The effect of pindolol on exercise-induced cardiac acceleration in relation to plasma levels in man The correlation between the beta receptor blocking activity of pindolol and plasma level was studied in 8 subjects after a JO-mg oral dose. Exercise tachycardia was markedly reduced over a period of at least 6 hr. Significant effects were recorded 30 min after the drug. For each individual there was a close correlation between log plasma level and beta blockade. The regression lines were parallel as shown by analysis of covariance; the intercepts, however, were significantly different. It can be concluded that there is a correlation between plasma level and beta adrenergic blockade by pindolol, but the data failed to establish in different individuals the blood levels necessary to achieve effective adrenergic blockade.

Roland Gugler, M.D., Werner Habel, M.D., Gunter Bodem, M.D., and Hans J. Dengler, M.D. Bonn, Germany

Medizinische Universitiitsklinik, 53 Bonn, und Institut fur Medizinische Dokumentation und Statistik, 63 Giessen, West Germany

For many drugs it is well established that pharmacologic effect is more closely correlated to plasma concentration than to oral or intravenous dose!' 4, 5, 6, 8-10, 15, 17, 18, 24, 25 provided that pseudodistribution equilibrium is achieved. Plasma concentrations of some commonly used drugs, however, differ over a wide range among people on the same dosage schedule. 18 A sevenfold and fourfold variation after the same oral dose has been shown with the beta receptor blocking agents propranolol24 and pindoloP2 respectively. In man the pharmacologic effect of beta adrenergic blocking agents can be assessed quantitatively by measuring the blockade of the tachycardia induced by exercise, passive tilting, and isoproterenol infusion. This paper describes the blockade of exerciseReceived for publication July 23. 1974. Accepted for publication Oct. 7. 1974. Reprint requests to: Roland Gugler. M.D .• Department of Medicine. University of Bonn. 53 Bonn- Venusberg. West Germany.

induced tachycardia by pindolol (Visken) and examines the correlation between plasma levels of the drug and beta receptor blocking activity. The beta receptor blocker pindolol, DL-4-(2hydroxy-3-isopropylaminopropoxy)-indole,differs in its pharmacodynamic properties from propranolol, particularly in its high intrinsic agonist activity. 2, 13, 14, 23 Methods

Exercise-induced tachycardia. The studies were performed in 6 male and 2 female healthy subjects 20 to 25 years of age. Identical ergometer tests were performed by each subject three times in the control period and after the administration of pindolol, at the times cited below, using an electrically braked bicycle ergometer. The individual exercise load was calculated with regard to the parameters of height and body weight. 16 The load varied between 540 and 720 kpm/min. The increase in heart rate was at least 50 bpm in the control period in all individuals. 127

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The heart rate was estimated from the ECG tracing at rest and during the last 15 sec of the 4 min test period. The subjects had a standard light meal 3 hr before the test and rested supine for 20 min before every exercise period. The exercise period was preceded by a 5-min rest sitting on the bicycle. After three control values were recorded the subjects received two 5-mg tablets of pindolol. Pharmacokinetic data12 previously obtained for pindolol in this laboratory prompted us to investigate the beta blocking effect at the following times: 0.5, I, 2, 3, 4, and 6 hr after ingestion of the drug. Plasma samples were obtained immediately after the period of exercise. The relative blockade of the exercise tachycardia was determined. The response (R) was calculated by subtracting the heart rate at rest from the heart rate at work. The individual response before administration of pindolol was considered to be the normal response (Ro) to the applied workload. The per cent blockade in each subject at various times was calculated as follows 15 : R -R Per cent blockade = ~ . 100. o

Determination of plasma levels of pindolol. Plasma pindolol was measured using a slightly

modified method described by Pacha. 20 Plasma, 5 ml, from blood containing 2 to 3 drops of heparin as an anticoagulant, was made alkaline by adding I ml of I N sodium hydroxide and extracted with 12 rnl of diethylether by shaking for 10 min on an automatic shaker. Pindolol was re-extracted out of one 10-ml aliquot of the organic layer into 2 ml of 0.1 N hydrochloric acid. A portion of the acidic layer, 1.5 ml, was reacted with 0.5 mg of o-phthalaJdehyde by heating the mixture in a water bath at 50° C for 30 min. The fiuorophor was stabilized by adding 5 mg of ascorbic acid and examined in an Aminco-Bowman spectrophotofiuorometer at excitation and emission maxima of 390 and 440 nm, respectively. Statistical methods. In Table I and Figs. I and 2 values are expressed as mean ± SEM. For the comparison of heart rates, per cent blockade, and plasma levels the paired t test was used. Regression lines were calculated by least square analysis and tested for parallelism. 7 Results

Resting heart rates. Heart rates at rest recorded before and after the administration of pindolol are shown in Fig. 1. Following a single dose of 10 mg pindolol to 8 subjects, the mean resting heart rate decreased from 76.9 to 73 bpm from the second to the sixth hour during the

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Table I. Pindolol plasma levels, changes in heart rate, and per cent blockade (x ± SEM, N = 8) at stated times after the oral administration of 10 mg of pindolol Heart rate Time (hr)

0 0.5 1 2 3 4 6

I

Rest

76.9 75.9 74.3 73.5 72.9 73.4 72.9

± ± ± ± ± ± ±

4.9 4.5 4.7 4.6 4.6 4.6 5.3

Pindolol Exercise

139.3 131.1 116.4 110.3 111.9 115.9 119.3

± ± ± ± ± ± ±

5.2 7.2 6.8 4.3 4.0 4.0 4.6

6-hr investigational period (Fig. I, Table I), which represents a reduction of 5%, an amount that is not significant. Exercise heart rates. The mean values for heart rate during exercise at different times after the administration of pindolol are presented in Fig. I. The mean heart rate during the exercise test before medication rose from the restingvalueof76.9 ± 4.9to 139.3 ± 5.2bpm (Table I), indicating an average increase of 62.4 bpm (8 I . I %). Exercise-induced tachycardia was reduced 30 min after administration of the drug by 8.2 bpm (Fig. I, Table I), which is equivalent to a blockade of 13.8 ± 3.9%. Significance tests for comparing pairs of dif-

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18.2 34.9 51.9 45.8 33.7 23.2

± ± ± ± ± ±

2.9 7.6 9.5 8.5 7.3 3.2

ferences showed a blocking effect at 30 min (p < 0.005), I hr (p < 0.001), and 6 hr (p < 0.005) after the drug. The maximum effect on exercise tachycardia was observed in 3 subjects at I hr, and in the other 5 subjects at 2 hr, after the ingestion of pindolol. The maximum per cent blockade ranged from 29.4 to 57%. Pindolol plasma levels. Mean plasma levels obtained the first 6 hr after oral administration of 10 mg pindolol to 8 subjects are shown in Fig. 2 and Table I. Individual plasma peaks were obtained after I hr (3 subjects) or 2 hr (5 subjects). Maximum plasma concentrations ranged from 22.8 to 104.8 ng/m!.

Relationship between plasma concentra-

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Table II. Statistical characteristics ofthe least square regression lines correlating log plasma levels to beta blocking effect Subject I 2 3 4 5 6 7 8

6 6 5 6 5 6 6 6

Per cent blockade (x)

Log plasma level (y)

Correlation coefficie nt

Regression coefficient

Regression constant

20.47 19.83 33.12 29.08 35.24 40.67 25.55 39.65

2.754 3.349 2.954 4.151 3.532 3.249 3.526 3.691

0.989 0.944 0.985 0.834 0.851 0.993 0.979 0.896

0.027 0.033 0.032 0.035 0.019 0.024 0.036 0.037

2.201 2.690 1.883 3.148 2.874 2.277 2.605 2.226

tion of pindolol and beta blockade. Fig. 2 compares the mean plasma levels of pindolol with the mean values of per cent blockade. Although very similar time curves were obtained, thus indicating a positive correlation, the principles of statistics do not allow one to correlate the mean value curves. Least square regression lines were calculated for the relationship between the logarithm of plasma pindolol concentration (corrected for kilogram body weight) and the effect on exercise tachycardia in each

subject (Fig. 3). The results clearly demonstrate a positive correlation for the individual subjects with correlation coefficients ranging from 0.83 to 0.99 (Table II). The regression lines were also tested for parallelism by an analysis of covariance. 7 The result of test (F = 1.08, degrees of freedom, 7.30), compared the single regression coefficients (Table II) and did not permit rejection of the null hypothesis, even allowing for the fact that the regression lines are not strictly parallel

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in Fig. 3. On the other hand, the analysis of variance shows a difference in the y axis intercept of the curves (F = 15.24; degrees of freedom, 6.32; p < 0.001). Therefore, a common regression line for all subjects participating in this study is not acceptable.

Discussion In our study, 10 mg of pindolol administered orally to 8 healthy volunteers had no significant effect on the resting heart rate during the first 6 hr after ingestion. For practoloPO and alpren010P5. 18 there were also no significant changes in resting heart rate reported after administration of a beta blocking dose. This lack of a negative effect on heart rate was assumed to be due to the intrinsic activity of these particular beta blockers. 3 Hicks and associates 13 found a small dose-related increase in resting heart rate after the intravenous administration of pind0101. Cardiac slowing was, however, recorded after an intravenous dose of propranolol, which is devoid of an intrinsic sympathomimetic activity.22 We chose to study the inhibition of exercise tachycardia in preference to other tests, depending either on the heart rate response to 80° head-up tilt or intravenous administration of isoproterenol, for the following reasons. Tilting was shown to be inadequate in some patients for assessing beta receptor blockade. 5 Isoproterenol increases heart rate by both a direct action at the cardiac receptors and an indirect mechanism induced by peripheral vasodilatation. Its actions have not yet been related to those of endogenous sympathetic stimulation. The exercise test offers a reproducible physiologic adrenergic stimulation provided that the workload is sufficiently high. Robinson and associates 22 showed that the increase in heart rate following mild exercise is mainly under parasympathetic control. At higher levels of exercise, however, vagal withdrawal has no effect upon the heart rate response, while beta receptor blockade attenuates the increase of heart rate to exercise. Individually calculated exercise loads 16 were used to study the effect of pindolol on exercise tachycardia; body weight and height (synonym to body surface) were regarded as ade-

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quate parameters for the determination of the individual exercise load. 1, 13 Pindolol reduced the exercise-induced tachycardia markedly over a period of at least 6 hr when it was administered orally in a dose of 10 mg. Significant changes were already present during the first 30 min. Pharmacokinetic studies with pindolol in man 12 have shown that in fasting subjects maximal blood levels develop at about 80 min after oral administration. After 2 hr they decline linearly on a semilogarithmic plot in most subjects, with a half-life of about 3.5 hr. The data presented here (Fig. 2) differ from the findings in our previous study in showing delayed absorption with mean peak plasma levels achieved at 2 hr. This is probably due to the altered conditions and the fact that the subjects had a standard light meal 3 hr before receiving the dose. We nevertheless viewed this procedure as more closely related to the clinical situation as well as enabling the elimination of many side effects caused by stressing fasting subjects. We administered 10 mg of pindolol orally, which is supposed to be safe, effective, and long-acting. According to a recent report,3 5 mg of pindolol given orally is equivalent to 100 mg of propranolol in reducing exercise tachycardia. Since we were particularly interested in relating the pharmacodynamic activity of beta blockade to plasma levels, we chose the lO-mg dose of pindolol, which is within the therapeutic range and also guarantees a reliable plasma concentration of the drug over a long period of time and a high degree of antagonist activity against exercise tachycardia. The pharmacokinetics of propranolol and pindolol cannot be compared unreservedly because there are differences in the volume of distribution and metabolism of each that, in the case of propranolol, leads to an active metabolite,21, 26, 27 whereas pindolol does not appear to be transformed to a comparable intermediate. 12 In our study the maximal per cent blockade of exercise tachycardia was recorded 2 hr after the oral administration of pindolol and averaged 40.6%. At that time the mean plasma level of pindolol was 51.9 ng/ mI. Our data indicate that a significant beta blocking effect was still pres-

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sent at plasma levels of 10 ng/ ml of pindolol or less. These results agree with recent reports of a beta blocking effect as long as 24 hr after a single therapeutic dose of pindolol. 19 All the individual plasma levels vs response curves exhibited a close relationship between the logarithm of the plasma concentration and the beta adrenergic receptor blocking activity as shown by the individual correlation coefficients (Table II). Attempts to correlate the data of all subjects in order to construct a common regression line failed, however, mainly due to the significant difference in the y axis intercept of the curves. The marked differences in the individual plasma levels obtained from identical doses of beta blocking agents, as shown by Shand, Nuckolls, and Oates 24 for propranolol and in this laboratory12 for pindolol, prompted us to assume that the plasma level, by analogy with a number of other drugs, would be the most reliable parameter to predict accurately the degree of activity of pindolol. The results presented here, however, prove this assumption to be correct only in a limited sense, since plasma levels of pindolol were associated with a great variation of beta blockade among subjects. This does not support the plasma level as a guide to pindolol therapy. Although the intercepts of the individual regression lines were different, there was no significant difference in the slopes of the curves. This means that a given concentration of pind0101 inhibits the exercise tachycardia in different subjects to different degrees, whereas a given increase in concentration is associated with a constant increase in the beta blocking effect. This set of parallel concentration-response curves located between +y+x is open to several interpretations. According to George and associates,11 there is a considerable difference in the heart rate response among various subjects after intravenous administration of identical doses of isoproterenol. Their log dose-response curves exhibited variations in the y axis (log dose) intercepts together with identical slopes of the individual curves. By analogy, a similar situation regarding the intrinsic beta mimetic action of pindolol might be assumed. An interindividual difference in the sympathetic ner-

vous system response to the ergometer test is, however, a more likely explanation for this effect. This study was supported by the Sandoz Foundation of Therapeutic Research. We thank Werner Herold for his skillful technical assistance and also all colleagues and students who participated in the investigations.

References I. Ablad, B., Ervik, M., Hallgren, J., Johnsson, G., and Si:ilvell, L.: Pharmacological effects and serum levels of orally administered alprenolol in man, Eur. J. Clin. Pharmacol. 5:44-52, 1972. 2. Aellig, W. H.: Isoprenaline antagonism and duration of action in exercise induced tachycardia of three ,a-adrenoceptor blocking drugs: Pindolol, LF 17-895 and propranolol, Br. J. Pharmacol. 47:621, 1973. 3. Barrett, A. M., and Carter, J.: Comparative chronotropic activity of beta adrenoceptive antagonists, Br. J. Pharmacol. 40:373-381, 1970. 4. Bigger, I. T., Jr., Schmidt, D. H., and Kutt, H.: Relationship between the plasma level of diphenylhydantoin sodium and its cardiac antiarrhythmic effects, Circulation 38:363-374, 1968. 5. Bodem, G., Brammell, H. L., Weil, J. V., and Chidsey, C. A.: Pharmacodynamic studies of beta adrenergic antagonism induced in man by propranolol and practolol, J. Clin. Invest. 52: 747-754, 1973. 6. Brodie, B. B., and Reid, W. W.: The value of determining the plasma concentration of drugs in animals and man, in LaDu, B. N., Mandel, H. G., and Way, E. L., editors: Fundamentals of drug metabolism and drug disposition, Baltimore, 1971, The Williams & Wilkins Company, pp. 328-339. 7. Brownlee, K. A.: Statistical theory and methodology in science and engineering, ed. 2, John Wiley & Sons, Inc., New York, 1969, pp. 376388. 8. Clevealand, C. R., and Shand, D. G.: Effect of route of administration on the relationship between ,a-adrenergic blockade and plasma propranolol level, CLiN. PHARMACOL. THER. 13: 181-185, 1972. 9. Coltart, D. J., and Shand, D. G.: Plasma propranolollevels in the quantitative assessment of ,a-adrenergic blockade in man, Br. Med. J. 3: 731-734, 1970. 10. Fitzgerald, 1. D., and Scales, B.: Effect of a new adrenergic ,a-blocking agent (lCI 50, 172) on heart rate in relation to its blood levels, Int. J. Clin Pharmacol. 1:467-474, 1968. II. George, C. F., Conolly, M. E., Fenyvesi, T., Briant, R., and Dollery, C. T.: Intravenously

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12. 13.

14.

15.

16.

17. 18. 19.

administered isoproterenol sulfate dose-response curves in man, Arch. Intern. Med. 130:361-364, 1972. Gugler, R., Herold, W., and Dengler, H. J.: Pharmacokinetics of pindolol in man. In press. Eur. J. Clin. Pharmacol. 7:17-24, 1974. Hicks, D. c., Arbab, A. G., Turner, P., and Hills, M.: A comparison of intravenous pindolol and propranolol in normal man, 1. Clin. Pharmacol. 12:212-216, 1972. Hill, R. C., and Turner, P.: Preliminary investigations of a new beta-adrenoceptive receptor blocking drug, LB46, in man, Br. J. Pharmacol. 36:368-372, 1969. Johnsson, G., Sjogren, J., and SOIvell, L.: Betablocking effect and serum levels of alprenolol in man after administration of ordinary and sustained release tablets, Eur. J. Clin. Pharmacol. 3:74-81, 1971. Kaltenbach, M.: Beurteilung der Leistungsreserven von Herzkranken mit Hilfe von Stufenbelastungen, Boehringer Mannheim GmbH, pp. 7188, 1968. Koch-Weser, J., and Klein, S. W.: Procainamide dosage schedules, plasma concentrations, and clinical effects, J.A.M.A. 215: 1454-1460, 1971. Koch-Weser, J.: Serum drug concentrations as therapeutic guides, N. Engl. J. Med. 287:227231, 1972. Olsson, B. S., and Varnauskas, E.: Duration of beta-receptor blockade after oral administration of LB-46, Eur. 1. Clin. Pharmacol. 5: 214-217, 1973.

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20. Pacha, W. L.: A method for the fluorometric determination of 4-(2-hydroxy-3-isopropylaminopropoxy)-indo1e, a J3-blocking agent, in plasma and urine, Experientia 25:802-803, 1969. 21. Paterson, J. W., Conolly, M. E., Dollery, C. T., Hayes, A., and Cooper, R. G.: The pharmacodynamics and metabolism of propranolol in man, Pharmacol. Clin. 2:127-133, 1970. 22. Robinson, B. F., Epstein, S. E., Beisa, G. D., and Braunwald, E.: Control of heart rate by the autonomic nervous system, Cir. Res. 19:410411,1966. 23. Saameli, K.: Die pharmakologische Charakterisierung J3-sympathikolytischer Substanzen, in Dengler, H. J., editor: Die therapeutische Anwendung J3-sympathikolytischer Stoffe, Stuttgart, 1972, F. K. Schattauer Verlag. 24. Shand, D. G., Nuckolls, E. M., and Oates, J.A.: Plasma propranolol levels in adults, with observations in four children, CLIN. PHARMACOL. THER. 11:112-120, 1970. 25. Vesell, E. S., and Passananti, G. T.: Utility of clinical chemical determinations of drug concentrations in biological fluids, Clin. Chern. 17: 851-866, 1971. 26. Walle, T., and Gaffney, T. E.: Propranolol metabolism in man and dog: Mass spectrometric identification of six new metabolites, J. Pharmacol. Exp. Ther. 182:83-92, 1972. 27. Zacest, R., and Koch-Weser, J.: Relation of propranolol plasma level to J3-blockade during oral therapy. Pharmacology 7:178-184, 1972.