Neurological, respiratory and cardiac effects of cardiac glycosids administered intracerebrally to conscious mice

Pharmacological Research Communications, Vol. 6, No. 5, 1974

NEUROLOGICAL, RESPiPJ~TORY J~D CARDIAC EFFECTS OF CARDIAC GLYCOSIDS ADI;~{ISTERED INTRACEREBRALLY TO C O N S C I O U S

I,IICE

BY A..~i. AFIFI* Ph.D. and E.M. A~.~R, Ph.D. Department of Pharn~cology, Faculty of ~'iedicine, University of Assiut, Assiut, Egypt. Received 1 October 1973

SUI,~,tARY

The toxic doses of ouabain, digitox~n and digoxin after their intravenous and intracerebral injections in mice arc obtained and compared. Neurological, respiratoz~ and ECG changes following their systemic and intracerebral administration are studied in conscious mice. ?~here as systemic aamqnistration leads primari]~ to cardiac toxicity and death the intracerebral administration leads to net~ological changes including convulsions, initial respiratory stimulation followed by depression and also certain ECG changes. Death from intracerebral injection of these drugs is primarily due to convulsions and respiratory failure° The implication of the central action of these glycosides is discussed. I N TRODUCTI ON Our knowledge of the frequency s~d types of non-cardiac signs during digitalis toxicity is much scarcer than the cardiac ones.

In a number of reports

(Chm~g, 1969; Lely, 1970;

Lely, 1972 and ~deyler, 1972) where the extra-cardiac siLgas of digitalis intoxication are mentioned, neurological signs are always included.

This has stimulated the interest of a number

of authors (Haley, 1955; Idelville, 1957; Doggett, 1970, 1971, and 1973) to study the direct effect of injecting or applying minute amounts of cardiac glycosides into the brain of experimental animals. Z]

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Of interest also is the demonstration of j

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* Communication to A. ~i. Afifi

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417

Pharmacological Research Commun/cat/ons, Vo/. 6, No. 5, 1974

418

producing aardiac arrhytl~nias when cardiac glycosides are injecteded into tl~e thi1~d and fot~th ventricles of dogs (Haley, 1955 o~d Bircher, 19G3), ¢he lateral ventricles of cats (I,~elville, 1957), and rabbits (Ammar). It is the aim of this work to study and compare the cardiac, respiratory and neurological manifestations of digitalis intoxication in mice injected either systemically or intracerebrally ~,~th tltree cardiac glycosides: ouabain, digitoxin and digoxin. The intracerebral injection was done to exclude certain factors as the cardiovascular effects of large doses injected systemically and the blood-brain barl-ier, which may contribute to or hinder the appearance of the central actions of these glycosides. iiATI';RIALS i~D I LETIt0DS Solutionsouabain,

In t!~is study

a]ao~,~n cmd d i g i t o ~ i n

0uabain were used

was u s e d

solution

were prepared

f~b_ite m i c e w e i g h i n g were injected

injection

or into

injection

using

for

D i g o x i n and d i ~ i t o x i n

15

the

lateral

tail

vein for

ventricles

for

microsyr~,ge

localizing

from which differ-

30% a l c o h o l .

1 8 - 2 5 gm w e r e u s e d .

into

the

contain

were used:

Paris).

i n 7CY% a l c o h o l

to

50 t u l - H ~ i l t o n

H a l e y e~t __~ ( 1 9 5 7 ) mice.

(Nativelle,

as aqueous solution.

as alcoholic

ent dilutions utions

tt~ree c&rdiac glycosides

The g l y c o s i d i c intravenous

sol-

(i.v.)

,

intracerebral

(i.c.)

and the method

of

the ventricles

in

conscious

The LD50 was determined after injecting the drugs both i.v. or i.e. by Pershins' method ( Pershin, 1950). At least 30 mice were used in each determination. Behavioral changes were observed in mice during the determination of the LD50. Electrocardio&~aphic and respiratory changes were recorded in a number of mice following the i.v. and i.c. injection of the respective LD50 dose of the glycosides. An E&~ Physiograph was used to record the ECG and respiratory changes.

419

Pharmacological Research Communicatfons, Vol. 6, No. 5, 1974

Standard lead II was recorded using hypodermic needles as electrodes

(Ammar, 1968).

Respiratory ehahges were recorded

by inserting t~o needle electrodes across the chest of the mouse and connected tkrough a pneumotransducer to one of the p1%ysiograph amplifiers. For i.v. injection a,volume of 0.2-0.5 ml per 20 gm mouse was used.

For i.¢. administration a volume of 20 ul per 20 gm

mouse was injected.

In both cases control animals were injected

i.v. or i.c. with the same volume

of the vehicle.

RESULTS Table 1 pl-esents the systemic(i.v.)

LD50 , the intracerebral

(i.c.) LD50 and the ratio of the two values (i.v./i.c.) for each of ouabain, digoxin and digitoxin. It is quite evident that the i.e. t o , city resulted from a much smaller dose c o m p a r e d w i t h the systemic toxicity dose. Table 1 The LD50 of Digitoxin,

Ouabain and Digoxin following their intra-

venous (i ~v._~ _and intracerebral ~i~o.l . . . . . ln3ectmon in conscious mice L " " "

.oru

......

)

Ratio

..... i ~ . O ,

i.v./i .c.

'c mg/kg

..v.

2.334

0.241 0.059

33.82

7.670

0.124

61.85

Digitoxin

6.900

Ouabain Digoxin

28.63

E_;mVIORAL OliAN~-7~S Intrac erebe ral admini s trati on The same type of behavioral chs~ges was observed with the three glycosides with only some differences in the severity of the toxic manifestations and the type of the eonvuisive fits. Immediately following the injection of the animals they remained quiet in a fixed position for about one minute.

The

same type of response was obta~led when control mJ ce were inject-

420

Pharmaco/ogica/ Research Communications, Vo/. 6, No. 5, 1P74

e d with saline or dilute alcohol. ~.liceinjected ~'~ith cardiac glycosides exhibited tachypnea, Straubs' tall and became highly excitable for any stimulus. After aboGt three~,znmautes they started a very peculiar phenomenon which was cons~'s\tant ~ all the tested anJ~n~So The mice started circling at f~r~st in narrow circles which becs me w i d e r and wider U n t i l it in~volved \ the whole laboratory space. Control auimals did not 'sho~\this phenomenon. I n t e m i t t e n t periods of excitation and depression were observed. Other chauges included licking, Sczatc~z[ng, catatonia, ptosis and loss of righting and pain reflexes. After about 5 minutes the nL'~ce started conv~Isive fits, tonic in case of ouabain and clonic ending with tonic convulsions in ease of digox~u and digitoxin. ~he number of convulsive fits-\were found to increase with increasing the ose of the .glycosides. Death was found to follow the convulsive ~its in about 5-7 minute s. ~[ice injected intracerebrally with saline o ~ l u t e ,a~cohol ( 2 ~ ) never exhibited auy of the mentioned b e h ~ v i o r a l ~ h a n g e s v~th the exception of some depression and bradypnea in ~~°~ase6f alcohol. No convulsive fits were oSserved in control a n i m a ~ and recovery was always complete. Intravenous a~m~ nistration Death occured after few minutes in =~ce injected intravenously v~th the three glycosides v~thout noticing any of the behavioral changes. However, excitation and convulsive fits occured occasionally in ouabain toxicity. E C G CHA~TGES I ntracerebral administration The intracerebral injection of lethal doses of ouabain resulted into an initial bradycardia and heart block followed by an increase in the heart rate with subsequent complete heart block and finally death of the mice within 6-8 m~uutes (Fig. 1). When alcoholic solution of digoxin and di~itoxin were injected intracerebrally with lethal doses initial brief tach~cardia followed by bradycardia and heart block preceeded the

Pharmacological Research Communications, Vol. 6, No, 5, 1974

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The effect .of~intrt',cerebral(fJ.c. )

ac~minfs,%ration of the lethal

inconScious mie,e on ECG and resl)ir&tion,, d~atA of the a n ! ~ I s Which eccured', ~vithin,.k,5-10\minutes. ~CG"o~l'~tl~h d 0 ~ s . Charac~erlswlo diglwa~Is,to "xiol~Y, was seen~leluding initial bradyeardia ~and nodalx rhythm followed by extrasystole, multifocal ven~ricular tachyc~rdi a and_,death of the: animals ~ " froT,~ c o m p l e t e

heart

block

a~'te

m~uu~'es

(Fig.

2).

Pharmacological Research Communications, Vol. 6, No. 5, 1974

422

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The effect of intrav~nous(i.v.) administration of the lethal dose ( i . v ) of ouabain, digoxin, dl~itOXln and alcohol in mice on E C G and respiration. •

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Control animals injected with amounts of alcohol equivalent to what is present in the vehicle of digoxin and digitoxin showed only b r i e f tachycardia followed by bradycardia and finally normalization

of heart rate.

R E S P I R A T O R Y CHANGES I ntracerebral administration '"

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Solutions of ouabain,

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digoxin and digitoxin injected intra-

cerebrally were found to lead to noticeable hyperpnea snd tachypnea followed by shallow respiration and finally re spirat or,# arrest

(Fig.l)

423

Pharmacological Research Communications, Vol.. 6, IVo. 5, 1974

Dilute alcohol (20%) injected into the ventricles of control mice lead only to a slit%it slJ xn~ and increase in the depth of l-espiration. Normalization of the respirator~r rate and delgth took place after about one minute.

Noz~al sal~nle injected in

~is" manner lead to brief si~alloY/ and irregular resplra"'~zon fol!o~zea by norT~alization. I ntravenous achuinistr~tion Lethal doses of ouaoa]n injected intravenously resumted into bradypnea and hyperpnea and finally respiratol~r arrest with the simultaneous heart block and death of the animals (Fig.2). Alcoholic solutions of digoxin and digitoxin injected intravenously lead to an initial increase in the rate of respiration followed by irregular rate and finally respiratory failure. p I_SCUS $ I OIj The intravenous injection of the "three cardiac glycosides in rmice leads to th@ characteristic digitalis toxicity seen in different species of animals.

Lethal doses of these drtkgs lead

to death of the snimals mainly due %o cardiac toxicity. ~h/rthermore, the intraeerebral injection of these cardiac glycosides leads to ECG changes similar to what have been reported in dogs (Bircher,1963) and in rabbits (Ammar) which 8,re exp!ained by these authors to be due to a central autononLic stimulation . Our finding in mice that the LDao on intracerebr~l injection of these glycosides are far sntaller than the intravenous LDaO may also implicate the central mechanism of action of these drugs. In favour of t}kis thesis is the absence of be]~avioral, resi~ii-story and neurological choruses on their systemic administration.

Hov¢-

evel-, cardiovascular toxicity w}~ich is the main cause of death follovTing their systemic ~dministration, may occur early enou~q to obscure the al~pearance of these central manifestations. On the other hand, the death of mice injected intracerebraliF is apparently the result of respirator-j arrest either due to the eonv~[Isant effect of these dru~'s or their direct effect on the respiratory center Or both.

Pharmacological Research Communications, Voi. g, IVo. 5, 7974

424

The work of Doggett (1970,1971)

indicate

that doses higher than 0.4 ~

injected intracerebrally produce

depression

e~t a_~l (1970) and Do~gett and Spencer of ouabain

are excitatory while doses below 0.4 ug

of the central nervous

a more recent investigation

Doggett

activity

and Spencer

ed the action

of centrally injected

gic mechanism

as well as to a component

of mice.

In

(1973) attribut-

ouabain to both a dopaminerof dopamine receptor

blockad ~ . Our results also indicate glycosides

a dual central action of three

when injected intracerebrally

Small doses show predominent

b e h a v i o r a l depression

large doses tend to ex/~ibit behavioral ion.

while

and neurological

stimulat-

~-~ven at the high dose levels intel~littent periods

of excit-

ation and depression r~bits

in mice.

are seen.

EEG and behavioral

(Ammar et a l) also indicated

excitation

when relatively large doses of ouabain ed into the rabbits' ventricles.

and depression

(5-10 t~g/kg) were inject-

The ECG changes which are recorded in mice injection may also indicate

changes in

the central origin

on intracerebral of these

changes.

Cardiac arrl~rthi~ias are also reported to occur when cardiac glycosides are injected into •~'~ -" ~.,~ '~_l±~ and fourth ventricles of dogs (Haley, 1955 anc~ ~ Bircher,1963), the lateral ventricles of cats (i~elville, 1957) ant: rabbits ~ a r ) . In conclusion

the ECG, respiratoz~j and behavioral

which are seen on intracerebral are t o

a great extent

ate that these central a common mechanism

injection

of the same t~pe.

cnan,,e" ~ s

of these glycosides Otu~ results also indic-

changes may be due to a specific and

of action intrinsic

The exact localization of the site ~ d need f ~ t h e r ~ v e s t i g a t i o n .

to this group of dru~s. mechanism

of action-still

Pharmacological Research Communications, Vol. 6, No. 5, 1974

425

REFER~NCES

Ammar, E. M., (In press) Ammar, E. ~il., Afifi, A. M. and Z ohdy A. M., (In press) Ammar, E. M. and Kudrin, A. N.,(19Ggl, Farmakol .i. Toxicol.

( USSR ), 5 , 566 Bircher, R. P., Kanai, ~. and Wang, S. 0.,(1963) J. Pharmaccl. Exper. Therap., 141,. G Chung, E. K. ,(1969) Digl~%-ails Intoxication, Excerpta ~ledica Foundation, Amsterdam, pp 44 Doggett, N. S. and Spencer, P. S. J.,(1971) Brit. J. Pharmacol., D ~ 42, 242 og~ett,--~. S. and Spencer, P. S. J.,(1973) Brit. J. Pharmacol., D ~ 47, 47 o~gett,~[. S., Spencer, P. S. J. and Turner, T. A. R.,(1970) Brit. J. Pharmacol., ,~.0, 138P Haley, T. J. and ~IcCormick, G.,(1957) Brit. J. I~srmacol., 12, 12 Haley, T. G. and Weinberg, S. J.,(1955) Proc. Soc. Exp. Biol. Med. ,a~84, 345 Lely, A . H . van Enter, 0. H. J.,(1970) Brit. }~led. Journal, , 737 Lely, A~ H. and van Enter, C. H. J. ,(1972) Amer. Heart Journal, 83, 149 ~lelville, K. I. and Shister, H. E.,(1957) Amer. Heart Journal, ~;leyler, and Peck, H. M., (1972) Drug Induced Diseases, Excerpta ~[edica Foundation, Amsterdam, vol. 4, PP 345 Pers1~in, G. N.,(1950) 2armakol .i. Toxicol. ( USSR ), ~, 53