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*Department of Anesthesiology and Critical Care Medicine, and the
Second Department of Surgery, Kagoshima University School of Medicine, Kagoshima, Japan
Address correspondence and reprint requests to Rumi Katai, MD, Department of Anesthesiology and Critical Care Medicine, Kagoshima University School of Medicine, 8–35–1 Sakuragaoka, Kagoshima 890–8520, Japan. Address email to tsune{at}m.kufm.kagoshima-u.ac.jp
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Abstract |
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1-adrenergic antagonist, prazosin. Results qualitatively similar to these were obtained in both GEA and IMA, although the contractile responses were far smaller. In RA, DA enhanced the norepinephrine (NE)-induced contraction, and this action of dopamine was enhanced by SCH23390. In GEA, small concentrations (<10-7 mol/L) of DA attenuated the NE-induced contraction but larger concentrations did not. In IMA, DA induced a vasorelaxation on the NE-contraction only at higher concentrations (10-6–10-5 mol/L). In both GEA and IMA, the dopamine-induced vasorelaxations on the NE contraction were completely inhibited by SCH23390. These results suggest that the affinities of DA for DA1- and 1-adrenergic receptors may explain its variable contractile and vasorelaxant effects among these arteries.
IMPLICATIONS: Differing affinities of dopamine for dopamine A1- and 1-adrenergic receptors may lead to it having variable contractile and vasorelaxant effects among the arteries supplying grafts for coronary bypass surgery.
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Introduction |
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Among the medical therapies used in revascularized patients, dopamine (DA) is often used to treat myocardial dysfunction (8,9). Despite the widespread use of DA in the treatment of cardiovascular disorders, only limited information is available concerning its mechanisms of action in human vascular smooth muscle. Moreover, little is known regarding the vasomotor effects of DA on arterial bypass grafts in vitro. Thus, the question arises as to whether this amine may affect the vascular functions of any grafts that are inserted.
The main purpose of this study was to characterize the effects and mechanisms of action of DA in three arteries commonly used to provide arterial bypass grafts, namely, RA, GEA, and IMA. In arteries obtained from patients undergoing coronary bypass graft surgery, we tested the effects of DA both on resting tone and on the contractions induced by the -adrenergic agonist, norepinephrine (NE). This study coincidentally provided valuable additional information about the different actions of DA on dopaminergic and adrenergic receptors among the three arteries.
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Methods |
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Before we began applying NE (10-6 mol/L) at intervals to obtain a stable contraction, the lack of a functional endothelium was confirmed by the absence of an acetylcholine-induced (ACh, 10-6 mol/L) endothelium-dependent relaxation of the NE-induced maximum contraction in such arteries. In the endothelium-intact rings, if the relaxation induced by ACh proved to be smaller than 30% of the initial maximal contraction, the segment was discarded as the endothelium was considered to be damaged. Conversely, if our efforts to remove the endothelium failed (i.e., if an ACh-induced relaxation of more than 5% was observed after attempted denudation), we discarded that ring. To exclude any residual effect of ACh still being present, the experimental interventions were performed 30 min after the ACh-trials.
The mechanical activity of the ring was measured using a strain gauge (UL-100GR; Minebea, Tokyo) in a tissue bath (volume, 1.0 mL) filled with Krebs solution continuously bubbled with 95% O2: 5% CO2. The temperature of the solution was maintained at 37°C. The resting tension was set at 20 mN, a value shown by the length-tension relationship to allow a maximal active tension to be induced by NE (10-6 mol/L). During a 2-h equilibrium period, Krebs solution was continuously infused at a rate of 2 mL/min by a pump (Perista pump SJ-1211; ATTO, Tokyo) from one end of the bath, and simultaneously aspirated from the other. During the experiment the infusion rate was increased to 10 mL/min so that the bath solution was exchanged quickly for new solution. In an initial series of experiments, control contractile responses were induced by adding the -adrenergic agonist NE (10-6 mol/L) to the Krebs solution for 7 min every 30 min. Preliminary experiments showed that the responses reached maximal levels within 30 s after application of DA and NE and that a 23-min interval was sufficient for the tension to return to the control level. Previously, we reported that in our experimental model, the amplitude of the contraction induced by 10-6 mol/L NE in endothelium-intact and endothelium-denuded human GEA rings remained stable for more than 12 h (10,11). Thus, in this study the vascular rings would be expected to react to the same extent to identical concentrations of NE or DA over the 4-h experimental period. After the amplitude of the control NE-induced contraction stabilized, the control contractile response in each experiment was determined as the maximum amplitude of the phasic response to 10-6 mol/L NE (normalized as a relative tension of 1.0 in each ring).
In the present study, we tested the effects of DA both on the resting tension (Fig. 1) and (in a separate series) on the contractions induced by NE (Fig. 2 and 3). For example, a concentration-response relationship for DA was determined by adding it either to the Krebs solution or to a 10-6 mol/L NE-containing solution for 7 min every 30 min. To examine the effects of a DA1-receptor antagonist (SCH23390) and an -adrenoceptor antagonist (prazosin) on the DA-induced contraction (Fig. 1), rings were exposed for 7 min to DA plus either 10-7 mol/L SCH 23390 or 10-6 mol/L prazosin. We used prazosin at 10-6 mol/L because our preliminary experiments showed that at this concentration it completely inhibited the phenylephrine-induced 1-adrenoceptor-mediated contraction in human mesenteric arteries. In addition, to clarify the roles played by DA1-receptors in the DA-mediated modulation of NE-induced contractions (Fig. 2 and 3), 10-7 mol/L SCH 23390 was added to the NE plus DA solutions. The reasoning behind the use of a 10-7 mol/L concentration of SCH 23390 in the above experiment is that this concentration seemed to be most effective at suppressing DA-mediated DA1-receptor activation in the vessels under study. (see Results).
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The results are expressed as mean ± SD (n = number of isolated vessels). Statistical analysis (Figs. 1, 3, 4) was performed by a two-factor analysis of variance for repeated measures, followed by Scheffé’s test for multiple comparisons (including those between endothelium-intact and endothelium-denuded groups) or a two-tailed, unpaired Student’s t-test (for comparison between the control and treated groups). Probability values <0.05 were considered significant.
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Results |
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DA (10-7 to 10-5 mol/L) produced concentration-dependent contractions in RA (P < 0.05 versus the resting tension). In IMA and GEA, the threshold concentrations of dopamine required to induce contraction were larger than in RA, and this amine elicited contractile effects only at the largest concentration (10-5 mol/L)(Fig. 1). The maximal contraction induced by DA (10-5 mol/L) was much larger in RA than in the other arteries (note different ordinate scales in Fig. 1). The concentration-dependent contractions of all three arterial grafts were unaffected by the presence or absence of a functional endothelium. A DA1-antagonist, SCH23390, and an 1-adrenoceptor antagonist, prazosin, did not directly modify the resting tone in any of the three arteries at the indicated doses. Treatment with 10-7 mol/L SCH23390 enhanced the contractile response to DA in all three arteries (Fig. 1). As also shown in Figure 1, the contractions induced by DA in the three arteries were reduced by treatment with prazosin (10-6 mol/L).
As can be seen from Figure 2 (representative traces) and Figure 3 (summarized data), the NE-contracted RA contracted further in response to DA and the effect was dose-related, the contractions being significant at concentrations of 10-6 mol/L and larger. In contrast 1) the NE-contracted GEA responded to dopamine at concentrations of up to 10-7 mol/L with dose-related relaxations, although this effect was not seen at larger concentrations, whereas 2) the NE-contracted IMA relaxed in response to DA only at concentrations of 10-6 mol/L or larger. Treatment with SCH23390 (10-7 mol/L) enhanced the contractile effect of DA in RA and abolished or reversed the relaxant responses to DA in GEA and IMA. These contractile and vasorelaxant actions of DA on NE-contractions were independent of the presence or absence of vascular endothelium.
As mentioned before, 10-5 mol/L DA elicited a contraction of GEA. SCH23390 produced a significant enhancement of this DA-induced contraction at a concentration of 10-7 mol/L (Fig. 4). However, larger concentrations (
10-6 mol/L) of SCH23390 showed no such enhancing effect. In the NE (10-6 mol/L)-constricted GEA, SCH23390 induced relaxation at concentrations of 10-6 mol/L (P < 0.05 or P < 0.01 versus the initial 10-6 mol/L NE-induced contraction). Results qualitatively similar to these were obtained for rings (with or without endothelium) obtained from RA and IMA.
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Discussion |
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-adrenergic receptors. Our main conclusion is that, in the arteries studied here, DA induced DA1-mediated vascular relaxant effects together with 1-agonistic vascular contractile effects, but that the balance between dopaminergic- and adrenergic- receptor activation differed considerably among the three vessels.
Dopamine primarily stimulates DA1- and DA2-receptors, but at larger doses it also stimulates -receptors. In 1972, Goldberg et al. (12), the first to report on the cardiovascular and renal actions of DA, predicted the existence of DA DA1-receptors in renal, coronary, and mesenteric vascular beds. Since then, the DA1-receptor has been characterized as a postjunctional receptor on vascular smooth muscle and has been linked to the enzymes adenylate cyclase and phospholipase C, with its stimulation leading to vasodilation via activation of adenylyl cyclase and an increase in the intracellular concentration of cyclic AMP (13). In addition, DA2-receptors modulate NE release from prejunctional sympathetic nerve endings in vascular smooth muscle, presumably resulting in a reduction in peripheral vascular tone (14). In addition, at large concentrations this amine activates the vascular adrenergic receptors that mediate vasoconstriction. Some years ago, in GEA isolated from humans, Toda et al. (15) demonstrated that the contractile responses to DA were reversed to relaxation by treatment with phentolamine, suggesting that 1-adrenoceptors are involved in mediating this response. Earlier, Shepperson et al. (16) had indicated that DA decreases mesenteric blood flow in the anesthetized dog through stimulation of postsynaptic 2-adrenoceptors. The overall vascular actions of DA may depend on its selectivities for dopaminergic and adrenergic receptors on the smooth muscle in various vascular beds.
Little is known about the direct actions of DA on arterial bypass grafts even though this drug is commonly used for inotropic support in patients undergoing coronary bypass surgery. In the arterial rings examined in the present study, DA substantially increased tone, especially in RA, an effect that was largely prevented by the 1-adrenoreceptor antagonist prazosin. However, the DA1-receptor antagonist SCH23390 potently enhanced the DA-induced contractile response. Both effects were independent of the presence of a functional endothelium. These results suggest that DA may directly contract the smooth muscle of human RA grafts via stimulation of 1-adrenoreceptors but that these vasoconstrictor effects may be significantly attenuated via activation of DA1-receptors. Results qualitatively similar to these were obtained in both GEA and IMA, although the contractile responses were far smaller than in RA. With regard to dopaminergic receptors, DA2-receptors are also reported to be present in presympathetic ganglia in vascular smooth muscle, and their activation inhibits neurotransmission through attenuation of the release of endogenous NE (17). However, because our facilities are limited we could not directly examine whether the exogenously-applied DA used in our in vitro study prevented NE release from sympathetic nerve endings in the arteries we examined. Therefore, caution should be exercised in extrapolating these results to patients with coronary bypass grafting.
In another series of experiments (Fig. 3), DA significantly relaxed rings contracted with the -adrenergic agonist NE, and these responses were completely suppressed by the DA1-receptor antagonist SCH23390. These data support a specific DA1-receptor playing a major role in the relaxant responses to DA in the arteries used to provide grafts (when they are exposed to NE). In this study, GEA responded to small concentrations of DA with relaxation, an effect lost at larger concentrations. In contrast, we found that in IMA contracted with NE, DA induced relaxation only at larger concentrations. These effects in GEA and IMA were prevented by SCH23390. This indicates that in GEA and IMA, DA may stimulate the DA1-receptor, but it has a relatively weak effect on the 1-adrenoceptor. However, in RA DA induced a dose-related enhancement of the NE-contraction, and a further enhancement was obtained with SCH23390. This suggests that in RA, DA acts through a DA1-receptor-coupled mechanism to attenuate the -adrenergic contraction induced by NE. Taken together, the above results suggest that different selectivities of DA for DA1/1-adrenergic receptors among the human arteries used for arterial bypass grafts may lead to this amine having different vasomotor effects among these arteries. In addition, the different wall-structures of these arteries (IMA being elastic, RA muscular, and GEA mixed) (18) may in part account for the differences among these arteries in the relaxant potencies of DA, although this remains to be clarified in future investigations.
DA has been reported to dilate femoral arteries and veins in dogs by releasing an endothelium-dependent vasodilator, nitric oxide (NO), via a stimulation of the 2-adrenoceptors located in the endothelium (19). However, the physiological significance of this action is unknown in human arterial coronary grafts. In our experiments, removal of the endothelium made no difference to the vasomotor responses, indicating that a vasodilator action of DA via an 2-adrenoceptor-mediated release of NO plays little or no role in human arterial grafts. The above discrepancy may be explained by a species difference, by the portion of the vessel used in the experiments, and/or by the differing distribution of -adrenoceptor subtypes among arteries.
In clinical settings, IV DA administration in normal adults has been reported to produce dose-dependent effects (8). At small concentrations (0.5–1 µg · kg-1 · min-1), DA activates adenylyl cyclase to increase the intracellular concentration of cyclic AMP through DA1-receptor stimulation and thus induce vasodilation. At somewhat larger concentrations (2–4 µg · kg-1 · min-1), DA increases cardiac contractility because of its direct effect on ß1-adrenergic receptors and the release of NE from sympathetic myocardial fibers. With dosages larger than 5 µg · kg-1 · min-1, arterial blood pressure increases because of the agonist effect of DA on resistance blood vessel 1-receptors (8). It has been reported previously that when DA is infused at 1, 5, and 10 µg · kg-1 · min-1 in human adults, the arterial plasma DA concentration reaches 
1 x 10-7, 5 x 10-7, and 1 x 10-6 mol/L, respectively (20). Taken together, the above data indicate that of these three plasma concentrations, the smallest should be vasodilator and the others should be vasoconstrictor. In support of this, we found that DA at 10-7 mol/L relaxed GEA but that this relaxation was not seen at larger concentrations. However, in the NE-constricted IMA the in vitro threshold for relaxation was at or more than the upper limit of the aforementioned range of DA therapeutic plasma concentrations (i.e., >10-6 mol/L), although admittedly more rapid rates of DA infusion (>10 µg · kg-1 · min-1) are occasionally used clinically, depending on the hemodynamic responses of the patient. To judge from our results, such large doses of DA may require caution because a vasoconstrictor response may occur in RA grafts.
In our experiments, the relaxant potency of DA in the NE-contracted GEA was very similar to that previously reported in the prostaglandin F2-contracted human GEA (isolated from the omentum during operations for stomach cancer) (15). Thus, we can predict that some (but not all) of the complex actions of DA in vessels contracted by an agent with a mechanism of action different from that of NE may be similar to those obtained in NE-constricted vessels. However, further pharmacological and biochemical experiments will be needed to test this hypothesis.
In summary, the responses to DA differed considerably among the human arteries commonly used to provide coronary arterial grafts, presumably because of dual -adrenergic and dopaminergic actions on smooth muscle that vary in magnitude among the vessels examined. The ability of DA to attenuate NE-mediated contraction indicates its clinical importance because NE is the major sympathomimetic amine at vascular nerve endings and high levels of NE are produced during coronary bypass surgery (21,22). Theoretically, this excess of NE may contribute to graft contraction (7,23), leading to early myocardial ischemia and consequent perioperative morbidity and mortality (21). Under such conditions, however, some of the vasodilator effects of DA, exerted through DA1-receptor activation, may be beneficial in preventing or reducing complications related to hypoperfusion.
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Acknowledgments |
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References |
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2-adrenoceptors in canine vessels: comparisons between femoral arteries and veins. Clin Exp Pharmacol Physiol 1998; 25: 669–75.[ISI][Medline]
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