Acidification of interstitial fluid in hippocampal formation caused by seizures and by spreading depression

Brain Reseurct~. 31, ~ 1984 | 1Sty- 1S~

186

[qse~ lcr BRE 20399

Acidification of interstitial fluid in hippocampal formation caused by seizures and by sprea~ng depression GEORGE G. SOMJEN Department o f Physiology, Duke University. Durham. NC ~U.S.A.

(Acceptcd May 22nd. 1984~ Key words: seizure

spreadin~ depression - - pH of tissuc

hippocampus

paroxysmal discharge

ion-selective ctectmde

Changes in the pH of interstitial fluid were measured with H+-selective double-barreled micropipene electrodes in fascia dentata of urethane-anesthetized rats. Paroxysmal afterdischarges provoked by repetitive stimulation of an afferent fiber tract brought in their wake acidification by 0.07 to 0.2 pH units. Spreading depression caused acidification by 0.2-0.5 pH units. Acid shifts were often preceded by transient alkalinization. Acidification is attributed to the production of COo and of other acid metabolites.

It has been known for some time that the extracellular fluid of the brain becomes acid during seizures 1-3,~3. In these earlier studies p H was m e a s u r e d with electrodes in contact with the surface of the brain. Recently it has b e c o m e possible to m a k e H +selective m i c r o e l e c t r o d e s of small enough tip size to be inserted into cerebral tissue enabling measurement of the actual m a g n i t u d e and the time course of p H changes in interstitial fluid. Urbanics et al. 14 used needle-shaped pH-sensitive glass electrodes to measure the changes of p H e v o k e d by direct stimulation of the cerebral cortex. L e h m e n k i i h l e r et al. 5 used antimony as p H - s e n s o r in the tip of micropipettes to record the responses of cerebral cortex during spreading depression. Kraig et a l ? used an H+-selective liquid m e m b r a n e to m e a s u r e the p H in the cerebellar cortex. W e have r e c o r d e d the p H of cortical interstitial fluid during acute acidosis and alkalosis H,12. using a similar liquid m e m b r a n e . This preliminary report concerns p H changes during and after paroxysmal discharges and spreading depression p r o v o k e d in the h i p p o c a m p a l formation, mainly in the fascia dentara, by repetitive stimulation of afferent nerve pathways. A d u l t male rats of 450-550 g weight were anesthetized with u r e t h a n e (1.5 g/kg b.wt., i.p.). The head of the animals was fixed in a stereotax~c frame. The

skull was o p e n e d to permit bipolar stimulating electrodes to be inserted into the p a r t o f white m a t t e r known as the "angular b u n d l e ' through which the fibertract connecting the entorhinal cortex to t h e hippocampal formation passes. A n o t h e r trephine hole permitted the insertion o f the d o u b l e - b a r r e l e d glass microetectrodes into the hilus o f the fascia d e n t a t a . The landmarks for the latter were 4.0 m m posterior to bregma and 2.1 or 2.3 mm lateral relative to the midline. The recording e l e c t r o d e was inserted to a de pth where the focal synaptic potential ( ' p o p u l a t i o n EPSP'); e v o k e d b y a single pulse of 0.1 m s duration and 0.6 m A intensity applied to the angular b u n d l e . was positive and of 10 m V or m o r e in amplitude. This was usually between 3.5 and 4.0 m m from the dorsal surface of the hemisphere. R e c o r d i n g s were started m this position in all experiments: s u b s e q u e n t l y the electrode was m o v e d to o t h e r positions as welt. The d o u b l e - b a r r e l e d microcapillary e l e c t r o d e s were made and filled as described earlier for K +- and CaZ+-selective electrodes 7,9. T h e liquid m e m b r a n e was W P I b r a n d IE-010. Changes in p H were recorded as the potential difference b e t w e e n the two microelectrode barrels, one with the liquid m e m b r a n e in the tip, the o t h e r serving as reference. Extracellular electric signals g e n e r a t e d by the tissue were r e c o r d e d from the reference barrel relative t o ground poten-

Correspondence: G. G. Somjen. Duke University Medical Center. Dept. of Physiology, Box 3709, Durham, NC 27710, U.S A.

0006-8993/84/$03.00 ~ 1984 Elsevier Science Publishers B.V.

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Fig. 1. The changesof pH in the granule cell layer of the fascia dentata caused by paroxysmal firing induced by stimulation of the angular bundle (6 Hz, 0.5 ms, 1.2 mA pulses). The tracing was produced with a polygraph by playing back the recording made on mag-

netic tape at 1/4 of the original speed to enhance the frequency response of the pen recorder. In the upper tracing, which shows the extracellular electric potential, the regularly repeated upward deflections of the pen are focal synaptic potentials ('population EPSPs'). The bursts of downward deflections, which begin halfway during the stimulus train and continue after cessation of stimulation, are negative compound action potentials ('population spikes'). In the lower trace, upward movement of the pen signifies increased positive potential of the H+-selective electrode, i.e. lowering of pH.

tial. Generally, single stimuli applied to the angular bundle evoked no detectable change of p H in the fascia dentata. However, stimulus trains that provoked paroxysmal firing invariably caused a change in the interstitial pH (Fig. 1). Stimulus trains that did not provoke seizure discharges did not always induce pH changes and, when they did, these did not exceed 0.04 pH units. Non-paroxysmal pH shifts could be either in the acid or in the alkaline direction, but the former were more common. Electrical stimulation can provoke two kinds of paroxysmal firing of dentate granule cells. One form consists of 'giant' negative compound action potentials8 10-40 mV in amplitude that are fired during delivery of a stimulus train, but timed independently

from the stimulus pulses. The other consists of bursts of such 'population spikes' fired after completion of the stimulus train. We termed the former type of firing intercurrentparoxysmal d&chargeor IPaD, while the latter are known as paroxysmal afterdischarges or PaAD (see also ref. 12). Paroxysmal firing of both types, either I P a D or P a A D , invariably brought in its wake acidification of the interstitial fluid. Stimulation strong enough to induce IPaD but not P a A D caused relatively mild changes of pH, ranging from less than 0.01 to 0.04 pH units. After intense P a A D pH could decrease by more than 0.2 units, but usually acidification amounted to 0.07-0.15 units. In some but not all cases a period of mild alkalinization preceded the acid shift (Fig. 1). Acidification was maximal 6 - 2 0 s

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Fig. 2. Changes of pH during spreading depression. At the mark 20 Hz stimulation was applied to the angular bundle. The oscillations following the stimulus train indicate paroxysmal afterdischarge; the frequency response was limited from DC to 3 Hz to emphasize slow changes. The large negative shift of the electric potential in the middle of the tracing indicates spreading depression.

188 after c o m p l e t i o n of t h e stimulus train, which was sev-

cess of H + ions is t h e p r o d u c t of e n h a n c e d cellular

eral s e c o n d s after cessation of the p a r o x y s m a l firing,

m e t a b o l i c activity. O x i d a t i o n of i n t r a m i t o c h o n d r i a l

A c i d shifts w e r e o b s e r v e d in all the c y t o a r c h i t e c t o n i c

e n z y m e s , m o n i t o r e d by optical m e t h o d s ,

layers of the fascia d e n t a t a ,

but they w e r e m o s t

similarly d e l a y e d t i m e c o u r s e < r T h e two b e s t - k n o w n

m a r k e d in the hilus of the fascia d e n t a t a , w h e r e the

acid products of c e r e b r a l m e t a b o l i s m are C O , and

p o s i t i v e focal synaptic p o t e n t i a l s and n e g a t t v e "pop-

lactic acid and b o t h m a y h a v e c o n t r i b u t e d to the acid-

ulation spikes' w e r e also m a x i m a l . T h e alkaline tran-

ification of the interstitial fltfid T h e fact thai non-

sients that s o m e t i m e s p r e c e d e acidification s e e m e d .

p a r o x y s m a l electrical activity was a s s o c i a t e d with

h o w e v e r , to be m a x i m a l slightly v e n t r a l and dorsal to

only mild c h a n g e s of p H . and s o m e t i m e s with n o n e al

the cell b o d y layers

all. indicates the p o w e r of i n t r a c e r e b r a l buffers.

In s o m e rats n o alkaline shifts

w e r e e v o k e d a n y w h e r e in the fascia d e n t a t a .

T h e transmnt a l k a l i n i z a t i o n that s o m e t t m e s pre-

A c i d i f i c a t i o n of the interstitial fluid was s e v e r e s t after e p i s o d e s of s p r e a d i n g d e p r e s s i o n sionally

followed

intensive

that occa-

paroxysmal

shows a

c e d e d the acid shift of p H is tess easily e x p l a i n e d . Possible c o n t r i b u t o r s to the d e c r e a s e of

[H*]o could

activity

be: ( 1 ) the release of alkaline p r o d u c t s f r o m cells into

(Fig. 2). T h e p H shifts a s s o c i a t e d with Lefio's de-

interstitial fluid: (2) i n c r e a s e d b l o o d flow t h r o u g h the

pression a m o u n t e d to 0 . 2 - 0 . 5 pH units. O f t e n t h e s e

tissue with c o n s e q u e n t w a s h o u t of C O , : (3) transient

acid p H shifts w e r e p r e c e d e d by an intense but brief

b r e a c h i n g of the b l o o d - b r a i n b a r r i e r and c o n s e q u e n t

p e r i o d of alkalinization. A c i d i f i c a t i o n was m a x i m a l

e q u i l i b r a t i o n of c e r e b r a l interstitial fluid with the

1 5 - 3 0 s after the s u d d e n n e g a t i v e shift of e x t r a c e l l u -

m o r e alkaline p H o f the b l o o d p l a s m a . O f t h e s e 3 al-

lar p o t e n t i a l that is the h a l l m a r k of n e u r o n a l d e p r e s -

ternatives I c o n s i d e r the s e c o n d to be the most likely,

sion of Lefio's type (see also ref. 10). T h e s e re-

but only f u r t h e r

sponses in the h i p p o c a m p a l f o r m a t i o n w e r e similar to

choice b e t w e e n t h e m .

research

will allow a definitive

those seen in c e r e b e l l u m by Kraig el al. 4. but different from the p r e d o m i n a n t l y alkaline shifts r e p o r t e d by L e h m e n k i i h l e r et al 5 for c e r e b r a l cortex. T h e d e l a y of acidification r e l a t i v e to the t i m i n g of electrical activity s u p p o r t s the s u g g e s t i o n that the ex-

1 Caspers. H. and Speckmann. E.-J., Cerebral pO 2, Pco2 and pH changes during convulsive acnvity and their significance for spontaneous arrest of seizures. Epitepsia. 13 ~1972) 699-725. 2 Dusser de Barenne, J. G.. Marshall, C. S., McCulloch W. S. and Nims. L. F.. Observations of the pH of arterial blood and the pH and electrical activity of the cerebral cortex. Arner. J. Physiol.. 124 (1938) 631-636 3 Jasper, H. and Erickson. T. C.. Cerebral blood flow and pH in excessive cortical discharge induced by metrazol and electrical stimulation, J. Neurophysiol.. 4 ( 1941 ) 333-347. 4 Kraig, R. P.. Ferreira-Filho. C. R. and Nicholson. C., Alkaline and acid transients in cerebellar microenvironment, L Neurophysiol.. 49 11983 b831-851"t, 5 Lehmenkiihler. A.. Zidek. W.. Staschen, M. and Caspers. H., Cortical pH and pCa in relation to DC potentials shifts during spreading depression and asphyxiation. In E. Svkov~,. P. Hnik and L. Vyklick~ {Eds.), Ion-Selective Electrodes and Their Use in Excitable Tissues. Plenum. New York. 1981. pp. 225-229. 6 Lewis. D V. and Schuette. W H., NADH fluorescence and [K+]o changes during hippocampal electrical stimulation. J, Neurophysiol., 38 [ 1975) 405-417. 7 Lothman. E.. LaManna, J.. Cordingley, G., Rosenthal, M. and Somjen. G., Responses of electrical potential, potassium levels and oxidative metabolic activity of cerebral neocortex of cats, Brain Research. 88 f1975l 15-36. 8 Pumura, D. P.. McMurtrv, J. G.. Leonard, C. F. and Mal-

I w o u l d like to t h a n k Mr. L. H a i t h for e x p e r t technical assistance. This w o r k was s u p p o r t e d by G r a n t s NS 17771 and NS 18670 of the N I H . U S P H S

liani, A.. Evidence of dendritic origin of spikes without depolarizing prepotentials in hippocampal neurons during and after seizure, J. Neurophysiol,. 29 ( 1966} 954-979. 9 Somjen. G. G.. The why and the how of measuring the activity of ions in extracellular fluid of spinal cord and cerebral cortex. In T. Zeuthen (Ed.), I'he Application o f IonSelective Microelectrodes. Elsevmr. Amsterdam, 1981, pp. 175-193, 10 Som]en. G. G.. Ionic and metabohc responses in neuronal depression of Leto's type An A c a d BrasiL Ci, Suppl., in press. 11 Somjen, G. G. and Allen. B. W.. Does hyperventtlatnm cause tetanv7 Proc. lnt. Union Phvsiol Sci . 15 1983l 14ft. 12 Somjen. G. G.. Aitken, P. G., Giacchino, J. L. and McNamara, J~ O., Interstitial ion concentrations and paroxysmal discharges in hippocampal formation and spinal Cord. In A. V. Delgado-Escueta. A. A. Ward and D. M. Woodbury (Eds.), Basic Mechanisms o f the Epilepsies. 12nd edn. ), Raven Press. New York, in press 13 Tschirgi, R D.. lnanaga, K.. Taylor, J. L.. Walker. R. M and Sonnenschein. R. R.. Changes in cortical pH and blood flow accompanying spreading cortical depressmn and convulsion, Amer. J. Physiol., 190 (1957) 557-562 14 Urbanics. R.. Leniger-Follert, E. and LObbers. D W.. Time course of changes of extracellular H + and K ~ activities during and after direct electrical stimulation of the brain cortex, Pfli~ger~s Arch ec~ Physiol., 37g (19781