Usichenko, Taras I. MD*; Foellner, Sebastian†; Gruendling, Matthias MD*; Feyerherd, Frank MD*; Lehmann, Christian MD*; Wendt, Michael MD*; Pavlovic, Dragan MD*
Akrinor (AKR), a mixture of theodrenaline (TDR) and cafedrine (CDR), is a sympathomimetic agent widely used to counter transitory hypotension in internal medicine and anesthesia.1–3 To satisfy the clinical criteria for a mild short-acting vasopressor agent, the theophylline was esterified with noradrenaline to yield TDR 7-2-2-(3,4-dihydroxy-phenyl-2-hydroxy-ethylamino)-ethyl-theophylline and norephedrine, yielding CDR (−)-7-2-(2-hydroxy-1-methyl-2-phenyl-ethylamino)-ethyl-theophylline.4 According to experimental data obtained in a dog model, AKR should increase the coronary blood flow,5 however, some cases of vascular complications in patients (myocardial and cerebral ischemia) associated with AKR have been reported.6,7 Investigating the clinical effects of the AKR constituents TDR and CDR separately, Schleusing and Bartsch8 described typical symptoms of angina pectoris accompanied by cardiac arrhythmia after the intravenous application of TDR in patients with coronary heart disease and arterial hypertension. As there are no experimental data about direct effects of AKR on coronary arteries, we tested its effect on pig coronary arteries using an in vitro model of isometric contraction of ring preparations.
We also studied the possible role of the endothelium, as well as dopamine, adenosine, and adrenergic receptors on the effects induced by AKR in pig coronary artery preparations. As AKR is a mixture of TDR and CDR, the effects of each substance on the coronary arteries in vitro were also studied. AKR is used only in Austria, Germany, South Africa, and countries of the former USSR. Therefore, we compared the effects of AKR, TDR, and CDR with the noncatecholamine sympathomimetic agent ephedrine (EDR), which is commonly used for treatment of transitory hypotension in other countries of Europe and in North America.9
Preparation of Arteries
The experiments were carried out using the preparations of the middle segment of porcine left anterior descending coronary artery. On the morning of each experiment day the arteries were carefully dissected out of the fresh porcine hearts that were obtained immediately after animal death in the local abattoir. The coronary arteries were cleaned of connective tissue and immersed in Krebs-Henseleit solution [KH (in mM): 113 NaCl, 4.8 KCl, 1.3 MgCl2×6H2O, 1.2 KH2PO4, 25 NaHCO3, 2.5 CaCl2, 5.7 glucose]. The cleaned segments of the coronary arteries (2 to 3 mm in diameter) were cut into 2 to 3 mm long rings and mounted between 2 stainless steel hooks, where the lower hook served as a fixed point and the upper hook was connected to the isometric force transducer (Entran, ELJ-S045C-35G, EMKA Technologies, Paris, France). The signal was amplified (STA 2808, EMKA Technologies, Paris, France) and displayed on a paper recorder (Rikadenki multipen recorder, R-50 series, Hugo Sachs Elektronik, March-Hugstetten, Germany). The mounted preparation was then immersed in the organ bath (20 mL) that was filled with KH solution. The KH solution (at 37°C, pH 7.4, oxygenated with 95% O2/5% CO2) was changed every 20 minutes. The force transducer was attached to a micromanipulator, which permitted displacement of the upper hook along a strict vertical axis and adjustment of the muscle length.
After a period of 60 minutes of stabilization, the coronary artery ring preparations were stretched to their optimal length which, according to our preliminary experiments, corresponded to a counterweight of 2 g. The relaxation experiments were then performed after precontraction with 20 mM KCl. In some preparations the endothelium was removed by gently rubbing the interior surface of the arterial ring with a cotton swab.
The effects of AKR alone, in concentration from 2×10−8 to 10−2 from the original solution (ORIG) for intravenous application (which contains 10.5 g% of active agents, see Substances), and in combination with unspecific β-adrenoreceptor antagonist propranolol (PROP, 130 mM), α1-adrenoreceptor antagonist prazosin (PRAZ, 100 mM), dopamine receptor D1 antagonist R(+)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrochloride-SCH 23390 (SCH), or adenosine receptor antagonist 9-chloro-2-(2-furanyl)-[1,2,4]triazolo[1,5-c]quinazolin-5-amine; 10-CGS 15943 (CGS) were studied. The substances were added to the organ bath after the preparations were precontracted with KCl and the tension reached a plateau. After the period of 30 minutes of preincubation with each substance, AKR was added to the bath solution in a cumulative manner. In the next set of tests the effects of AKR constituents TDR and CDR, and the other vasopressor EDR in concentrations of 2×10−8 to 10−2 M were studied alone and in combination with PROP or PRAZ in pig coronary artery preparations precontracted with KCl. The effects of AKR, TDR, CDR, and EDR were also examined at basic tension, that is, not precontracted with KCl.
AKR, CDR (−)-7-2-(2-hydroxy-1-methyl-2-phenyl-ethylamino)-ethyl-theophylline, and TDR 7-2-2-(3,4-dihydroxy-phenyl-2-hydroxy-ethylamino)-ethyl-theophylline original substances were obtained from AWD.pharma GmbH, Dresden, Germany. EPH original substance, dopamine receptor antagonist R(+)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrochloride (SCH 23390) and adenosine receptor antagonist 9-chloro-2-(2-furanyl)-[1,2,4]triazolo[1,5-c]quinazolin-5-amine; 10 (CGS) were purchased from Sigma-Aldrich Chemie, Deisenhofen, Germany. All substances were dissolved in distilled water on the day of the experiments and later diluted with KH solution.
The results are expressed as mean±SEM. Relaxation or contraction is expressed as percentage of precontraction induced by 20 mM KCl, which was taken as 100% from basic tension. The results were analyzed with a Student t test for paired observations. A probability level of less than 0.05 was considered significant. N indicates the number of animals.
Concentration-response curves obtained in preparations precontracted with 20 mM KCl with various compounds are shown in Figure 1. The derived values of corresponding EC50 are given in Table 1.
AKR Relaxes Ring Preparations of Pig Coronary Arteries Precontracted With KCl
AKR induced concentration-dependent relaxation of the preparations precontracted with 20 mM KCl (Figs. 1A, 2A), beginning with the concentration of 2×10−3 ORIG (N=7, 8.3±11.3%). Removal of the endothelium facilitated the AKR-induced relaxation of coronary artery rings, which was already noticeable at an AKR concentration of 4×10−6 ORIG (N=8, −29±9.9%). Incubation either with 10−6 M SCH or with 10−6 M CGS did not influence relaxation induced by AKR (EC50, Table 1).
AKR Contracts Ring Preparations of Pig Coronary Arteries Pretreated With PROP
AKR induced transient contraction of preparations precontracted with KCl, which were pretreated with 130 mM PROP (Figs. 1B, 2B). The contraction of vascular rings reached its peak at an AKR concentration of about 4×10−4 ORIG (N=10, 164±9%). The higher concentration of AKR relaxed the contracted preparations to 16±4.8%. AKR-induced transient contraction of pig coronary ring pretreated with PROP was prevented by incubation with 100 mM PRAZ (N=5).
TDR and EDR but not CDR Contract Ring Preparations of Pig Coronary Artery Pretreated With PROP
TDR (Fig. 1C) induced dose-dependent relaxation of the pig coronary artery preparations precontracted with KCl (N=10). Similar to the effect of AKR, TDR induced transient contraction of preparations pretreated with 130 mM PROP (N=6, 180±14%). CDR (Fig. 1D) relaxed the preparations of pig coronary arteries in a dose-dependent manner alone (N=7) and after pretreatment with 130 mM PROP (N=6). EDR (Fig. 1E) increased the tonus of the vascular rings (N=6, 126±8%). Pretreatment of the pig coronary ring preparations with 130 mM PROP slightly enhanced the effect of EDR (N=6; 143±22%). In higher concentrations, EDR did not produce relaxation of the coronary arteries of the pig.
Effects of Tested Substances on the Basic Tension of Pig Coronary Artery Ring Preparations
Tested substances AKR, TDR, CDR, and EDR in concentrations of 2×10−8 to 10−2 did not influence the basic tension of coronary artery ring preparations (results not shown).
The results of the present study demonstrate that AKR, a mixture of TDR and CDR, relaxed pig coronary artery preparations with and without endothelium. In the preparations pretreated with the β-adrenoreceptor antagonist PROP, AKR induced transitory contraction. This contraction could be prevented by application of the α1-adrenoreceptor antagonist PRAZ. Further, separate examination of the AKR constituents TDR and CDR demonstrated that both of them also relaxed pig coronary artery preparations. However, only TDR seemed to be responsible for transitory contraction of coronary artery ring preparations pretreated with PROP.
Both animal and human experimental studies from the 1960s demonstrated that AKR-induced increase of blood pressure, cardiac output, and stroke volume could be attributed to the stimulation of β-adrenoreceptors.1,2,4,8,10–14 AKR-induced relaxation of pig coronary artery preparations in our study indirectly confirms the presence of the possible β-adrenergic mechanism. Indeed, porcine coronary arteries are rich in β-adrenergic receptors and their stimulation produces coronary dilation.15 The α-adrenoreceptors, mediating the contraction of the smooth muscles of the arterial wall, seem not to be present to a significant extent in pig tissue.15,16 This is also true of human coronary arteries, where the relaxation mediated by β-adrenoreceptors is greater in absolute magnitude than α-adrenoreceptor-mediated contraction.17,18 The dopamine and adenosine receptor systems do not seem to be involved in the relaxation mechanism of AKR, as neither the dopamine receptor D1 antagonist SCH nor the adenosine receptor antagonist CGS affected AKR-induced relaxation.
Although some experimental data indicated that the inhibition of NO synthesis enhances the noradrenergic vasoconstriction in canine coronary arteries,19 we did not observe diminished sensitivity to AKR-induced relaxing effects of pig coronary arteries after removing the endothelium in our study. On the contrary, the removal of endothelium partially enhanced the relaxing effect of AKR. This might be due to the poor content of endothelium-derived relaxing factor in pig coronary arteries.20 Another reason may be the precontraction with KCl, which was shown to reduce the vasodilation potency of endothelium-derived relaxing factors and prostacyclin released from endothelial cells.21
Our finding of AKR-induced and TDR-induced transitory contraction in coronary artery ring preparations pretreated with PROP are in agreement with previous data, indicating that nonspecific β-blockade preceding the adrenergic stimulation results in coronary vasoconstriction.16,17,21–23 The mechanism of coronary contraction, induced by the β-adrenoreceptor antagonist in this case, might be the unmasking of α-adrenergic tonus by blocking the dilation mediated through the β-adrenoreceptors.22 As β-adrenoreceptors mediate the dilation of epicardial coronary arteries, the β-adrenoreceptor blocking agents would be expected to facilitate vasoconstriction and vasospasm.16,18 Studying the functional distribution of α-adrenergic receptors in the coronary arteries, Chilian24 demonstrated that the α1-adrenoreceptors, dominating in large epicardial arteries, are responsible for coronary vasoconstriction. Our finding, where the α1-adrenoceptor antagonist PRAZ prevented contraction of coronary arteries pretreated with PROP, permitting AKR to produce relaxation, confirmed the finding of this previous study. This further confirmed that if the β-adrenoceptor is blocked, AKR stimulation of the α1-adrenergic receptor is in fact responsible for transitory coronary contraction.
The experimental animal and human studies on the effects of TDR and CDR separately showed that TDR alone produced α-adrenergic contraction in various types of vessels.25,26 These contracting effects were attenuated or completely antagonized by simultaneous application of CDR, suggesting that it stimulates β-adrenoreceptors.26 In our experiments we could not confirm that CDR had important β-adrenoreceptor–stimulating effects, as both CDR concentration-response curves with and without PROP pretreatment were identical (Fig. 1D).
It was found that EDR produced a continuous increase of the tonus of pig coronary artery preparations, which was slightly enhanced after preincubation with PROP. EDR is both an α-adrenergic and β-adrenergic agonist, which in addition enhances the release of norepinephrine from sympathetic neurons.27 The moderate increase of the pig coronary ring tonus after incubation with EDR might be due to direct stimulation of α-adrenoreceptors.28 The further increase in tension after application of PROP, similar to that induced by AKR, might have been caused by enhanced stimulation of α-adrenergic receptors; the relaxing effect mediated through β-adrenoreceptors, however, was absent. In contrast to AKR, in higher concentrations, EDR did not produce a relaxation of the coronary artery preparations, indicating that EDR-induced release of norepinephrine produces mainly α-adrenergic stimulation, whereas β-adrenergic receptors, responsible for the relaxation effect, are weakly stimulated.
Our findings are confined to the in vitro investigation of porcine epicardial coronary artery rings, focusing on the neurohumoral factors influencing the contractility of smooth muscles. We cannot conclusively extrapolate the results of our study to human physiology, as there is at least a considerable genetic difference in receptor distribution between the species. The pathophysiology and pathopharmacology of human coronary arteries in vivo are much more complex. Quantitative coronary angiography combined with Doppler and PET techniques did not reveal the α-adrenergic coronary constrictor tone in humans with intact coronary arteries.18,29 But the α-adrenergic coronary vasoconstriction, enhanced during exercise and coronary interventions in patients with coronary artery disease may indeed cause myocardial ischemia and restrict myocardial function.30 Therefore, the role of β-adrenoreceptor antagonists is controversial as recently reviewed reports show that β-adrenoreceptor antagonists aggravate coronary spasm.31
In conclusion, we demonstrated that the sympathomimetic agent AKR induces endothelium-independent relaxation of pig coronary artery preparations precontracted with KCl. This mechanism may be partly mediated through β-adrenergic receptors, as the pretreatment with the nonselective β-adrenoreceptor antagonist PROP produces contraction of the preparations, unmasking the α-adrenergic receptors for stimulation. Further investigation will be required to study the effects of AKR and its constituents in the animal model in vivo and also in vitro on human coronary arteries. Considering the results of the present study, it is obvious that AKR should be used with caution in patients receiving β-blocker therapy. Regarding the compound structure of AKR and complex, partly reciprocal antagonizing effects of its constituents,26 the expedience of AKR application, at least to treat hypotension during anesthesia in the era of universal preoperative β-blockade,32–34 is questionable.
The authors thank the AWD.pharma GmbH, Dresden, Germany, for providing us with the samples of original substances of the drug used in our investigation.
1. Schieffer H, Heinz H, Sternitzke N, et al. Effect of Akrinor on the cardiac and circulatory dynamics as well as on pulmonary circulation in patients with and without heart diseases. Verh Dtsch Ges Inn Med. 1971;77:948–952.
2. Heller A, Grosser KD. Hemodynamics in patients with myocardial infarct following intravenous administration of Akrinor. Med Welt. 1974;25:1890–1892.
3. Dubova MN. Correction of cardiovascular hypodynamics in the early postoperative period with the adrenergic beta receptor agonist acrinor. Anesteziol Reanimatol. 1981;5:21–24.
4. Hutschenreuter K, Schneider M. On a circulatory analeptic with a new type of mechanism of action. Anaesthesist. 1962;11:129–131.
5. Schlepper M, Witzleb E. Coronary circulation and O2
consumption in mammalian hearts under the influence of a circulatory analeptic with a new type of activity. Arzneimittelforschung. 1962;12:841–842.
6. Kulka PJ, Scheu C, Tryba M, et al. Coronary artery plaque disruption as cause of acute myocardial infarction during cesarean section with spinal anesthesia. J Clin Anesth. 2000;12:335–338.
7. Conde Lopez VJ, Ballesteros Alcalde MC, Blanco Garrote JA, et al. Cerebral infarction in an adolescent girl following an overdose of paroxetine and caffedrine combined with theodrenaline. Actas Luso Esp Neurol Psiquiatr Cienc Afines. 1998;26:333–338.
8. Schleusing G, Bartsch C. The effect of synthetic theophylline derivatives with molecule groups having circulatory activity on blood pressure and pulse rate as well as ECG-recordings of persons with and free from circulatory disorders. Arzneimittelforschung. 1963;13:470–474.
9. Morgan P. The role of vasopressors in the management of hypotension induced by spinal and epidural anaesthesia. Can J Anaesth. 1994;41:404–413.
10. Fischer F, Weis KH. Experimental circulatory tests and clinical experiences with 2 theophylline derivatives. Anaesthesist. 1965;14:147–153.
11. Mehmel H, Schmidt HD, Schmier J. Effects of 2 theophylline derivatives and their combination on heart-lung-preparation of dog before and after a blockade of beta-receptors and after a catecholamine depletion. Arzneimittelforschung. 1967;17:1407–1411.
12. Eichhorn O. Testing of Akrinor in neuroleptic-related hypotensive circulatory disorders in a double-blind process. Med Welt. 1969;21:1250–1252.
13. Sakai K, Hashimoto K. Studies on the pressor response to aminoalkyl derivatives of theophylline, noradrenalinetheophylline and norephedrinetheophylline. Arzneimittelforschung. 1972;22:693–697.
14. Sternitzke N, Schieffer H, Bette L. Effect of Akrinor on cardiovascular-dynamics before and after blockade of adrenergic beta-receptors by propranolol. Z Kardiol. 1975;64:419–430.
15. Ito Y, Kitamura K, Kuriyama H. Effects of acetylcholine and catecholamines on the smooth muscle cell of the porcine coronary artery. J Physiol. 1979;294:595–611.
16. Horst MA, Robinson CP. Action of agonists and antagonists on adrenergic receptors in isolated porcine coronary arteries. Can J Physiol Pharmacol. 1985;63:867–871.
17. Ginsburg R, Bristow MR, Harrison DC, et al. Studies with isolated human coronary arteries. Some general observations, potential mediators of spasm, role of calcium antagonists. Chest. 1980;78:180–186.
18. Hodgson JM, Cohen MD, Szentpetery S, et al. Effects of regional alpha- and beta-blockade on resting and hyperemic coronary blood flow in conscious, unstressed humans. Circulation. 1989;79:797–809.
19. Woodman OL, Pannangpetch P. Enhancement of noradrenergic constriction of large coronary arteries by inhibition of nitric oxide synthesis in anaesthetized dogs. Br J Pharmacol. 1994;112:443–448.
20. Christie MI, Griffith TM, Lewis MJ. A comparison of basal and agonist-stimulated release of endothelium-derived relaxing factor from different arteries. Br J Pharmacol. 1989;98:397–406.
21. Noll G, Buhler FR, Yang Z, et al. Different potency of endothelium-derived relaxing factors against thromboxane, endothelin, and potassium chloride in intramyocardial porcine coronary arteries. J Cardiovasc Pharmacol. 1991;18:120–126.
22. Turlapaty PD, Altura BM. Propranolol induces contractions of canine small and large coronary arteries. Basic Res Cardiol. 1982;77:68–81.
23. Tilmant PY, Lablanche JM, Thieuleux FA, et al. Detrimental effect of propranolol in patients with coronary arterial spasm countered by combination with diltiazem. Am J Cardiol. 1983;52:230–233.
24. Chilian WM. Functional distribution of alpha 1- and alpha 2-adrenergic receptors in the coronary microcirculation. Circulation. 1991;84:2108–212.
25. Sakai K, Shioya A, Hashimoto K. Effect of combining noradrenalinetheophylline and norephedrinetheophylline in various ratios on renal circulation. Arzneimittelforschung. 1972;22:698–701.
26. Sternitzke N, Schieffer H, Rettig G, et al. Hemodynamic effects of the theophylline derivatives cafedrine and theodrenaline as single agents in comparison with their combination. Herz Kreislauf. 1984;16:401–412.
27. Hoffmann BB. Catecholamines, sympathomimetic drugs, & adrenergic receptor antagonists. In: Hardmann JG, Limbird LE, eds. Goodman & Gilman's The Pharmacological Basis of Therapeutics. New York: McGraw-Hill; 2001:237–240.
28. Cohn JN. Comparative cardiovascular effects of tyramine, ephedrine and norepinephrine in man. Circ Res. 1965;16:174–182.
29. Aptecar E, Dupouy P, Benvenuti C, et al. Sympathetic stimulation overrides flow-mediated endothelium-dependent epicardial coronary vasodilation in transplant patients. Circulation. 1996;94:2542–2550.
30. Heusch G, Baumgart D, Camici P, et al. Alpha-adrenergic coronary vasoconstriction and myocardial ischemia in humans. Circulation. 2000;101:689–694.
31. Konidala S, Gutterman DD. Coronary vasospasm and the regulation of coronary blood flow. Prog Cardiovasc Dis. 2004;4:349–373.
32. Auerbach AD, Goldman L. Beta-blockers and reduction of cardiac events in noncardiac surgery: scientific review. JAMA. 2002;287:1435–1444.
33. London MJ, Zaugg M, Schaub MC, et al. Perioperative beta-adrenergic receptor blockade: physiologic foundations and clinical controversies. Anesthesiology. 2004;100:170–175.
34. Lindenauer PK, Pekow P, Wang K, et al. Perioperative beta-blocker therapy and mortality after major noncardiac surgery. N Engl J Med. 2005;353:349–361.
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