antagonized the effect o f piribedil. After haloperidol the frequency response curve to lumbar chain stimulation was not significantly different from the control one.
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Frequence of nerve stimulation Fig. 1. F r e q u e n c y d e p e n d e n t increase in t h e perfusion pressure in the hindlimb (7 e x p e r i m e n t s ) and in the mesenteric artery (7 experiments) elicited by stimulat i o n o f the s y m p a t h e t i c nerves, o, Effect after piribedil (1 m g / k g i.v.); A, reversal effect o f haloperidol (0.5 m g / k g i.v.). The significance o f differences f r o m the c o n t r o l is r e p r e s e n t e d by asterisks (P < 0.01). e, Control.
7 5
3.4. Effects o f piribedil on the responses to stimulations o f the cardiac sympathetic nerves In dogs with transection of the spinal cord and with both vagus nerves cut, stimulation of the right anterior ansa induced a frequency dependent increase in heart rate of 19 + 4 beats/min at 0.3 Hz and 130 + 5 beats/min at 10 Hz. Piribedil (2 mg/kg i.v.) did not change the basal heart rate but significantly reduced the tachycardia induced by stimulation of the cardiac sympathetic nerve at low frequencies stimulation (0.3--3 Hz) but not at the higher frequency of 10 Hz (fig. 2--5 experiments). Piribedil did not change the dose dependent increase in heart rate induced by noradrenaline and tyramine (5 experiments; table 1). After haloperidol (0.5 mg/kg i.v.}, the heart rate response curve to nerve stimulation was not significantly different from the control one at any o f the frequencies investigated. Stimulation of the left anterior ansa produced a marked inotropic response with a minimal chronotropic effect. The maximal rate of rise in left ventricular pressure (max. dp/dt) increased by 110 + 20 mm Hg/sec. at 0.3 Hz and by 3,300 mm Hg/sec at 10 Hz.
DOPAMINE RECEPTORS AND ADRENERGIC MECHANISMS I
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Fig. 2. Left panel: increase in heart rate elicited by stimulation of the right anterior ansa. D, Piribedil (2 mg/kg i.v.) attenuated the response between 0.3-3 Hz but did not change the cardioaccelerator effect at 10 Hz. A Haloperidol (0.5 mg/kg i.v.)antagonized the inhibitory effect of piribedil (5 experiments). e, Control. Right panel: increase in max. dp/dt induced by stimulation of the left anteror ansa. El After piribedil ( 2 mg/kg i.v.); A, reversal effect of haloperidol (0.5 mg/kg i.v.) (6 experiments). The significance of differences from the control is represented by asterisks (P < 0.01). e, Control.
Piribedil (2 mg/kg i.v.) did not change the basal maximal rate of rise in left ventricular pressure b u t significantly reduced the effects of the nerve stimulation at all the frequencies investigated; the frequency response curve was shifted to the right. A subsequent administration of haloperidol (0.5 mg/kg i.v.) antagonized the inhibitory effect of piribedil. After haloperidol the frequency response curve to sympathetic stimulation was not significantly different from the control one (fig. 2 - 6 experiments).
3.5. Stimulation of the splanchnic nerve In the anaesthetized dogs with the spinal cord transected and b o t h vagus nerves cut, stimulation of the splanchnic nerve increased the systemic blood pressure in a frequency dependent manner. The increase in blood pressure was 22 -+ 1 mm Hg at 1 Hz and 98 + 7 mm Hg at 8 Hz. Piribedil (1 mg/kg i.v.) significantly reduced the pressor responses to splanchnic nerve stimulation at all the frequencies used. In contrast piribedil did not
0
1 2 4 8Hz Fig. 3. Systemic hypertensive response elicited by stimulation of the splanchnic nerve in spinal cordtransected dogs. D, PiribedLl (1 mg/kg i.v.) shifted the frequency response cuwe to the right; ~, pimozide (0.2 mg/kg i.v.) antagonized the effects of p~ibedll (6 experiments). The significance of differences from the control is represented by asterisks (P < 0.01 ). e, Control.
change the pressor dose response to noradrenaline and to tyramine (table 1). Pimozide (0.2 mg/kg i.v.) administered subsequently could antagonize the effect of piribedil. The pressor response curves to splanchnic nerve stimulation were not significantly different before piribedil and after pimozide (fig. 3--6 experiments).
3.6. Perfusion o f the mesenteric artery Stimulation of the postganglionic sympathetic nerves increased the perfusion pressure of the mesenteric artery b y 28 + 4 mm Hg at 4 Hz and 106 + 9 mm Hg at 16 Hz. Piribedil (1 mg/kg i.v.) induced a transient increase in the perfusion pressure of the mesenteric artery (43 + 8 mm Hg). Stimulation of the mesenteric nerves was performed after the recovery of the perfusion pressure (20 min). Piribedil reduced the response to stimulation of the sympathetic nerves at all frequencies of stimulation investigated; the frequency response curve was shifted to the right. In contrast piribedil did not significantly change
104
M. L A U B I E , H. S C H M I T T
the increase in the perfusion pressure (70 + 9 mm Hg) induced by injection of noradrenaline (0.3 pg) into the mesenteric artery. The subsequent administration of pimozide (0.2 mg/kg i.e.) induced the recovery of the pressor response to stimulation of the mesenteric sympathetic nerves (fig. 1--7 experiments).
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3.7. Stimulation nerves
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renal sympathetic
Stimulation of the renal sympathetic nerves caused a frequency d e p e n d e n t reduction in renal blood flow. Renal blood flow decreased from 143 + 20 ml/min to 128 + 14 ml/min at a stimulation frequency of 1 Hz and from 141 + 24 ml/min to 72 -+ 10 ml/min at a frequency of 4 Hz. Piribedil (1 mg/kg i.e.) did not change the renal blood flow but the drug significantly reduced the vasoconstrictor responses to renal sympathetic nerves stimulation (fig. 4--6 experiments). Noradrenaline (0.1 pg/kg i.e.) decreased the renal b l o o d flow from 153 + 21 ml/min to 108 + 14 ml/min and noradrenaline (0.3 pg/ kg i.e.) decreased the renal blood flow from 149 + 22 ml/min to 70 +- 14 ml/min. These renal vasoconstrictor responses to noradrenaline were n o t altered by piribedil (1 mg/kg i.e.) (6 experiments. The subsequent adminis-
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Fig. 4. R e d u c t i o n in renal artery b l o o d flow elicited by postganglionic s y m p a t h e t i c nerve stimulation, c~, A f t e r piribedil (1 m g / k g i.e.), the renal vasoconstrictor responses were a t t e n u a t e d ; ~, p i m o z i d e (0.2 rag/ kg i.e.) antagonized the effects of piribedil (6 experiments). The significance of differences from the control is represented by asterisks (P < 0.01). $, Control.
tration of pimozide (0.2 mg/kg i.e.) induced a recovery of the vasoconstrictor responses to sympathetic stimulation and the frequency response curves to sympathetic stimulation obtained in the control period and after injection of pimozide were not significantly different.
TABLE 2 Effects of haloperidol (0.5 m g / k g i.e.) and p i p e r o x a n (0.5 m g / k g i.e.) on the inhibitory effects of clonidine (15 }Jg/kg i.e.) on the cardioacceleration elicited by cardiac s y m p a t h e t i c stimulation in dogs with spinal cord transe c t e d and b o t h vagus nerves cut (5 experiments). Mean values -+ S.E.M. are given. F r e q u e n c y of stimulation (Hz)
0.3 1 3 10
Increase in heart rate (A b e a t s / m i n +- S.E.M.) Before i.e. injection
A f t e r clonidine 1
After haloperidol 2
After p i p e r o x a n 1
11+ 4 33-+ 7 81 -+ 11 116 _+ 13
2 -+ 1 3 11_+ 3 3 50 -+ 10 3 92 -+ 11 3
2-+ 1 11-+ 5 52 -+ 11 93 -+ 10
10-+ 3 32-+ 6 83 -+ 13 123 -+ 18
1 C o m p a r e d with before clonidine. 2 C o m p a r e d with after clonidine. a p < 0.01.
DOPAMINE RECEPTORS AND ADRENERGIC MECHANISMS 3.8. Action o f haloperidol on presynaptic aadrenoceptors
In dogs with the spinal cord transected and both vagus nerves cut, clonidine (15 pg/kg i.v.) significantly reduced the tachycardia elicited by stimulation of the cardiac sympathetic nerve. The subsequent injection of haloperidol (0.5 mg/kg i.v.) did not change the inhibitory effect of clonidine. In contrast piperoxan (0.5mg/kg i.v.) injected after haloperidol antagonized the effects of clonidine and the frequency responses curves in the control period and after injection of piperoxan were not significantly different (table 2--5 experiments).
4. Discussion
Piribedil has been suggested to cause femoral vasodilatation by an action on presynaptic dopamine receptors, probably by reducing noradrenaline release from the postganglionic sympathetic fibres (Laubie et al., 1977; Buylaert, 1977). The present experiments extend these findings and show an impairement of sympathetic transmission by piribedil in several sympathetic nerves. These experiments are compatible with a reduction of noradrenaline release by an action of piribedil on dopamine receptors. An effect of piribedil on postsynaptic sites as the cause of the reduction of the effects of sympathetic stimulation could be ruled out. In fact, piribedil did not change the pressor dose-response curve to noradrenaline in any of the experiments. An action of piribedil on the postganglionic sympathetic fibres is likely. In fact the mesenteric and renal sympathetic nerves are known to be composed of postganglionic fibres and piribedil reduced the effects of sympathetic stimulation. These experiments suggest that piribedil reduced noradrenaline release. Two other findings also support this view. Piribedil was shown to be more effective to reduce the effects of sympathetic stimulation at low frequencies than it
105
was at high frequencies. It is well known (Langer, 1977; Starke et al., 1977; Westfall, 1977) that the activation of presynaptic ~adrenoceptors or dopamine receptors reduced the noradrenaline release induced by low frequencies of stimulation more efficiently than they did that induced at high frequencies. In addition, in contrast to its action on sympathetic nerve stimulation, piribedil did not change the pressure and heart rate increases in response to tyramine. Tyramine is known to release noradrenaline from postganglionic sympathetic nerves by a calcium non-dependent process in contrast to noradrenaline release elicited by nerve stimulation (Lindmar et al., 1967). The splanchnic nerve and the lumbar sympathetic chain contain preganglionic sympathetic fibres and in the experiments performed by stimulating these nerves, piribedil may also have impaired sympathetic transmission by an action on the ganglia. In fact dopamine is known to impair ganglionic transmission (Bogaert et al., 1977) and it remains to be established that piribedil acts on glanglia. The effects of piribedil appear to be due to a dopaminergic mechanism. In fact, the classical dopamine receptor blocking agents, haloperidol and pimozide abolished the inhibitory effects of piribedil on sympathetic transmission, It seems unlikely that these agents, in the doses used in our experiments, blocked presynaptic a-adrenoceptors. They did not change the inhibitory effects of clonidine, an ~-adrenoceptor agonist, on the cardioacceleration elicited by cardiac sympathetic stimulation. Therefore, the present results suggest that piribedil reduced noradrenaline release by stimulating presynaptic dopamine receptors. Besides, piribedil reduced the effects of stimulation of seven sympathetic nerves suggesting the generality of the process. In fact, there are conflicting results on the existence and on the role of dopamine receptors. The existence of postsynaptic dopamine receptors in the femoral vascular bed has thus b e e n : ~ u t e d . McNay and Goldberg (1966) reported that dopamine injected into the
106
femoral artery of dogs treated with an aadrenoceptor blocking agent induced vasodilation which was abolished by f~-adrenoceptor blockade, suggesting the absence of specific dopamine receptors. In contrast Bell et al. (1975) found an increase in femoral blood flow in some dogs after a- and fladrenoceptor block with phentolamine and propranolol respectively; they concluded therefore that specific dopamine receptors may be present in the femoral bed. Our previous studies (Laubie et al., 1977) using apomorphine and piribedil however failed to detect postsynaptic dopamine receptors in this vascular bed. Similar conflicting results have been published concerning the presence of presynaptic dopamine receptors. Dopamine infused into the autoperfused hindlegs of cats did not change the increases in perfusion pressure induced by stimulation of the lumbar sympathetic chain (Haeusler 1976). However, dopamine presynaptic receptors have been detected in the femoral bed of dogs (Laubie et al., 1977; Buylaert et al., 1977). Apomorphine and- piribedil injected into the femoral artery induced a dose-dependent increase in femoral blood flow. This effect was abolished by section of the motor nerves, transection of the spinal cord and ganglionic blockade. The dilator effect of piribedil and apomorphine was also antagonized by haloperidol and pimozide. The present experiments indicate that piribedil impaired the pressor responses to sympathetic nerve stimulation of the perfused hinlimb of the dog and preferentially, the responses to low frequencies of stimulation; in addition haloperidol and pimozide antagonized the effects of piribedil. Similar results have been obtained with the cyclic analog of dopamine GJH 166 (Sharabi et al., 1977). Therefore the experiments reported are compatible with the view that pix-ibedil reduced noradrenaline release by stimulating presynaptic dopamine receptors during spontaneous or evoked sympathetic discharges. Previous studies have suggested the presence of presynaptic dopamine receptors
M. LAUBIE, H. SCHMITT
located on the postganglionic terminals of the cardiac nerves. In fact, dopamine in presence of cocaine, dopamine analogs or apomorphine {Long et al., 1975; Ilhan et al., 1976a) have been shown to reduce the chronotropic response to 2 Hz stimulation of the'right cardioaccelerator nerve in cats and dogs. In the present study piribedil reduced the tachycardia induced by stimulation of the right anterior ansa at low frequencies (0.3--3 Hz) but was ineffective when the stimulation frequency was 10 Hz. Piribedil also reduced the inotropic effect induced by stimulation of the right anterior ansa at all frequencies of stimuJ lation used. The effect of piribedil was abolished by haloperidol. There are only two reports describing a presynaptic inhibition of neurally mediated vasoconstriction in the splanchnic area under in vivo conditions. In anaesthetized cats, infusion of noradrenaline or dopamine into the superior mesenteric artery, reduced the decrease in mesenteric blood flow induced by periarterial nerve stimulation (Sanders and Ross, 1975). In dogs, the dopamine agonist GJH 166 attenuated the vasoconstrictor response to postganglionic sympathetic nerve stimulation in the spleen (Sharabi et al., 1977). In our experiments, piribedil was shown to reduce the increase in perfusion pressure of the mesenteric artery induced by postganglionic nerve stimulation at all the frequencies studied. As this effect was antagonized by low doses of haloperidol the involvement of dopamine receptors is suggested. Similarly piribedil reduced the pressor response to splanchnic nerve stimulation. As this response is mainly due to the release of catecholamines into the systemic circulation the reducing effect of piribedil suggests that dopamine receptors may also be present on the membrane of the chromaffin cells of the adrenal medulla. The intravenous infusion of dopamine reduced the renal vasoconstrictor response to sympathetic nerve stimulation (Lokhandwala and Buckley, 1977). This inhibitory effect was significantly potentiated by desipramine
DOPAMINE RECEPTORS AND ADRENERGIC MECHANISMS probably by the inhibition of the neuronal u p t a k e o f d o p a m i n e and was a n t a g o n i z e d b y p i m o z i d e . In t h e p r e s e n t e x p e r i m e n t s , piribedil clearly a n t a g o n i z e d t h e v a s o c o n s t r i c t o r response t o renal postganglionic nerve stimulat i o n a n d p i m o z i d e inhibited t h e response t o piribedil. In c o n c l u s i o n , these e x p e r i m e n t s provide evidence t h a t piribedil reduces s y m p a t h e t i c t r a n s m i s s i o n a n d t h e results are c o m p a t i b l e with t h e c o n c e p t o f a r e d u c t i o n o f n o r a d r e n a line release b y an a c t i o n o f piribedil o n presynaptic dopamine receptors.
References Bell, C., E.L. Conway, W.J. Lang and R. Padanyi, 1975, Vascular dopamine receptors in the canine hindlimb, Brit. J. Pharmacol. 55,167. Bogaert, M.G., A.F. de Schaepdryver and J.L. Willems, 1977, Dopamine-induced neurogenic vasodilatation in the intact hindleg of the dog, Brit. J. Pharmacol. 59, 283. Buylaert, W.A., 1977, Femoral vasodilatation produced by piribedil (ET 495) and its metabolite S 584 in the hindleg of the dog, Naynyn-Schmiedeb. Pharmacol. 299, 101. Buylaert, W.A., J.L. Willems and M.G. Bogaert, 1977, Vasodilation produced by apomorphine in the hindleg of the dog, J. Pharmacol. Exptl. Therap. 201,738. Enero, S. and S. Langer, 1975, Inhibition by dopamine of H3 noradrenaline release elicited by nerve stimulation in the isolated cat's nictating membrane, Naunyn-Schmiedeb. Arch. Pharmacol. 289, 179. Haeusler, G., 1976, Studies on the possible contribution of a peripheral presynaptic action of clonidine and dopamine to their vascular effects under in vivo conditions, Naunyn-Schmiedeb. Arch. Pharmacol. 295,191. Hope, W., M. Law, M.W. Mc Culloch, J.M. Rand and D.F. Story, 1975, Effects of some catecholamines on noradrenergic transmission in the rabbit ear artery, Clin. Exptl. Pharmacol. Physiol. 3, 15. Ilhan, M., J.P. Long and J.G. Cannon, 1976a, Effects of some dopamine analogs and haloperidol on response to stimulation of adrenergic nerves using cat atria in vitro, Arch. Intern. Pharmacodyn. 219,193.
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Ilhan, M., J.P. Long and J.G. Cannon, 1976b, The ability of pimozide to prevent inhibition by dopamine analogs of cardioaccelerator nerves in cat hearts, Arch. Intern. Pharmacodyn. 222, 70. Langer, S.Z., 1977, Presynaptic receptors and their role in the regulation of transmitter release, Brit. J. Pharmacol. 60, 481. Laubie, M., H. Schmitt and J.C. Le Douarec, 1969, Cardiovascular effects of the 1-(2"-pyrimidyl)-4piperonyl piperazine (ET 495), European J. Pharmacol. 6, 75. Lauhie, M., H. Schmitt and E. Falq, 1977, Dopamine receptors in the femoral vascular bed of the dog as mediators of a vasodilator and sympathoinhibitory effects, European J. Pharmacol. 45,307. Lindmar, R., K. LSffelholz and E. Muscholl, 1967, Unterschiede zwischen Tyramin and Dimethylphenyl piperazin in der Ca2÷ Abh/ingigkeit und im zeitlichen Verlauf der Noradrenalin-Freisetzung am isolierten Kaninchenherzen, Experientia, 23, 933. Lokhandwala, M.F. and J.P. Buckley, 1977, Presynaptic dopamine receptors as mediators of dopamine induced inhibition of neurogenic vasoconstriction, European J. Pharmacol. 45, 305. Long, J.P., S. Heintz, J.G. Cannon and J. Kim, 1975, Inhibition of the sympathetic nervous system by 5,6-dihydroxy-2-dimethylaminotetralin (M7), apomorphine and dopamine, J. Pharmacol. Exptl. Therap. 192, 336. McCulloch, M.W., M.J. Rand and D.F. Story, 1973, Evidence for a dopaminergic mechanism for modulation of adrenergic transmission in the rabbit ear artery, Brit. J. Pharmacol. 49, 41P. McNay, J. and L. Goldberg, 1966, Comparison of the effects of dopamine isoproterenol, norepinephrine and bradykinin on canine renal and femoral blood flow, J. Pharmacol. Exptl. Therap. 151, 23. Norris, J.E. and M.C. Randall, 1977, Responses to the canine myocardium to stimulation of thoracic cardiac nerves, Amer. J. Physiol. 232, H 485. Sanders, K.M. and G. Ross, 1975, Inhibition of in vivo neural vasoconstriction by exogenous catecholamines, Blood Vessels 12, 13. Sharabi, F.M., J.P. Long and J.G. Cannon, 1977, Inhibition of canine adrenergic transmission by an analog of dopamine: GJH 166, J. Pharmacol. Exptl. Therap. 202, 97. Starke, K., H.D. Tauhe and E. Borowski, 1977, Presynaptic receptor systems in cetecholaminergic transmission, Biochem. Pharmacol. 26, 259. Westfall, T.C., 1977, Local regulation of adrenergic neurotransmission, Physiological Rev. 57, 659.
99
© Elsevier/North-Holland Biomedical Press
INHIBITORY EFFECTS OF PIRIBEDIL ON ADRENERGIC NEUROTRANSMISSION MICHEL LAUBIE a:ad HENRI SCHMITT
Department of Cardiovascular Pharmacology, I.R.S., 14 Rue du Val d'Or, 92150 Suresnes, France Received 17 March 1978, revised MS received 4 July 1978, accepted 6 July 1978
M. LAUBIE and H. SCHMITT, Inhibitory effects of piribedil on adrenergic neurotmnsmission, European J. Pharmacol. 52 (1978) 99--107. The effects of piribedil on responses to sympathetic stimulation were investigated in anaesthetized dogs. Piribedil (1 mg/kg i.v.) impaired the vasoconstrictor responses to lumbar sympathetic chain stimulation of the perfused hindlimb without changing the effects of noradrenaline. Piribedil (2 mg/kg i.v.) depressed the chronotropic responses to stimulation of the right anterior ansa and the inotropic response to stimulation of the left anterior ansa. Stimulation of the splanchnic nerve induced frequency dependent increases in systemic blood pressure. Piribedil antagonized this effect. Piribedil (1 mg/kg i.v.) attenuated the constrictor responses of the perfused mesenteric artery to postganglionic sympathetic stimulation and reduced the decreases in renal blood flow caused by stimulation of sympathetic renal nerves. The inhibitory effects of piribedil were preferential on responses induced by low frequency stimulation of nerves. The hypertensive, vasoconstrictor and tachycardic effects of noradrenaline and tyramine were not affected. The effects of piribedil were reversed by haloperidol (0.5 mg/kg i.v.) or pimozide (0.2 mg]kg i.v.). Adrenal medulla
Piribedil
Presynaptic dopamine receptors
1. Introduction Piribedil was shown to increase femoral blood flow when injected in small doses into the femoral artery, likely by an action on dopamine presynpatic receptors (Laubrie et al., 1977). This effect of piribedil was abolished by inhibition of the sympathetic tone (Laubie et al., 1969, 1977) and in addition it was antagonized by previous administration of the dopamine receptor blocking agents, haloperidol and pimozide (Buylaert, 1977; Laubie et al., 1977). It was therefore suggested that piribedil induced femoral vasodilatation by inhibiting the release of noradrenaline probably by an action on dopamine presynaptic receptors. However, it is not yet known whether piribedil could inhibit sympathetic transmission in other cardiovascular areas. Dopamine is known to impair peripheral sympathetic transmission by reducing the
Sympathetic nerve stimulation
release of noradrenaline from postganglionic sympathetic nerves and by an action on sympathetic ganglia. Due to the experimental conditions it was not possible to distinguish the part played by each mechanism in all the experiments reported here. However it is clearly shown that piribedil could reduce the effect of sympathetic nerve stimulation in all the cardiovascular areas investigated. Dopamine and the dopaminergic agonistic agent, apomorphine have been shown to reduce the release of noradrenaline induced by stimulation of sympathetic nerves in many in vitro preparations e.g., isolated rabbit ear artery (Mc Culloch et al., 1973; Hope et al., 1975), isolated nictating membrane of the cat (Enero and Langer, 1975). In addition, haloperidol has been shown to increase the response of the guinea pig atria to transmural stimulation (Ilhan et al., 1976a,b) suggesting a negative feedback mechanism. This mechanism has been shown to operate
100 in vivo. Do p a m i ne and t he dopaminergic agonistic agents, a p o m o r p h i n e and 5,6-dihyd r o x y - 2 - m e t h y l a m i n o t e t r a l i n e reduced the cardioacceleration elicited by stimulation of the cardiac nerves in cats and dogs (Long et al., 1975), the c o n t r a c t i o n o f t he spleen induced by stimulation of t he splenic nerves in dogs (Sharabi et al., 1977) and the decrease in renal blood flow induced by stimulation of the renal nerves (Lokhandwala and Buckley, 1977). These experiments suggested t hat the inhibition o f noradrenaline release by dopaminergic agonists m a y be a general phenomenon. F u r t h e r evidence for this view is provided by th e present experiments on the effects o f a dopaminergic agonist, piribedil on the response o f different cardiovascular regions to s y m p a t h e t i c stimulation.
2. Materials and m e t h o d s
2.1. General procedures Mongrel dogs of either sex weighing 15--21 kg were anaesthetized with pentobarbital (30 mg/kg i.e.). The trachea was i nt ubat ed and the dogs were ventilated with a Mark VII Bird respirator. Blood pressure was recorded by means o f a c a t he t e r i n t r o d u c e d into the abdominal aorta via the right femoral artery and c o n n e c t e d to a Statham P23 Db pressure transducer on one channel o f a Brush 440 recorder. Mean blood pressure was obtained by electrical integration of t he blood pressure.
2.2. Perfusion o f the hindlimb After transection of the spinal cord (C~-C2) a p o l y e t h y l e n e c a t he t e r was i n t r o d u c e d into the abdominal aorta via t he left femoral artery; the caudal end o f the cat het er was inserted into the distal end o f the femoral artery. Blood was infused f r om the aorta to the femoral artery by means of a Sigmamotor pump. The perfusion pressure was measured by means o f a Statham P23 Db transducer and was adjusted to 100 m m Hg at t he
M. LAUBIE, H. SCHMITT beginning of the experiment. The left lumbar sym pat het i c t runk was exposed at the level of L3 and placed on a bipolar platinum electrode. The nerve was stimulated at supramaximal voltage (7 V) for 20 sec at frequencies ranging from 0.5 to 16 Hz and a pulse duration of 0.5 msec.
2.3. Stimulation of the sympathetic cardiac nerves In these experiments, the spinal cord was transected at the level of C1-C: and bot h vagus nerves cut. The chest was opened on the right or the left side at the level of the second intercostal space. Either the left or the right anterior ansa was isolated, placed on a pair of platinum electrodes and stimulated at supramaximal voltage (7 V), at frequencies from 0.3 to 10 Hz with a pulse width of 0.5 msec for 20 sec. Stimulation of t he right anterior ansa induced mainly an increase in heart rate as c o u n t e d on the pulse record. In the group o f dogs in which the left anterior ansa was stimulated, a cat het er was i nt roduced into the left ventricle via the left carotid artery. The maximal rate of rise of the left ventricular pressure ( d p / d t max) was recorded by differentiating the signal of the left ventricular pressure. Electrical stimulation of the left anterior ansa induced a m arked inotropic response with a minimal increase in heart rate (Norris and Randall, 1977).
2.4. Perfusion o f the mesenteric artery The mesenteric artery was isolated, its distal end cannulated and infused with bl ood taken from the abdominal aorta. A Si gm am ot or pum p was used to infuse the blood at a constant flow rate and the perfusion pressure was measured by means o f a Statham P23 Db transducer. The perfusion pressure was adjusted at 100 m m Hg at the beginning of the experiment. The mesenteric sympathetic postganglionic nerves were ligated and cut distally to the coeliac ganglion. The nerves were stimulated at supramaximal voltage (7 V)
DOPAMINE RECEPTORS AND ADRENERGIC MECHANISMS
at frequencies from 4 to 16 Hz, a pulse width of 1 msec for 30 sec.
2.5. Stimulation of the splanchnic nerve The spinal cord was transected at the level of CI-C2, the splanchnic nerve was isolated at its exit from the diaphragm, cut and its distal end placed on a pair of platinum electrodes. The nerve was stimulated at supramaximal voltage (7 V) at frequencies ranging from 1 to 8 Hz, a pulse width of 0.5 msec for 30 sec.
2.6. Stimulation nerves
of
101
3. Results
3.1. Effects of piribedil, haloperidol and pimozide on blood pressure and heart rate Piribedil (1--2 mg/kg i.e.) was administered slowly; under these conditions, blood pressure and heart rate did not change significantly. Haloperidol (0.5 mg/kg i.e.) and pimozide (0.2 mg/kg i.e.) induced a transient decrease in blood pressure; stimulation of the sympathetic nerves was always performed after the blood pressure had recovered.
the renal sympathetic
The left kidney was exposed b y a flank incision, the renal artery was dissected and an electromagnetic flow probe (Statham, internal diameter 3 mm) was placed on the renal artery near the aorta. The renal nerves were isolated and a ligature placed at their central end. The renal plexus was placed on a pair of platinum electrodes and stimulated at supramaximal voltage (7 V) at frequencies from 1 to 4 Hz, a pulse width of 1 msec for 20 sec. The renal blood flow was reduced at frequencies of stimulation greater tha 1 Hz.
2.7. Drugs used The following drugs were used: piperoxan hydrochloride (synthetized b y Dr. R6gnier, Servier), piribedil hydrochloride (Servier), haloperidol hydrochloride (Janssen Pharm.), pimozide hydrochloride (Janssen Pharm.), (--)noradrenaline bitartrate (RhSne Poulenc) and tyramine hydrochloride (RhSne Poulenc). All solutions were made in saline except for pimozide which was dissolved in a solution of tartaric acid (10% W/V) and diluted to 5 ml with 5% glucose solution. The doses of the drugs are given as the base.
2.8. Statistical analyses Statistical analyses were performed using Student's t-test for paired comparisons.
3.2. Effect o f piribedil on the pressor response and the tachycardia induced by noradrenaline and tyramine In dogs with both vagus nerves cut, piribedil (2 mg/kg i.e.) did n o t change the pressor responses to noradrenaline (0.5 to 2 pg/kg i.e.) and tyramine (100/~g/kg i.e.) (5 experiments). In dogs with spinal cord transection and both vagus nerves cut, piribedil (2 mg/kg i.e.) did not change the pressor response and the tachycardia after noradrenaline and tyramine (5 experiments; table 1).
3.3. Effects of piribedil on the responses o f the perfused hindlimb to stimulation of the lumbar sympathetic chain In dogs with transection of the spinal cord, electrical stimulation of the lumbar sympathetic chain induced a frequency dependent increase in the perfusion pressure of the hindlimb. The average increase in perfusion pressure ranged from 25 + 3 mm Hg at 0.5 Hz to 120 + 11 mm Hg at 16 Hz. Piribedil (1 mg/ kg i.e.) induced a transient increase in blood pressure from 90 + 4 to 120 + 10 mm Hg and in the perfusion pressure of the hindlimb from 1 0 4 + 5 to 1 1 5 + 7 m m H g . After recovery, the increases in perfusion pressure induced b y stimulation of the lumbar sympathetic nerve were reduced (fig. 1--7 experiments). This reduction was greater at low frequencies of stimulation (0.5--4 Hz) that at
102
M. L A U B I E , H. S C H M I T T
TABLE 1 Effects of piribedil (2 m g / k g i.v.) on the tachycardia and hypertensive responses to noradrenaline and t y r a m i n e in dogs with spinal cord transection and b o t h vagus nerves cut (5 experiments). Each value is the mean ± S.E.M. Increase in b l o o d pressure (A m m Hg * S.E.M.)
Increase in heart rate (A b e a t s / m i n ± S.E.M.)
Before piribedil
After piribedil
Before piribedil
A f t e r piribedil
Noradrenaline (p g/kg i.v.) 0.1 0.3 1.0
23+_ 5 35+_ 8 66 _+ 16
26_+ 6 34_+ 7 65 _+ 16
10+_4 20+_5 40 +_ 9
8+_ 1 20_+ 6 44 + 13
Tyramine (pg/kg i.v.) 50 100
32+_ 9 74+_15
34_+ 7 69+10
19+_6 38_+6
21+_ 39+_
high frequencies. In contrast, piribedil did not change the increase in perfusion pressure (60 + 4 mm Hg) induced by injection of noradrenaline (3 pg) into the femoral artery. Haloperidol (0.5 mg/kg i.v.) antagonized the effect o f piribedil. After haloperidol the frequency response curve to lumbar chain stimulation was not significantly different from the control one.
hindlimb,~ E E
i
mesenteric, artery
100
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t
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(Hz)
Frequence of nerve stimulation Fig. 1. F r e q u e n c y d e p e n d e n t increase in t h e perfusion pressure in the hindlimb (7 e x p e r i m e n t s ) and in the mesenteric artery (7 experiments) elicited by stimulat i o n o f the s y m p a t h e t i c nerves, o, Effect after piribedil (1 m g / k g i.v.); A, reversal effect o f haloperidol (0.5 m g / k g i.v.). The significance o f differences f r o m the c o n t r o l is r e p r e s e n t e d by asterisks (P < 0.01). e, Control.
7 5
3.4. Effects o f piribedil on the responses to stimulations o f the cardiac sympathetic nerves In dogs with transection of the spinal cord and with both vagus nerves cut, stimulation of the right anterior ansa induced a frequency dependent increase in heart rate of 19 + 4 beats/min at 0.3 Hz and 130 + 5 beats/min at 10 Hz. Piribedil (2 mg/kg i.v.) did not change the basal heart rate but significantly reduced the tachycardia induced by stimulation of the cardiac sympathetic nerve at low frequencies stimulation (0.3--3 Hz) but not at the higher frequency of 10 Hz (fig. 2--5 experiments). Piribedil did not change the dose dependent increase in heart rate induced by noradrenaline and tyramine (5 experiments; table 1). After haloperidol (0.5 mg/kg i.v.}, the heart rate response curve to nerve stimulation was not significantly different from the control one at any o f the frequencies investigated. Stimulation of the left anterior ansa produced a marked inotropic response with a minimal chronotropic effect. The maximal rate of rise in left ventricular pressure (max. dp/dt) increased by 110 + 20 mm Hg/sec. at 0.3 Hz and by 3,300 mm Hg/sec at 10 Hz.
DOPAMINE RECEPTORS AND ADRENERGIC MECHANISMS I
100
300O
C 15C
103
E (n
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.,1= 10( oJ .c ._c
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Fig. 2. Left panel: increase in heart rate elicited by stimulation of the right anterior ansa. D, Piribedil (2 mg/kg i.v.) attenuated the response between 0.3-3 Hz but did not change the cardioaccelerator effect at 10 Hz. A Haloperidol (0.5 mg/kg i.v.)antagonized the inhibitory effect of piribedil (5 experiments). e, Control. Right panel: increase in max. dp/dt induced by stimulation of the left anteror ansa. El After piribedil ( 2 mg/kg i.v.); A, reversal effect of haloperidol (0.5 mg/kg i.v.) (6 experiments). The significance of differences from the control is represented by asterisks (P < 0.01). e, Control.
Piribedil (2 mg/kg i.v.) did not change the basal maximal rate of rise in left ventricular pressure b u t significantly reduced the effects of the nerve stimulation at all the frequencies investigated; the frequency response curve was shifted to the right. A subsequent administration of haloperidol (0.5 mg/kg i.v.) antagonized the inhibitory effect of piribedil. After haloperidol the frequency response curve to sympathetic stimulation was not significantly different from the control one (fig. 2 - 6 experiments).
3.5. Stimulation of the splanchnic nerve In the anaesthetized dogs with the spinal cord transected and b o t h vagus nerves cut, stimulation of the splanchnic nerve increased the systemic blood pressure in a frequency dependent manner. The increase in blood pressure was 22 -+ 1 mm Hg at 1 Hz and 98 + 7 mm Hg at 8 Hz. Piribedil (1 mg/kg i.v.) significantly reduced the pressor responses to splanchnic nerve stimulation at all the frequencies used. In contrast piribedil did not
0
1 2 4 8Hz Fig. 3. Systemic hypertensive response elicited by stimulation of the splanchnic nerve in spinal cordtransected dogs. D, PiribedLl (1 mg/kg i.v.) shifted the frequency response cuwe to the right; ~, pimozide (0.2 mg/kg i.v.) antagonized the effects of p~ibedll (6 experiments). The significance of differences from the control is represented by asterisks (P < 0.01 ). e, Control.
change the pressor dose response to noradrenaline and to tyramine (table 1). Pimozide (0.2 mg/kg i.v.) administered subsequently could antagonize the effect of piribedil. The pressor response curves to splanchnic nerve stimulation were not significantly different before piribedil and after pimozide (fig. 3--6 experiments).
3.6. Perfusion o f the mesenteric artery Stimulation of the postganglionic sympathetic nerves increased the perfusion pressure of the mesenteric artery b y 28 + 4 mm Hg at 4 Hz and 106 + 9 mm Hg at 16 Hz. Piribedil (1 mg/kg i.v.) induced a transient increase in the perfusion pressure of the mesenteric artery (43 + 8 mm Hg). Stimulation of the mesenteric nerves was performed after the recovery of the perfusion pressure (20 min). Piribedil reduced the response to stimulation of the sympathetic nerves at all frequencies of stimulation investigated; the frequency response curve was shifted to the right. In contrast piribedil did not significantly change
104
M. L A U B I E , H. S C H M I T T
the increase in the perfusion pressure (70 + 9 mm Hg) induced by injection of noradrenaline (0.3 pg) into the mesenteric artery. The subsequent administration of pimozide (0.2 mg/kg i.e.) induced the recovery of the pressor response to stimulation of the mesenteric sympathetic nerves (fig. 1--7 experiments).
E
0 25 O 0
e-
~
3.7. Stimulation nerves
of
the
renal sympathetic
Stimulation of the renal sympathetic nerves caused a frequency d e p e n d e n t reduction in renal blood flow. Renal blood flow decreased from 143 + 20 ml/min to 128 + 14 ml/min at a stimulation frequency of 1 Hz and from 141 + 24 ml/min to 72 -+ 10 ml/min at a frequency of 4 Hz. Piribedil (1 mg/kg i.e.) did not change the renal blood flow but the drug significantly reduced the vasoconstrictor responses to renal sympathetic nerves stimulation (fig. 4--6 experiments). Noradrenaline (0.1 pg/kg i.e.) decreased the renal b l o o d flow from 153 + 21 ml/min to 108 + 14 ml/min and noradrenaline (0.3 pg/ kg i.e.) decreased the renal blood flow from 149 + 22 ml/min to 70 +- 14 ml/min. These renal vasoconstrictor responses to noradrenaline were n o t altered by piribedil (1 mg/kg i.e.) (6 experiments. The subsequent adminis-
0
5o
c
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1
2
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Fig. 4. R e d u c t i o n in renal artery b l o o d flow elicited by postganglionic s y m p a t h e t i c nerve stimulation, c~, A f t e r piribedil (1 m g / k g i.e.), the renal vasoconstrictor responses were a t t e n u a t e d ; ~, p i m o z i d e (0.2 rag/ kg i.e.) antagonized the effects of piribedil (6 experiments). The significance of differences from the control is represented by asterisks (P < 0.01). $, Control.
tration of pimozide (0.2 mg/kg i.e.) induced a recovery of the vasoconstrictor responses to sympathetic stimulation and the frequency response curves to sympathetic stimulation obtained in the control period and after injection of pimozide were not significantly different.
TABLE 2 Effects of haloperidol (0.5 m g / k g i.e.) and p i p e r o x a n (0.5 m g / k g i.e.) on the inhibitory effects of clonidine (15 }Jg/kg i.e.) on the cardioacceleration elicited by cardiac s y m p a t h e t i c stimulation in dogs with spinal cord transe c t e d and b o t h vagus nerves cut (5 experiments). Mean values -+ S.E.M. are given. F r e q u e n c y of stimulation (Hz)
0.3 1 3 10
Increase in heart rate (A b e a t s / m i n +- S.E.M.) Before i.e. injection
A f t e r clonidine 1
After haloperidol 2
After p i p e r o x a n 1
11+ 4 33-+ 7 81 -+ 11 116 _+ 13
2 -+ 1 3 11_+ 3 3 50 -+ 10 3 92 -+ 11 3
2-+ 1 11-+ 5 52 -+ 11 93 -+ 10
10-+ 3 32-+ 6 83 -+ 13 123 -+ 18
1 C o m p a r e d with before clonidine. 2 C o m p a r e d with after clonidine. a p < 0.01.
DOPAMINE RECEPTORS AND ADRENERGIC MECHANISMS 3.8. Action o f haloperidol on presynaptic aadrenoceptors
In dogs with the spinal cord transected and both vagus nerves cut, clonidine (15 pg/kg i.v.) significantly reduced the tachycardia elicited by stimulation of the cardiac sympathetic nerve. The subsequent injection of haloperidol (0.5 mg/kg i.v.) did not change the inhibitory effect of clonidine. In contrast piperoxan (0.5mg/kg i.v.) injected after haloperidol antagonized the effects of clonidine and the frequency responses curves in the control period and after injection of piperoxan were not significantly different (table 2--5 experiments).
4. Discussion
Piribedil has been suggested to cause femoral vasodilatation by an action on presynaptic dopamine receptors, probably by reducing noradrenaline release from the postganglionic sympathetic fibres (Laubie et al., 1977; Buylaert, 1977). The present experiments extend these findings and show an impairement of sympathetic transmission by piribedil in several sympathetic nerves. These experiments are compatible with a reduction of noradrenaline release by an action of piribedil on dopamine receptors. An effect of piribedil on postsynaptic sites as the cause of the reduction of the effects of sympathetic stimulation could be ruled out. In fact, piribedil did not change the pressor dose-response curve to noradrenaline in any of the experiments. An action of piribedil on the postganglionic sympathetic fibres is likely. In fact the mesenteric and renal sympathetic nerves are known to be composed of postganglionic fibres and piribedil reduced the effects of sympathetic stimulation. These experiments suggest that piribedil reduced noradrenaline release. Two other findings also support this view. Piribedil was shown to be more effective to reduce the effects of sympathetic stimulation at low frequencies than it
105
was at high frequencies. It is well known (Langer, 1977; Starke et al., 1977; Westfall, 1977) that the activation of presynaptic ~adrenoceptors or dopamine receptors reduced the noradrenaline release induced by low frequencies of stimulation more efficiently than they did that induced at high frequencies. In addition, in contrast to its action on sympathetic nerve stimulation, piribedil did not change the pressure and heart rate increases in response to tyramine. Tyramine is known to release noradrenaline from postganglionic sympathetic nerves by a calcium non-dependent process in contrast to noradrenaline release elicited by nerve stimulation (Lindmar et al., 1967). The splanchnic nerve and the lumbar sympathetic chain contain preganglionic sympathetic fibres and in the experiments performed by stimulating these nerves, piribedil may also have impaired sympathetic transmission by an action on the ganglia. In fact dopamine is known to impair ganglionic transmission (Bogaert et al., 1977) and it remains to be established that piribedil acts on glanglia. The effects of piribedil appear to be due to a dopaminergic mechanism. In fact, the classical dopamine receptor blocking agents, haloperidol and pimozide abolished the inhibitory effects of piribedil on sympathetic transmission, It seems unlikely that these agents, in the doses used in our experiments, blocked presynaptic a-adrenoceptors. They did not change the inhibitory effects of clonidine, an ~-adrenoceptor agonist, on the cardioacceleration elicited by cardiac sympathetic stimulation. Therefore, the present results suggest that piribedil reduced noradrenaline release by stimulating presynaptic dopamine receptors. Besides, piribedil reduced the effects of stimulation of seven sympathetic nerves suggesting the generality of the process. In fact, there are conflicting results on the existence and on the role of dopamine receptors. The existence of postsynaptic dopamine receptors in the femoral vascular bed has thus b e e n : ~ u t e d . McNay and Goldberg (1966) reported that dopamine injected into the
106
femoral artery of dogs treated with an aadrenoceptor blocking agent induced vasodilation which was abolished by f~-adrenoceptor blockade, suggesting the absence of specific dopamine receptors. In contrast Bell et al. (1975) found an increase in femoral blood flow in some dogs after a- and fladrenoceptor block with phentolamine and propranolol respectively; they concluded therefore that specific dopamine receptors may be present in the femoral bed. Our previous studies (Laubie et al., 1977) using apomorphine and piribedil however failed to detect postsynaptic dopamine receptors in this vascular bed. Similar conflicting results have been published concerning the presence of presynaptic dopamine receptors. Dopamine infused into the autoperfused hindlegs of cats did not change the increases in perfusion pressure induced by stimulation of the lumbar sympathetic chain (Haeusler 1976). However, dopamine presynaptic receptors have been detected in the femoral bed of dogs (Laubie et al., 1977; Buylaert et al., 1977). Apomorphine and- piribedil injected into the femoral artery induced a dose-dependent increase in femoral blood flow. This effect was abolished by section of the motor nerves, transection of the spinal cord and ganglionic blockade. The dilator effect of piribedil and apomorphine was also antagonized by haloperidol and pimozide. The present experiments indicate that piribedil impaired the pressor responses to sympathetic nerve stimulation of the perfused hinlimb of the dog and preferentially, the responses to low frequencies of stimulation; in addition haloperidol and pimozide antagonized the effects of piribedil. Similar results have been obtained with the cyclic analog of dopamine GJH 166 (Sharabi et al., 1977). Therefore the experiments reported are compatible with the view that pix-ibedil reduced noradrenaline release by stimulating presynaptic dopamine receptors during spontaneous or evoked sympathetic discharges. Previous studies have suggested the presence of presynaptic dopamine receptors
M. LAUBIE, H. SCHMITT
located on the postganglionic terminals of the cardiac nerves. In fact, dopamine in presence of cocaine, dopamine analogs or apomorphine {Long et al., 1975; Ilhan et al., 1976a) have been shown to reduce the chronotropic response to 2 Hz stimulation of the'right cardioaccelerator nerve in cats and dogs. In the present study piribedil reduced the tachycardia induced by stimulation of the right anterior ansa at low frequencies (0.3--3 Hz) but was ineffective when the stimulation frequency was 10 Hz. Piribedil also reduced the inotropic effect induced by stimulation of the right anterior ansa at all frequencies of stimuJ lation used. The effect of piribedil was abolished by haloperidol. There are only two reports describing a presynaptic inhibition of neurally mediated vasoconstriction in the splanchnic area under in vivo conditions. In anaesthetized cats, infusion of noradrenaline or dopamine into the superior mesenteric artery, reduced the decrease in mesenteric blood flow induced by periarterial nerve stimulation (Sanders and Ross, 1975). In dogs, the dopamine agonist GJH 166 attenuated the vasoconstrictor response to postganglionic sympathetic nerve stimulation in the spleen (Sharabi et al., 1977). In our experiments, piribedil was shown to reduce the increase in perfusion pressure of the mesenteric artery induced by postganglionic nerve stimulation at all the frequencies studied. As this effect was antagonized by low doses of haloperidol the involvement of dopamine receptors is suggested. Similarly piribedil reduced the pressor response to splanchnic nerve stimulation. As this response is mainly due to the release of catecholamines into the systemic circulation the reducing effect of piribedil suggests that dopamine receptors may also be present on the membrane of the chromaffin cells of the adrenal medulla. The intravenous infusion of dopamine reduced the renal vasoconstrictor response to sympathetic nerve stimulation (Lokhandwala and Buckley, 1977). This inhibitory effect was significantly potentiated by desipramine
DOPAMINE RECEPTORS AND ADRENERGIC MECHANISMS probably by the inhibition of the neuronal u p t a k e o f d o p a m i n e and was a n t a g o n i z e d b y p i m o z i d e . In t h e p r e s e n t e x p e r i m e n t s , piribedil clearly a n t a g o n i z e d t h e v a s o c o n s t r i c t o r response t o renal postganglionic nerve stimulat i o n a n d p i m o z i d e inhibited t h e response t o piribedil. In c o n c l u s i o n , these e x p e r i m e n t s provide evidence t h a t piribedil reduces s y m p a t h e t i c t r a n s m i s s i o n a n d t h e results are c o m p a t i b l e with t h e c o n c e p t o f a r e d u c t i o n o f n o r a d r e n a line release b y an a c t i o n o f piribedil o n presynaptic dopamine receptors.
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