phenomenon has been referred to as the "cocaine paradox" [Burn and Our own interest in the actions of cocaine and tyramine lay in the

Transcription

1 : : : : THE POTENTIATION OF HISTAMINE AND TYRAMINE EFFECTS BY THE COMBINED ACTION OF ERGOTOXINE AND COCAINE. By I. DE BURGH DALY, P. FOGGIE,1 and G. VON LUDAiNY. From the Physiology Department, University of Edinburgh. (Received for publication 29th October 1936.) DALE and Dixon [1909] have shown that the actions of adrenaline and tyramine are similar in many respects, although the potency of adrenaline is many times that of tyramine, if the pressor substance is taken as the criterion of comparison. They found that tyramine while exerting its main action peripherally also possesses some nicotine-like action. A fundamental distinction between the actions of adrenaline and tyramine has been described by Tainter and Chang [1927] since cocaine which potentiates the action of adrenaline [Froblich and Loewi, 1910] depresses the action of tyramine on the anasthetised cat. This phenomenon has been referred to as the "cocaine paradox" [Burn and Tainter, 1931]. Burn [1930] in an exhaustive analysis of the effect of adrenaline and of tyramine on perfused organs made the important discovery that while the presence of adrenaline greatly enhances the tyramine pressor effect, cocaine still diminishes or abolishes it. The same investigator in collaboration with Tainter [1931] showed that the enhancement of adrenaline action by cocaine on the isolated heart only occurs when there is a sudden application of the cocaine dose, a steady concentration of cocaine diminishing the adrenaline response. Our own interest in the actions of cocaine and tyramine lay in the hope that they might be useful as diagnostic agents to determine the nature of pulmonary vasomotor nerves. Functionally active pulmonary vasoconstrictor nerves in isolated perfused lungs have been demonstrated by Daly and Euler [1932], and since it was apparent that some of these fibres were adrenergic [Dale, 1933] in character, it became desirable to examine the effects of their stimulation in cocainised preparations. Furthermore, Burn and Tainter [1931] and Burn [1933] had expressed the view that the enhancement of tyramine pressor effects by the presence of adrenaline in the circulating blood of perfused 1 Nee Alcock.

2 236 Daly, Foggie, and Luda'ny organs depended upon the fact that tyramine stimulated sympathetic nerve endings and released the transmitter which then exerted its action on the cell. In perfused organs the transmitter store was considered to be exhausted so that tyramine had little or no effect. When, however, the infusion of adrenaline replenished the store, tyramine administration was then able to release it and produce a pressor effect. This view and the work of Frohlich and Loewi [1910] suggested that isolated perfused lungs into which adrenaline was being steadily infused would be highly responsive to sympathetic nerve stimulation following an injection of cocaine, but irresponsive to stimulation if cocaine was withheld and the transmitter store exhausted by the previous administration of tyramine. As a preliminary to testing this hypothesis, we investigated the actions of adrenaline and tyramine on perfused lungs in the presence of cocaine and ergotoxine; we also tested the effect of adrenaline infusions on the tyramine response. The results of this research are now presented because we have encountered a curious phenomenon relating to the action of tyramine. In order to clarify the account of the experiments this phenomenon will be described forthwith. It has been found that the rise in pulmonary arterial pressure which tyramine always produces in isolated perfused lungs is not significantly altered by a previous injection of cocaine alone or of ergotoxine alone. If, however, cocaine is added to the circulation of ergotoxinised preparations, the pulmonary arterial pressure rise to a subsequent dose of tyramine is markedly potentiated. This combined action of cocaine and ergotoxine in potentiating the pressor action of tyramine is not confined to the pulmonary vascular system, because a similar phenomenon has been observed on the contractile response of the non-pregnant guinea-pig uterus. ISOLATED PERFUSED LUNGS. The isolated lung preparations of the dog were perfused through the pulmonary artery only at constant inflow or constant pressure under negative pressure ventilation by the methods described by Alcock, Berry, Daly, and Narayana [1936], the venous outflow and blood volume changes of the lungs being measured by a differential method [Daly, 1928]. Most of the animals were bled under local anaesthesia and the lungs perfused with the animal's own blood. The total amount of blood in circulation was c.c. The drugs used throughout the investigation were crystalline adrenaline (P. D. & Co.), cocaine hydrochloride (B. D. H.), ergotoxine ethane sulphonate (B. D. H.), ergamine acid phosphate (B. W. & Co.), nicotine (B. D. H.), and tyramine hydrochloride (B. W. & Co.). The doses are given in terms of the base in each case. Most of the experiments

3 Potentiation of Histamine and Tyramine Effects 237 to be described relate to the effect of one drug upon the subsequent response of another. For this purpose it is necessary to maintain e(}ual intervals of time between successive injections of the drug the response of which is being tested. The procedure is illustrated by the tvramine tests in fig. 5., where in each experiment after a preliminary trial of the dosage required for a suitable response, the injection intervals are kept the same. A;drenaline.-The rise in pulmonary arterial pressure which is pro(luced by single injections of adrenaline (1.0 y and upwards) is Fic.. 1.-Dog 3-0 kg. 19/6/33. I.P.L. Constant inflow~. T.A. =tidal air; TP.A.p. pulmonary arterial pressure; VYR.=venous reservoir volume. Time 10 sec. Effects of adirenaline (cryst.) 4-0 BiIetween 'a" and " b" cocaine (300;y) xwas inijected into the P.A. I.P.L. =isolated perfuised lungs. An -upwar(1 nioxveiiienit of the VYR. tracing (lenotes anl increase in outflow. poteutiated by a previous injection of cocaine (0-3 to 30() mug.). In sonle experiments not only the height of the pressure response was augmented but also the (Iuration. In 7 experiments, 5Showed a lpotentiation effect, I a slight reduction in the pressor response and I 111 change. Fig. I illustrates the most miarked p~otentiation obtained. The increase iu venous outflown whiich adlrenaline generally produces reimains approximately time same after cocaine. Inm ergotoxinised p)reparations, it has already been show-n that adrenaline causes a fall in puilmonary- arterial pressure [Daly andl Euler. 1932], and ax e rmoxx find that, the pressuire fall takes place after fuill (loses of atropine. We have also examinedl the action of adrenaline

4 238 Daly, Foggie, and Luda'ny upon the venous outflow after ergotoxine. Our previous experience that adrenaline generally augments the venous outflow [Alcock, Berry, and Daly, 1935] in normal preparations is confirmed. In 98 further tests on 15 lung preparations, an increase in venous outflow always occurred. In ergotoxinised lungs, adrenaline produced no change in outflow in 25 tests, a diminution in 7 and an increase in 2. Thus ergotoxine may suppress or reverse the usual adrenaline effect on the outflow. Cocaine produced no significant alteration in the adrenaline depressor response of ergotoxinised preparations. Owing to the rapidly declining effect of successive doses of adrenaline it was difficult to make a comparison before and after cocaine, but by plotting against time the fall in pressure due to successive doses of adrenaline and injecting cocaine between two adrenaline doses, it was possible to show that cocaine produced no action on the subsequent expected adrenaline response. Thus cocaine potentiates the pulmonary arterial pressor action of adrenaline, but in ergotoxinised preparations has no modifying effects on the adrenaline depressor response. Tyramine.-The effects of tyramine are remarkably constant from one preparation to another. At constant pressure perfusion tyramine (1-10 mg.) injected into the pulmonary artery causes a diminution in inflow and an augmentation in outflow. At constant inflow perfusion the pulmonary arterial pressure rises and the outflow increases (fig. 2). These effects occurred in nicotinised, ergotoxinised or atropinised lungs (figs. 4 and 5). In 12 experiments the tyramine response was noted before and after the injection of cocaine (0.3 to 30 0 mg.). The normal response to successive doses of tyramine falls off until, as a rule, the second and third injections produce the same quantitative effect (fig. 3). After a constant response has been obtained, the cocaine is added to the venous reservoir, and the effect of tyramine again observed after a period of minutes. The result appears to have no relation to the strength of cocaine dosage, since we have observed a potentiation as well as a depression of the tyramine effect with 30 mg. of cocaine. None of the changes observed was very large and no obvious influence on the venous outflow took place. In 4 experiments the effect on the tyramine response of ergotoxine (2-3.5 mg.) was tested. With the exception of one experiment in which a slight potentiation was obtained (fig. 3 c, d) ergotoxine did not affect the tyramine response. It appears therefore that cocaine or ergotoxine have no constant effect on the action of tyramine on the pulmonary vascular bed. The bronchi for the most part remained unaffected by tyramine in the cocainised or ergotoxinised preparations, except that a slight tyramine broncho-

5 1 I=_ Potentiation of Histamine and Tyrainine Effects.2.39 dilatation occurred in 3 out of 9 ergotoxinised preparations. The two tracings (figs. 2, a, b and 3, c, d) illustrate the maximal extent to which potentiation of the tyraminine response occurred after cocaine alone or A. Fig. 2, a, b. Dog 7-5 kg. 5/7/35. I.P.L. Constant inflow. Effects of tyrainine 2-0 wig. Between "a" and "b" cocaine (300 y) was injected. FiG. 2, c, d. Dog 16-0 kg. 18/7/35. I.P.L. Constant inflow. Effects of 3-0 mg. tyramine. Cocaine (30 0 mg.) injected between "c" and "d" pq- L( V L LL FiG. 3. Dog 7-5 kg. 12/7/35. I.P.L. Constant inflow. At signals marked T, 1-0 mg. tyrarnine injected. At E, ergotoxine (2-0 mg.). after ergotoxine alone. It is to be noted that these effects are excep tional, but the tracings are given for comparison with fig. 4, which illustrates quite a different phenomenon, namely, the marked degree of potentiation of the tyramine response by cocaine in ergotoxinised preparations.

6 240 Daly, Foggie, and Ludany In 13 ergotoxinised preparations cocaine ( mg.) potentiated the pressor effect of tyramine on the pulmonary vascular bed (figs. 4 and a, Expts. 1 and 3), but had no certain effect on the increase in blood outflow which tyramine always produces (fig. 5). One experiment only showed no such enhancement, and in this the dose of cocaine was 05 mg. It will be seen from fig. 4 that the response to 10 mg. of tyramine which in the ergotoxinised preparation is just effective on the pulmonary arterial pressure-the rise is 3*0 mm. blood is increased after 30 mg. cocaine to approximately 20 times this value. tta. 10 FiG. 4.-Dog 1335 kg. 26/7/35. Constant inflow. Ergotoxinised and atropinised (2-0 Ing.) I.P.L. 1st signal, tyramine,.30 mg.; 2nd and 3rd signals, tyramine, 10 0 mg. Between "a" and "b", cocaine, 30 mng. In 3 experiments the tyramine response was enhanced slightly by cocaine (5 mg.) and ergotoxine (2 mg.) given simultaneously. Onthe other hand ergotoxine had no enhancing effect on the tyramine response in cocainised preparations. How far these differences are to be ascribed to fixation of the drugs in the tissues or to their changing concentration in the cell, we have not attempted to investigate. We are reminded. however, of the work of Burn and Tainter [1931] who observed a depression of adrenaline action on the cat's perfused heart by cocaine in steady concentration, but an adrenaline enhancement by cocaine in changing concentrations. The experimental conditions were not favourable in order to discover how far the ratio between cocaine and tyramine dosage determined the suppression or potentiation of the tyramnine effect by cocaine. Tile animals from which the preparations were made varied widely in weight and in sensitivity to tyramine, and the doses used were not calculated in respect of body-weight but rather to give a pulmonary arterial pressure rise of from 1-3 cm. of blood.

7 Potentiation of Histamine and Tyramine Effects 241 Taking all the experiments together, the only significant effect observed was the potentiation of the tyramine response by cocaine over a wide range of doses in the preparations which had been ergotoxinised. Adrenaline and Tyramine.-Single injections of adrenaline have no certain effect on the subsequent tyramine response, but in confirmation of Burn we find that an adrenaline solution steadily infused into the FIG. 5.-Pulmonary arterial pressure changes plotted from portions of continuous records of three isolated perfused lung preparations. Each ordinate division represents a pressure change of 2 cm. of blood. A=adrenaline; C= cocaine; E = ergotoxine; N = nicotine; T = tyramine; Inf. = continuous infusion of drug. The numerals adjacent to the letters denote single injection dose in mg. In Experiments 1 and 3 the constant infusions were started 30 min. before points indicated by arrows at right angles. In Experiments 1 and 2, A = 20 y/min.; in 3, A = 15 y/min. + N = 1-7 mg./min. circulation (15-25 y/min.) may cause a marked enhancement of the tyramine response (fig. 5, Expt. 2). This effect, although not invariable, requires min. adrenaline infusion before it manifests itself, and we have yet to discover the correct infusion dose to obtain the tyramine enhancement in every experiment. The pressor response to tyramine generally falls off as the experiment progresses, although in a few experiments it was well maintained up to one hour. We have never observed a significant potentiation of the response due to successive doses of tyramine alone. In Expt. 1 of fig. 5, 1 0 mg. of tyramine injected before the adrenaline infusion was started gave a pressure

8 242 Daly, Foggie, and Ludainy rise of 05 cm., and 10 0 mg. was required to give a similar rise after 60 min. of adrenaline infusion. We do not know whether this loss of sensitivity to tyramine was due to the adrenaline infusion or was an unusually rapid natural decline, but it is remarkable that the potentiation of tyramine following ergotoxine and cocaine was greater than that generally observed. The ratio of adrenaline to tyramine dosage to produce the same pulmonary arterial pressure rise depends upon a variety of factors. In perfusion experiments lasting over several hours, adrenaline injections given at 1-1-hour intervals produce approximately the same FIG. 6. Dog 18-0 kg. Constant inflow perfusion. I.P.L. a =adrenaline (cryst.), 10 y; b = tyramine, 1-0 mg.; c =adrenaline, 100 y; d =tyramine, 5 0 mg.; e =adrenaline, 1-0 mg.; f=tyramine, 5 0 mg.; g =adrenaline, 10-0 mg.; h =tyramine, 10-0 mg. The injections at a, b, c, d, e, f, g, and h were made 10, 20, 65, 75, 125, 135, 175, and 185 min. respectively after commencement of perfusion. Time marker = 30 sec. response. If, however, repeated doses of adrenaline are given at shorter intervals (10-20 min.), a condition of adrenaline insensitivity develops. Repeated doses of tyramine at short intervals do not affect very greatly the subsequent responses to tyramine. When tyramine and adrenaline are injected alternately at short intervals of time, the adrenaline response diminishes much more rapidly than the tyramine, so that finally equal doses of adrenaline and of tyramine give approximately similar pressor responses (fig. 6). Such results are difficult to correlate with the suggestion of Burn [1933] that tyramine acts by liberating adrenaline from the store at the nerve ending, a point which will be discussed later. The fact that the tyramine response remains after the adrenaline pressor response has practically disappeared rather suggests that the tyramine effect is on the muscle cell which has become insensitive to adrenaline. Moreover, tyramine and adrenaline, as has been shown, both increase the pulmonary arterial pressure or diminish the inflow of blood, yet after ergotoxine the qualitative action of tyramine

9 Potentiation of Histamine and Tyramine Effects 243 remains the same, whereas that of adrenaline is reversed (fig. 7). The method used in the experiment illustrated by this figure is that described on pp in the paper by Alcock, Berry, and( Daly [1935]. The tracing sho-ws that tyramnine diminishes the inflow, wnhereas adrenaline increases it. The corresponding changes in outflow are for the most part due to the effects upon the inflow. In assessing the value of this argument it shoull be pointed out that more recent work by Bilbring and Burn [1936] has led them to conclude that adrenaline action hasm no necessary relation to the innervation of the tissue. A crucial experimnent, to perform w-ould be one to determine the effect of FiG. 7. Dog 16-0 kg. Constant pressure perfusion. I.P.L. Ergotoxinised preparation. Top tr. =venouis outflow, a downward movement indicates diimninution in outflow. Second tr. inflow, an upward movement denotes a (liminut ion in inflow. Time = 15 sec. For explanation see text. sympathetic nerve stimulation after the preparation has become insensitive to adrenaline. The blood-vessels in perfused lung preparations usually lose their sensitivity to nerve stimulation before that to. adrenaline and the opportunity for making this test has not yet arisen. In view of the close similarity between histamine and tyramline actions [Burn and TUainter, 1931], we tested the effects of cocaine anud of ergotoxine on the histamine response of the pulmonary vascular bed, also of cocaine on the histamine response in ergotoxinised preparations. The results were not consistent and the synergistic action of cocaine and ergotoxine in potentiating the histainine response was not observed (4 expts.). ISOLATED GUINEA-PIG UTERUS. The uterus taken from young non-pregnant animals was iiunnediately set up in a bath of 20-0 c.c. kept at C. and containing

10 244 Daly, Foggie, and Ludany Ringer-Locke of the following composition: NaCl, 8 5 g.: KCl, 0 2 g.; CaCl2 (cryst.) 0 4 g.; MgCl2 0.1 g.; NaHCO3 0 5 g.; NaH2PO2 0 0"5 g.; distilled H20 to 1000 c.c. Air was bubbled through the solution and the ph was approximately 7*6. The results to be described were obtained over a range of ph 7*4 to 8*4, and were more easily reproduced if adrenaline (1 0 y) were added to the bath before testing the response to tyramine. The presence of adrenaline, however, was not essential. although it was added to the bath in the majority of experiments in order to inhibit the spontaneous contractions. Histamine and Tyramine. Since both histamine and tyramine influenced the ergotoxinised or cocainised uterus in a similar manner, Fie. 8.-Isolatted noni-pregniant guinea-pig uterus. The numbers of the curves indicate the order in which the tests were made. T3= tyramine, 3 mg.; T5 tyramine, 5 mg.; C =cocaine, S- mg.; IE= ergotoxine, 01 mg. The fluid in the bath was changed between each test. their actions may be considered together. Both ergotoxine alone and -cocaine alone slightly potentiated the contraction response of histamine and of tyramine in some preparations, but not in all. In 1 out of 9 experim-ents cocaine caused an enormous enhancement of the tyramine contraction. Moreover, a subliminal (lose of tyramnine would sometimes become effective wh~len repeated after the addition of cocaine to the bath. In all ergotoximiised preparations cocaine greatly enhanced the histamine and the tyramnine responses. These effects are illustrated by figs The most marked results AA-ere obtained wnith preparations made from very young animals,. The absence of contraction of the guinea-pig uterus following the addition of ergotoxine to the bath fluid is not, a unique experience, although -unusual. Dale and Spiro [1922] obtained contraction of p~reparationls made from young animals wiith doses of 1 part in 125 millions, whereas Tate and Clark [1922] observed no action or else relaxation of fresh preparations both wnith ergot oxine and with tyramnine.

11 Potentiation of Histamine and Tyramine Effects Thompson [1929] found ergotamine to contract the uterus of the virgin guinea-pig, but to have no action on the uterus removed from mature 245 FIG. 9.-Isolated non-pregnant guinea-pig uterus. A= adrenaline (1-0 y). Other drugs as for experiment in fig. 8. E..E I FIG. 10.-Isolated non-pregnant guinea-pig uterus. A =adrenaline, 1 0 y; H =Histamine, 0-25 y in tracing 1, and 05 yin tracing 2. E =ergotoxine, 0-1 mg.; C=cocaine, 0.1 mg. parous animals. The animals from which our own preparations were made were taken at random from stock, and we have made no special investigation of the conditions which lead to irresponsiveness of the uterus to ergotoxine. CAT'S NICTITATING MEMBRANE. The animals were prepared according to the method of Rosenblueth [1932] and anoesthetised with intraperitoneal injections of c.c. of dial (Ciba) per kg. body-weight. In the majority of experiments both cervical vago-sympathetic nerves were cut and both suprarenal glands clamped off from the circulation. A lever magnifying fifteen times and loaded with a 10 g. weight was employed. All drugs were injected into the right femoral vein, a period of 5 seconds being adopted as the standard time for each injection. Tyramine.-We confirm other workers in finding that tyramine contracts the nictitating membrane [Bacq and Lefebvre, 1935] and ergotoxine, which itself produces a prolonged contraction of the membrane [Rosenblueth, 1932], suppresses the response of subsequent doses of tyramine (fig. 11). In confirmation of Bacq [1934] we find

12 246 Daly, Foggie, and Ludany that cocaine also diminishes the contraction response to subsequent doses of tyramine. The injection of cocaine in ergotoxinised animals, or of ergotoxine in cocainised animals has no effect in potentiating the action of tyra- WEm -_-U FIG. 11. Cat 2-6 kg. Dial 0 7 c.c./kg. body-wt. Both cervical sympathetic and vagi cut; both suprarenal glands tied off. Top tracing =B.P.; 2nd =nictitating membrane response (mag.=15); 3rd 30 sec. int.; A=effect of 0 75 mg. tyramine. Between A and B, 0-5 mg. ergotoxine injected. B =tyramine, 0-75 mg. 8' after ergotoxine injection. Between B and C, cocaine 10 0 mg. injected. C =tyramine, 0-75 mg. D =tyramine, 4-0 mg. FIc. 12. Cat 2-4 kg. Dial 0-7 c.c./kg. body-wt. Both cervical sympathetic and vagi cut; both suprarenal glands tied off. A, B and C =effects of histamine, 200 y; between A and B, ergotoxine, 0-5 mg.; between B and C, cocaine, 10 0 mg. mine on the inembrane; on the contrary, a definite diminution in the tyramine response occurred (fig. 11). In 6 experiments we failed to encounter the phenomenon described by Bacq and Lefebvre [1935] that tyramine in 30 per cent. of experiments sensitises the membrane to its own action.

13 Potentiation of Histamine and Tyramine Effects 247 Histamine.-The nictitating membrane is relatively insensitive to histamine, intravenous injections of 200 y being necessary to produce a significant response. In two experiments we obtained a slight relaxation; in 3, a well-marked contraction. In those experiments in which histamine contracted the membrane, ergotoxine had the effect of reversing the histamine response. If histamine initially relaxed the membrane, then after ergotoxine the membrane relaxed more fully to histamine or the response remained unchanged. In ergotoxinised animals, cocaine did not influence the relaxation response of the membrane to histamine. These results are illustrated in fig. 12. Preparations of isolated guinea-pig's gut showed no potentiation of the tyramine or histamine response by cocaine after ergotoxinisation. DISCUSSION. The results presented throw some light upon the action of tyramine on the pulmonary blood-vessels. A dilator action on the systemic circulation has been described by Handovsky and Pick [1913], Amsler and Pick [1920], and Burn [1930], but since we have never seen a fall in pulmonary arterial pressure following pulmonary intra-arterial tyramine injections we will confine the discussion solely to its pressor action. That the nicotine-like action of tyramine [Dale and Dixon, 1909] has little if any responsibility for raising the pulmonary arterial pressure by stimulation of the pulmonary nerve ganglia is evident from the experiments in which the tyramine response remained after large doses of nicotine were injected or continuously infused. The potentiation of tyramine by cocaine in ergotoxinised preparations was also observed after nicotinisation. As regards the suggestion put forward by Burn [1933] that tyramine acts by stimulation of sympathetic nerve endings with liberation of the neurohumoral transmitter, certain of our observations appear to militate against this view. If we assume, with Burn, that the transmitter is adrenaline, then injections of adrenaline should produce at least as large a pressor response as that following quantitatively similar doses of tyramine. It has already been noted that doses of tyramine may yield a larger pressor response than quantitatively similar doses of adrenaline; indeed, the tyramine effect may occur in a preparation insensitive to adrenaline. If intra-arterial injections of adrenaline are assumed to have the same effect on the cells as adrenaline liberated by stimulation of sympathetic nerve endings, it is reasonable to infer from our results that tyramine does not produce its action wholly by stimulating the nerve endings. It is true that we have insufficient knowledge as to the precise mode of action of adrenaline when liberated by nerve stimulation to assume categorically that it is similar to that produced by adrenaline injections, even so it seems to us that the argument as

14 248 Daly, Foggie, and Ludany presented somewhat weakens the hypothesis that tyramine liberates adrenaline. A further difficulty in accepting this hypothesis arises from the experiments with ergotoxinised preparations. Whether we visualise a paralytic action of ergotoxine on the sympathetic nerve endings [Dale, 1913] or a direct action on the cell [Navratil, 1927; Bauer, 1928], on Burn's hypothesis the pressor effect of tyramine should be suppressed in ergotoxinised preparations. For, if tyramine acts wholly by stimulating the nerve endings, the paralysing action of ergotoxine should cause a failure of subsequent doses of tyramine to release adrenaline, or alternatively, the action of ergotoxine on the cell should suppress the action on the cell of the released adrenaline. We find, however, that doses of ergotoxine, sufficient to reverse the pressor effect of adrenaline have little or no effect on the tyramine pressor response. In view of the fact that Burn and Tainter [1931] have reported a suppression of tyramine effects by ergotoxine in the perfused hind legs of the dog and that ergotoxine, as we have shown here, does not suppress the tyramine contractile response of the guinea-pig uterus or of the lungs, the question arises as to whether the discrepancies in the reactions of these preparations are to be attributed to differences in the selective site of action of tyramine, that is on the nerve endings or on the cell, or to differences in the qualitative response of the specific tissue cells to tyramine. Although we are unable to answer this question, our results suggest that the use of tyramine as an agent for producing an exhaustion of the sympathetic transmitter store is unreliable; indeed, as far as the pulmonary vascular bed responses are concerned we are inclined to interpret our results as indicating that tyramine exerts its action on the muscle cells alone, a view in agreement with that of Tainter [1926] and one which we will develop in discussing the significance of the potentiation of the tyramine response by cocaine in ergotoxinised preparations. We are unable to put forward any hypothesis based upon the recognised actions of histamine, tyramine, cocaine and ergotoxine to account for the cocaine potentiation of the tyramine and of the histamine responses which occurs in some ergotoxinised tissues but not in others. Such evidence as we have presented, coupled with the knowledge accumulated by other investigators of the individual actions of these drugs, suggests there are more grounds for provisionally interpreting the observed phenomena in terms of an altered responsiveness of the muscle cell itself due to a direct action of the drugs on the cell, than for considering it to be the result of two distinct mechanisms-the release of the sympathetic transmitter by tyramine or histamine which then exerts an action on the cell under the influence of ergotoxine and cocaine. In ergotoxinised lung preparations cocaine potentiates the tyramine

15 Potentiation of Histamine and Tyramine Effects 249 response but not the histamine; in the isolated non-pregnant ergotoxinised guinea-pig uterus cocaine potentiates both the tyramine and histamine response, whereas in the isolated guinea-pig gut and cat's nictitating membrane no potentiation of histamine or of tyramine effects by cocaine after ergotoxinisation is observed. Unless the preparations are ergotoxinised the potentiation of tyramine or of histamine effects by cocaine does not take place except in rare cases (guineapig's uterus). Such variations in reactivity might be due to fundamental differences in the innervation of the tissues tested, but this appears to us to be improbable because the ergotoxinised pulmonary blood-vessels which exhibit a potentiated tyramine response after cocaine, and the nictitating membrane which does not, both contract to adrenergic fibre stimulation. Moreover, the pulmonary vessels respond by contraction to cholinergic fibre stimulation [Daly and Ludainy, 1936], and there is some evidence that cholinergic fibres play some part in causing contraction of the nictitating membrane [Bacq and Fredericq, 1934 a]. Thus, two tissues which exhibit similar responses to nerve stimulation behave differently as regards the potentiation of tyramine effects by cocaine. The available evidence indicates that cocaine in sensitising to sympathetic nerve stimulation or to adrenaline acts directly on the muscle [Rosenblueth, 1932; Rosenblueth and Cannon, 1932; Bacq and Fredericq, 1934 b], and that ergotoxine produces its adrenaline reversal effect also by virtue of its action on the muscle [Rosenblueth, Leese and Lambert, 1933]. It is also significant that adrenaline may simulate sympathetic nerve stimulation effects on cardiac tissue in the absence of sympathetic innervation [Matsumori, 1929; Markowitz, 1931]. It is not improbable therefore that the adrenaline pulmonary pressor potentiation by cocaine is due to changes in the response of the smooth muscle of the blood-vessels, as also the reversal of the adrenaline pulmonary pressor effect by ergotoxine. If tyramine is also exerting its action directly on the muscle of the pulmonary vascular bed, a reasonable hypothesis in view of our experiments, then the ergotoxinecocaine-tyramine phenomenon might be due to changes in the response of the muscle alone. Until a similar phenomenon has been observed with denervated tissues the participation of the nerve endings cannot be ruled out. The similarity in some respects between the actions of adrenaline, tyramine and histamine has been pointed out by Burn and Tainter, and it is interesting to note that the enhancement of histamine and of tyramine actions by cocaine needs the presence of ergotoxine, whereas the enhancement of adrenaline by cocaine only takes place in the absence of ergotoxine. There is one other point to which we wish to draw attention. Burn [1930] has stressed the importance of tyramine concentration in VOL. XXVI., NO

16 2 50 Daly, Foggie, and Ludany comparing its action on one tissue with another. The results on isolated perfused lungs have been obtained with a wider range of doses of tyramine and cocaine than have been used for the experiments on the isolated guinea-pig's uterus and intestine or for the cat's nictitating membrane. Notwithstanding, there appears to be a close resemblance between the responses of the ergotoxinised lungs and ergotoxinised uterus to tyramine after the exhibition of cocaine. The isolated gut of the guinea-pig and the cat's nictitating membrane appear to be quite different in this respect a difference so marked that we feel the discrepancies cannot be due to a failure on our part to select the correct dosage of the drugs favourable for the appearance of the potentiation effects described. SUMMARY. In isolated perfused lungs- 1. Cocaine enhances to a small extent the pressor action of adrenaline, but has no constant action on the pressor action of tyramine. 2. Adrenaline steadily infused into the circulation for half an hour is sometimes effective in potentiating the pressor action of tyramine. 3. The pressor response to tyramine may be present at a time when quantitatively similar doses of adrenaline are ineffective. 4. Ergotoxine reverses or suppresses the pressor action of adrenaline, but has no significant effect on that of tyramine. 5. In ergotoxinised preparations cocaine has no action on the adrenaline depressor response, but produces a marked potentiation of the tyramine pressor response. 6. The histamine pressor response is not potentiated by cocaine in ergotoxinised preparations. 7. Reasons are given for the opinion that tyramine exerts its action directly on the muscle of the pulmonary vascular bed. In the ergotoxinised non-pregnant guinea-pig uterus, cocaine enhances the contractions produced by tyramine or by histamine. No such enhancement takes place in the ergotoxinised guinea-pig intestine or ergotoxinised cat's nictitating membrane. The expenses of this research have been defrayed in part by a grant to one of us (I. de B. D.) from the Government Grant Committee of the Royal Society, to whom we express our thanks.

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