Clinical Science and Molecular Medicine (1976) SO, 81-90.
SHORT COMMUNICATION
Effects of halothane on pulmonary inactivation of noradrenaline and prostaglandin E, in anaesthetized dogs Y. s. BAKHLE A N D A. J. BLOCK'^) Department of Pharmacology, Institute of Basic Medical Sciences, Royal College of Surgeons of England, London (Received 2 October 1975)
Y S 1. The inactivation of noradrenaline and pro-
staglandin E2 was studied in the pulmonary circulation of anaesthetized dogs. 2. Under chloralose anaesthesia and ventilation with air, the inactivation of noradrenaline was 20% and that of prostaglandin E2 was 91 %. These values are in agreement with results from previous work. 3. When the dogs were ventilated with halothaneair mixtures, the inactivation of prostaglandin Ez was unaffected but that of noradrenaline was significantly reduced. 4. This ef€ect of halothane is probably due to an interaction with the transport system for noradrenaline associated with cell membranes. 5 . Analogous changes in pulmonary noradrenaline inactivation could occur in patients anaesthetized with halothane. Key words: halothane, metabolism, noradrenaline, prostaglandin, pulmonary circulation.
Introduction The ability of the lung to modify the biological activity of substances passing through the pulmonary circulation is now well established (for references see Bakhle & Vane, 1974). Two dissimilar substrates which are inactivated on passage through the pulPresent address: Department of Pharmacology, Astra Pharmaceuticals Ltd, Worcester, Mass., U.S.A. Correspondence: Dr Y.S. Bakhle, Department of Pharmacology, Institute of Basic Medical Sciences, Royal College of Surgeons of England, Lincoln’s Inn Fields, London WC2A 3PN.
87
monary circulation are noradrenaline (Ginn & Vane, 1968; Hughes, Gillis &Bloom, 1969; Alabaster & Bakhle, 1973) and prostaglandin El (Piper, Vane & Wyllie, 1970). For both substrates, the initial step in the inactivation process is uptake into cells, probably endothelial cells, and it has been suggested (Bakhle & Vane, 1974) that gaseous anaesthetics could affect cell membranes of endothelial cells and thus the transport mechanisms associated therewith. We decided to investigate the effect of halothane on the pulmonary inactivation of noradrenaline and PGEItZ) in viuo, using the blood-bathed organ technique (Vane, 1964) in dogs.
Methods General Mongrel dogs of either sex were anaesthetized with a-chloralose (322 pmol/kg; intravenous injection) after induction with thiopentone sodium (1 10 ,umol/kg; intravenous injection). The trachea was cannulated and all animals were artificiallyventilated with air. Polyethylene cannulae were inserted centrally into the following vessels: left and right external jugular veins, left and right femoral arteries, and ascending aorta via the right common carotid artery. The tip of the cannula in the ascending aorta lay just distal to the aortic valve, as was verified at autopsy. Systemic arterial blood pressure was measured from the right femoral artery. Rectal temperature was maintained at approximately 37°C by a thermostatically controlled heating unit c2) Abbreviation: PGE2, prostaglandin E2.
Y. S. Bakhle and A . J. Block
88
beneath the operating table. After all surgery was completed and immediately before placement of the aortic cannula (the last to be inserted), heparin (500-1000 i.u./kg) was administered intravenously.
Bioassay The pulmonary removal of noradrenaline and PGE2 was determined by the blood-bathed organ technique (Vane, 1964; Ginn & Vane, 1968; Ferreira &Vane, 1967). After heparinization, blood withdrawn from the left femoral artery, at 10 ml/min, was reoxygenated and made to superfuse several rat stomach strips. Thereafter the blood collected in a reservoir and returned to the animal by gravity into the left external jugular vein. When PGE, was to be assayed, the stomach strips were superfused for at least 1 h with Krebs solution containing a mixture of antagonists [final concentrations: mepyramine, 350 nmolll; hyoscine, 330 nmol/l ; phenoxybenzamine, 330 nmol/l; propranolol, 7 pmolll ; methysergide, 560 nmol/l; indomethacin, 2.8 ,umol/l (Eckenfels & Vane, 197211 to increase specificity and sensitivity of the assay. No antagonists were used when noradrenaline was assayed. The resting load on the tissues was 1-3 g and their contractionsweremeasured with Harvard smooth muscle transducers connected to a pen recorder. Inactivation was determined initially during chloralose anaesthesia as follows : while the assay tissues were bathed in femoral arterial blood, infusions of PGEz or noradrenaline were made alternately into the right external jugular vein (intravenous) and the ascending aorta (intraarterial). Dogs received either PGE2or noradrenaline infusions only. The infusion rates were chosen so that the contraction of the assay organs produced by the two intra-arterial infusions bracketed that produced by the intravenous infusion. For example,
if an intravenous infusion of 20 nmol/min was bracketed by intra-arterial infusions of 1 and 2 nmol/min and was equivalent to 1.6 nmol/min, this was interpreted as showing that 1-6 nmol/min of the agonist had survived passage through the pulmonary circulation. The percentage removal would then be calculated as (20- 1.6)/20 x 100 or 92%. Infusions were maintained until the assay tissues reached a plateau (3-4 min). Halothane was then administered with a Goldman vapourizer connected to the tracheal cannula via the inflow respiratory tube. After about 20 min, during which systemic arterial blood pressure had equilibrated, usually at a lower value, pulmonary inactivation was determined again. In five of the eight dogs, samples of femoral arterial blood were withdrawn and sealed in glass capillary tubes for subsequent measurement of blood halothane concentrations. Halothane was measured with a Perkin-Elmer F 11 Gas Chromatograph utilizing a flame-ionization detector equipped with a 2m x 3.1 mm(outsidediameter)column of 10% Carbowax 400 on Chromosorb W (80-100 mesh) at 110°C. We thank Mr W. C. Tilsley of the Department of Anaesthetics at the Royal College of Surgeons for carrying out these measurements.
Results Pulmonary removal of prostaglandin Ez and noradrenaline during non-gaseous as well as during halothane-augmented anaesthesia is summarized in Table 1. Control values for removal of each substance are similar to values reported previously (Ginn & Vane, 1968; Ferreira & Vane, 1967). During halothane-augmented anaesthesia, the assay tissues relaxed slightly and their responses to intra-arterial infusion of each substance were depressed slightly. Despite its direct effect on the
TABU1. Effectof halothane on pulmonary removal ofprostaglandin-Ez andnoradrenaline Values shown are meanf SEM; numbers in parentheses indicate number of animals. Removal (%)
Drug Prostaglandin Ez (3) Noradrenaline (5) (')
Control 91-Of4.1 20.0f 5.2
Halothad') 90.4f4.4 6.2f4*1'z)
Infusion rates (nmol/min) Intra-arterial Intravenous 14-55 1.5-6
55-110 3-6
Halothane concentration was 91Of 17 pmol/1(18*0+_ 3-4 mg\lOO ml). Differs significantly (P
SHORT COMMUNICATION
Effects of halothane on pulmonary inactivation of noradrenaline and prostaglandin E, in anaesthetized dogs Y. s. BAKHLE A N D A. J. BLOCK'^) Department of Pharmacology, Institute of Basic Medical Sciences, Royal College of Surgeons of England, London (Received 2 October 1975)
Y S 1. The inactivation of noradrenaline and pro-
staglandin E2 was studied in the pulmonary circulation of anaesthetized dogs. 2. Under chloralose anaesthesia and ventilation with air, the inactivation of noradrenaline was 20% and that of prostaglandin E2 was 91 %. These values are in agreement with results from previous work. 3. When the dogs were ventilated with halothaneair mixtures, the inactivation of prostaglandin Ez was unaffected but that of noradrenaline was significantly reduced. 4. This ef€ect of halothane is probably due to an interaction with the transport system for noradrenaline associated with cell membranes. 5 . Analogous changes in pulmonary noradrenaline inactivation could occur in patients anaesthetized with halothane. Key words: halothane, metabolism, noradrenaline, prostaglandin, pulmonary circulation.
Introduction The ability of the lung to modify the biological activity of substances passing through the pulmonary circulation is now well established (for references see Bakhle & Vane, 1974). Two dissimilar substrates which are inactivated on passage through the pulPresent address: Department of Pharmacology, Astra Pharmaceuticals Ltd, Worcester, Mass., U.S.A. Correspondence: Dr Y.S. Bakhle, Department of Pharmacology, Institute of Basic Medical Sciences, Royal College of Surgeons of England, Lincoln’s Inn Fields, London WC2A 3PN.
87
monary circulation are noradrenaline (Ginn & Vane, 1968; Hughes, Gillis &Bloom, 1969; Alabaster & Bakhle, 1973) and prostaglandin El (Piper, Vane & Wyllie, 1970). For both substrates, the initial step in the inactivation process is uptake into cells, probably endothelial cells, and it has been suggested (Bakhle & Vane, 1974) that gaseous anaesthetics could affect cell membranes of endothelial cells and thus the transport mechanisms associated therewith. We decided to investigate the effect of halothane on the pulmonary inactivation of noradrenaline and PGEItZ) in viuo, using the blood-bathed organ technique (Vane, 1964) in dogs.
Methods General Mongrel dogs of either sex were anaesthetized with a-chloralose (322 pmol/kg; intravenous injection) after induction with thiopentone sodium (1 10 ,umol/kg; intravenous injection). The trachea was cannulated and all animals were artificiallyventilated with air. Polyethylene cannulae were inserted centrally into the following vessels: left and right external jugular veins, left and right femoral arteries, and ascending aorta via the right common carotid artery. The tip of the cannula in the ascending aorta lay just distal to the aortic valve, as was verified at autopsy. Systemic arterial blood pressure was measured from the right femoral artery. Rectal temperature was maintained at approximately 37°C by a thermostatically controlled heating unit c2) Abbreviation: PGE2, prostaglandin E2.
Y. S. Bakhle and A . J. Block
88
beneath the operating table. After all surgery was completed and immediately before placement of the aortic cannula (the last to be inserted), heparin (500-1000 i.u./kg) was administered intravenously.
Bioassay The pulmonary removal of noradrenaline and PGE2 was determined by the blood-bathed organ technique (Vane, 1964; Ginn & Vane, 1968; Ferreira &Vane, 1967). After heparinization, blood withdrawn from the left femoral artery, at 10 ml/min, was reoxygenated and made to superfuse several rat stomach strips. Thereafter the blood collected in a reservoir and returned to the animal by gravity into the left external jugular vein. When PGE, was to be assayed, the stomach strips were superfused for at least 1 h with Krebs solution containing a mixture of antagonists [final concentrations: mepyramine, 350 nmolll; hyoscine, 330 nmol/l ; phenoxybenzamine, 330 nmol/l; propranolol, 7 pmolll ; methysergide, 560 nmol/l; indomethacin, 2.8 ,umol/l (Eckenfels & Vane, 197211 to increase specificity and sensitivity of the assay. No antagonists were used when noradrenaline was assayed. The resting load on the tissues was 1-3 g and their contractionsweremeasured with Harvard smooth muscle transducers connected to a pen recorder. Inactivation was determined initially during chloralose anaesthesia as follows : while the assay tissues were bathed in femoral arterial blood, infusions of PGEz or noradrenaline were made alternately into the right external jugular vein (intravenous) and the ascending aorta (intraarterial). Dogs received either PGE2or noradrenaline infusions only. The infusion rates were chosen so that the contraction of the assay organs produced by the two intra-arterial infusions bracketed that produced by the intravenous infusion. For example,
if an intravenous infusion of 20 nmol/min was bracketed by intra-arterial infusions of 1 and 2 nmol/min and was equivalent to 1.6 nmol/min, this was interpreted as showing that 1-6 nmol/min of the agonist had survived passage through the pulmonary circulation. The percentage removal would then be calculated as (20- 1.6)/20 x 100 or 92%. Infusions were maintained until the assay tissues reached a plateau (3-4 min). Halothane was then administered with a Goldman vapourizer connected to the tracheal cannula via the inflow respiratory tube. After about 20 min, during which systemic arterial blood pressure had equilibrated, usually at a lower value, pulmonary inactivation was determined again. In five of the eight dogs, samples of femoral arterial blood were withdrawn and sealed in glass capillary tubes for subsequent measurement of blood halothane concentrations. Halothane was measured with a Perkin-Elmer F 11 Gas Chromatograph utilizing a flame-ionization detector equipped with a 2m x 3.1 mm(outsidediameter)column of 10% Carbowax 400 on Chromosorb W (80-100 mesh) at 110°C. We thank Mr W. C. Tilsley of the Department of Anaesthetics at the Royal College of Surgeons for carrying out these measurements.
Results Pulmonary removal of prostaglandin Ez and noradrenaline during non-gaseous as well as during halothane-augmented anaesthesia is summarized in Table 1. Control values for removal of each substance are similar to values reported previously (Ginn & Vane, 1968; Ferreira & Vane, 1967). During halothane-augmented anaesthesia, the assay tissues relaxed slightly and their responses to intra-arterial infusion of each substance were depressed slightly. Despite its direct effect on the
TABU1. Effectof halothane on pulmonary removal ofprostaglandin-Ez andnoradrenaline Values shown are meanf SEM; numbers in parentheses indicate number of animals. Removal (%)
Drug Prostaglandin Ez (3) Noradrenaline (5) (')
Control 91-Of4.1 20.0f 5.2
Halothad') 90.4f4.4 6.2f4*1'z)
Infusion rates (nmol/min) Intra-arterial Intravenous 14-55 1.5-6
55-110 3-6
Halothane concentration was 91Of 17 pmol/1(18*0+_ 3-4 mg\lOO ml). Differs significantly (P
