Toxicon, Vol ., 18 pp . 389-391 .

0041-0101/80/0501-0389$02 .00/0© Pergamon Press Ltd . 1980 . Printed in England.

EFFECT OF INTRAVENTRICULAR ADMINISTRATIONOF STREPTOLYSIN O ON THE ELECTROCARDIOGRAM

OF DOGS

SUSHMA GUPTA* and R. K. GUPTAtDepartments of "Physiology and tPathology, Maharani Laxmi Bai Medical College,

Jhansi-284001, India

(Acceptedfor publication 6 September 1979)

STREPTOLYSIN O is a potent streptococcal exotoxin released during recurrent infections withgroup A haemolytic streptococci . Besides its haemolytic and cytotoxic effects (THOMPSONet al ., 1970), this toxin also produces cardiac damage (GUPTA et al., 1973, 1976a), derange-ment of cardiac activity (GUPTA et al., 1976b, 1976c ; GUPTA and GUPTA, 1977a) andneuroexcitation (GUPTA, 1979 ; GUPTA and SRIMAL, 1979 ; RASKOVA and VANECEK, 1957) inexperimental animals . Recently, intracerebroventricular administration of streptolysin Ohas also been shown to produce electrocardiographic changes in rabbits (GUPTA andGUPTA, 1977b) . In the present study the role of the central nervous system in mediating thecardiotoxic effects of streptolysin O in dogs has been investigated .The experiments were performed on 21 healthy mongrel dogs of either sex weighing

5-6 kg . Under ether anaesthesia a metal cannula was implanted into either of the lateralcerebral ventricles (BHARGAVA and TANGRI, 1959) . Three weeks later the animals wereanaesthetized with i.p . pentobarbitone sodium (35 mg/kg) . Half an hour after induction ofanaesthesia the electrocardiogram (ECG) of these animals were recorded using needleelectrodes . The effect of 1, 2, 5 and 10 haemolytic units (HU) per kg of streptolysin O, in0-5 ml sterilized isotonic phosphate buffered (pH 7-2) saline (1 HU is equivalent to 1minimum haemolytic dose using a 50% haemolysis end point, i .e . MHD50), injectedintraventricularly at 30 min intervals on the ECG was studied . Similar experiments werealso performed in dogs subjected to bilateral cervical vagotomy and/or spinal transectionat C1. In a parallel set of experiments the effect of the same doses of streptolysin O injectedi .v . in intact, vagotomized and/or spinalectomized animals was also investigated . Animalssimilarly receiving streptolysin O inactivated by prior treatment with various doses ofantistreptolysin O (Wellcome Reagents Ltd., England), cholesterol (37 °C, 30 min) or heat(60°C, 30 min) served as control . Three animals were used for each test . Animals weremaintained under standard conditions and were never reused earlier than 3 weeks.

Partially purified, reduced and lyophilized streptolysin O and antistreptolysin O (Well-come Reagents Ltd., England) were reconstituted in sterile isotonic phosphate bufferedsaline (pH 7-2) just before use . This streptolysin O contained 3500 HU/mg protein and wasapproximately 96 % in the reversibly oxidised state . Polyacrylamide disc gel electrophoresisrevealed a major protein band (streptolysin O, 92%) followed by a second minor band(8 %) . Antistreptolysin O revealed a single precipitin band against streptolysin O in immune

389

390

Short Communications

FIG. 1 . ELECrROCARDIOGRAPHIC CHANGES AFTER INTRAVENTRICULAR INIECnON OF STREPTOLYSINO (10 HU/kg) IN ADOG.

Essentially similar changes were recorded after i .v . injection of streptolysin O. Bilateral

cervical vagotomy and/or spinal transaction at C, did not alter these changes .

precipitation electrophoresis . The cholesterol suspension was prepared by the method ofCOHEN et al. (1937) ; 0 " 1 Vg of cholesterol completely inhibited 1 HU of streptolysin O .The intraventricular administration of streptolysin O in doses of 1 and 2 HU/kg did not

produce any ECG changes . However, in all animals a dose of 5 HU/kg produced, within60-90 sec, a reduction of 34 ± 3 % in the heart rate ; this effect was transient and com-pletely reversible within 2-3 min after the injection . In higher doses (10 HU/kg) it producedprogressively increasing bradycardia and idioventricular rhythm. Elevation of ST segmentwas observed in all the animals ; cardiac stand-still ensued within 6-8 min after the injection .These changes are shown in Fig. 1 . Intravenous injection of streptolysin O (10 HU/kg) indogs produced, within 45-60 sec, marked sinus bradycardia (^60% decrease in heart rate)followed by other ECG changes as observed after intraventricular administration ofstreptolysin O . Bilateral cervical vagotomy and/or spinal transaction per se had no effecton the ECG of dogs, and the ECG changes produced by intraventricular or i.v . admini-stration of streptolysin O were not altered by these procedures . Animals receiving anti-streptolysin O and cholesterol treated streptolysin O injected intraventricularly or i.v.showed cardiotoxic effects in proportion to the residual haemolytic activity of these pre-parations ; heat inactivated streptolysin O had no effect .

Similar ECG changes have also been reported after intraventricular administration ofstreptolysin O in rabbits (GUPTA and GUPTA, 1977b) . Since bilateral cervical vagotomyand/or spinal transaction failed to alter these effects, the cardiotoxic effects ofstreptolysin Oinjected intraventricularly do not appear to be centrally mediated . They may be due to adirect cardiotoxicity subsequent to peripheral leakage of this toxin . Demonstration ofessentially similar ECG changes after i .v . injection of streptolysin O in intact, vagotomizedand/or spinalectomized animals is consistent with this suggestion . Streptolysin O pre-treated with antistreptolysin O, cholesterol or heat (agents which inhibit streptolysin O,

Time Electrocardiogram Rate P-R Interval ST Segment Remarks

(see) (lead I) (per "in) (Sec)

:: ._ ._ .- ._ ..t. Pseudo ST.. . .

214 .04 depression Normal EC6j r (due to high pattern

heart rate)

90 . .. - y -- 60 .05 Slightly Sinuselevated bradycardia

150 40 .05 Elevated Sinusbradycardia

Idio-

240 - 26 - - ventricularrhythm

O _ _ Cardiac360 - -- stand-still

Short Communications

391

THomPSON et al., 1970) failed to produce any changes in the ECG suggesting that thecardiotoxicity observed in animals receiving active streptolysin O, wasprobably due to thistoxin and not to other streptococcal components contaminating streptolysin O.

Acknowledgement-We are grateful to Indian Council of Medical Research, New Delhi, India for financialsupport.

REFERENCES

BHARGAVA, K. P. and TANGRi, K. K. (1959) Central vasomotor effects of 5-hydroxytryptamine. Br. J.Pharmac. Ther. 14, 411 .

COHEN, B., SCHWACHMAN, H. and PERKINS, M. E. (1937) Inactivation of pneumococcal haemolysin bycertain sterols. Proc. Soc. exp. Mol. Med. 35, 586.

GuPTA, R. K. (1979) Effect of intracerebroventricular administration of streptolysin O on the behaviour ofunanaesthetized rabbits and dogs . Toxicon 17, 167.

GuPTA, S. and GuPTA, R. K. (1977a) Studies on toxic effects of streptolysin O : electrocardiographic changesin mammals. Indian J. Physiol. Pharmac. 21, 199.

GuPTA, R. K. andGuPTA, S. (1977b) Effect of intracerebroventricular administration of streptolysin O onECG of rabbits. I.R .C.S . Med. Sci. Lib. Compend. 5, 430.

GuPTA, R. K. and SRIMAL, R. C. (1979) Effect of intraventricular administration of streptolysin O onelectroencephalogram of rabbits. Taxicon 17, 321.

GuPTA, R. K., MEHRoTRA,R. M. L. and CHATURVEDI, U. C. (1973) Cardiotoxicity of streptolysin Oin rats .Indian J. med. Res. 61, 1584.

GuPTA, R. K., GuPTA, S., VARMA, D. R. and ATAL, P. R. (1976a) Studies on toxic effects of streptolysin 0 :production of experimental pancarditis in rats . Indian J. exp. Mot. 14, 242.

GuPTA, S., GuPTA, R. K. andVARMA, D. R. (1976b) Effect of streptolysin Oon myocardial contractility ofmammalian heart. I.R.C.S. Med. Sci. Lib. Compend. 4,120.

GuPTA, S., GuPTA, R. K. andVARMA, D. R. (1976c) Studies on toxic effects of streptolysin O: effect on thecontractility of isolated and intact mammalian and amphibian heart. Indian J. Physiol. Pharmac. 20, 164.

RASKOVA, H. and VANECEK, J. (1957) Effect of intraventricular administration of streptolysin O in un-anaesthetized cats . Science 126, 700.

THOMPSON, A., HALBERT, S. P. and SMrrH, U. (1970) The toxicity of streptolysin O for beating mammalianheart cells in tissue culture. J. exp. Med. 131, 745.