And Angiotensin II- Induced Changes by Calcium Antagonists in the Peripheral Circulation Ofanaesthetized Rabbits Robert P
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Br. J. Pharmac. (1985), 85, 75-87 Modification ofvasopressin- and angiotensin II- induced changes by calcium antagonists in the peripheral circulation ofanaesthetized rabbits Robert P. Hof Cardiovascular Unit of Preclinical Research, Sandoz Ltd, CH-4002 Basel, Switzerland 1 Investigations into the site of vasodilator and antivasoconstrictor activity of calcium antagonists previously performed in cats were extended to a second species, barbiturate-anaesthetized rabbits, and a second vasoconstrictor agent, vasopressin. 2 The dihydropyridine derivative darodipine (code name PY 108-068; 10, 30 and 100 pg kg-' i.v.) showed systemic haemodynamic effects comparable to those seen in cats at half these doses. Darodipine effected regional vasodilatation (measured with tracer microspheres) in the heart, brain and skeletal muscles as in cats. Only the vessels of the adrenals (dilated in rabbits but not in cats), and the kidneys and skin (constricted in rabbits but not in cats) responded differently to darodipine. 3 Angiotensin II (A II; 0.15 and 1.5 yg kg- min ')constricted the same vascular beds in rabbits as in cats, namely the heart, kidneys, small intestine, pancreas, spleen, skin and arterio-venous shunts (inferred from microspheres reaching the lungs), the only exceptions being the vessels of the stomach and liver (constriction only in cats) and the adrenals (constriction only in rabbits). 4 Darodipine (30 and 100 yg kg-') attenuated the A II-induced vasoconstriction in the same vascular beds in rabbits as in cats including the kidneys, which were constricted after administration of the antagonist alone. 5 These results indicate surprisingly small species differences for the vasodilator effects of darodipine as well as the attenuation of the vasoconstrictor effects of A II. 6 Lysine-vasopressin (2 and 50 mu kg' min-I) did not increase blood pressure in anaesthetized rabbits but dose-dependently lowered heart rate, cardiac output, total peripheral conductance and myocardial contractile force (measured with a strain gauge). Vasopressin constricted all peripheral vascular beds dose-dependently, except for those of the kidney and liver. 7 The effects of vasopressin persisted in the animals infused with placebo solution. Darodipine (30 and 100 pg kg- ), but not verapamil (300 and 1000 pg kg-') reversed the vasopressin-induced cardiac depression and decrease in cardiac output. This probably also explains most ofthe apparent differences between the effects of the two calcium antagonists on the peripheral circulation. 8 Both calcium antagonists diminished the vasopressin constriction in most vascular beds except those of the spleen, skin and arterio-venous shunts. Most of the effects were dose-related but not strictly competitive, as far as this can be judged based on two doses of agonist and antagonist. 9 As with A II the effects of vasopressin were diminished in vascular beds not normally dilated by calcium antagonists. 10 Calcium antagonists display two typical patterns of activity. The vasodilator pattern consists of dilatation of the vesels of the heart, brain and, to a degree varying with the agents, skeletal muscle. The antivasoconstrictor effects occur in some but not all of the vessels constricted by the constrictor agent, vasoconstriction of the spleen, skin and arterio-venous shunts being resistant to the action of calcium antagonists. The pattern of antivasoconstrictor activity appears to depend on the constrictor compound used, inasmuch as such agents constrict different vascular beds. Introduction Activation of various receptors on blood vessels may contributing to it (Bolton, 1979). The interaction of open receptor operated channels admitting calcium calcium antagonists with effects mediated by receptor into smooth muscle cells thus eliciting contraction or stimulation varies considerably depending on the © The Macmillan Press Ltd 1985 76 R.P. HOF vessel and on the species studied (Cauvin et al., 1983). blood gases were checked regularly. The anaesthesia Most ofthese experiments were carried out on isolated was then deepened by a further 50 mg kg- I phenobar- vessels in vitro. In earlier experiments in whole animals bitone. Catheters were placed in the lower abdominal we have searched for vascular beds where calcium aorta, the inferior vena cava and the right atrium. The antagonists could abolish or diminish angiotensin II left atrium was cannulated by means ofa thoracotomy (A II)-induced vasoconstriction. We found that the in the left 3rd intercostal space. The aortic root was antivasoconstrictor effects of the calcium antagonists cleaned of connective tissue and a flowprobe (Narco, were seen even in some vascular beds which were not 4.0- 5.5 mm) fitted onto it. The probe was calibrated in dilated when calcium antagonists were administered situ by the reference flow method at the time ofthe last to cats not pretreated with A II (Hof, 1983; 1984). We microsphere injection. The phasic flow signal was have speculated that the anatomical site of action of integrated to obtain mean aortic flow, and differen- calcium antagonists might change depending on the tiated to obtain dQ/dt, i.e. acceleration of blood in the mechanism contracting the vessel at the time of aorta, which we use as an ejection phase parameter of observation. myocardial function (Hof & Hof, 1981b). A The main aim of the present experiments was to thoracotomy in the fifth right intercostal space was extend our investigations to a second species, the used to sew a Walton-Brodie strain-gauge onto the rabbit, and to a second vasoconstrictor agent, right ventricle parallel to the superficial muscle fibres. vasopressin, in order to strengthen or to reject the The output of all devices was amplified and recorded hypothesis mentioned above. These experiments on a Beckman R612 8-channel recorder. should also provide a better basis for eventual ex- trapolations to further species, especially man. Microspheres As in the previous series of experiments we inves- tigated whether the vasoconstrictor effects of All The use of the microsphere method in our laboratory were antagonized specifically in the vascular beds has been reported in detail previously (Hofet al., 1980; where vasodilatation was observed upon administra- Hof & Hof, 1981a; 1982; Hof, 1983). tion of a calcium antagonist or whether the attenua- In brief: for each determination of regional blood tion occurred in different vascular beds. The same flow we injected about 1.5 x 105 microspheres labelled experiments were then carried out using vasopressin as with one ofthe following isotopes: 1251, 141Ce, 51Cr, 85Sr a constrictor agent. This endogenous vasoconstrictor or 46Sc. In order to avoid systematic errors due to small exerts an action on blood vessels which depends on differences between different batches ofmicrospheres, extracellular calcium (Altura & Altura, 1977). spheres with different labels were rotated, so that each Therefore, an interaction with calcium antagonists label was used for each measuring period. The spheres similar in mechanism to that of A II, yet occurring in were injected into the left atriumwith 1 ml of0.9% w/v different parts of the vascular system would be NaCl solution (saline). This procedure had no effect expected if the hypothesis, that the anatomical site of on blood pressure, heart rate or aortic flow. The action ofcalcium antagonists depends on the mechan- reference sample was withdrawn through the catheter ism of action, is correct. in the aorta at a rate of 6 ml min-' at least 30 s after finishing the injection and flushing of the left atrial catheter. The theoretical accuracy of the determina- Methods tion ofcardiac output (CO) approximates ± 2% with this protocol (Hof & Hof. 1981a). Experimental animals At the end ofthe experiment the animals were killed Large mongrel rabbits (body weight 2.8-4.5 kg) were with an overdose ofpentobarbitone and the organs to anaesthetized by injection into an ear vein of be counted were dissected and weighed. Samples of 25 mg kg-' pentobarbitone followed by 50 mg kg-' skeletal muscle were obtained from the hindlegs. All phenobarbitone a few minutes later. The animals were other organs mentioned were counted in toto. The tracheotomized and ventilated with a Loosco (Ams- heart was dissected to obtain samples of the free wall terdam) MK2 infant ventilator. Room air was used of the left ventricle, which was then divided into 3 and a positive end-expiratory pressure of2 mmHg was layers as described in detail elsewhere (Hof & Hof, applied as soon as the thorax was opened. The 1982). The papillary muscles were weighed and coun- ventilation was started with a rate of 45, a volume of ted together with the subendocardial layer. The 250 ml kg-' min-' and an inspiratory time of 40% of radioactivity of the samples was counted in a Packard the respiratory cycle; however, the rate was im- gamma counter (Mod 5921) and the spectra processed mediately adjusted to the animals own rate and the on an OKI if-800 Mod. 30 microcomputer according volume set to keep the end-expiratory CO2 between 4.0 to the method of Rudolph & Heymann (1967) with and 4.5 volume % (measured continuously with a modifications of the calculations described by Schos- Gould-Godart capnograph). In addition the arterial ser et al. (1979). VASOPRESSIN - CALCIUM ANTAGONIST INTERACTION 77 Experimental protocols calcium antagonists was infused: plus 70 Af kg-' darodipine (total cumulative dose 100tgkg-) and After the preparative procedures were complete the plus 7001tg kg-' verapamil (total cumulative dose rabbits were allowed to stabilize for approximately 1000 tig kg- ). Just before terminating the experiments 60 min while fresh drug solutions were prepared. the infusion of vasopressin was accelerated to 50 mu Synthetic vasopressin (Lysine-vasopressin, Sandoz) kg- I min-' in order to assess whether or not the effects was diluted in 5% glucose to give a dose of ofthe calcium antagonist could be reversed by a higher 100 mu kg'- ml -' solution. Darodipine was dissolved dose of vasopressin and the last set of measurements in a mixture of ethanol and polyethyleneglycol 400, was obtained after 2 min of infusion at this rate.