Constant Volume Inflow Research Articles

Methylprednisolone attenuates airway and vascular responses induced by reactive oxygen species in isolated, plasma-perfused rat lungs.

The effects of methylprednisolone (MP) on the acute airway and pulmonary vascular responses induced by reactive oxygen species (ROS) were investigated in isolated, plasma-perfused rat lungs. ROS were generated by adding xanthine oxidase and hypoxanthine to the perfusate. MP was administered in 3 different ways: 1. Added to the perfusate (1 mg*ml-1) 5 min prior to xanthine oxidase and hypoxanthine, 2. Given as intraperitoneal injections (40 mg*kg-1) to lung donor rats 12 and 2 hours prior to the experiments, or 3. Combining 1 and 2. The lungs were perfused at constant volume inflow (15 ml*min-1). Pulmonary arterial pressure and transpulmonary pressure were followed for 30 min after addition of xanthine oxidase and hypoxanthine. ROS induced a powerful, acute broncho- and vasoconstriction, which was inhibited by addition of MP to the perfusate. Pretreatment with MP also inhibited the vascular and airway responses. Adding MP to the perfusate of pretreated lungs further reduced the ROS-induced smooth muscle constriction. In conclusion, MP inhibits vasoconstriction and bronchoconstriction induced by ROS in isolated rat lungs.

Constant flow‐ vs. constant pressure‐perfusion for studies of pulmonary vasoactive responses

We have compared the pulmonary vascular responses to a standardized hypoxic vasoconstrictor stimulus (FIO2 = 0.02) obtained during 1) constant volume inflow, with pulmonary arterial pressure as the dependent variable, and 2) constant inflow pressure, with flow as the dependent variable. Isolated rat lungs were perfused at different baseline transvascular pressures. The experimental arrangement allowed changes between the two types of perfusion. Hypoxia at constant pressure perfusion gave a higher percentage rise in pulmonary vascular resistance (PVR) at all pressure levels. This advantage was however, more than offset by the finding that a) vascular closure (total or partial) often occurred, particularly below arterial pressure of 3 kPa, making detection of graded responses impossible, and b) the control situation was rarely regained. Responses obtained during constant flow were less reduced by elevations in baseline transvascular pressure, and the control situtaion was rapidly and completely regained. The observation that hypoxic vascular closure may occur in the pulmonary vascular bed supports the hypothesis that high altitude edema is caused by precapillary occlusion of a major part of the vascular bed, thereby subjecting still perfused regions to very high pressure and flow.

Prostaglandin content in blood and lung tissue during alveolar hypoxia

The aim of the present work was to investigate whether prostaglandins (PGs) are synthetized and released from isolated blood-perfused rat and cat lungs secondary to vasoconstriction induced by alveolar hypoxia. The lungs were perfused with autologous blood with constant volume inflow via the pulmonary artery in a recirculating system. They were ventilated with constant volume positive pressure, and acute alveolar hypoxia was induced by ventilation with a gas containing 2% O2. A superfusion bioassay technique was used to measure PG-like activity in the perfusate from the lungs, the blood being re-oxygenated before reaching the assay tissues. The oxygenator prevented the perfusate hypoxia induced by ventilation hypoxia to affect the bioassay tissues. The assay tissues were rat stomach strip, rat colon and chick rectum. They were sensitive to calibrating doses of 0.5--1 ng/ml PGE2 and 1--2 ng/ml PGF2alpha. In another series of experiments PGs of the F-series were measured in lung tissue from normoxic and hypoxic lungs with radioimmunoassay technique. No increase in PG-like activity could be detected in the venous effluent by means of bioassay during hypoxia, nor was the lung tissue content of immunoactive PGF increased by hypoxia. The present findings indicate that alveolar hypoxia does not stimulate PG-synthesis in lungs, refuting that PGs are important mediators of the pulmonary vasoconstrictor response to alveolar hypoxia. It is concluded that PGs play no significant role in producing the pressor response to alveolar hypoxia.

Pulmonary vasomotor nerve responses in isolated perfused lungs of Macaca mulatta and Papio species.

1. Lung lobes of Macaca mulatta and Papio species were isolated from the body and perfused by a pump delivering a constant volume inflow. The left atrial pressure was kept constant and therefore any recorded change in pulmonary arterial pressure reflected a change in pulmonary vascular resistance. 2. In five Macaca mulatta preparations stimulation of the upper thoracic sympathetic chain, the stellate ganglion, the middle cervical ganglion and the thoracic vagosympathetic nerve caused a small increase in calculated pulmonary vascular resistance usually followed by a larger decrease. Evidence is produced which suggests that the depressor response is mediated by adrenergic beta-receptors. In three preparations no change in pulmonary vascular resistance occurred. 3. In four Papio preparations stimulation of similar nerves invariably caused an increase in calculated pulmonary vascular resistance. In one animal no change in vascular resistance occurred. 4. A regression analysis of the results showed an inverse relationship between the magnitude of the pulmonary vascular response to nerve stimulation and the degree of excitement of the animals during capture, restraint and anaesthesia (P less than 0.01).

Plasma-kinins and adrenergic vasodilatation in the submandibular salivary gland of the cat.

AbstractThe submandibular salivary gland of the cat has been perfused in situ at constant volume inflow and in such a way that perfusion periods with kininogen‐containing and with kininogenfree perfusates alternated. The gland was stimulated with close arterial injections of nor‐adrenaline and the secretory and vascular responses recorded. Doses of noradrenaline which caused vasodilatation or no vascular effects when cat kininogen was present in the perfusate induced brisk vasoconstrictor responses in the same gland when a kininogen‐free perfusate was used. The vasoconstrictor responses could be gradually reduced and finally abolished as the gland was reloaded with cat kininogen. Glands which had undergone repeated stimulations with noradrenaline had a markedly lower content of kallikrein than had the contralateral, unstimulated organ. It is concluded that during adrenergic stimulation of the submandibular salivary gland, kallikrein is released into the interstitial space where it induces formation of plasmakinins. The plasma‐kinins formed cause vasodilatation, and they probably play an important role in the vasodilator response observed after a period with activation through the sympathetic nerves.

The vasomotor component of the carotid sinus baroreceptor reflex in the cat during stimulation of the hypothalamic defence area.

1. The effect of moderate intensities of stimulation of the hypothalamic defence area on the baroreceptor reflex has been investigated in the cat by comparing the responses of arterial blood pressure and perfusion pressure of the isolated hind-limb muscle bed perfused at constant volume inflow, when the isolated carotid sinus was subjected to a series of non-pulsatile pressures with and without simultaneous hypothalamic stimulation.2. In the absence of hypothalamic stimulation the characteristic sigmoid curves relating sinus pressure to blood pressure or muscle perfusion pressure were obtained.3. With simultaneous stimulation of the hypothalamus a similar sigmoid relationship was found. There was no evidence of any reduction in the over-all power or maximum sensitivity of the baroreceptor reflex.4. However, in those cats which had been atropinized to abolish the cholinergically mediated muscle vasodilatation, the curves obtained during hypothalamic stimulation were displaced in such a manner as to suggest that, while baroreceptor modulation of vasoconstrictor tone continued during defence area stimulation, the blood pressure regulating mechanism had been ;reset' so that, within the physiological range of sinus pressures, any given level of sinus pressure was associated with a greater vasoconstrictor tone.5. In non-atropinized cats there was little displacement of the curves relating sinus pressure to blood pressure, while the curves relating sinus pressure to muscle perfusion pressure were displaced in the opposite direction so that over-all muscle vascular resistance was less than normal at each level of sinus pressure.

The effect of sympathetic nerve stimulation on pulmonary blood volume in isolated perfused lungs.

AbstractAarseth, P. G. Nicolaysen, and B. A. Waaler, The effect of sympathetic nerve stimulation on pulmonary blood volume in isolated perfused lungs. Acta physiol. scand. 1971. 81. 448–454.Stimulations of the vago‐sympathetic nerve trunk at 5–10 imp/sec have been carried out in isolated dog lung preparations perfused at constant volume inflow. When the left atrial pressure was kept at 3–4 cm of water, the predominant response to these stimulations was a weight reduction of the preparation. This response is interpreted as being due to a reduction in pulmonary blood volume. Nerve stimulations caused only small weight reductions, or even weight augmentations, when the left atrial pressure was at or near zero. The blood volume reductions resulting from nerve stimulations appeared to be mediated via sympathetic nerve fibres and an α‐receptor mechanism.

Consumption of kininogen in the submandibular salivary gland when activated by chorda stimulation.

1. A method is described for determination of kininogen 1 (substrate mainly for plasma kallikrein) and kininogen 2 (substrate for glandular kallikrein) independently in cat plasma.2. In anaesthetized cats the arterial inflow to, and venous outflow from, the submandibular salivary gland were isolated: a roller pump giving constant volume inflow was interposed in the arterial circuit.3. Venous blood was collected at rest, during and after stimulation of the chorda tympani, and its content of kininogens 1 and 2 were estimated. Kininogen 2 was reduced up to 60% by chorda stimulation, whereas the level of kininogen 1 was unchanged.4. On close arterial infusion of bradykinin or histamine in amounts which produce large vascular effects, including increased capillary permeability, the venous blood levels of both kininogens 1 and 2 were unchanged.5. It is concluded that the selective loss of kininogen 2 on chorda stimulation results from the release of kallikrein into the tissue spaces and reflects the extent of kinin formation within the gland.

The cite of action of nerves in the pulmonary vascular bed in the dog.

1. The effects of stimulation of the thoracic vagosympathetic nerve or upper thoracic sympathetic chain on the pulmonary vascular resistance have been studied in atropinized, isolated, ventilated lung lobes under various conditions of pulmonary circulation perfusion. Throughout the nerve-stimulation tests bronchial circulation perfusion was maintained or temporarily interrupted.2. The pulmonary vascular resistance increase evoked by nerve stimulation (a) occurred in the absence of tidal air changes; (b) did not consistently differ during predominantly ;sluice' and ;non-sluice' conditions of pulmonary circulation perfusion; (c) was approximately one and a half times greater during constant pressure than during constant volume inflow perfusion of the pulmonary circulation; and (d) was greater during reverse than during forward perfusion.3. In lung lobes perfused in either direction at constant volume inflow nerve stimulation produced an increase in inflow pressure and a diminution in total lung blood volume reflected by a temporary increase in blood outflow.4. In lung lobes in which neither the pulmonary nor the bronchial circulations were perfused and the capillaries were completely blocked by high intratracheal pressures, thus isolating the pulmonary arterial system from the venous system, nerve stimulation produced a diminution in the blood volume of both systems.5. Nerve stimulation produced a rise in bronchial arterial pressure in the absence of pulmonary circulation perfusion.6. Further evidence is adduced that pulmonary vasomotor nerve responses do not depend upon the transfer of transmitter substances from the bronchial to the pulmonary circulation.7. The possible significance of these observations in relation to the site of action of pulmonary vasomotor nerves is discussed.

The interaction of two different vasodilator mechanisms in the chorda-tympani activated submandibular salivary gland.

AbstractThe vasodilatation caused by stimulation of the chordo‐lingual nerve has been studied in the cat submandibular salivary gland perfused under conditions of constant volume inflow, at a rate equal to the flow existing in the resting organ.When a perfusate of red cells suspended in normal plasma was used at perfusion pressures of 55–120 mm Hg, a typical two‐phasic and long‐lasting vasodilatation was observed as a response to nerve stimulation. This vasodilator pattern was maintained when repeated nerve stimulations were carried out during a single perfusion period. Wlien the gland was perfused continuously with red cells suspended in a kininogen‐free solution then one of two different response patterns could be seen, When a low perfusion pressure prevailed (below 55 mm Hg), then the chorda‐mediated vasodilatation was reduced or abolished on repeated nerve stimulation. At conditions with a higher perfusion pressure (60–120 mm Hg) there developed on chorda stimulation a one‐phasic, short‐lasting vasodilatation, which was not reduced on repeated stimulations.In another series of experiments the kinin‐destroying activity of a perfusate of red cells in normal plasma was increased by addition of carboxypeptidase‐B. Chorda stimulation then caused a short‐lasting vasodilatation of the type seen when a red cell perfusate containing no kininogen was used.It is concluded from these findings that functional vasodilatation in the submandibular salivary gland is probably initiated by the effect of vasodilator nerve fibres and supported and maintained by the action of kinins formed in the activated gland.

Studies on kinin formation in functional vasodilatation of the submandibular salivary gland in cats.

AbstractThe submandibular salivary gland in cats has been perfused undcr conditions of constant volume inflow with red blood cells suspended in normal cat plasma. Such a method is favourable for studying the time course of changes in glandular vascular resistance. A 30 sec supra‐maximal stimulation of the chordo‐lingual nerve caused the vascular resistance to fall markedly within 1–6 sec. When the nerve stimulation was finished, a further decrease in perfusion pressure was usually observcd, whereafter a slow increase in vascular resistance took place.The rapidity with which the vascular response occurred indicates an initial direct action of true vasodilator nerve fibres.The second phase of the vasodilatation, outlasting the chorda stimulation itself, is claimed to be mediated by kinin formation in the activated gland:a) When the gland was perfused with normal cat plasma, so that resting perfusion pressure was low (under 55 mm Hg), then the vasodilator response to chorda stimulation changed, developing gradually and outlasting the nerve stimulation.b) Reduction of kininogen 2 in the plasma perfusate resulted then in a reduced vasodilator response to chorda stimulation.c) With no kininogen 2 in the cell‐free perfusate, stimulation of the chordo‐lingual nerve caused only a very small vasodilatation or no vasodilatation at all.d) Addition of partly purified kininogen 2 to such perfusate did then restore the chorda‐induced vasodilatation.

The involvement of the complement system in the pressor response to the injection of Forssman antibody in the guinea-pig.

1. When the hind limbs of the guinea-pig were perfused at constant volume inflow with undiluted pooled blood, an intra-arterial injection of the Forssman antibody caused a rapid and long-lasting pressor response.2. In the whole animal the pressor response was accompanied by a fall in the number of circulating platelets, but perfusion experiments suggest that serotonin was not responsible for the pressor response.3. When decomplemented serum was perfused through the hind-limb preparation the Forssman antibody failed to produce a rise in pressure, but a pressor response developed when unheated serum was subsequently introduced. This suggests that the actual combination of antibody with its tissue antigen is not the immediate cause of the pressor reaction.4. The cause of the pressor response is still unknown but apparently depends on the activation of the complement system. The effectiveness of serum shows that polypeptides split off from fibrinogen are not involved.

Role of histamine in hypoxic pulmonary hypertension in the rat. II. Depletion of histamine, serotonin, and catecholamines.

The present investigation was undertaken to see if any of the naturally occurring vasoactive substances was likely to act as a mediator for the pulmonary vasoconstrictor response to acute alveolar hypoxia. Rats were treated with agents that deplete local stores of vasoactive amines or inhibit their synthesis. Lungs from these animals were isolated, ventilated, and perfused with homologous blood at constant volume inflow. Their pressor responses to 2- to 3-minute periods of ventilation hypoxia (2% O 2 ) were observed. The histamine-releasing agent 48/80 or the histidine decarboxylase inhibitor NSD 1055, or both, depleted 73% of the histamine in the lung but had no consistent effect on the pressor responses to hypoxia. When 48/80 (0.2 to 1 mg) was given in vitro, histamine in the lung was reduced to 10% of normal, or less, and the pressor response to alveolar hypoxia was completely abolished. Reserpine, guanethidine, or alpha-methyl-tyrosine reduced catecholamine stores in heart tissue (as an index of general tissue changes) by a maximum of 90% and reserpine decreased serotonin in the lung by 93% without inhibiting the hypoxic pressor responses. The findings strengthen the concept that histamine mediates the pressor response to acute alveolar hypoxia in the rat.

PULMONARY VASOMOTOR RESPONSES OF ISOLATED PERFUSED CAT LUNGS TO ANOXIA AND HYPERCAPNIA

1. A method for perfusing isolated cats' lungs with the animal's own heparinized blood is described, with some observations on the condition of the lungs during and after perfusion.2. The pulmonary vasomotor responses to inhalation of various gas mixtures have been studied during perfusion of the pulmonary vessels at constant volume inflow.3. Inhalation of gas mixtures containing 5–10 per cent. CO2 produces pulmonary vasoconstriction, as shown by an increase of pulmonary arterial pressure.4. Inhalation of gas mixtures containing less than 15 per cent. O2 also produces a pressor response.5. Ventilation of the lungs with pure O2 has no effect under the conditions of the experiments.6. Ventilation of the lungs with pure neon or H2 produces similar effects to that produced by pure N2.7. The pulmonary vasomotor responses to CO2 and O2 lack are not abolished by dihydroergotamine or atropine.

THE ACTION OF CARBON DIOXIDE ON ISOLATED PERFUSED DOG LUNGS

1. Isolated dog lungs under negative or positive pressure ventilation have been perfused with blood through the pulmonary artery at constant volume inflow. The response of the pulmonary vessels to various concentrations of CO2 in the ventilating gas has been observed.2. The capacity of the pulmonary blood‐vessels is markedly diminished by increasing the concentration of CO2 inhaled from the small percentage present in atmospheric air to 5 or 10 per cent., and the pulmonary arterial pressure frequently shows a 5 to 20 per cent. increase.3. The lung blood‐volume shows an inverse relationship to the CO2 content of the arterial blood.4. The effect of adrenaline on the pulmonary vessels is not altered during ventilation of the lungs with 5 per cent. CO2 in air.5. The response to CO2 is not inhibited by ergotoxine or by atropine.6. The possible site of action of CO2 is discussed.

ACETYLCHOLINE ACTION ON THE PULMONARY VASCULAR BED OF THE DOG AND ITS MODIFICATION BY ADRENALINE AND BY ERGOTOXINE

In isolated lungs of the dog under negative pressure ventilation and perfused at constant volume inflow with defibrinated blood, it has been found that:1. In non‐nicotinised preparations adrenaline tends to suppress or reverse the rise in pulmonary arterial pressure produced by large doses of acetylcholine.2. In fully nicotinised preparations, ergotoxine partially or wholly suppresses, or reverses the pulmonary arterial pressure rise produced by large doses of acetylcholine.I wish to express my thanks to Professor I. de Burgh Daily and to Miss Catherine O. Hebb for helpful advice, and to the Moray Endowment Committee for defraying part of the cost of the research.

AN EXPERIMENTAL ANALYSIS OF THE ACTION OF ADRENALINE AND HISTAMINE ON DIFFERENT PARTS OF THE PULMONARY VASCULAR BED

In isolated dog's lungs under negative pressure ventilation and perfused at constant volume inflow with defibrinated blood—1. The increase in pulmonary arterial pressure and the augmentation in venous outflow produced by adrenaline are suppressed or reversed by ergotoxine.2. Evidence is produced which favours the view that the blood‐vessels responsible for the adrenaline rise in pulmonary pressure differ from those which cause the increase in venous outflow.3. Histamine in small doses tends to increase and in large doses to diminish the venous outflow. The venous outflow response to histamine is also determined by other experimental conditions; the presence of ether in the lungs and a low venous pressure favour a histamine diminution in outflow.Experiments have been reported which suggest that, by injection of drugs into the pulmonary vascular bed by way of the bronchial arteries, it is possible to confine the site of drug action chiefly or almost exclusively to the pulmonary veins in lungs perfused in the normal direction, and in all probability chiefly or almost exclusively to the pulmonary arteries in lungs perfused in the reverse direction. Using this method, results have been obtained which indicate that adrenaline in small doses constricts both pulmonary arteries and veins, and in large doses probably constricts the pulmonary capillaries.The authors express their thanks to the Government Grants Committee of the Royal Society for a grant to one of them (I. de B. D.) and to the Lewis Cameron Research Fund which defrayed the cost of animals.

THE ACTION OF DRUGS ON THE ISOLATED PERFUSED LUNGS OF THE PIG

The lungs of the pig isolated and perfused with the animal's own blood are not suitable for testing bronchial responses to the injection of drugs. Sooner or later the preparations exhibit œdema and prevent measurement of the tidal air. This statement applies to preparations made from animals bled to death under Pernocton and local anæsthesia. Vascular responses to drugs are easily obtainable and, in respect of adrenaline, exhibit a remarkable uniformity from one preparation to another. Adrenaline invariably reduces the pulmonary arterial pressure in freshly perfused lungs, and augments it during later stages of perfusion. It has been reported that a similar phenomenon may on rare occasions occur in the dog [Alcock, Berry, and Daly, 1935]. How far the adrenaline pulmonary effect depends upon the maintenance of vessel tone in some parts of the vascular bed, or to physicochemical changes in the blood or lung tissues under the artificial conditions of persusion, there is insufficient evidence to decide.In isolated pig's lungs under negative pressure ventilation and perfused at constant volume inflow with defibrinated blood:1. Adrenaline reduces the pulmonary arterial pressure in freshly perfused preparations and augments it during later stages of perfusion; ergotoxine reverses the pressor effect. Adrenaline dilates the bronchi.2. Small doses of A.Ch. tend to lower, and larger doses to raise, the pulmonary arterial pressure in ergotoxinised preparations. The bronchi constrict.I wish to express my thanks to Professor I. de Burgh Daly for his advice and encouragement during this work, and to the Moray Endowment Committee, who defrayed the expenses of the research.

OBSERVATIONS ON THE BLOOD‐PERFUSED LUNGS OF THE DOG, GUINEA‐PIG, AND MACACUS RHESUS, WITH SPECIAL REFERENCE TO “SPONTANEOUS” LUNG MOVEMENTS

The interpretation of venous outflow changes in organs perfused at constant volume inflow is discussed.The recording system itself may be responsible for changes in venous outflow when a physiological alteration in inflow pressure takes place. The main error introduced is due to capacity changes in the manometer used for measuring the inflow pressure, and is such that if a pressor response is accompanied by a venous outflow augmentation, both being due to physiological events, the latter may be masked, or even reversed by reason of the transfer of a portion of the circulating fluid to the manometer.