BRAIN DAMAGE DUE TO ASPHYXIA: MECHANISM OF CAUSATION

B R A I N DAMAGE DUE TO A S P H Y X I A ; MECHANISM OF CAUSATION Ronald E. Myers, M . D . / Ph.D.

Why do the nerve c e l l s of fetuses or newborns d i e or become destroyed äs a consequence of t h e f r exposure to oxygen d e f i c i e n c y states? The oldest and s t i l l most w i d e l y accepted I n t e r p r e t a t i o n of the mechanlsms that u n d e r l l e both the loss of nervous System f u n c t l o n and the development of b r a l n I n j u r y w l t h oxygen d e p r i v a t Ion i s a presumed d e f i c i e n c y In energy

a v a i l a b i l i t y to the c e l l s r e q u l r e d to support v i t a l c e l l u l a r processes ( 2 / 13). A s l m i l a r mechanism has been poslted to e x p l a l n the death of c e l l s In other parenchymal organs ( 3 ) . However/ recent s t u d l e s from a number of l a b o r a t o r l e s have ralsed serlous doubts that a d e f l c i t of energy a v a i l a b i l i t y can account for the loss of nervous system f u n c t l o n w i t h

oxygen d e f l c i e n c y (l, HO w h l l e other results obtained in our own laboratory have demonstrated a lack of c o r r e l a t l ö n between

the a v a i l a b i l i t y of h i g h energy phosphate d u r i n g exposure to a v a r l e t y of oxygen d e f l c i e n c y states and the development of b r a l n pathology. Rather/ our r e s u l t s e s t a b l i s h a close

c o r r e l a t i o n between the tissue content of l a c t i c acid at the end of exposure and whether or not b r a i n i n j u r y w i l l develöii (8/ 9/ IQ, 12). These results are now described.

A reduced oxygen a v a i l a b i l i t y to an a n i m a l or to man causes redox changes in the b r a i n äs a consequence of an

i m p a i r e d o x i d a t i o n of cytochrome oxidase - the f i n a l l i n k in the electron transport c h a i n . An i m p a i r e d o x i d a t i o n of

cytochrome oxidase/ in t u r n / leads to an i m p a i r e d o x i d a t i o n of a l l t h e various antecedent l i n k s i n t h e electron transport c h a i n i n c l u d i n g n i c o t i n a m i d e a d e n i n e d i n u c l e o t i d e / t h e water s o l u b l e o x i d a t i o n - r e d u c t i o n co-factor essential for many enzymatic t r a n s f o r m a t i o n s that take place in many m e t a b o l i c pathways important for c e l l u l a r bioenergetics. Thus/ the

a l t e r e d redox state brought about by oxygen d e f i c i e n c y reduces the a c t i v i t y of the electron transport c h a i n I t s e l f and

increases both the absolute concentration and the Proportion of a v a i l a b l e n i c o t i n a m i d e adenine d i n u c l e o t i d e that is present

in its reduced state. An i m p a i r e d electron transport is associated w i t h major reductions in ATP production and/ thus/

major d e f i c i t s in energy a v a i l a b i l i t y to the c e l l . The malntenance of major portions of a v a i l a b l e

n i c o t i n a m i d e adenine d i n u c l e o t i d e and the electron transport c h a i n in a reduced state also leads to major impairments i n c i t r i c acid cycle f u n c t i o n because of a lack of hydrogen

acceptors at the various dehydrogenase Steps äs orte progresses from i s o c i t r ä t e / a-ketoglutarate/ succinate/ and malate. The maintenance of large amounts of n i c o t i n a m i d e adenine

d i n u c l e o t i d e i n t h e reduced state also i n h l b i t s t h e o x i d a t i v e decarboxylation of pyruvate thereby I n t e r f e r l n g w i t h the

f o r m a t i o n of acetyl-coenzyme A and f u r t h e r reducing c t t r l c . acid cycle f u n c t i o n . R a t h e r / the presence of n i c o t i n a m i d e a d e n i n e d i n u c l e o t i d e p r i m ä r ! l y in the reduced state favors the reduction of p y r u v i c a c i d to l a c t i c a c i d . As a consequence of a l l these a l t e r a t i o n s i n b i o c h e m i c a l f u n c t i o n / a l l a v a i l a b l e carbohydrate i n the tissue i n c l u d i n g that present äs free

0300-5577/81/0091-0021 $ 2.00 Copyright by Walter de Gruyter & Co.,

glucose and that present äs glucose polymerlzed to glycogen l s converted to l a c t l c a c l d . T h l s process (of anaeroblc

g l y c o l y s i s ) I s g r e a t l y accelerated by the loss of the b r e a k l n g effect that f s normal1y exerted by the c e l l u l a r ATP when

present at h f g h concentratIons actlng at m u l t i p l e p o f n t s along the Embden-Meyerhoff pathway. The major consequence of t h i s Pasteur effect Is to Increase the rate of carbohydrate

breakdown to l a c t i c acld by a factor äs great äs seven ( f c ) . The o v e r a l l effect of a l l these processes Is to lead to a near-total breakdown of all a v a l l a b l e free glucose and of glucose e q u i v a l e n t s present In the form of glycogen to l a c t t c acld over the f l r s t few mlnutes of exposure to c l r c u l a t o r y a r r e s t or to another form of anoxia,

The b r a i n t l s s u e of monkeys n o r m a l l y contalns free

glucose at a concentratlon of 3 to 5 umoles/g and polymerlzed glucose I n the form of glycogen also at a concentratlon of 3

to 5 umoles/g. Taklng Into account the l to 3 umoles/g of l a c t i c acid a l r e a d y present In the ttssue, the converslon of a l l a v a i l a b l e tissue carbohydrate to l a c t i c acld leads to a

local a c c u m u l a t i o n of l a c t i c a c i d to concentratIons of Ik to 25 umoles/g (11, 15).

The complete o x i d a t t o n of one molecule of glucose to 6 molecules of carbon d i o x t d e and 6 molecules of water leads to a -686.0 k i l o c a l o r l e s per mole free energy change. In

contrast to t h i s / the breakdown of the same glucose molecule to 2 molecules of l a c t i c acid is associated w l t h a free energy change of o n l y -U7.0 k i l o c a l o r i e s per mole. Thus, convertlng a v a i l a b l e glucose to l a c t i c acid rather than o x l d l z l n g it completely to carbon d i o x t d e and water does lead to a marked energy d e f i c i t despite the considerable Increase In rate of g l y c o l y s i s brought about by the decreased a v a l l a b l l l t y of ATP and the d i s i n h i b i t l o n of s p e c l f l c enzymes at a v a r l e t y of c r i t i c a l control points.

These consideratIons reveal that a reduced oxygen

a v a i l a b i l i t y to tissue causes 1) a major reductlon in energy a v a i l a b i l i t y to tissue to support v i t a l c e l l u l a r processes and 2) an a c c u m u l a t i o n of l a c t i c a c i d both In the tissue and in the c i r c u l a t i n g blood. The f i r s t of these Inferences is

c o n f i r m e d by f i n d l n g marked reductions in the tissue Contents of ATP and phosphocreatine äs a r e s u l t of exposure to any one of the oxygen d e f i c i e n c y states. Those I n v e s t l g a t o r s who have

raised doubts whether such reduced energy a v a i l a b i l i t y can be i n c r i m i n a t e d äs the cause for f u n c t i o n a l central nervous

System d i s t u r b a n c e s developing d u r i n g exposure to oxygen d e f i c i e n c i e s p o i n t out these disturbances can develop under circumstances where the energy a v a i l a b l e to the b r a i n in the form of h i g h energy phosphate compounds has declined only to a m i n o r degree ( o f t e n after a reduction of less than 2 Q % ) ( 1 ,

l**). However, those tissue ci rcumstances that lead to

Pathologie change are r e a d i l y and r e p r o d u c i b l y separable and d l f f e r e n t from those that lead to d l s t u r b e d f u n c t l o n . Thus/

i t is q u i t e p o s s i b l e to restore a normal nervous System f u n c t i o n and to l a t e r demonstrate an i n t a c t b r a i n In a n i m a l s f o l l o w i n g exposures to periods to c i r c u l a t o r y arrest or anoxia

that far exceed those d u r a t i o n s that are r e q u i r e d to produce a complete but temporary loss of nervous System a c t l v l t y (5/ 6/ 7)

Studies we have c a r r l e d out over the last seypral years i n c o l l a b o r a t l o n w l t h Dr. M i c h i o Yamaguchi have polnted out a lack of c o r r e l a t i o n between the behavior of the h i g h energy Phosphate compounds äs e x e m p l l f l e d by the levels of ATP

present In the b r a i n tlssue and the subsequent development of b r a i n pathology whether or not the b r a i n subsequently develops pathology or not. In a f l r s t study, rhesus monkeys were

exposed to 10 m i n u t e episodes of c i r c u l a t o r y arrest (11).

Some of the a n i m a l s were p r e v i o u s l y infused w i t h p h y s f o l o g i c a l s a l i n e S o l u t i o n s w h i l e others were infused w i t h 2 5 % gtucose Solutions leading to elevations of t h e i r serum glucose

concentratIons to values äs h i g h äs 650 mg %. P r i o r studies have c l e a r l y i n d i c a t e d that monkeys that have been

food-deprived for 2k hours and infused w i t h s a l i n e Solutions can undergo exposure to periods of c i r c u l a t o r y arrest that

last for äs long äs 1k m i n u t e s w i t h o u t developing any

neurologic abnormal I t i e s or any gross or microscopic f i n d i n g s o f b r a i n damage w h i l e s i m i l a r a n i m a l s i n f u s e d w i t h glucose S o l u t i o n s l e a d i n g to o n l y s l i g h t elevations of t h e i r serum glucose concentrations undergo a grave neurolpgical

d e t e r i o r a t i o n beginn!ng several hours after they are

resuscitated and t e r m i n a t i n g in t h e i r deaths in 100% of cases many hours later (12). The bar graphs depicted in FTgure l

ATP CONTENT OF CEREBRAL CORTEX AFTER 10 MINUTES OF CIRCULATORY ARREST

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CIRCULATORY CIRCULATORY ARREST ARREST

B r a i n t i s s u e concentrations of adenosine t r i p h o s p h a t e (ATP) in food-deprl ved control a n i m a l s and in a n i m a l s exposed to 10 m i n u t e s of c i r c u l a t o r y arrest f o l l o w i n g pretreatment w i t h s a l i n e or glucose i n f u s i o n s (Myers and Yamaguchi/ 1976).

i n d l c a t e the food-deprIved monkeys that were infused e i t h e r w i t h s a l i n e or w t t h glucose s o l u t t o n s before they were exposed

to c i r c u l a t o r y arrest both experlenced major reductlons in t h e i r cerebral c o r t i c a l ATP contents s measured at the t e r m i n a t i o n of exposure· The reductlons i n ATP were

s l g n i f I c a n t l y greater i n t h e a n i m a l s pretreated w i t h s a l i n e than in those pretreated w i t h glucose i n f u s i o n s even though the former would be expected to s u r v l v e b r a i n - i n t a c t w h i l e the l a t t e r , in 100% of cases, would be expected to undergo a grave neurologic d e t e r i o r a t i o n and die w i t h major widespread b r a i n pathology. These f i n d i n g s show a lack of c o r r e l a t i o n between the m a g n i t u d e of reduction in the ATP content of the cerebral cortex and whether or not the a n i m a l s subsequently develop b r a i n pathology.

A s i m i l a r lack of c o r r e l a t i o n between the extent of

reductions in h i g h energy phosphate d u r i n g exposure to oxygen d e f i c i e n c y and appearance of b r a i n pathology is i l l u s t r a t e d in

the bar graphs of F i g u r e 2. Food-deprived rhesus monkeys were

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F i g . 2: Adenosine t r i p h o s p h a t e ( A T P ) contents of cerebral cortex of young a d u l t control monkeys and of monkeys a f t e r exppsure to 10 m i n u t e s of tracheal o c c l u s i o n ( a n o x i a ) / 10 m i n u t e s of c i r c u l a t o r y arrest ( a n o x i a ) , and 10 and 25 m i n u t e s of 3.5% oxygen b r e a t h i n g (marked h y p o x i a ) (Yamaguchi and Myers, 1976).

exposed to k d i f f e r e n t c o n d l t l o n s of oxygen d e p r l y a t i o n

I n c l u d l n g : 1) 10 m i n u t e s of tracheal occlusion, 2) 10 minutes of c i r c u l a t o r y arrest, 3) 10 m i n u t e s of b r e a t h i n g 3*5% oxygen

in n i t r o g e n and k) 25 m i n u t e s of b r e a t h i n g the same hypoxic gas m i x t u r e (11). P r i o r work in our laboratory has

demonstrated that o n l y the a n i m a l s of category k exposed to 25 m i n u t e s of marked hypoxia w i l l develop neurologic

abnormal i t i e s and many w i l l d i e several hours later w i t h b r a i n edema or s u r v i v e but w i l l show focal b r a i n i n j u r y . Exposure to 10 m i n u t e s of tracheal occlusion or of c i r c u l a t o r y arrest both of w h i c h are w e l l tolerated by the food-deprIved a n i m a l s depresses the c o r t i c a l tissue ATP contents far more than does exposure to e i t h e r 10 or 25 m i n u t e s of marked hypoxia. Thus, once again, the development of b r a i n pathology f a l l s to f o l l o w the b e h a v i o r of h i g h energy phosphate compounds. Furthermore, the a n i m a l s exposed to marked hypoxia for 10 and 25 minutes, though they show no s i g n i f i c a n t d i f f e r e n c e s in t h e i r cortical ATP contents d u r i n g exposure and at the t e r m i n a t i o n of

exposure/ they do show marked d i f f e r e n c e s in b r a i n pathologic outcome. The a n i m a l s exposed for 10 m i n u t e s of hypoxia

s u r v i v e and do w e l l w h i l e those exposed for 25 m i n u t e s develop marked b r a i n i n j u r y and die in large Proportion. Thus, in the

number of circumstances depicted in F i g u r e s l and 2 no

c o r r e l a t i o n could be demonstrated between the ATP content of the cerebral cortex d u r i n g exposure to oxygen d e f i c i e n c y and whether or not the a n i m a l s would develop c o r t i c a l i n j u r y .

In contrast to t h i s lack of c o r r e l a t i o n between the

behavior of tissue ATP and the development of b r a i n pathology is the circumstance of l a c t i c acid and its a c c u m u l a t i o n in the b r a i n d u r i n g exposure to the oxygen d e f i c i e n c y states« The extents to w h i c h l a c t i c acid accumulates in the b r a i n tissue

in the a n i m a l s e a r l i e r described w h i c h were food-deprived for 2k hours, g i v e n i n f u s i o n s of s a l i n e or glucose Solutions, and then exposed to 10 m i n u t e s of c i r c u l a t o r y arrest is described in the bar graphs of F i g u r e 3 (11). The a n i m a l s pretreated wi th s a l i n e i n f u s i o n s and exposed to 10 m i n u t e s of c i r c u l a t o r y arrest ( a l l of which would s u r v i v e b r a i n - i n t a c t ) accumulated

l a c t i c a c i d to an average concentration of 12 umoles/g, tissue.

In contrast, t h e a n i m a l s pretreated w i t h glucose i n f u s i o n s ( a l l of w h i c h would later undergo a grave neurologic

d e t e r i o r a t i o n and d i e w i t h b r a i n edema and widespread

necrosis) accumulated l a c t i c acid to concentrations in excess of 30 umoles/g. The basis for t h i s marked d i f f e r e n c e in

behavior w i t h respect to l a c t i c acid a c c u m u l a t i o n relates to the carbohydrate state of the a n i m a l s and t h e i r p r i o r h i s t o r y of food intake or of i n f u s i o n of glucose. The I n f u s i o n of glucose S o l u t i o n s or the recent I n g e s t i o n of a meal h i g h i n carbohydrates s i g n i f i c a n t l y increases the b r a i n content of free glucose. The exposure to c i r c u l a t o r y arrest or to total a s p h y x i a causes all a v a i l a b l e free glucose and also a l l

glycogen in the b r a i n tissue to break down to l a c t i c a c i d . . The extent to which l a c t i c acid accumulates in the b r a i n under these circumstances is s t o i c h i o m e t r i c a l l y related to the

a v a i l a b i l i t y of carbohydrate in all these forms in the b r a i n . For t h i s reason, the p r i o r f e e d i n g of carbohydrate or the i n f u s i o n of glucose S o l u t i o n s into the a n i m a l s p r o p o r t i o n a l l y increases the extent to w h i c h l a c t i c a c i d accumulates in the tissue.

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ARREST ARREST F l g . 3: B r a i n tissue concentrations of lactate i n

food-deprived control a n i m a l s and in a n i m a l s exposed to 10 m i n u t e s of c i r c u l a t o r y arrest f o l l o w f n g

pretreatnent w i t h s a l i n e or glucose I n f u s l o n s (Myers and Yamaguchi, 1976).

The behavfor of l a c t i c acid In the b r a l n tissue in the a n i m a l s exposed to the 2 types of anoxia or the 2 d u r a t i o n s of marked hypoxia äs descrtbed above is i l l u s t r a t e d i n the bar graphs of F i g u r e k (16). > T h e exposure of food-deprlved monkeys to 10 minutes of c i r c u l a t o r y arrest or of tracheal occlusion both of w h i c h is w e l l tolerated causes l a c t i c acid

to accumulate in the cerebral cortex only to 10 to 12

umoles/g. Likewise, the exposure of a n i m a l s to 10 m i n u t e s of marked hypoxia 03.5% oxygen b r e a t h i n g ) also causes l a c t i c acid

to accumulate I n the cortex to mean values close to 10

umoles/g and such an exposure is also w e l l tolerated by these a n i m a l s . However, the exposure of a n i m a l s to 25 m i n u t e s of

to a mean v a l u e of 2U umoles/g. Thus/ In 2 q u i t e separate circumstances, e.g., exposure of glucose-infused a h l m a l s to anoxla and exposure of any a n l m a l s to a prolonged hypoxia, the accumulation of l a c t i c acid in b r a i n tlssue at h i g h

concentrations i s associated w l t h the development of a marked b r a i n pathology.

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F i g . l·: Lactate contents of cerebral cortex of young a d u l t control monkeys and of monkeys a f t e r exposure to 10 m i n u t e s of tracheal occluslon ( a n o x i a ) / 10 rninutes of c i r c u l a t o r y arrest ( a n o x i a ) , and 10 and 25 minutes of 3.5% oxygen b r e a t h i n g (marked h y p o x i a ) ( Y a m a g u c h i and Myers, 1976).

C o n c l u s l o n s : The results of our present s t u d l e s f a l l to show any c o r r e l a t l o n between the magnltude of the energy

d e f i c i t of the b r a l n tissue äs d e f l n e d by I t s ATP contents at the t e r m l n a t l o n of exposure to some form or other of oxygen d e p r i v a t Ion and the later behavlor of the b r a l n w l t h respect to appearance of edema or development of pathology. R a t h e r / our s t u d l e s p o i n t out a close c o r r e l a t l o n between the

a c c u m u l a t i o n of l a c t i c acid In the b r a l n at h i g h

concentratIons (>18 to 20 umoles/g) and the occurrence of b r a i n edema/ widespread t i s s u e necrosts/ and death of the

a n i m a l s . I t is I n f e r r e d that t h e c r l t l c a l d e t e r m l n e r o f b r a l n P a t h o l o g i e outcome in r e l a t l o n to exposure to the v a r l o u s

oxygen d e f i c i e n c y states is the extent to w h i c h l a c t i c a c i d accumulates in the b r a l n t i s s u e . The extent to w h i c h l a c t i c a c i d accumulates i n these clrcumstances/ I n t u r n / i s

d e t e r m i n e d by the a n i m a l s¹ carbohydrate state or t h e l r h l s t o r y of food i n t a k e at the t i m e of exposure.

REFERENCES

1. D u f f y / T . E . / N e l s o n / S . R . / and Lowry/ O . H . : Cerebral c a r b o h y d r a t e m e t a b o l l s m d u r i n g acute h y p o x i a and recovery. J. Neurochem. 19: 9S9-977/ 1972.

2. F e i n / J . M . : B r a l n energetlcs and cerebral death. From:

B r a i n Death: I n t e r r e l a t e d Medlcal and Social Issues.

Ann. N . Y . Acad. Sei. 315: 97-10if/ 1978.

3 . J e n n i n g s / R . B . / H a w k i n s / H . K . / Löwe/ J . E . / H i l l / M . L . / K l o t m a n / S. and R e i m e r / K . A . : R e l a t l o n between h i g h

energy phosphate a n d l e t h a l I n j u r y I n m y o c a r d l a l i s c h e m i a in the dog. Amer. J. P a t h . 92: 187-207/ 1978.

k. Lowry/ O . H . / Passonneau/ J . V . / Massel berger, F . X . / and S c h u l z / D . W . : E f f e c t of i s c h e m i a on known Substrates and cofactors of the g l y c o l y t i c pathway I n b r a l n . J. B I o l . Chem. 239: 18-30/ 196U.

5. M i l l e r / J . R . and Myers/ R . E . : N e u r o l o g i c a l effects of systemic c i r c u l a t o r y arrest In the monkey. N e u r o l . 20:

715-721*/ 1970.

6. M i l l e r / J . R . and Myers/ R . E . : Neuropathology of systemic c i r c u l a t o r y arrest I n a d u l t monkeys. N e u r o l . 2 2 :

888-90U/ 1972.

7. Myers/ R . E . : Neuropathology of total oxygen lack ( a n o x i a ) I n rhesus monkey. In: Cervos-Navarro/ J.

( E d . ) : "Pathology of Cerebral M I c r o c I r c u l a t i o n^{1 1}. B e r l i n / W a l t e r de G r u y t e r / 197l*/ pp. 299-306.

8. Myers/ R . E . : A n o x l c b r a l n pathology and blood glucose.

N e u r o l . 26: 3^5/ 1976.

9. Myers/ R . E . : E x p e r i m e n t a l models of p e r i n a t a l b r a i n damage: Relevance to human pathology. In: G l u c k / L.

( E d . ) : " l n t r a u t e r l n e A s p h y x l a and The D e v e l o p i n g Fetal Brain^{1 1}. New York/ Year Book P u b l . Co./ 1977/ pp. 37-97.

10. Myers, R , E . : L a c t i c acid accumulatlon äs cau?e of b r a i n edema and cerebral necrosls r e s u l t i n g from oxygen

d e p r i v a t i o n . In: Korobkin, R. and G u l l l e m l n a u l t / C.

( E d s . ) : "Advances i n P e r i n a t a l Neurology11, New York, Spectrum Publ., 1979, pp. 85-11U.

11. Myers, R . E . and Yamaguchi, M.: Effects of serum glucose concentration on b r a l n response to c l r c u l a t o r y ärrest.

J. Neuropathol. Exp. N e u r o i . 35: 301, 1976.

12. Myers, R . E . and Yamaguchi, S.: Nervous System effects pf cardiac ärrest in monkey: Preservatlon of v i s f o n . Arch.

Neurol. 3/*: 65-7**, 1977.

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Sauerstoffversorgung des G e h i r n s und den Mechanismus der M a n g e l w i r k u n g e n . Ergebn. P h y s i o l . kB: 126-260, 1950.

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MYERS, Ronald E., M.D., Ph.D.

Chief

Laboratory of P e r i n a t a l Physiology N a t i o n a l I n s t i t u t e s of H e a l t h

Park B u i l d i n g , Room i*51G

Bethesda, M a r y l a n d 20205 / USA