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Cardiovascular effects of nitroglycerin given sublingually and intravenously in unanesthetized and unrestrained dogs
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P.we.ll6 ] Guanosi~e and GTP augment glyceryl trinitrate and sodium nitrite induced vascular smooth muscle relaxation and cyclic GMP accumulation P0rsti, I., Vucrinen, P,, Laustiola *, K.E. Mets~i-Ketel~i, T., Arvola, P., Manninen *, V. and Vapaatalo, H. Department of Biomedical Sciences. Universi.tvof Tampere, Tampere and * Wihuri Research Institute, Helsinki, Finland Nitrovasodilators, including both organic and inorganic nitrogen compounds, induce smooth muscle relaxation by stimulating soluble guanylate cyclase with subsequent formation of cyclic OMP. We studied the effects of guanosine and GTP on glyceryl trinitrate and sodium nitrite induced smooth muscle relaxation and cyclic GMP accumulation in endothelium denuded rat mescnteric a,"te~al preparations. The mesenteric arteries of Spragne-Dawley rats were excised and cut to 3 mm long tings, and the endothelium was removed by gently rubbing. The rings were placed between two hooks and mounted in an organ bath chamber in Krebs bicarbonate buffer solution. Contractions were elicited by 0.5 /tM norepinephrine and 60 mM potassium chloride and measured with isometric transducers and registered on a polygraph. Successful removal of the endothelium was confirmed with 1 pM acetylcholine and if any relaxation was observed, the endothelium was further rubbed. After precontraction with either norepinephrine or potassium chloride the effects of glyceryl trinitrate and sodium nitrite on the relaxation of mesenteric arterial rings were significantly augmented by 100 pM guanosine and GTP, i.e. the dose-response curves were clearly shifted to the left. The simultaneous rises in cyclic GMP levels were also increased. E.g. after precontraction with norepinephrine, 100/LM GTP increased the relaxatica to 33 nM glyceryl trinitrate and 330 pM sodium nitrite from 22~ to 48~ and from 34 to 58~, respectively. Cyclic GMP levels increased nearly two-fold from 0.50 to 0.98 pmol/mg protein with glyceryl trinitrate and from 0.47 to 0.83 pmt~!/mg protein with sodium nitxite. Different pre-incubation times (0 to 30 min) did not affect the potentiating effect of guanosine and GTP on smooth muscle relaxation and cyclic GMP accumulation. The results indicate that guanosine and GTP augment the nitrova-sodilator induced stimulation of guanylate cyclase and smooth muscle relaxation. However, GTP is also the substrate for the production of cyclic GMP by guanylate cyclase. In order to enter a cell GTP must first undergo hydrolysis to guanosine by ectonucleotidases. Yet, the potentiating effect of guanosine and GTP on smooth muscle relaxation was not affected by different pre-incubation times, thus the results can unlikely be explained by a pure substrate effect. The possibility of a cell surface receptor linked to soluble guanylate cyclase is raised by the present results. P.we.ll7 1
Cardiovascular effects of nitroglycerin given sublinguaily and intravenously in unanesthetized and unrestrained dogs N o n a k a , K. and Ueno, A. Dept. of Pharmacology and Experimental Therapeutics, Nagasaki University School of Medicine, 12.4 Sakamoto-machi, Nagasaki 852, Japan The effects of nitroglycerin (GTN) 5-25/tg/kg given sublingually and intravenously on the cardiovascular system were studied systemically in unanesthetized and unrestrained dogs implanted electromagnetic flow sensors (Skalar, Delft, Holland), sonomicrometer elements and pressure tubings at least three weeks before the experiment. The examined parameters were the blood flow in the aorta (CO), coronary artery (CBF) and vena cava (BFV), diameters of the aorta, coronary, carotid, mesenteric, iliac, internal thoracic and pulmonary arteries and vena cava, internal diameter of the left ventricle (LVID), the pressures in the left ventricle, abdominal aorta and pulmonary arteries, organ
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thickness of the spleen, liver and sartorius muscle, and the rate of rise in arterial pressure pulses in the caudal portion of abdominal aorta (Adp/dt). GTN given sublingnally (s.i.), did not effectively dilate the venous system which revealed by calibrating the diameter of the vena cava and the thickness of the liver, spleen and sartorius muscle. This was also supported indirectly by the observation that GTN never reduced CO and BFV. GTN s.l. essentially produced no effect on coronary resistance vessels, e.g. CBF was not increased, but efficiently dilated the large coronary artery. This action was apparent not only in coronary artery but also in all artery tested. It is therefore, proposed that this action of GTN can be called "selective conductance a~erial dilation". GTN s.l. did not essentially reduce the preload, as far as changes in heart rate (HR). and CO concerned. Reduction of preload, decrease of LVID with reduction of LVEDP, was noted only in cases exhibiting a moderate fall in systolic pressure with concomitant increase in HR and CO with a moderate large dose. Thus, the reduction of preload by GTN is a secondary feature resulting from shift of the blood in heart, lung and vein to the arterial side by the baroreceptor reflexes mediated cardio-accerelation in response to hypotension in normal subjects. Intravenous GTN caused increases in HR, CO and CBF and reductions in BP, LVEDP and LVlD transiently, suggesting participation of dilation of resistance vessels exposed to high concentration of GTN, and increases in arterial diameters lasted several minutes with decreasing Adp/dt and systolic pressure indicating increase in the volume of Windkessel. The present results indicate that the prime vascular action of GTN s.l. in a certain limited dose range is solely conductance arterial dilation. Other actions such as reduction of LVID and LVEDP and increase of HR and CO are mainly derived from the vasoreceptor reflexes and suggest that the therapeutically important effects of GTN in angina pectoris involve selective dilation of conductance vessels, including both coronary and general systemic arteries. The effect of the former would act directly on stenotic lesions in the epicardiai large coronary artery, improving blood flow to the ischemic part of the myocardium, whereas the latter might be beneficial for reducing cardiac work load and lowering systolic pressure by increasing the arterial compliance. Therefore, the heart regains its ventricular performance and propels stagnated blood in the heart, lungs and veins to the arterial side, resulting in a reduction of the elevated preload in anginal patients. This work was supported by a research grant from the Japanese Ministry of Education.
IpwoHsl Study of the mechanism behind the relaxing effect of furosemide on vascular smooth muscle Tian, R., Aalkjeer, C. and Andreasen, F. Institute of Pharmacology, University of Aarhus, DK-8000, Aarhus C, Denmark
A relaxing effect of furosemide on vascular smooth muscle in vitro has been observed although the mechanism responsible is only partly understood. It has been suggested by Deth et al. (1987) and Kreye et al. (1981) that furosemide via the inhibition of the Na ÷, K ÷, Cl--cotransport causes membrane hyperpolarization and consequent suppression of contraction. The present study was intended to evaluate the importance of the endothelium, the membrane potential and the transmembrane ion movement for the in vitro effect of furosemide. Se~,nients of the rabbit central ear artery (CEA) were mounted in a tissue bath with physiological saline solution (PSS) gassed with carbogen at 37°C and p H - 7.4. The effect of furosemide on isometric contraction was studied. Results are expressed as mean :l: s.e.m.(number of segments). Furosemide (0.06 mM-1.0 raM) caused a dose-dependent inhibition of contraction (8.7 :t: 1.5~ (10) to 33.3 :t: 7.7~ (8)) to electrical field stimulation. Removal of the endothelium did not affect the force development nor the relaxing effect of furosemide (p > 0. 05). A constant depression of contraction by furosemide was seen over the entire range of the K ÷ depolarization dose-response curve (p < 0.05). In fully depolarized segments ([K ÷] - 1 2 0 mM) furosemide (0.12 mM or 1.0 mM) did not alter the sensitivity of the vessel segments to external [Ca +*] a|though the maximal
P.we.ll6 ] Guanosi~e and GTP augment glyceryl trinitrate and sodium nitrite induced vascular smooth muscle relaxation and cyclic GMP accumulation P0rsti, I., Vucrinen, P,, Laustiola *, K.E. Mets~i-Ketel~i, T., Arvola, P., Manninen *, V. and Vapaatalo, H. Department of Biomedical Sciences. Universi.tvof Tampere, Tampere and * Wihuri Research Institute, Helsinki, Finland Nitrovasodilators, including both organic and inorganic nitrogen compounds, induce smooth muscle relaxation by stimulating soluble guanylate cyclase with subsequent formation of cyclic OMP. We studied the effects of guanosine and GTP on glyceryl trinitrate and sodium nitrite induced smooth muscle relaxation and cyclic GMP accumulation in endothelium denuded rat mescnteric a,"te~al preparations. The mesenteric arteries of Spragne-Dawley rats were excised and cut to 3 mm long tings, and the endothelium was removed by gently rubbing. The rings were placed between two hooks and mounted in an organ bath chamber in Krebs bicarbonate buffer solution. Contractions were elicited by 0.5 /tM norepinephrine and 60 mM potassium chloride and measured with isometric transducers and registered on a polygraph. Successful removal of the endothelium was confirmed with 1 pM acetylcholine and if any relaxation was observed, the endothelium was further rubbed. After precontraction with either norepinephrine or potassium chloride the effects of glyceryl trinitrate and sodium nitrite on the relaxation of mesenteric arterial rings were significantly augmented by 100 pM guanosine and GTP, i.e. the dose-response curves were clearly shifted to the left. The simultaneous rises in cyclic GMP levels were also increased. E.g. after precontraction with norepinephrine, 100/LM GTP increased the relaxatica to 33 nM glyceryl trinitrate and 330 pM sodium nitrite from 22~ to 48~ and from 34 to 58~, respectively. Cyclic GMP levels increased nearly two-fold from 0.50 to 0.98 pmol/mg protein with glyceryl trinitrate and from 0.47 to 0.83 pmt~!/mg protein with sodium nitxite. Different pre-incubation times (0 to 30 min) did not affect the potentiating effect of guanosine and GTP on smooth muscle relaxation and cyclic GMP accumulation. The results indicate that guanosine and GTP augment the nitrova-sodilator induced stimulation of guanylate cyclase and smooth muscle relaxation. However, GTP is also the substrate for the production of cyclic GMP by guanylate cyclase. In order to enter a cell GTP must first undergo hydrolysis to guanosine by ectonucleotidases. Yet, the potentiating effect of guanosine and GTP on smooth muscle relaxation was not affected by different pre-incubation times, thus the results can unlikely be explained by a pure substrate effect. The possibility of a cell surface receptor linked to soluble guanylate cyclase is raised by the present results. P.we.ll7 1
Cardiovascular effects of nitroglycerin given sublinguaily and intravenously in unanesthetized and unrestrained dogs N o n a k a , K. and Ueno, A. Dept. of Pharmacology and Experimental Therapeutics, Nagasaki University School of Medicine, 12.4 Sakamoto-machi, Nagasaki 852, Japan The effects of nitroglycerin (GTN) 5-25/tg/kg given sublingually and intravenously on the cardiovascular system were studied systemically in unanesthetized and unrestrained dogs implanted electromagnetic flow sensors (Skalar, Delft, Holland), sonomicrometer elements and pressure tubings at least three weeks before the experiment. The examined parameters were the blood flow in the aorta (CO), coronary artery (CBF) and vena cava (BFV), diameters of the aorta, coronary, carotid, mesenteric, iliac, internal thoracic and pulmonary arteries and vena cava, internal diameter of the left ventricle (LVID), the pressures in the left ventricle, abdominal aorta and pulmonary arteries, organ
1273
thickness of the spleen, liver and sartorius muscle, and the rate of rise in arterial pressure pulses in the caudal portion of abdominal aorta (Adp/dt). GTN given sublingnally (s.i.), did not effectively dilate the venous system which revealed by calibrating the diameter of the vena cava and the thickness of the liver, spleen and sartorius muscle. This was also supported indirectly by the observation that GTN never reduced CO and BFV. GTN s.l. essentially produced no effect on coronary resistance vessels, e.g. CBF was not increased, but efficiently dilated the large coronary artery. This action was apparent not only in coronary artery but also in all artery tested. It is therefore, proposed that this action of GTN can be called "selective conductance a~erial dilation". GTN s.l. did not essentially reduce the preload, as far as changes in heart rate (HR). and CO concerned. Reduction of preload, decrease of LVID with reduction of LVEDP, was noted only in cases exhibiting a moderate fall in systolic pressure with concomitant increase in HR and CO with a moderate large dose. Thus, the reduction of preload by GTN is a secondary feature resulting from shift of the blood in heart, lung and vein to the arterial side by the baroreceptor reflexes mediated cardio-accerelation in response to hypotension in normal subjects. Intravenous GTN caused increases in HR, CO and CBF and reductions in BP, LVEDP and LVlD transiently, suggesting participation of dilation of resistance vessels exposed to high concentration of GTN, and increases in arterial diameters lasted several minutes with decreasing Adp/dt and systolic pressure indicating increase in the volume of Windkessel. The present results indicate that the prime vascular action of GTN s.l. in a certain limited dose range is solely conductance arterial dilation. Other actions such as reduction of LVID and LVEDP and increase of HR and CO are mainly derived from the vasoreceptor reflexes and suggest that the therapeutically important effects of GTN in angina pectoris involve selective dilation of conductance vessels, including both coronary and general systemic arteries. The effect of the former would act directly on stenotic lesions in the epicardiai large coronary artery, improving blood flow to the ischemic part of the myocardium, whereas the latter might be beneficial for reducing cardiac work load and lowering systolic pressure by increasing the arterial compliance. Therefore, the heart regains its ventricular performance and propels stagnated blood in the heart, lungs and veins to the arterial side, resulting in a reduction of the elevated preload in anginal patients. This work was supported by a research grant from the Japanese Ministry of Education.
IpwoHsl Study of the mechanism behind the relaxing effect of furosemide on vascular smooth muscle Tian, R., Aalkjeer, C. and Andreasen, F. Institute of Pharmacology, University of Aarhus, DK-8000, Aarhus C, Denmark
A relaxing effect of furosemide on vascular smooth muscle in vitro has been observed although the mechanism responsible is only partly understood. It has been suggested by Deth et al. (1987) and Kreye et al. (1981) that furosemide via the inhibition of the Na ÷, K ÷, Cl--cotransport causes membrane hyperpolarization and consequent suppression of contraction. The present study was intended to evaluate the importance of the endothelium, the membrane potential and the transmembrane ion movement for the in vitro effect of furosemide. Se~,nients of the rabbit central ear artery (CEA) were mounted in a tissue bath with physiological saline solution (PSS) gassed with carbogen at 37°C and p H - 7.4. The effect of furosemide on isometric contraction was studied. Results are expressed as mean :l: s.e.m.(number of segments). Furosemide (0.06 mM-1.0 raM) caused a dose-dependent inhibition of contraction (8.7 :t: 1.5~ (10) to 33.3 :t: 7.7~ (8)) to electrical field stimulation. Removal of the endothelium did not affect the force development nor the relaxing effect of furosemide (p > 0. 05). A constant depression of contraction by furosemide was seen over the entire range of the K ÷ depolarization dose-response curve (p < 0.05). In fully depolarized segments ([K ÷] - 1 2 0 mM) furosemide (0.12 mM or 1.0 mM) did not alter the sensitivity of the vessel segments to external [Ca +*] a|though the maximal