i 4-3' ‘p—J-r' '7“ v." I: "~ SOME REWIM’TQRY $11 MULANTS EN SEVERAL COMBINATION!» IN DEEFLY BARE‘ITALIZED was flaws for flu bag!” ed M. S. MlfiHSGAN STATE UNIVERSITY 3M Naraén Géri WM 6:7 hols .‘J‘ I \j (f'r‘\}.\ * v _ . | . ' .' ~£:u‘ruuV' ' V \ «1“ ”I ‘ OVERDUE FINES: 25¢ per day per item RETURNING LIBRARY MATERIALS: Place in book return to remove charge from circuhtion records SOME RESPIRATORY STIMULANTS IN SEVERAL COMBINATIONS IN DEEPLY BARBITALIZED DOGS BY SHRI NARAIN GIRI A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Physiology and Pharmacology 1961 ,6%«{77fl" Respectfully dedicated to my father Prasidh Narain Giri. ii iii ACKNOWLEDGMENT I wish to express my sincerest gratitude and appreciation to Dbctor Clyde F. Cairy, Professor of Pharmacology and Physiology for the help and encouragement manifested in numerous ways, for the excellent spirit of guidance and the invaluable aid which helped me at every stage in the planning and preparation of this manuscript. I am sincerely grateful to Dr. B. V. Alfredson, Professor and Head of the Department of Pharmacology and Physiology, for his interest and sympathetic attitude. I am deeply indebted to Doctor William D. Collings, Pro- fessor of Physiology and Pharmacology, and Doctor P. O. Fromm, Associate Professor of Physiology and Pharmacology for their valuable suggestions and encouragement. Thanks are due to my sincere colleagues Jack R. Hoffert and Robert Shone for their delightful and helping nature and also to others who contributed in making my stay in this uni- versity a pleasant experience. INTRODUCTION LITERATURE REVIEW Amphetamine Metaraminol Bitartrate Methetharimide Pentylenetetrazol MATERIAL AND METHODS . RESULTS . . DISCUSSION . Pentylenetetrazol-Amphetamine Combination Metaraminol Bitartrate-Amphetamine Combination Methetharimide-Amphetamine Combination SUMMARY . . BIBLIOGRAPHY TABLE OF CONTENTS iv Page 16 23 3O 37 41 60 6O 63 65 68 7O INTRODUCTI ON In 1954, 391 people committed suicide in England and wales by taking an overdose of barbiturate--eight times as many as in 1945. There was no corresponding increase in the total number of suicides. So, it is clear that the barbi- turates are increasingly preferred as a means of self- destruction and that the treatment of intoxication by these agents has consequently become an important medical problem. Thus, the increasing popularity of the barbiturate as a suicidal agent has created a worlddwide problem. Over the past decade the amount of barbiturate used on both sides of the Atlantic has more than trebled, while the incidence of barbiturate coma has increased fivefold (Nilsson, 1951; Locket and Angus, 1952; Clemmensen, 1954; Goldstein, 1947; Koppanyi and Fazekas, 1950, 1952, 1954; Moller, 1954; Alwall andILunderquist, 1951, 1953; Goodman and Gilman, 1947). The widespread use of many barbituric acid derivatives has focused increasing attention upon drugs and other measures which may counteract the effects of an overdose with these agents. This problem has been particularly acute in the field of veterinary medicine, where certain barbiturates, notably pentobarbital, are so commonly employed as hypnotics, narcotics, and anesthetics. The more extensively a drug is used for these purposes, the more are the chances of accidents by its delib- erate or accidental improper use and overdose. There is a lack of unanimity on recommendation for the most effective treatment of barbiturate poisoning. The tradi- tional use of pharmacological analeptics in treating patients depressed to the point of coma has been more often employed to counteract the depressant effect. But in the absence of adequate information concerning the mode of action of central analeptics, some dangers are involved in their use (Nilsson, 1951; Locket and Angus, 1952; Clemmensen, 1954). Some workers do not favor their use on the grounds that the convulsions they produce may cause irreparable damage to the brain through anoxia by increasing the central oxygen demand in excess of the available oxygen. Others who stand in their favor condemn this objection (Miller and Miller, 1956) by the reasoning that analeptics by their awakening effects and respiratory and cardiovascular stimulation potentially can save the patients from barbiturate poisoning. They account for the inability to awaken the patient fully by suggesting a concomitant cerebral hypoxia and disordered cerebral metabolism resulting from the prolonged action of the barbiturate (Bailey £3 11., 1953; Bain, 1952, Brody and Bain, 1954) and naturally, time must be allowed for restitution to normal. 3 However, the analeptic therapy in barbiturate poisoning is accompanied by the following advantages: 1. It removes the immediate risk to the patient's life and prevents the onset of complications often associated with prolonged barbiturate coma. Prolonged endotracheal intubation is not needed. It is valuable from the viewpoint of hospital economy in that it affords relief from prolonged and strict nursing. Analeptic therapy in intoxication other than barbiturate provides a differential diagnosis in planning further therapy for the patient. With the idea that two analeptics when used together may enhance the therapeutic value by synergising each other, the author was led to determine the efficiency of analeptics on respiration of several combinations of pentylenetetrazol and amphetamine, methetharimide and amphetamine, metaraminol bitartrate and amphetamine. Amphetamine and metaraminol were chosen because they were found to be useful in a previous study (Sisodia, 1960). CHAPTER 1 LITERATURE REVIEW Amphetamine (Benzedrine, Amfetasul) ‘ChemiCally Racemic B phenyl isopropyl amine or a-phenyl B-amino propane NHz CH - CH - CH Amphetamine is a sympathomimetic amine which exists in three isomeric forms: levo, dextro and d1 or racemic form. The levo isomer is the least potent of all the isomers. The dextro form is the most potent and commonly known as dextro- amphetamineiwhereas the racemic form is commonly known as amphetamine. Pharmacological Action Although amphetamine is a synthetic sympathomimetic amine related more or less to epinephrine and ephedrine, it differs from them in the following properties, viz: l. Cocaine synergises the action of epinephrine by inacti- vating the amine oxidase and phenol oxidase which destroy epinephrine; but cocaine shows no synergism with amphetamine and ergotamine shows no reversal effect with amphetamine. 2. Amphetamine is effective orally as it cannot be destroyed by enzymatic action in the gastrointestinal tract. Amphetamine is a more powerful stimulant to the central nervous system than any other sympathomimetic amine. It stimu- lates the cerebro-spinal axis, especially the brain stem and the cortex. It stimulates the respiratory and other medullary centers effectively. It compares well with all other drugs in analeptic effectiveness and finds considerable use in counteracting overdepression caused by anesthetics, narcotics, hypnotics, etc. Animals receiving sufficiently large amounts of amphetamine exhibit tremors, restlessness and increased motor activity. This is due to the cortical stimulation by the drug, but they may also result in part, from excitation of the brain stem. It does not produce seizures or subcon- vulsive dysrhythmia in a normal animal. Indeed, the drug can obtund the maximal electroshock seizure and prolong the re- covery period after such seizures. These properties may be related to the usefulness of amphetamine in certain cases of epilepsy. Amphetamine is a strong inhibitor of amine oxidase and according to Goddum and Kwaitkowski (1938), it would act by preventing the oxidation of epinephrine by amine oxidase and by competing with epinephrine for the receptor substance in the effector cells. However, many unsolved problems prevent the full acceptance of this theory and further investigation is needed to elucidate the mechanism of the sympathomimetic action of emphetamine on the CNS. Amphetamine produces an "arousal reaction" in animals under anesthesia and as an analeptic finds wide use in abolishing or shortening the duration or decreasing the intensity of anesthesia. The psychic effects of amphetamine have been studied in great detail in man; the response elicited depends upon the mental state and dose administered. The main results seen are wakefulness, alertness, increased initiative and elevation of mood, enhanced confidence, euphoria, lessened sense of fatigue, increased vasomotor and speech activity and increased ability to concentrate. It is generally agreed that amphetamine inhibits production of fatigue. Diminished sense of fatigue is purely subjective and central in origin. Amphetamine does not enable subjects doing rapidly exhausting work to perform longer or to recover more quickly. It is said to inhibit pro- duction of fatigue, particularly in monotonous skilled tasks and somewhat to restore performance in fatigued individuals. The wakeful psychologic effects are related to some unknown control stimulation (Myerson, 1940). Amphetamine by its central action exerts a direct anal- gesic effect. Amphetamine has been found to enhance and pro- long the analgesic action of morphine in man while it decreases the drowsiness, dizziness, and weakness caused by morphine. However, amphetamine largely eliminates the analgesic action of nitrous oxide. Amphetamine facilitates monosynaptic and polysynaptic transmission in the spinal cord and enhances decrebrate rigidity. It improves reflex activity and recovery response even in the decrebrate and decorticate animals. This may be due to an increased internuncial activity which compensates for the loss of facilitation from centers destroyed. Respiratornyffects Amphetamine affects respiration in two ways? by stimula- ting the medullary respiratory center and by dilating bronchi- oles. (The latter effect is not so marked.) The effect is weaker but much more prolonged than with epinephrine (Alles and Prinzmel gt _1., 1933). The respiration is first depressed (mostly in amplitude), probably reflexly, with the rise of blood pressure; then it soon comes to the preinjection level and then is further stimulated with a marked rise in both rate and amplitude to increase the ventilation considerably (Alles, 1933). Detrick, Millikan, Modern, and Thienes (1937) observed that with the first dose of benzedrine (0.25 - 4 mg/kg) in dogs and cats anesthetized with nembutal, there was a marked increase in rate and depth of respiration. Subsequent doses produce successively smaller increases and finally often a decrease in rate. The actual mechanism for stimulation of respiration is not determined. As the anesthetized animal shows the symptoms of central stimulation, it may be inferred that the mechanism is central in origin. Due to stimulation of the medullary respiratory center both rate and depth of respiration are increased. This effect is marked when the respiratory center has been depressed chemically but not so marked in normal animals. However, amphetamine has no ability to increase the respiration of a brain inhibited by anesthetics or to increase oxygen consumption of normal brain tissues. The stimulant action of amphetamine on a normal or anbsthetized brain is not yet clear and it is doubtful whether its peripheral sympathomimetic action can be profitably correlated at present with its excitatory effects on the CNS. Repeated injections of amphetamine may cause depression of respiration. Hyperventilation is reported as another risk associated with the use of these amines. Small doses should be preferred since they are said to relax the bronchial muscle and slow and deepen the respiration. Bronchodilator action however, is weak. Cardiovascufilar Action Cardiovascular action of amphetamine is comparatively weak, as are all its sympathomimetic actions (Hahn, 1960). Neverthe— less, the peripheral action may represent an undesirable com- plication if the central action of high doses is required. Cardiovascular responses of animal to amphetamine are charac- terized by considerable variability. Hewever, adequate doses usually cause a rise in both systolic and diastolic pressures and an increase in cardiac output and work. The effects are apparently accomplished by a direct myocardial action and by peripheral constriction of the arterioles (Goodman and Gilman, 1955). Therapeutic doses of amphetamine in normal Subjects do not increase cardiac output, pulmonary circulation time, vital capacity, BMR, and respiratory dynamics were not changed (Altschule and Iglauer, 1940; Goodman and Gilman” 1955). Amphetamine given causes a rise in mean blood pressure. This rise in blood pressure was accompanied by a decrease in cerebral blood flow and cerebral oxygen utilization. Amphe- tamine causes vasoconstriction centrally by stimulating the medullary vasoconstrictors center and locally by stimulating the sympathetic receptive substance in the muscle cell of the 10 blood vessel. Amphetamine applied locally to mucous membranes causes vasoconstriction and shrinkage of congested tissue by the same mechanism. Amphetamine sulphate has no marked effect on cardiac muscle. The electrocardiogram is not greatly altered in the vast majority of cases (Meyerson, 1940). In several instances a transitory reflex slowing of pulse occurred in man at the onset of the rise of arterial pressure. In some cases a transitory slight increase in cardiac output was also detected (Altschule and Iglauer, 1940). The rise in blood pressure in man is marked and lasts for 1-2 hours. The effects tend to lessen and disappear after the drug is used over an extended period (Meyerson, 1940). Amphetamine in ordinary clinical doses, 5 - 10 mg in man has no significant effect on the cardiovascular dynamics (Altschule and Iglauer, 1940). Arrhythmias of different types, palpitation and pre— cordial pain may occasionally occur after amphetamine in the normal individual as well as in patients with heart disease. Detrick, Millikan, Modern and Thienes (1937) found an irregular effect of amphetamine on the blood pressure in dogs. Blood pressure varied over a wide range from decrease to an increase in normal pressure of the blood. They also noticed that ergotamine tartrate when used before could increase, decrease, and even abolish the effects of the following amphe- ll tamine and that cocaine decreased the pressor effect of amphetamine. Addiction to Amphetamine There appears to be very little danger of serious habitua- tion with this drug. Although addiction to amphetamine occurs it is probably infrequent. However, as reported (Meyerson, 1940), the drug is neither habit-forming nor does it have any untoward symptoms. Connel (1950) states that addiction causes "delusion of persecutions and auditory and visual halucinations indistinguishable from those occurring in paranoid schizophrenia." A characteristic abstinence syndrome does not develop when amphetamine is abruptly withdrawn but depression, tremors, weakness and gastrointestinal symptoms may have been observed in some individuals. Prolonged use of amphetamine in orthostatic hypotension to maintain blood pressure in normal limits leads to insomnia which is rather difficult to overcome even by full doses of barbiturate and lasts for 24 - 72 hours (Korns and Randall, 1930). Fate and Excretion According to Goodman and Gilman 50 percent of amphetamine is destroyed in the body by deamination and the rest is excreted unchanged in the urine. Amphetamine resists oxidative 12 deamination by amine oxidase which accounts in part for its long duration of action. It is a potent inhibitor of amine oxidase. However, other enzymes e.g. phenol oxidase destroy it. Less than 50 percent of amphetamine is excreted in 48 hours following ingestion (Beyer and Skinner, 1940). In man the percentage excreted of a given dose generally paralleled the volume output of urine. With smaller doses percentages excreted were greater. In order to account for the fate of the remainder of the drug dose experiments were conducted to show that: l. Probably all the drug is absorbed from the gastrointes- tinal tract. 2. Hydrolysis of urine does not result in greater yield of amine. 3. Amine oxidase does not activate deamination of amphetamine. The drug is apparently slowly and partially inactivated in the body by a loose combination with some agents normally contained therein. Whether the drug is then partially destroyed or excreted slowly over a period of several days either free or so loosely combined as though not apparently conjugated is yet a problem.. 13 Barbiturate-Amphetamine Antagoni§_m_ The injection of amphetamine uniformly causes a marked increase in blood pressure reaching a maximum within a few minutes after injection. This may be due to peripheral vaso- constriction (Sollmann, 1948). Alles (1933) noticed that amphetamine when given iv "excited a considerable effect in waking the animal from barbital anesthesia." In man Meyerson §t_§1, (1936) reported that benzedrine sulphate subcutaneously could not affect the depth although it definitely shortened the duration of nembutal narcosis and stated that hypertension produced by benzedrine sulphate subcutaneously could be reduced by soluble amytal intravenously and also the hypotensions produced by soluble amytal intravenously could be elevated by benzedrine sulphate subcutaneously. Reifenstein and Davidoff (1930) within an hour aroused ten patients from deep narcosis of soluble amytal after 1, 2 or 3 injections of benzedrine sulphate (10 mg) in each indi- vidual. Fourteen barbiturate poisoned subjects were treated by Frereich and Landsberg (1946) with intravenous injection of amphetamine. Thirteen patients recovered without any ill effects except for some headache. The one who died was probably due to lack of drug. 14 Lee and Alfredson (1952) observed that a dose level of 2.5 mg of amphetamine per kg would be ample to combat the depressant effect of large doses of the barbiturate on blood pressure. Although the pressure dropped slowly after the maximum was reached it was still above any previous level in all subjects at the termination of experiments one-half to one hour following injection. They also observed that deep pentobarbital depression raised the threshold of respiratory response to sciatic stimulation enormously. After amphetamine injection the threshold was decreased to almost its previous level. The central depressant effects of pentobarbital on respiration have been reported. The marked improvement in the respira- tory response of these dogs to sciatic stimulation follow- ing amphetamine would then indicate that both of these drugs acted on the central respiratory mechanism. Amphetamine has been found to possess central nervous stimulation. However, Sollmann has reported that some stimulation in respiration from acute amphetamine injection was by way of the carotid sinus reflex. There is a reciprocal relationship of amphetamine sulphate to barbiturate which is generally referred to as "reciprocal pharmacology." If one desires to obtain a sedative effect without the ill effect of hangover and depression, then the addition of small doses of amphetamine sulphate will be of considerable value. 15 l6 Metaraminol-Bitartrate (Aramine) Chemically 1-(m-hydroxyphenyl) 2-amino l-propano1 gfhydrogentartrate or a (1 amino ethyl) m hydroxybenzyl alcohol hydrogen d- tartrate 0H CH - CH - NH2 O\\ $ ?H 1’0 | | c - c - c - c / I l I OH CH3 Ho H H OH Pharmacological Action Aramine is a synthetic sympathomimetic amine used as vasopressor. It has found extensive use to combat the hypo- tension of spinal analgesia and its effects compared with those of methoxamine and ephedrine. Since metaraminol has peripheral arteriolar action, so much of the studies have been centered around its effect on the renal vasculature as well as on cardiac hemodynamics of normal subjects. 0n Blood Pressure The cardiovascular response to metaraminol has been extensively studied in dogs by Sarnoff and Associates. In normal animals aramine administration intravenously resulted in an elevation of the mean arterial blood pressure with an 17 associated bradycardia (Mayerand Beazley, 1955) . Metaraminol bitartrate with a potent vasopressor effect elevates both systolic and diastolic pressure due to increased blood pressure. The cerebral, renal and coronary blood flow also improve (N. N. R., 1959). However, as the blood pressure increased there was a parallel increase in peripheral resis- tance without a significant effect on cardiac output (Mayer and Beazley, 1955). Increased pulse pressure was the usual finding, and it does not produce a primary or secondary fall in blood pressure under experimental conditions studied so far (Poe, 1954). The pharmacological property of aramine appears to be a safe vasopressor action employed both prophylactically and therapeutically to combat the hypotensions commonly associated with spinal anesthesia. No complications attributable to the drug were noted after administration (Poe, 1954). However, metaraminol,primarily a vasopressor, does not include anorexi- genic or cardioaccelerator effects (weil, 1957). Aramine as a potent vasopressor has the advantage of t 31. (1955) reported prolonged duration of action. Mayer that in hypotension metaraminol bitartrate increases cerebral blood flow and relieves hypoxia, if any. But on the other hand, in normal dogs aramine decreases cerebral blood flow by causing hypotension. 18 Aramine is usually effective in restoring and maintaining blood pressure in clinical shock due to numerous causes. It appears to have some advantage over norepinephrine in that its action is not so evanescent as norepinephrine and thereby permits maintenance of a more stable blood pressure. It is non-irritant to tissue and is effective by oral as well as parenteral routes. The drug seems to be a valuable agent against hypotension, but treatment of a patient suffering from chronic orthostatic hypotension of the ideopathic type and postural hypotension related to neurogenic disturbance was of less value (weil, 1957). It was reported that because of a more gradual onset and prolonged duration of action, maintenance of blood pressure with metaraminol bitartrate is generally more smooth and is subject to fewer of the abrupt variations and excessive responses sometimes observed with other pressor agents (Council on Drugs, 1957). Although on local application of Aramine to the nasal mucosa vasoconstriction is prolonged but not to that extent to cause endothelial anoxia. It does not cause secondary vaso- dilation. On the Heart The total effect of Aramine is one of general vasoconstric- tion with little or no change in cardiac output (Livesay t 31., 19 1954). The administration of Aramine to the vagotomized dog was followed by an increase in cardiac output, left main coronary artery flow and aortic and pulmonary artery pressure. Right and left arterial pressure fell. Larger doses of Ara- mine had little effect further on cardiac output though it increased the blood pressure by its vasopressor effect (Mayer and Beazley, 1955 and Sarnoff _§ al., 1954). Unlike most pressor amines Aramine slows the heart rate. This effect appears to be predominantly a reflex response to the increase in blood pressure (Poe, 1954). It has no toxic effect on the heart. On the other hand according to New and Non-official Drugs (1959), "Aramine exerts a moderately positive inotropic effect on the heart and does not appear to provoke cardiac arrhythmias in the sensitized myocardium." weil is of the opinion (1955) that metaraminol bitartrate increases coronary circulation and has no specific effects on cardiac output. Sarnoff _t_§l, (1954) observed the following effect of Aramine in hemodynamics: 1. In the normal anesthetized dog meteraminol produces a slight to moderate elevation of arterial pressure and bradycardia. When the bradycardia is abolished, either after vagotomy or during hemorrhagic hypotension, metar- aminol produces marked elevation of arterial pressure. 20 2. In small doses, metaraminol produces a striking improve- ment in the ventricular function. Subsequent doses do not further improve myocardial contractibility but do increase peripheral resistance and tone. 3. The myocardium does not require a greater coronary flow per unit of work after the administration of metaraminol, nor did it produce significant arrhythmia in the presence of severe myocardial hypoxia. weil (1955) showed that bradycardia prominent in normotensive volunteers was unknown in hypotensive patients when treated with metar- aminol. Mechanism of Action Metaraminol bitartrate constricts the peripheral vascular beds, increases the venous return and coronary blood flow, and acts directly on the heart muscle to increase its contractility (weil, 1957). Aramine tends to increase the blood pressure in hypotensive and normotensive as well as hypertensive animals by its pressure and cardiac effects, but in normotensive ani- mals the blood pressure rise to hypertensive levels is opposed by a reflex vagus stimulation. Atropine can block the vagal effect without affecting the pressure effect. In the vagotomized animal, the blood pressure does go to the anticipated hyper- tensive levels and there is no bradycardia or any blood vascular reflex effect (Sarnoff t al,, 1954). 21 Metaraminol bitartrate is long acting, presumably because of its structural insusceptibility to the action of phenol and amine oxidases (weil, 1955). On CNS (Poe, 1954) Metaraminol has little or no stimulant action on the CNS. It is said to produce (N. N. R., 1959) less stimulation than ephedrine to patients with shock. After administration of Aramine, cerebral blood flow is increased Mayer and Beazley, 1955) as manifested by improvement in the state of consciousness. Mayer 2E.§l- (1955) also reported the same thing that in hypotension metaraminol increases cerebral blood flow and relieves hypoxia and so tends to improve the state of consciousness. 0n the Respiratory_System Livesay §£_al, (1940) showed an increase in respiratory rate, but this was inconstant. However, several individuals showed an increase in respiratory rate as blood pressure was elevated. Several of the patients complained of a sense of constriction in the chest without pain. The pulmonary artery pressure increased. The arteriovenous oxygen difference in- creased from 4.5 volumes percent to 5.0 volumes percent. Total oxygen consumption was found increased. 22 In normotensive volunteers metaraminol bitartrate caused subjective shortness of breath and tachypnea. 0n the Kidney Moyer and associates (1954) demonstrated that dogs pre- viously rendered hypotensive by phlebotomy showed improved renal function when the blood pressure was restored to normo- tensive levels. This was manifested by increased urinary output, glomerular filtration rate and renal blood flow. However, in the normal subject metaraminol causes renal vasoconstriction andfreduction in renal blood flow and glo- merular filtration rate (Livesay gt 21., 1954). This is suggestive evidence that the renal hemodynamics during the administration of vasopressor agents will be different in patients with shock and other hypotensive states as compared to normal subjects with normal blood pressure (Livesay _E‘al., 1954). 23 Methetharimide (Mikedimide, Megimide) Chemically B,B-methyl ethyl glutarimide or 3,3-methyl ethyl glutarimide CH3 \JC/CHZ - CO CH CH / \CH co 3 2 2 ', NIH \/ Pharmacological Action Regarding the compound methetharimide Shulman.and.his assoCiates (1955) fee1_there may be direct antagonism to the barbiturate itself rather than a physiological antagonism.’ Banica and Wilson (1950) and Shaw _E_§1; (1954) are othhe opinions that 3,3-methyl ethyl glutarimide is.a CNS stimulant in both barbiturized and normal animals. The clinical impres- sion remains that methetharimide provides the most effective means of reversing barbiturate anesthesia. Full animal investigation (Shaw and Bentley, 1952; Shaw _tfa1., 1954) has indicated that methetharimide possesses a high therapeutic index. 3,3-methyl ethyl glutarimide, a barbiturate antagonist, seems to be a respiratory stimulant. It greatly reduces the risk of toxic manifestation and in therapeutic doses appears to cause a slight rise in blood pressure, but a large dose (75 mg/kg) given intravenously to a barbiturized patient has 24 produced a large rise in blood pressure and sweating, sug- gesting a direct effect on the autonomic ganglion (Shulman gt 1., 1955) . Excretion Much of the 3,3-methyl ethyl glutarimide is excreted in t al,, 1955). the urine unchanged (Shulman, Methetharimide and Barbiturate Antagonism Shulman ;§._1. (1955) emphasized the value of 3,3-methyl ethyl glutarimide, not only as a barbiturate antagonist but also as a potent, non-specific respiratory stimulant. It appears to possess a specific respiratory stimulation effect only in the barbitalized animals and routinely its use is suggested as the effective agent to terminate barbiturate anesthesia, shorten the sleeping time, restore reflex activity and minimize the need for prolonged medical attention. The drug appears to be a specific barbiturate antagonist on almost a milligram for milligram basis (Baker and Englewood, 1956). Oriordan and Breward (1958) compared the effectiveness of methetharimide with piCrotoxin, nikethamide and a combination of methetharimide and deplazole for their analeptic properties on 55 patients divided into five groups. Each patient has been anesthetized with thiopentanone for minor procedures and 25 found the most promising results with methetharimide and its combination with deplazole. According to the finding in the more complicated field of barbiturate intoxication, Oriordan and Breward (1950) held the view of Nilsson (Nilsson, 1951) that this condition is analogous to one of deep and prolonged anesthesia and that its morbidity and mortality are due largely to the complication of such anesthesia. They said "Whatever the exact pharmacologic action of methetharimide will prove to be, its specificity for the barbiturateseries is marked." In support of this theory, they further found the drug to be of little value as an analeptic when the main anesthetic agente wemacyclopropane, trichlorethylene, vi or ethylether The use of methetharimide and deplazole recommended by Shulman and associates would seem to be the most effective way of overcoming barbiturate intoxication and provides the follow- ing noteworthy advantages: 1. It removes the necessity for prolonged intubation. 2. It minimizes the immediate risk to the patient's life and the remote risk which may follow the onset of com- plication, often accompanying prolonged barbiturate coma. 3. It is valuable from the hospital's economical point of View and it avoids prolonged and strict nursing. 4. It may serve to distinguish barbiturate from other types of poisoning and differentiate the coma from other causes. 26 Boyan _t-_l. (1958) found by several controlled cross over experiments that methetharimide is 1.73 times as potent as metrazol in reversing the electroencephalographic effects of thiopental in man. Mechanism of Antagonism Methetharimide bears a definite resemblance in its chemi- cal structure to the barbiturate ring system; that its mode of action is one of competitive inhibition, however, would now seem doubtful (Oriordan and Breward, 1958) and that methethari- mide, like the barbiturates, depresses the respiration (Jalling, 1956). Therapeutic dose may exert a direct antag- onism to the barbiturate (Shulman £2 31., 1955) as indicated by the fact that the rabbits can be put to sleep or awakened almost at will by alternate intravenous administration of thiopentanone sodium and with methetharimide. Present evi- dence suggests that methetharimide CNP13 along with diamino— phenylthiazole is the best substance yet used in the treatment of barbiturate coma, which insures a quick safe recovery without the risk of convulsion and secondary depression which often follow treatment with central analeptics (Shulman gt__1., 1955). Shulman _t.al. (1955) observed that "if after the patient has been brought to the 'safe state,‘ his condition regresses, further, small treatment may be given as required. Regression 27 is more likely to occur when the coma has lasted a long time before treatment is started or if the barbiturate concerned is a long acting one (e.g. phenobarbitone)." This suggests that methetharimide does not directly chemically antagonize the barbiturate, as then this regression would not occur. The work of Boyan gt a1. (1957) attempts to indicate a new explanation for the antagonism. A patient under deep narcosis of barbiturate for 57 hours was given 5500 mgms. of methetharimide. The patient was Clinically improved without any sign of overdose of this drug. The electroencephalogra- phic pattern changed from a deeper to a lighter level of anesthesia and when the electroencephalogram stopped improving despite continued treatment with methetharimide it was con- cluded that the drug had achieved its maximum therapeutic value at this time, and its administration was discontinued. The drug antagonized the circulatory and respiratory depres- sion successfully. A transient increase of oxybarbiturate level of the plasma was found after methetharimide administration, though the electroencephalograph level was improving towards a lighter level of anesthesia. This was explained by Butler and walell; that undissociated barbiturate is absorbed into the brain while dissociated barbiturate remains largely in the plasma. Thus there exists an equilibrium between the 28 plasma and the brain barbiturate concentration. When plasma pH is increased dissociation of barbiturate is also increased. This shifts the equilibrium in such a way that the plasma concentration of barbiturate is increased, whereas the brain concentration of barbiturate is decreased. This may explain why the patient was more responsive even though the oxybar- biturate level of the blood had increased. J. Pederson (1956) was of the opinion that the drug neither shortens coma nor hastens the rate of elimination of barbiturate from the blood. "The experimental and clinical data" from the laboratory, as observed by Plum and Swanson (1957), indicate that respon- siveness to 3,3-methyl ethyl glutarimide as a stimulant is inversely related to the depth of narcosis. Plum and Swanson (1957) expressed their opinions in their experiment, most of which do not stand in favor of the drug. 3,3-methyl ethyl glutarimide showed little evidence of true barbiturate antagonism, but appeared to possess non-specific analeptic properties. The analeptic effects were more rapid in onset than other analeptics. There were fewer undesirable side reactions. 3,3-methyl ethyl glutarimide, however, failed to stimulate spontaneous breathing in an animal receiving more than 1.4 times the lethal dose in untreated animals. The pressor 29 response of the drug was more reliable in lightly anesthe- tized animals than in deeply comatose animals. The properties of CNS stimulation appeared similar to, but less intense than, those of picrotoxin. There was little convincing evidence that 3,3-methyl ethyl glutarimide shortened coma, although muscle spasms were pro- duced with ease as was accentuation of stretch reflexes. Megimide and Mikedimide are preparations of methetharimide hence, chemically the same differing in their solvents and concentrations. Megimide is a 0.5 percent solution in water and Mikedimide is 3 percent in propyleneglycol of 3,3-methyl ethyl glutarimide. The author used only Mikedimide. The way of excretion of propyleneglycol is the same as that of 3,3 methyl ethyl glutarimide, and in larger doses it has a CNS depressant action. However, propyleneglycol seems to be devoid of any demonstrable toxicity when administered in smaller doses. So it is hard to say what effect, if any, propyleneglycol will have on the effect of 3,3 methyl ethyl glutarimide in such small doses. 30 Pentylenetetrazol (Metrazol, Cardiazol) Chemically HZC -- CH2 -- C§2\ ‘>N - N -- H -- C/J Hzc C 2 \ | N — N Pharmacological Action Pentylenetetrazol is very specifically a central nervous system stimulant. Its actions are highly complex and apparent- ly occur at all levels of the cerebrospinal axis. Applied locally to sensory neurons, it causes motor activity reflexly. Applied to particular motor neurons, it causes responses in muscles supplied by these nerves. This drug causes generalized clonic convulsions when given parenterally. It'can stimulate both the motor as well as sensory apparatus of the nervous system in the body. The medulla is also stimulated by pentylenetetrazol to increase ventilation. The analeptic action of pentylene- tetrazol on the higher centers of the CNS is evident from its ability to counteract the depression produced by barbiturates: tribromoethanol, paraldehyde, chloralhydrate etc. Its medullary stimulation effect is a direct one, for typical effects on blood pressure and respiration can be obtained even after the carotid 31 sinus and carotid body have been completely denervated. It produces significant stimulation in normal animals only with doses approaching the convulsive doses, but in anesthetized animals the subconvulsive therapeutic doses are quite effec- tive. The effect of pentylenetetrazol on the medulla is more prominent when the region is depressed than when it is func- tioning. In cases of acute depression of barbiturates, pentylene- tetrazol not only restores motor activity but failing respira- tion and circulation are stimulated through the medullary action of the drug. Circulatory effects of pentylenetetrazol are due entirely to its action on the medulla. When circula- tion has been depressed by a hypnotic, a marked rise in blood pressure can be noticed after the injection oprentylene- tetrazol as a result of central vasomotor stimulation. How- ever, when the drug is given to a normal individual or to a patient whose circulation is depressed by other than a central mechanism no such responses are obtained. The pressure may fall due to the bradycardia as a result of central vagal stimulation. Sometimes the vagal effect is very prominent and this causes bradycardia and hypotension. Reflex activity of the spinal cord is increased by pentylene- tetrazol. This action on the spinal cord is especially evident if the reflex activity of the cord has been depressed chemically. 32 Koll (1937) observed that maximum reflex activity of the spinal cord obtained by pentylenetetrazol could be further enhanced by strychnine and vice versa. This indicates that both drugs act on different centers in the brain. Pentylenetetrazol exerts characteristic effects on the electroencephalogram. With large doses the drug causes marked restlessness, excitement, increased motor activity and clonic convulsion. It is observed that the convulsive dose is increased at the lower level of transection of the brain. Character of the convulsions indicate their origin in the midbrain. Direct injection of pentylenetetrazol into the hypothalamus causes autonomic stimulation, cyclodilation, piloerection, urination, defecation, marked emotional excite- ment, and increased response to faradic stimulation of the region (Masserman, 1938). Pentylenetetrazol convulsions result from the summation of a greater number of stimuli acting locally in the brain and spinal cord. Thus pentylene- tetrazol convulsions are accompanied by central excitation of both divisions of the autonomic nervous system, especially the sympathetic and this occurs also if the convulsions are prevented by curare (Gellhorn and Darrow, 1939). Convulsive responses of the animal are decreased by starvation, by calcium deficiency, by dehydration, and by repeated injections of metrazol. However, as reported by 33 Kaslein (1937), convulsive response is increased by water retention. During the time of convulsions brain metabolism is diminished by anoxia (Himwich gt_§1,, 1939) and the content of lactic acid thereby increased (Stone, 1938). Depressing Effect of Pentylenetetrazol In addition to the stimulant action of pentylenetetrazol, there occurs depression following the convulsions. With pen- tylenetetrazol no depression was observed with any subconvul- sive dose. With convulsive dose depression was present but remained shorter than with picrotoxin (Dille and Hazelton, 1939). There is a critical level with pentylenetetrazol when used as an analeptic, above which there is prolongation of recovery of the cortical placement reaction, but decrease in the recovery of the righting reflex. Dose levels immediately below this decrease the recovery of both these responses, but still lower doses prolong their recovery (Dille and Hazelton, 1939). Pentylenetetrazol, besides producing central rhythmic discharge and convulsions, also stimulates the neuromuscular junction (Eyzaguirre and Lilienthal, 1949). Pentylenetetrazol has negligible action on the heart and blood vessels. It has no definite cardiac effect nor does it improve the coronary circulation to a significant degree. It has no constrictor 34 action on the musculature of blood vessels. It may cause splanchnic and cerebral vasodilation. Due to skeletal muscle contraction at the time of a convulsion, arterial pressure may be considerably raised. Conduction disturbances e.g. sinus arrhythmia, alteration of the impulses between S.A. and A.V. nodes, and rarely heart block occurs, but these have been attributed to the central effects of the drug on autonomic nuclei. Fate and Excretion (Goodman and Gilman, 1955) Pentylenetetrazol is rapidly absorbed from the sites of administration and stored temporarily in the liver as an unidentified compound. Esplin gt a1. (1954) say 75 percent of the absorbed drug is excreted in the urine in unchanged form. An entirely different view is held by Tatum, Kozelka and Nelson (1938), that pentylenetetrazol after intravenous injec- tion, leaves the blood rapidly and maintains equal concentra- tion in muscle, brain, liver and blood. A very small quantity is excreted with the urine and the bile and only a part can be demonstrated in the feces of guinea pigs by bioassay but not chemically (Hinsberg, 1939). 35 PentylenetetrazolgBarbiturate Antagonism Pentylenetetrazol is one of the most potent, effective analeptics in saving the lives of acutely barbiturized dogs and rabbits. Besides the medullary actions, which are largely responsible for the life-prolonging or life-saving action, this has a denarcotizing effect, which is due to cortical t 21,, 1936), a "definite respiratory stimulation (Koppanyi stimulant effect" (Alfredson, 1941). Pentylenetetrazol does not remove pentobarbital from the blood. There are three suggested mechanisms by which pentylene- tetrazol may exert its protective effects against barbiturates: 1. Physiological antagonism. This phenomenon is not fully understood yet. However, Torda (1954) is of the opinion that pentylenetetrazol increases the concentration of acetycholine in the brain, thereby inducing central stim- ulation. There is no doubt about the antagonism of pentylene- tetrazol against barbiturate. 2. Induction of vomiting. Pentylenetetrazol in convulsive doses acts as a stimulant to the vomiting center. Schwartz (1920) proved that pentylenetetrazol denarcotizes the emetic mechanism depressed by barbiturate. This denar- cotizing effect further emphasizes the physiological antagonism. In combination with barbiturate, pentylene- tetrazol can induce vomiting as long as seven hours and 36 so, in such cases, acts as a long acting drug. Fezekas and Koppanyi (1954) showed that metrazol may act even up to 24 hours after oral administration. Delayed absorption. Emptying time of the stomach is decreased by 20 percent by sodium amytal (Vanliere and Northrup, 1941). These barbiturates are quickly absorbed from the intestine, where pentylenetetrazol counteracts this effect and tends to slow down passage to the intes- tine. It has been observed that barbiturates administered along with pentylenetetrazol attain much lower level in the blood than by the same doses of barbiturates if administered alone. Moreover, barbiturate is less effec- tive in the presence of pentylenetetrazol. 37 CHAPTER II MATERIALS AND METHODS All the dogs used in this project were obtained from a city dog pound. These animals were not allowed to live to- gether but were kept in individual cages and maintained on a commercial dog food and water ad libitum. As large num- bers of dogs were required in this project, no discrimina- tion was made between the weights of dogs. Dogs weighing as low as 7 kg and as high as 21 kg were used. Every effort was made not to use any dog operated or anesthetized for any other experimental work in at least the preceding seven days. Breed type was not at all taken into consideration. Three percent sodium pentobarbital solution in ten per- cent ethyl alcohol was employed to anesthetize the dogs. Amphetamine sulphate (Amfetasul) was a five percent aqueous solution (Pitman-Moore Co., Indianapolis, Ind.). Methetharimide was a three percent solution in propylene- glycol (Mikedimide, Parlam Corporation, Englewood, N. J.). Metaraminol bitartrate was an aqueous solution of one _percent (Aramine, Merck, Sharp, and Dohme, Philadelphia). Pentylenetetrazol was a ten percent aqueous solution (Metrazol, Knoll Pharmaceutical Co., Orange, N. J.). 38 Experimental Procedures In the beginning the dog was weighed and sodium pento- barbital was injected intravenously--40 mg/kg body weight to induce deep anesthesia and to slow down the respiratory rate markedly. A rubber tracheal tube with an inflation cuff around its distal end was then passed into the trachea through the mouth. The cuff was then inflated from the outside. The degree of inflation was indicated by a pilot balloon. This procedure made the tracheal tube fit snugly into the trachea and thus allowed the air to pass freely into the trachea and out only through the tracheal tube. The latter was connected to a meter to record the respiratory volume in liters and respiratory rate. A wet test gas meter connected with tracheal tube was employed to measure the expired air to find out the value for respiratory volume. The gas meter was so constructed as to record each 250 cc of air by means of an electrically operated signal magnet. A small tube from the air valves was attached to a tambour which had one electrode attached to its diaphragm and the other just above it, so that with each expiration and inspiration, there was an inflation and collapse and so a make and a break of the circuit to another signal magnet took place. A third signal magnet was used as a timer for every five seconds. All the signal magnets were attached to ink writing 39 points in order to write on a white kymograph paper belt. Now minute respiratory volume was observed for at least 10 minutes. In some of the dogs it was reduced to 0.5 liters per minute and in others to 2.75. In the latter case addi- tional sodium pentobarbital, 5 mg/kg body weight was injected intravenously to cut down the respiratory volume to 1.5 liters per minute. The plan was to inject the analeptic when respiratory volumes per minute were 1.5 liters or below, but sometimes even additional doses failed to reduce the respira- tory volume to 1.5 liters per minute. (This was observed mostly in large sized dogs but small sized dogs were not an exception to this.) After reducing the respiratory minute volume to approxi— mately 1.5 liters per minute, the analeptic combination was administered intravenously and its antagonistic effects were observed at least for 60 to 70 minutes. Analeptic combinations in part were tried at various dose levels. Combinations were prepared by taking 3/4 of the therapeutic doses of two analeptics or 3/4 of one analaptic and 1/2 of the other analeptic, and vice versa constituted the third dose level combination. In most of the cases both the analeptics were mixed to- gether in the syringe and injected intravenously. Only Mikedimide, due to its solvent propyleneglycol, could not be 40 mixed with aqueous solutions of other analeptics and so one was injected after the other by the same needle, intravenously. Mikedimide was injected after the first analeptic was in. In this way all the possible nine combinations from four analeptics were tried. Care was taken strictly to try each combination on at least six dogs. The intravenous injections were confined to the radial vein. Quite often ventilation of the anesthetized dog was raised in the process of manipulation and needle insertion, evenbefore the analeptics combination was injected. This effect is accounted for by the reflex induced by mechanical I stimulation at the site of injection. 41 CHAPTER III RESULTS All the nine combinations used in the project and three taken by courtesy from Dr. Sisodia's work have presented six different degrees of response on respiration as depicted below in the following descending order of efficiency, and also shown in Figures 5, 10 and 15. 1. Methetharimide 3/4, Amphetamine 1/2 2. Metaraminol bitartrate 3/4, Amphetamine 1/2 3. Metaraminol bitartrate 3/4, Amphetamine 3/4 4. Methetharimide 1/2, Amphetamine 3/4 5. Methetharimide 3/4, Amphetamine 3/4 Methetharimide 1/2, Amphetamine 1/2 Pentylenetetrazol 3/4, Amphetamine 3/4 Metaraminol bitartrate 1/2, Amphetamine 1/2 6. Metaraminol bitartrate 1/2, Amphetamine 3/4 Pentylenetetrazol 1/2, Amphetamine 1/2 Pentylenetetrazol 3/4, Amphetamine 1/2 Pentylenetetrazol 1/2, Amphetamine 3/4 Three-fourths therapeutic dose of methetharimide and 1/2 of amphetamine made the best combination in all of the series to arouse the barbitalized dog. Methetharimide seemed to exert a direct antagonistic action against barbiturate 42 anesthesia as evidenced by quite frequent return of paw pinch, corneal, palpebral and cough reflexes. The superior awakening property of methetharimide was further evidenced by fast heart beat, increase in respiratory rate, violent running movements, struggling, resistance to intubation, extreme stage of excitement, excessive salivation, and vomiting almost in every case--sometimes in the early part of the experiment and sometimes in the later part. Vomiting was invariably observed in the early part of the experiment when methetharimide was coupled in 3/4 therapeutic dose. This suggests that methetharimide stimulates the vomiting center directly in larger doses. Dogs receiving methetharimide combinations exhibited opisthotonus and.whining even after the termination of the experiment. The best combination, methetharimide 3/4 and Amphetamine 1/2, showed a very favorable response on ventilation. In the beginning minutes respiratory volume reached the peak; later, there was some regression followed by a gradual uniform increase. Metaraminol 3/4 and Amphetamine 1/2 made the next best combination. In the beginning little improvement in ventila- tion was seen but later it increased smoothly and was main- tained at a respectable level for a considerable period of time. 43 Pentylenetetrazol and Amphetamine, coupled with each other at any dose level other than 3/4 of each made the poorest com- bination and maintained the respiration at a very poor level. Abbreviations in the Graphs A Amphetamine P Pentobarbital Ar Artificial respiration ( )+ Arousal after analeptics and no more ventilation record due to struggling and excitement ( )- Died Single arrow with no sign on it indicates that two analeptics were mixed together in the same syringe and injec- ted at the same time; otherwise, arrows indicating other injections bear abbreviation for the drug injected. VENTILATION PER MINUTE IN LITERS Fig. l - 7-5 mg/kg )- - 1 dog - 18 kg _ k P - 40 mg/kg _ .1- T ' ~ 3 dog - 12.5 kg t P - 40 mg/kg L. ' 5 dog - 9.25 kg ' p - 40 mg/kg P “NJ/v - e r - - (Average of 6 dogs) ' T 60 min. Percent Increase 300% 100% 44 PENTYLENETETRAZOL - AMPHETAMINE 3 mgdkg 8 6- /' 4.- r— 2.. 2 dog - 7.75 kg ' P - 40 mg/kg 0L..- 1 1 I 111 r-c 6r ’ 4 dog - 13.75 kg 41~ P - 40 mg/kg O [J l n L 4 l l _1 6!- 4 - 6 dog - 15 kg _ ' P - 40 mg/kg 2 .. “M 0 L I g I l l 1 l V 200% ‘ 0% t F I I 1 l J l J_ 0 20 40 60 min. VENTILATION PER MINUTE IN LITERS Fig. 2 7.5 mg/kg 1 dog - 21.5 kg P - 40 mg/kg ‘ 1‘ 3 dog - 11.25 kg ’ P - 40 mg/kg 5 dog - 17 kg P - 45 mg/kg (Average of 6 dogs) Fig. Percent Increase 45 PENTYLENETETRAZOL - AMPHETAMIN 2 mg/kg 6 r— 2 dog - 16 kg ‘ P - 40 mg/kg 0 ’ 1 L 1 1 1 J 4 - 4 dog - 9 kg - ‘ P - 40 mg/kg 6 dog - 18.25 kg P - 40 mg/kg 300%? 200% I 100% 0% 1% -10 0 20 40 60 min. A“! VENTILATION PER MINUTE IN LITERS 46 Fig. 3 PENTYLENETETRAZOL - AMPHETAMINE - 5 mg/kg % m9/k9 4 — 4 L 1 dog - 11.5 kg ~ _ P — 41.75 mg/kg 2 - 2 k 2 dog - 13 kg \CJ " _ P — 42.7 mg/kg O LIIIIIII 0 llllllll 6 f 6 F ' “ 4 dog - 9 kg 4 ” 4 ~ P - 42.7 mg/kg 2 2 _ 0 0 LIIIIIII 6f 6r— r 5 dog - 10.5 kg . 4 F P - 40 mg/kg 4 _ - L. 2 ”‘Jr\"f/J\\—_—_- 2 ' 6 dog - 8 kg ' ' P - 40 mg/kg O I I | | | ' ' ' O I l l I I I 1 1 6~ $200% — m - g " 4 _ (Average for 6 days) 2100% - H n 2 o 2 ‘ m 0A " FA” . - a; r O I 1 I 1 | I J ‘ Q4 4 L l J | l l L J -10 0 20 40 60 min. 0 20 40 60 min. Taken by courtesy from Dr. Sisodia VENTILATION PER MINUTE IN LITERS 47 Fig. 4 PENTYLENETETRAZOL - AMPHETAMINE 5 mg/kg 3 mg/kg 4 L 4 L I. b 2 - 2 .\£\———\_/ 1 dog - 9.5 kg - 2 dog - 9 kg 0 h k . P - 40 mg/kg 0.- P - 40 mg/kg 4%- 4- 4 dog - 8.5 kg ' P - 40 mg/kg 2 - 3 dog - 11 kg 2 - _ P - 40 mg/kg - [\——— 4— 9. 0 11411411 0 jJJJlllI 8 F L e» 6- + _ 6 dog - 8.75 kg P - 40 mg/kg 4 r 4 _ 2. 2). 5 dog - 9.25 kg .4 e.’/”” Fk P-'-40mg/kg hi 0 1Lg11111 0 1111.444 (Average for 6 dogs) N l T 1 P.N’]\\ Percent Increase H c> o c: X. 32 I 1 I 0 -10() 20 40 60 min. -10 0 20 40 60 min. 48 Fig. 5 AVERAGE PERCENT INCREASE IN VENTILATION AT FOUR DIFFERENT DOSE LEVELS OF PENTYLENETETRAZOL AND AMPHETAMINE Percent Increase 400% ' 300%- 200%- 0 0 ° 0 0 O u 0 Q 0 (’ 100% 0 o :”,, V”/, 0%fl o 10 20 30 4o 50 60 Minutes 0—0—o Pentylenetetrazol 3 /4 Amphetamine 3 /4 “”9 Pentylenetetrazol 1/2 Amphetamine 3/4 wPentylenetetrazol 1/2 Amphetamine 1/2 "V~*'Pentylenetetrazol 3/4 Amphetamine 1/2 VENTILATION PER MINUTE IN LITERS Fig. 6 0.15 mg/kg 4% 2 _ 1 dog - 13 kg ‘ P - 40 mg/kg 0 1111111111 ‘ 6 I 3 dog - 10 kg ' P - 40 mg/kg 4. 2 h 5 . . . O Ill] 3 . 61-. 4 . I . 2 b 5 dog - 12.25 kg ’ . P - 42 mg/kg 011111111 '8 P 6 r L 4 - 2 5 (Average of 6 dogs) P ‘R 0_-|_JIIIIJ 0 20 40 60 min. Percent Increase 49 METARAMINOL BITARTRATE - AMPHETAMINE 3 mg/kg 4 0 2r- - 2 dog - 10 kg I F749 “lg/3‘9 4 dog - 11.5 kg P - 40 mg/kg 6 dog - 8.75 kg P - 40 mg/kg 200%” 100%” 0%' 0 20 40 60 min. VENTILATION PER MINUTE IN LITERS METARAMINOL BITARTRATE - 0.15 mg/kg P - 40 mg/kg ' n- l O 5 dog - 11 kg P - 40 mg/kg 1 I l .I l I _L (Average of 6 dogs) I l I #1 l l l 20 40 60 min. Percent Increase AMPHETAMINE 200% 100% 0%- 2 mg/kg 4 . 6 dog - 8.5 kg 50 dog - 8.5 kg P -. 4? 9949 4 dog - 8.75 kg P - 40 mg/kg P - 40 mg/kg 4 4 1 L 1 VENTILATION PER MINUTE IN LITERS Fig. 8 0.1 mg/kg 1 dog - 7 kg “ P - 40 mg/kg 3 dog - 16.5 kg . P - 36.6 mg/kg 1 J I L l L I L 5 dog - 13.5 kg P - 40 mg/kg 1 1 1 _1 l L L L - (Averages for 6 dogs) b ‘9‘» L19“ 5. I L 1 I 1 I L I Percent Increase 51 METARAMINOL BITARTRATE - AMPHETAMINE 2 rug/kg 2 dog - 14 kg P - 45 mg/kg 4.. 2 _ + 4 dog - 13.5 kg P - 40 mg/kg 0 1 1 1 1 1 l I 1 2 . v 6 dog - 14 kg . P - .40 rug/k9 O | L J 1 L L I L 200% - 3m - L (fig 100% " 0%,. I I I 1 I l L L -10 0 20 40 60 min. Taken by courtesy from Dr. Sisodia VENTILATION PER MINUTE IN LITERS Fig. 8 0.1 mg/kg 1 dog - 7 kg ’ P - 40 mg/kg I ’U l 36.6 mg/kg - 13.5 kg 40 rug/k9 1 1 1 I 4 I I L L - (Averages for 6 dogs) b GR \Q x #- Percent Increase 51 METARAMINOL BITARTRATE - AMPHETAMINE 2 rug/kg 2 dog - 14 kg 2 P - 45 mg/kg 0 1 J L I I _l 1 1L 6 I- 4 p 2 _ * 4 dog - 13.5 kg F P - 40 mg/kg 0 1 1 1 1 1 I I I 6 I- 4 r 2 ' I 6 dog - 14 kg L P -.40 rag/kg O I I J J I L I L 200%1- 3 ’ ‘Tk 133 100%" L 0%“- I I I 1 I I L 1 -10 0 20 40 60 min. Taken by courtesy from Dr. Sisodia 53 Fig. 10 AVERAGE PERCENTAGE INCREASE IN VENTILATION AT FOUR DIFFERENT DOSE LEVELS OF METARAMINOL BITARTRATE AND AMPHETAMINE Percent Increase " 500% /’/” ’l’/ 1” I, .. 400% ’I’ [I ' I /’ / / z'-’ I I ’ I / l' I, // / I- l’ / I 300% , / / / /_/ x’ ,/ l/ ’ z/ I, ’l’ I, I / ”’ / 7 I’ // - 200% ,I ,/ I / / // I’/ I ,l” \ x , 15 [I I,’ 00055 0“ "” . .0 \$\ 0 -—a—- m ... 0‘ W °‘o. O... I 1 1 I I I 1 0 10 20 30 40 50 60 70 Minutes "-‘“'Metaraminol Bitartrate 3/4 Amphetamine 1/2 “=*= Metaraminol Bitartrate 3/4 Amphetamine 3/4 “““WKMetaraminol Bitartrate 1/2 Amphetamine 3/4 m'Metaraminol Bitartrate 1/2 Amphetamine 1/2 52 VENTILATION PER MINUTE IN LITERS 40 mg/kg ;7 m I -’,/\;‘§;;“. 8.5 kg L I‘ P - 40 mg/kg l I l I j I I I m m P m _ (Average of 6 dogs) 3 U s ' l-l , u s v 8 3 , N a. I 1 I I I I l 1 Fig. 9 METARAMINOL BITARTRATE - AMPHETAMINE 0-1 m9/k9 3 mg/kg I 6 I . 4 _ ,r 2 dog - 11.75 kg ' 2 I' _ _ R 1 dog - 7.25 kg _ N P 40 mg/kg P - 40 mg/kg I j l 1 l l I 4 0 I l I L l I I l b 6 p 4.- 6 dog - 10 kg - P - 40 mg/kg 2+\[ PR 0 LI 1 I [J I 1 P 200%" F' 100%)' 0%- 2 1 l IILLJA 0 20 40 60 min. VENTILATION PER MINUTE IN LITERS 0.1 mg/kg T L . R ldog-7.25 kg . l lP‘- 40lmg/kg ,7 m I 40 rug/kg P " } 5dog-8.5kg N - P - 40 mg/kg m m P m _ (Average of 6 dogs) a o G " H , p a m ' U s ’ m I L I I J I I I 52 METARAMINOL BITARTRATE - AMPHETAMINE 3mg/kg 6%- L ,_ 4, j 2 dog - 11.75 kg 2 I P - 40 mg/kg I_ l l I 4‘- 6 dog - 10 kg 40 mg/kg "U I 200% P 100% F 0% - O 20 4O 60 min. 54 VENTILATION PER MINUTE IN LITERS O -10 O 20 40 60 min. Fig. 11 METHETHARIMIDE - AMPHETAMINE (As given below) 3 mg/kg 7 6 “2 dog - 10.5 kg r L P " 40 mg/kg P 1 dog - 10.5 kg 4 F _ P - 40 mg/kg ’ 2 I é'Nmi30 mg/kg INA ME 30 mg/kg - l§ I I I I l I I I O I AL I I _L_ I I I - 3 d°9 ‘ 9 kg 6 - 4 dog - 8.25 kg . P - 40 mg/kg _ P - 40 mg/kg . 4 .. + ’ ME 30 mg/kg 2 ' . NIA ME 30 mg/kg 7 ItA I I I I I I I I 0 I l I I P I l I 5 dog - 7.75 kg 6 6 dog - 8.75 kg P - 40 mg/kg r P - 40 mg/kg . 4 . ' F . Q—ME 30 mg/kg 2 ' (.ME 30 mg/kg I- NA .- A I I I I I I I I O I I I I_ I I I _I o Average of 5 dogs. 1 dog 3 P struggled before ME was in 8 200% ' - no further “1‘ recording. 2 - (53 - S 100% ' 3) ME g -— é——-ME ' A H 0% ' A L L l I I I I I 8),. I I I I I I I VENTILATION PER MINUTE IN LITERS 55 Fig. 12 METHETHARIMIDE - AMPHETAMINE (As given below) * 1 dog - 12.5 kg 6 - P - 40 mg/kg 4 P 2 n b ME 30 mg/kg 0 l 1A1 L l I l I l 61L 3 dog - 9 kg 4 . k 2 P ME 30 mg/kg ’ A O I I I I I I I I I 6 - 1 5 dog - 13 kg ' P - 40 mg/kg 4 , I. + 2 - 30 m ~I§€-ME g/kg . R A 0 I I I I I I I I_I 6 (Average of 6 dogs) .- ar ’\ (93 k“) 4 I- 2 ' ME I- ”A I L I I L I I I # 2 mg/kg 6 4. 2 dog - 11.75 mg/kg P - 40 mg/kg 2 ME 30 mg/kg 0 I_LAI L l I I I 1+ 6 300% 200% 100% Percent Increase 0% 4 dog - 13 kg {6 dog - 12.5 kg P - 40 mg/kg (— ME 30 mg/kg VENTILATION PER MINUTE IN LITERS Fig. 13 (As given below) L 1 dog - 7.5 kg P - 40 mg/kg X 2" A P K*ME 20 mg/kg I I I .I I I 3 dog - 9 kg P - 40 mg/kg If 2 h A * ME 20 mg/kg 0 I I I I I I I I (Average for 5 dogs) 4 2 0 L I _I_ I I I J‘ I -100 20 40 60 min. Taken by courtesy from Dr. “ P - 50 56 METHETHARIMIDE - AMPHETAMINE 2mg/kg 2 dog - 12 kg 4 I A. P - 40 mg/kg 2r- ME 20 mg/kg 0 11 I LA I I I 5 dog - 11 kg A ' ME 25 mg/kg I I I I I I_ I I o 3 3 200%- g . i. H +, lOOA’ G . 8 3 (”6' 1‘ O4 L I I I I I Sisodia 57 VENTILATION PER MINUTE IN LITERS Fig. 14 METHETHARAMIDE - AMPHETAMINE (As given below) 3 mg/kg 6 t 6 ,g P 1 dog - 8.5 kg _ 2 dog - 11.75 kg P - 40 mg/kg P - 40 mg/kg 4 I 4 - b . 2 L 2 * ME 20 k I ME 20 mg/kg .r “‘3/ g A A O I I I I I I I I J— O I I I L I I 4 I 4 3 dog - 12.25 kg 6 ‘ — 6 _ P 40 mg/kg I 4 dog - 1 kg * ' P - 40 mg/kg 4 - 4 . ME 20 tug/kg ~ 2 " 2 .. —— L. . ME 20 mg/kg A ‘ A I_ I I. _I I I I I I L —1 O ' I ‘ 1 0 I 6 I 5 dog - 8.5 kg 6 ’ 6 d°g ‘ 20kg /k ‘ P - 40 mg/kg - mg g 4 P 1 4 I- 2 b 2 - QME 20 mg/kg ME 20 tug/kg It I A _ A 0 4 I I I I .I L I . 0 I I I I I I I I I 6 m (Average for 6 dogs) 3 * . a 200% ' I \ k - 4 . ax g k 100% ‘ E 2 F QME 20 mg/kg 8 e ME 20 mg/kg .. NA 8 0% A O I I I I I I I I L a. L I I I I I I I_ I ~10 0 20 4O 60 min. 0 20 4O 60 min. (+) Awakened no further recording 58 Fig. 15 AVERAGE PERCENTAGE INCREASE IN VENTILATION AT FOUR DIFFERENT DOSE LEVELS OF METHETHARIMIDE AND AMPHETAMINE Percent Increase P500% '400% r3oo% I ' 13")“ I V M ,.,,,-:<%\I+e——vx\/ ( x it?“ 4.. W ____ ..___...- ,Q___... 3 k W)“- X (A) IJSCJI‘ * *‘QK , ' « W“ MIN‘m‘“ ImK Xx x «xxx xxxxxx who“ X X“ ’0‘“ x XW— __ “ W K “11 WI“ \\ “NW “\“x‘ILW; ,r“ W (W. I {W l .l . I J I I AIL ---- 0 10 20 30 40 50 60 70 Minutes #fit‘fiéMethetharimide 3/4 Amphetamine 1/2 WMethetharimide 1/2 Amphetamine 1/2 (Average of 5 dogs) Xxxmeethetharimide 3/4 Amphetamine 3/4 (Average of 5 dogs) x._;¢.x_xMethetharimide 1/ 2 Amphetamine 3/4 58 Fig. 15 AVERAGE PERCENTAGE INCREASE IN VENTILATION AT FOUR DIFFERENT DOSE LEVELS OF METHETHARIMIDE AND AMPHETAMINE. Percent Increase ~500% {400% r3oo% + ' 23.2... fl! ) ' \ ‘kw-A/ 1* \ {23‘5” )9 .200% ’ 0 o k “_m_ ') * V”)? X ’1'“ (HI , k”’”’ _. x‘kflxw‘fifixmxukx . x wot“ HIK— ‘* 0 ’ xxx “ Ix ukwwmmv ”Mug" (xx“‘*****“**““‘ x *xxxxm ‘” l * ' \ x * \ i = X “Hm \\ “WWW” N M2244 ’-'\& arms“ 9‘ ,_ ~ — ‘ 1‘:ka {I c> 32 l J I J _ I I I - N--- 10 20 30 4O 50 6O 70 Minutes a¢€fitfi§Methetharimide 3/4 Amphetamine L/2 uwmwwubthetharimide l/2 Amphetamine 1/2 (Average of 5 dogs) ”*XxxIMethetharimide 3/4 Amphetamine 3/4 (Average of 5 dogs) xrxfivaethetharimide 1/2 Amphetamine 3/4 Fig. 16 THREE BEST COMBINATIONS OF THREE PAIRS OF DRUGS STUDIED Percent Increase 59 “”'-Metaraminol Bitartrate 3/4 Amphetamine 1/2 {fioquethetharimide 3/4 Amphetamine 1/2 e—e—oPentylenetetrazol 3/4 Amphetamine 3/4 L500% // / -4oo% //’ // / / / / / / // - 300% / // / O ’x ‘\ ’K‘A) o o I \6‘ ’oo 0 a . o x’ z o r- 00% /” 0 Q / I / 0‘ [ll 1 I‘ , . I, WV ‘. 0 ’// gr,,ycr’/’ 1 I” o o \l 0% . I I I J I I 10 20 30 40 50 6O 70 Minutes 60 Fig. 17. AVERAGE PERCENTAGE INCREASE IN VENTILATION 10 PER MINUTE IN DEEPLY BARBITALIZED DOGS WITH NO ANALEPTICS' (9 Trials) 20 3O 4O 50 60 Minutes Four Trials Taken by Courtesy from Dr. Cairy Note: At 0% respiration was 1.35 61 CHAPTER IV DISCUSSION Pentylenetetrazol and Amphetamine Combination Pentylenetetrazol and amphetamine were tested at four different dose level combinations against barbiturate depres- sion to see the behavior of each combination on ventilation. we ended up with only two degrees of response, as indicated in Fig. 5. Pentylenetetrazol 3/4, amphetamine 3/4, turned out to be the most effective combination. All the others listed below showed more or less the same degree of response: Pentylenetetrazol 1/2, Amphetamine 1/2 Pentylenetetrazol 3/4, Amphetamine 1/2 Pentylenetetrazol 1/2, Amphetamine 3/4 Pentylenetetrazol stimulates the medulla directly to in- crease the ventilation. The effect of pentylenetetrazol on the medulla is much more prominent when this region is depres- sed by barbiturate intoxication or others. Thus in a barbitalized dog at higher dose level of pentylenetetrazol, ventilation may be pushed to a higher minute respiratory volume than with smaller doses. 62 Amphetamine is a sympathomimetic amine, stimulating both the central nervous system and the cardiovascular system. Barbiturates not only depress the respiratory system, but also the cardiovascular system considerably, even under con- trolled anesthetic conditions (Daniel _thgl., 1956). Amphe- tamine thus antagonizes barbiturates at the central nervous system as well as at the cardiovascular system. Amphetamine is a long-acting drug due to slow destruction and thus it maintains a good ventilation. Pentylenetetrazol in higher doses coupled with higher doses of amphetamine showed an immediate increase in ventila- tion by 150 percent; later, it dropped to 100 percent; then it rose gradually and was increased by 210 percent at the end of 60 minutes. The rest of the combinations are not so effective as the one described above. This suggests that pentylene- tetrazol and amphetamine exhibit additive synergism and do Thelp each other in producing their effects on ventilation by 'two ways: one by the quantitative stimulation of the respira- 'tory center of pentylenetetrazol; secondly by quantitative immmovement in cardiovascular system by amphetamine to over- <30me the barbiturate depression at the cardiovascular system. 13113 favors the opinion of weaver and Bunde (1960) that "the cardio-vascular system, as well as the respiratory, should be