Adenosine and the endothelium-dependent modulation of 3H-noradrenaline release in the canine pulmonary artery.

This study aimed at characterizing the influence of endothelium on noradrenaline release from the canine pulmonary artery. Tritium overflow from intact or endothelium-free vessels preloaded with 0.2 mumol.l-1 3H-noradrenaline was evoked by electrical stimulation (1 Hz, during 5 min) or potassium (25-100 mmol.l-1). The fractional release of tritium evoked by electrical stimulation was increased by removing the endothelium [from 1.7 (1.2; 2.4) to 2.7(2.3; 3.2) x 10(-5).pulse-1, n = 10; P < 0.05]. Neither NG-nitro-L-arginine methyl ester (L-NAME) (up to 300 mumol.l-1) nor indomethacin (up to 30 mumol.l-1), nor endothelin-1 (up to 30 nmol.l-1), nor suramin (up to 300 mumol.l-1) changed tritium release evoked by electrical stimulation. In contrast, the selective A1-adenosine antagonist 1,3-dipropyl-8-cyclopentylxanthine (DPCPX) (3.3-33 nmol.l-1) concentration-dependently increased, and the selective A1-adenosine agonist N6-cyclopentyladenosine (CPA) (3.3-100 nmol.l-1) concentration-dependently decreased the evoked release of noradrenaline. Since the effects of DPCPX were observed in endothelium-intact tissues only, it may be concluded that adenosine secreted by the endothelium activates prejunctional release-inhibiting A1-receptors. Tetraethylammonium (TEA) (3.3-33 mmol.l-1) enhanced tritium overflow evoked by electrical stimulation more in endothelium-free than in endothelium-intact vessels, indicating that some K(+)-channel opener is involved in the inhibitory role of endothelium on noradrenaline release. Since it had been previously shown that A1-adenosine receptors are coupled to K(+)-channels, it is suggested that adenosine may inhibit noradrenaline release through the opening of K(+)-channels. In conclusion, the results show that in the canine pulmonary artery, adenosine is a good candidate for the endothelium-dependent inhibitory factor which is responsible for the reduction of noradrenaline release evoked by electrical stimulation.

Long-term administration of 1,3-dipropyl-8-sulphophenylxanthine (DPSPX) alters alpha 2-adrenoceptor-mediated effects at the pre- but not at the postjunctional level.

The present investigation was undertaken to see whether a long-term inhibition of adenosine receptors--leading to hypertension--interferes with alpha 2-adrenoceptor-mediated modulation of noradrenaline release. Rat tail arteries were removed from normal and from hypertensive animals obtained by chronic treatment with intraperitoneally infused DPSPX (1,3-dipropyl-8-sulphophenylxanthine) or orally administered L-NAME (NG-Nitro-L-arginine methyl ester). To study prejunctional effects, the influence of UK-14,304 (5-bromo-6(imidazoline-2-ylamino)-quinoxaline) and yohimbine on the overflow of tritium evoked by electrical stimulation (100 V; 1 Hz; 2 ms; 5 min) from tissues preloaded with 3H-noradrenaline was analysed. To study postjunctional effects, concentration-response curves to UK-14,304 were determined. In DPSPX-treated rats there was an enhancement of the prejunctional effects of UK-14,304: its Ec30% was reduced from 381 (250; 579) to 85 (73; 99) nmol.l-1 (n = 5; P < 0.05) and its maximal effect--expressed as percent reduction of tritium overflow-increased from 45 +/- 5% to 61 +/- 5% (n = 6; P < 0.05). In L-NAME-treated rats there was no change in either of these two parameters. At the postjunctional level, there was no change in the sensitivity to UK-14,304 in tissues from either DPSPX- or L-NAME-treated rats. Yohimbine (10-1000 nmol.l-1) caused a concentration-dependent increase of tritium overflow evoked by electrical stimulation in both control and hypertensive animals (either DPSPX- or L-NAME-treated).(ABSTRACT TRUNCATED AT 250 WORDS)

Postnatal development of vascular beta-adrenoceptor-mediated responses and the increase in the adrenaline content of the adrenal gland have a parallel time course.

The present study was undertaken to analyse the relationship between postnatal development of vascular beta 2-adrenoceptor-mediated responses and the content of adrenaline in the adrenal gland and its concentration in plasma. Dog saphenous vein tissue from newborn, two-weeks old and adult animals were either preloaded with 3H-noradrenaline (or 3H-adrenaline) to study prejunctional beta-adrenoceptor-mediated effects or mounted in organ baths to determine isoprenaline-induced relaxation of preparations contracted by phenylephrine to about 65% of the maximum. The adrenal glands and samples of blood from the same animals were taken for estimation of adrenaline and noradrenaline. At birth, there were no beta-adrenoceptor-mediated effects pre- or postjunctionally. At two weeks, while the results at the prejunctional level were not significantly different from those obtained in newborns, at the postjunctional level there was a relaxant response to isoprenaline, which antagonised about 35% of the previous contraction to 1.75 mumol.l-1 phenylephrine. In adults, isoprenaline (50 nmol.l-1) increased by 24% tritium overflow evoked by electrical stimulation of tissues preloaded with 3H-noradrenaline but not that of tissues preloaded with 3H-adrenaline. On the other hand, propranolol (1 mumol.l-1) reduced by 21% the overflow of tritium evoked by electrical stimulation of tissues preloaded with 3H-adrenaline but not that of tissues preloaded with 3H-noradrenaline; postjunctionally, the maximal response to isoprenaline antagonised 70% of the previous contraction to 1.75 mumol.l-1 phenylephrine.(ABSTRACT TRUNCATED AT 250 WORDS)

Lack of pre- and postjunctional beta adrenoceptor-mediated effects in the canine saphenous vein at birth.

The present study was undertaken to compare the relevance of alpha and beta adrenoceptor-mediated responses at pre- and postjunctional level in the canine saphenous vein of neonates and adults. To quantify prejunctional action, the effect of drugs on the neurogenic outflow of tritium from the vessel loaded previously with [3H]norepinephrine or [3H]epinephrine was measured. The selective alpha-2 adrenoceptor agonist UK-14,304 [5-bromo-6-(2-imidazoline-2-ylamino)-quinoxaline; 10-1000 nM reduced and the selective alpha-2 adrenoceptor antagonist yohimbine (30-300 nM) enhanced the overflow of tritium evoked by electrical stimulation (1 Hz; 2 msec; 100 V; 300 pulses) in both adult and neonate tissues. However, in strips preloaded with [3H]norepinephrine, the beta adrenoceptor agonist isoproterenol (50 nM) increased the overflow of tritium in strips of adults but had no effect in strips of neonates; and in the strips preloaded with [3H]epinephrine, the beta adrenoceptor antagonist propranolol (1 microM) reduced the overflow of tritium in adults but had no effect in neonates. Postjunctionally, phenylephrine (0.1-50 microM) caused concentration-dependent contractions of the saphenous vein rings from adults and neonates but isoproterenol, which caused concentration-dependent relaxations on rings contracted previously by phenylephrine in adults, had no effect in neonates. In contrast to isoproterenol, forskolin (0.05-5 microM), under the same conditions, caused concentration-dependent relaxations of rings of both adults and neonates.(ABSTRACT TRUNCATED AT 250 WORDS)

Release and disposition of 3H-noradrenaline in the saphenous vein of neonate and adult dogs.

Release of 3H-noradrenaline and formation of 3H-metabolites were studied in the saphenous vein of newborn (mean age, 18 h) and adult dogs. Vein strips were incubated with 0.23 mumol/l of 3H-noradrenaline during 1 h and washed out for 110 min; thereafter, the perifusion fluid was collected in 5-min samples. Electrical stimulation was applied at 120 min (1 Hz, 2 ms, 100 V, for 5 min). In some experiments the tissues were preincubated with 1 mmol/l pargyline (to inhibit monoamine oxidase). In these experiments, 12 mumol/l cocaine (to inhibit uptake1), 41 mumol/l hydrocortisone (to reduce uptake2) and 50 mumol/l U-0521 (to inhibit COMT) were present during the perifusion. 3H-noradrenaline, 3H-DOPEG, 3H-NMN, 3H-DOMA and 3H-OMDA were separated by column chromatography. The noradrenaline content of the tissue was estimated by HPLC followed by electrochemical detection. A morphological study was also carried out by light and electron microscopy. The endogenous noradrenaline content of the saphenous vein was 4.3 times higher in adults than in neonates. The number of varicosities was similar in adults and newborns but the number of vesicles per varicosity profile was 5 times higher in adults. Hence, the endogenous noradrenaline content per vesicle was about the same in adults and newborns. The accumulation of 3H-noradrenaline per vesicle was about 5 times higher in newborns than in adults. On the other hand, the vein wall media of neonates was about 3 times thinner than that of adults. The evoked fractional release of tritium was about 10 times higher in neonates than in adults, whether the inactivation pathways were blocked or not.(ABSTRACT TRUNCATED AT 250 WORDS)

Involvement of alpha 1- and alpha 2-adrenoceptors in the responses of proximal and distal segments of the canine saphenous vein to exogenous and endogenous noradrenaline.

In the present study the relationship between adrenergic nerve terminals and postjunctional alpha-adrenoceptors mediating the responses to the endogenous transmitter was compared at proximal and distal levels of the canine saphenous vein. Concentration-response curves to noradrenaline and to tyramine as well as frequency-response curves to electrical stimulation were compared at both levels of the vessel, in the absence and presence of either prazosin (100 nmol.l-1) or yohimbine (100 nmol.l-1). The influence of inhibition of neuronal uptake by cocaine (12 mumol.l-1) on the responses to noradrenaline in the presence of prazosin (56 nmol.l-1) or yohimbine (20 nmol.l-1) was compared at the proximal level. The results show that, at the proximal level, the maximal responses to electrical stimulation and tyramine reached 80.1 +/- 2.2 (n = 18) and 74.2 +/- 1.9 (n = 18)%, respectively, of the maximal responses to noradrenaline, and 70.3 +/- 0.8 (n = 15) and 53.1 +/- 1.2 (n = 14)%, respectively, at the distal level. Furthermore, the proximal strips were more sensitive to electrical stimulation than the distal ones. Prazosin had a much greater inhibitory effect on the contractile responses to noradrenaline than on those to electrical stimulation, at both levels. At proximal level, the shifts (to the right) of the concentration (frequency)-response curves (at EC50) amounted to 0.58 +/- 0.02 (n = 16) and 0.18 +/- 0.02 (n = 8) log units, respectively (P less than 0.05), but, at the distal level, to 1.12 +/- 0.03 (n = 16) and 0.28 +/- 0.08 (n = 8) log units, respectively (P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Alpha 1- and alpha 2-adrenoceptors at different levels of the canine saphenous vein.

Presynaptic alpha2- and postsynaptic alpha1-adrenoceptors were compared at the distal and proximal parts of the dog saphenous vein. The results obtained show that: (1) yohimbine is more effective against postsynaptic responses to phenylephrine distally than proximally. On the contrary, WB-4101 is more effective proximally; (2) phenylephrine increases inositol monophosphate production at both levels, but the increase is more pronounced distally; (3) UK-14, 304 and adrenaline reduce and yohimbine and phentolamine increase the release of 3H-noradrenaline caused by electrical stimulation at both levels. However, while adrenaline as well as the antagonists are equipotent at the two levels, UK-14,304 is more potent distally than proximally. In conclusion, we suggest that: more alpha 1A-adrenoceptors exist distally than proximally; imidazoline sites can exist at the distal level which contribute to the higher potency of UK-14,304 distally.

Alpha-adrenoceptor reserve in the canine cephalic vein.

In the canine cephalic vein, the pD2 for the selective alpha 1-agonist, phenylephrine, was 6.08 +/- 0.08 (n = 14) and that for the selective alpha 2-agonist, UK-14,304, was 8.32 +/- 0.06 (n = 10). The pA2 values for the antagonism exerted by the selective alpha 1-antagonist, prazosin, against phenylephrine and UK-14,304 were 7.74 +/- 0.05 (n = 14) and 6.28 +/- 0.03 (n = 8), respectively, while those for the antagonism exerted by the selective alpha 2-antagonist, yohimbine, against phenylephrine and UK-14,304 were 7.40 +/- 0.02 (n = 14) and 8.93 +/- 0.05 (n = 14), respectively. Furthermore, the concentration-response curve for UK-14,304 was typically biphasic, the first phase being antagonized by yohimbine and the second phase by prazosin and phenoxybenzamine. These results show that there are postsynaptic alpha 1- and alpha 2-adrenoceptors in the canine cephalic vein. In such a preparation, only one concentration of phenoxybenzamine (1 nM) shifted the concentration-response curves for noradrenaline, adrenaline and isoprenaline to the right without reducing the maximum. However, at the concentrations tested, phenoxybenzamine did not shift the concentration-response curve for phenylephrine to the right without depressing its maximum. It is concluded that: (1) the canine cephalic vein is a suitable preparation to study postsynaptic alpha 1- and alpha 2-adrenoceptors; (2) according to the original definition of 'spare receptors', there is no alpha 1-adrenoceptor reserve in canine cephalic vein; (3) UK-14,304 is a partial agonist at alpha 1-adrenoceptors.

An unusually important role of O-methylation in the disposition of noradrenaline and adrenaline by the dog renal artery.

Using the oil immersion technique, the role of neuronal uptake, monoamine oxidase and COMT in the inactivation of 2 concentrations (0.23 and 2.3 mumol/l) of noradrenaline and adrenaline was determined by the prolongation of the inactivation time caused by cocaine (12 mumol/l), pargyline (1 mmol/l) and U-0521 (50 mumol/l), respectively. The results obtained allow us to conclude that: 1) as previously shown, for the saphenous vein and mesenteric artery, noradrenaline is inactivated more rapidly than adrenaline; 2) in all tissues and for both concentrations of noradrenaline and adrenaline, neuronal uptake is more important for the inactivation of noradrenaline than for that of adrenaline, while O-methylation is more important for the inactivation of adrenaline than for that of noradrenaline. The only exception is that in the renal artery, O-methylation is very clearly the most important pathway of inactivation for both concentrations of both amines.