422
BIOCHEMICAL SOCETY TRANSACTIONS
Table 2. Effectof reoxygenation on enzyme release by anoxicperfused mouse, rat, guineapig and rabbit hearts The release of creatine kinase from anoxic hearts (mean release during the 15min period of anoxia before reoxygenation) was compared with the release during reoxygenation (mean release during the first 15min after reoxygenation). Eight hearts were studied for each species. Enzyme released (units/min per g dry wt. of heart) c
Species Mouse Rat Guinea pig Rabbit
\
Before reoxygenation
After reoxygenation
1.57 1.26 1.69 3.10
7.67 56.7 1.90 3.80
In previous studies (Hearse et al., 1973, 1975) we have shown that reoxygenation of the rat heart during the early stages of phase 2 release greatly exacerbates ultrastuctural damage and enzyme release. The reoxygenation phenomenon appears to represent the compression of cellular damage and enzyme release, which would normally take several hours, into a few minutes. Hearts of each species were reoxygenated during the early stages of phase 2 release (rat, t = 150min; mouse, t = 95min; guinea pig, t = 170rnin; rabbit, t = 170min). The results (Table 2) indicate a marked species difference in the resistance of the myocardium to reoxygenation damage. The rat shows the greatest susceptibility, with a 45-fold increase in the rate of enzyme release. With the mouse heart there was also a marked exacerbation of release. In contrast, the rabbit heart and guinea-pig heart exhibited only very small increases. These results stress the marked species differences that exist in the response of different mammalian hearts to various stimuli. This work was supported with the aid of a grant from the British Heart Foundation. The technical assistance of Mr. M. Stoll and Miss R. Baig is gratefully acknowledged. BrachfeId, N., Ohtaka, Y., Klein, I. & Kawade, M. (1972) Circ. Res. 31, 453-467 Hearse, D. J. & Humphrey, S. M. (1975) J. Mol. Cell. Cardiol. in the press Hearse, D. J., Humphrey, S. M. & Chain, E. B. (1973) J. Mol. Cell. Cardiol. 5 , 395-407 Hearse, D. J., Humphrey, S. M., Nayler, W. G., Slade, A. & Border, D. (1975) J. Mol. Cell. Cardiol. in the press Lesch, M., Teichholz, L. E., Soeldner, J. S. & Gorlin, R. (1974) Circulation 49, 1028-1037 Medical Research Council (1968) Lancet ii, 1355-1360 Opie, L. H. (1970) J. Mol. Cell Cardiol. 1, 107-115 Sodi-Pallares, D. (1969) Ann. N. Y. Acud. Sci. 156, 603-619
The Effect of P-Adrenergic Agonists on Enzyme Release from the Hypoxic Isolated Berfused Rat Heart DAVID J. HEARSE, PAMELA B. GARLICK and JOHN P. SHILLINGFORD Cardiovascular Research Unit, Royal Postgraduate Medical School, Du Cane Road, London W12 OHS, U.K. The appearance in the serum of specific cardiac enzymes has been used in the diagnosis and assessment of myocardial infarction (Ewen & Griffiths, 1971;Sobel &Shell, 1972). 1975
555th MEETING, ABERYSTWYTH
423
The extent of enzyme leakage from the cell is related to the severity of both the biochemical and the morphological damage. From studies of myocardial enzyme depletion in experimentally infarcted dogs it has been suggested (Shell et al., 1971) that enzyme release may be used to quantify accurately the extent of cellular damage. Drugs designed to protect the myocardium against damage would therefore be expected to decrease or delay enzyme release. In recent studies (Hearse & Humphrey, 1975) with 02-deficient rat hearts we have illustrated how the availability of a variety of compounds (glucose, 02,methylprednisolone) can substantially decrease or delay the release of myocardial enzymes. After the onset of O2deprivation the myocardium enters a phase of reversible celluIar and metabolic damage; if the O2supply is not restored this becomes irreversible and leads to tissue necrosis. We have suggested (Hearse &Humphrey, 1975) that the onset of major enzyme leakage may reflect the transition from reversible to irreversible injury. In addition, it may well be the diminution of cellular energyreserves and the consequent inability to maintain metabolic and morphological integrity that initiates the onset of this irreversible damage. The known ability (Hearse &Chain, 1972; Hearse etal., 1974,1975) of exogenous glucose to maintain cellular reserves of ATP, to decrease and delay enzyme release and to preserve myocardial ultrastructure under these conditions supports this view. It is therefore possible that compounds known to increase cellular energy consumption and thereby deplete available energy reserves may well increase or advance enzyme release and thus extend tissue damage. Human hearts damaged by coronary artery thrombosis and associated ischaemia have sometimes been treated with 8-adrenergic agonists. Through their stimulation of the myocardial 8-receptors cardiac output and heart work are increased. Usually one observes both an inotropic (i.e. increased contractility) and chronotropic (i.e. increased heart rate) response. This increase in work and therefore energy consumption may possibly accelerate both cellular damage and enzyme release. It is known that, although 8-adrenergic agonists are able to exert an inotropic effect during conditions of cardiac O2shortage, the effect is often short-lived and eventually cardiac failure is accelerated. In addition, from clinical and experimental studies (Braunwald & Maroko, 1974) it is known that these agents can increase myocardial damage. Isoprenaline is a classical 8-adrenergic agonist that exerts strong inotropic and chronotropic effects. This compound is in routine clinical use as an inotrope, but its
Table 1. Znotropic and chronotropic responses of the isolatedperfused rat heart to various doses of 8-adrenergic agonists The volume of the recirculating perfusate was 100ml. Six hearts were studied for each group and the S.E.M. values are indicated. Dose Drug Control (+)-Isoprenaline
(+)-Dobutamhe
(,uM)
Heart rate (beats/min)
dP/dt (max.) (cmH,O/s)
0.001 0.01 0.1 1 .o 0.03 0.3 3.0
278 f 15 301 f 17 323 f 17 366f 13* 361 +11* 273 f5 308 f 5 359 9*
7108 f299 6435f451 8025 f406 10292 f430* 13 250? 576* 7000 f 325 11161+706* 15221 k 740*
* Greater than control value with P
BIOCHEMICAL SOCETY TRANSACTIONS
Table 2. Effectof reoxygenation on enzyme release by anoxicperfused mouse, rat, guineapig and rabbit hearts The release of creatine kinase from anoxic hearts (mean release during the 15min period of anoxia before reoxygenation) was compared with the release during reoxygenation (mean release during the first 15min after reoxygenation). Eight hearts were studied for each species. Enzyme released (units/min per g dry wt. of heart) c
Species Mouse Rat Guinea pig Rabbit
\
Before reoxygenation
After reoxygenation
1.57 1.26 1.69 3.10
7.67 56.7 1.90 3.80
In previous studies (Hearse et al., 1973, 1975) we have shown that reoxygenation of the rat heart during the early stages of phase 2 release greatly exacerbates ultrastuctural damage and enzyme release. The reoxygenation phenomenon appears to represent the compression of cellular damage and enzyme release, which would normally take several hours, into a few minutes. Hearts of each species were reoxygenated during the early stages of phase 2 release (rat, t = 150min; mouse, t = 95min; guinea pig, t = 170rnin; rabbit, t = 170min). The results (Table 2) indicate a marked species difference in the resistance of the myocardium to reoxygenation damage. The rat shows the greatest susceptibility, with a 45-fold increase in the rate of enzyme release. With the mouse heart there was also a marked exacerbation of release. In contrast, the rabbit heart and guinea-pig heart exhibited only very small increases. These results stress the marked species differences that exist in the response of different mammalian hearts to various stimuli. This work was supported with the aid of a grant from the British Heart Foundation. The technical assistance of Mr. M. Stoll and Miss R. Baig is gratefully acknowledged. BrachfeId, N., Ohtaka, Y., Klein, I. & Kawade, M. (1972) Circ. Res. 31, 453-467 Hearse, D. J. & Humphrey, S. M. (1975) J. Mol. Cell. Cardiol. in the press Hearse, D. J., Humphrey, S. M. & Chain, E. B. (1973) J. Mol. Cell. Cardiol. 5 , 395-407 Hearse, D. J., Humphrey, S. M., Nayler, W. G., Slade, A. & Border, D. (1975) J. Mol. Cell. Cardiol. in the press Lesch, M., Teichholz, L. E., Soeldner, J. S. & Gorlin, R. (1974) Circulation 49, 1028-1037 Medical Research Council (1968) Lancet ii, 1355-1360 Opie, L. H. (1970) J. Mol. Cell Cardiol. 1, 107-115 Sodi-Pallares, D. (1969) Ann. N. Y. Acud. Sci. 156, 603-619
The Effect of P-Adrenergic Agonists on Enzyme Release from the Hypoxic Isolated Berfused Rat Heart DAVID J. HEARSE, PAMELA B. GARLICK and JOHN P. SHILLINGFORD Cardiovascular Research Unit, Royal Postgraduate Medical School, Du Cane Road, London W12 OHS, U.K. The appearance in the serum of specific cardiac enzymes has been used in the diagnosis and assessment of myocardial infarction (Ewen & Griffiths, 1971;Sobel &Shell, 1972). 1975
555th MEETING, ABERYSTWYTH
423
The extent of enzyme leakage from the cell is related to the severity of both the biochemical and the morphological damage. From studies of myocardial enzyme depletion in experimentally infarcted dogs it has been suggested (Shell et al., 1971) that enzyme release may be used to quantify accurately the extent of cellular damage. Drugs designed to protect the myocardium against damage would therefore be expected to decrease or delay enzyme release. In recent studies (Hearse & Humphrey, 1975) with 02-deficient rat hearts we have illustrated how the availability of a variety of compounds (glucose, 02,methylprednisolone) can substantially decrease or delay the release of myocardial enzymes. After the onset of O2deprivation the myocardium enters a phase of reversible celluIar and metabolic damage; if the O2supply is not restored this becomes irreversible and leads to tissue necrosis. We have suggested (Hearse &Humphrey, 1975) that the onset of major enzyme leakage may reflect the transition from reversible to irreversible injury. In addition, it may well be the diminution of cellular energyreserves and the consequent inability to maintain metabolic and morphological integrity that initiates the onset of this irreversible damage. The known ability (Hearse &Chain, 1972; Hearse etal., 1974,1975) of exogenous glucose to maintain cellular reserves of ATP, to decrease and delay enzyme release and to preserve myocardial ultrastructure under these conditions supports this view. It is therefore possible that compounds known to increase cellular energy consumption and thereby deplete available energy reserves may well increase or advance enzyme release and thus extend tissue damage. Human hearts damaged by coronary artery thrombosis and associated ischaemia have sometimes been treated with 8-adrenergic agonists. Through their stimulation of the myocardial 8-receptors cardiac output and heart work are increased. Usually one observes both an inotropic (i.e. increased contractility) and chronotropic (i.e. increased heart rate) response. This increase in work and therefore energy consumption may possibly accelerate both cellular damage and enzyme release. It is known that, although 8-adrenergic agonists are able to exert an inotropic effect during conditions of cardiac O2shortage, the effect is often short-lived and eventually cardiac failure is accelerated. In addition, from clinical and experimental studies (Braunwald & Maroko, 1974) it is known that these agents can increase myocardial damage. Isoprenaline is a classical 8-adrenergic agonist that exerts strong inotropic and chronotropic effects. This compound is in routine clinical use as an inotrope, but its
Table 1. Znotropic and chronotropic responses of the isolatedperfused rat heart to various doses of 8-adrenergic agonists The volume of the recirculating perfusate was 100ml. Six hearts were studied for each group and the S.E.M. values are indicated. Dose Drug Control (+)-Isoprenaline
(+)-Dobutamhe
(,uM)
Heart rate (beats/min)
dP/dt (max.) (cmH,O/s)
0.001 0.01 0.1 1 .o 0.03 0.3 3.0
278 f 15 301 f 17 323 f 17 366f 13* 361 +11* 273 f5 308 f 5 359 9*
7108 f299 6435f451 8025 f406 10292 f430* 13 250? 576* 7000 f 325 11161+706* 15221 k 740*
* Greater than control value with P
