Synaptosomal Na, K-ATPase during forebrain ischemia in Mongolian gerbils

�9 1996 by Hurnana Press lnc. All rights of any nature whatsoever reserved. 1044-7393/9612901 --0067 $08.00

Synaptosomal Na,K-ATPase During Forebrain Ischemia

in Mongolian Gerbils

M. MATEJOVI(~OV,~, 1 S. MACH,~(~,' J . LEHOTSK~', t J . JAKUS, 2'3 AND V. M~:ZEgOVA *,t

1Department of Biochemistry; eDepartment of Physiology; and 3Department of Biophysics, Jessenius Medical Faculty, Comenius

University, Malk Hora 4, SK-036-01 Martin, Slovak Republic

Received May 17, 1995; Revised September 28, 1995; Accepted October 25, 1995

ABSTRACT

We studied the activity and kinetic parameters of synaptosomal Na, K-ATPase during 15 rain of forebrain ischemia and following 60 min of reperfusion produced by reversible common carotid occlusion in Mongolian gerbils. A synaptosomal fraction was obtained by both differential centrifugation of brain tissue homogenate and centrifugation of crude mitochondrial fraction at a discontinual sucrose density gradient. We found two components of ATP concentration dependence of ATP hydrolysis that represent two types of ATP-binding sites: high affinity and low affinity. Neither ischemia nor reperfusion affected kinetic parameters of a high-affinity site. However, low- affinity site parameters were affected by both ischemia and ischemia followed by reperfusion. Maximal velocity (Wmax) decreased by 43 and 42% after ischemia and after ischemia/reperfusion, respectively. The apparent K,, for ATP decreased by 52% after ischemia and by 47% after ischemia/reperfusion. The apparent affinities for K + and Na + were determined from the ATP hydrolysis rate as a function of Na § and K + concentrations. We found the half-maximal activation constant for K + (KaK +) increased by 60% after ischemia and by 146% after ischemia/reperfusion. On the other hand, we found that KaNa + decreased significantly after ischemia/reperfusion (16%). We concluded

*Author to whom all correspondence and reprint requests should be addressed.

Molecular and Chemical Neuropathology 6 7 Vol. 29, 1996

68 Matejovi~ov~ et al.

that it is the dephosphorylation step of the ATPase reaction cycle that is primarily affected by both ischemia and ischemia/reperfusion. This might be caused by alteration of the protein molecule and/or its sur- roundings subsequent to ischemia.

Index Entries: Forebrain ischemia; Mongolian gerbils; synaptosomal Na,K-ATPase; kinetic parameters.

INTRODUCTION

Neuronal damage during brain ischemia is associated with an im- balance in ionic homeostasis. Disordered ion homeostasis is thought to be primarily the result of decreased energy metabolism. A reduction in the ATP level results in disturbance of ATP-coupled ion pumps, and other energy-consuming membrane-bound enzymes. Na,K-ATPase (Na+,K +- activated MgR+-dependent ATP phosphohydrolase), an energy-utilizing enzyme, maintains Na+ and K + electrogenic gradients across the plasma membrane required for regulation of cell volume and, with regard to ner- vous tissue, for development of action potentials and synaptic processes. This enzyme is a member of the EJE2 type of ATPases, which alternates between those two states (Carafoli and Chiesi, 1992; Jorgensen, 1992; Bamberg et al., 1993). The Na,K-ATPase is a complex of two polypep- tides, o~ and fl, incorporated into the lipid bilayer. The large o~ (catalytic) subunit forms a large intracellular loop that contains the nucleotide binding site. The Na § +-binding sites, which are involved in cation transport, are localized in the transmembrane domains of the o~ subunit. It is known that minimally three isoforms of the o~ subunit exist with different affinities to ouabain and perhaps different kinetic parameters (Horisberger et al., 1991; Lingrel, 1992). The o~3 subunit with high sensitivity to ouabain has been found preferentially in neurons (Decollgone et al., 1993). The minimal functional unit of Na, K-ATPase appeared to be the (o~,fl) hetero- dimer. However, the function of the fl subunit still remains unknown.

Na,K-ATPase may be pivotal in the development of ischemia and post- ischemic neuronal injury. Possible alterations in Na, K-ATPase activity result in disruption of cellular ionic homeostasis and membrane depolarization. This can lead to excessive neurotransmitter release, calcium overload, and further to secondary ischemic damage, e.g., activation of phospho- lipases, lipases, proteases, endonucleases, uncontrolled phosphorylation, degradation of membranes, and brain edema.

Changes in Na,K-ATPase activity during brain ischemia or hypoxia have been studied previously, but with contradictory results. Most authors demonstrate decrease of brain Na, K-ATPase activity in various animal models (Bertoni and Sprenkle, 1988; Stojanovic and Mrsulja, 1990; Marzatico et al., 1990, 1991; Nagafuji et al., 1992; Goplerud et al., 1992). Similarly, decreased activity of sarcolemmal Na,K-ATPase owing to myocardial infarction has been observed (Dixon et al., 1992). However, some data

Molecular and Chemical Neuropathology Vol. 29, 1996

Na, K-A TPase During Forebrain lschemia 69

showed enhanced activity of cerebral Na,K-ATPase during ischemia and ischemia followed by reperfusion (MacMillan, 1982). It seems that changes in Na,K-ATPase activity depend on the animal species and ischemic model, as well as on the sampling and assay method used. Some authors explained a decrease in Na,K-ATPase activity as being the result of protein degradation during a time when synthesis is turned off (Schwartz et al., 1976) and/or phospholipid degradation, mainly during period of reperfusion (Mishra and Delivoria-Papadopoulos, 1988; Goplerud et al., 1992). To date, Na,K-ATPase activity had been measured largely in brain tissue homogenates or crude mitochondrial fractions. Recent literature has not adequately explained the kinetic parameters of synaptosomal Na,K-ATPase reaction in ischemia and reperfusion.

The aim of the present work was therefore to study the influence of ischemia on both the activity and the kinetic parameters of Na,K-ATPase in a synaptic membrane fraction and endeavor to explain possible mechanisms of ischemic/reperfusion damage to the enzyme.

MATERIALS AND METHODS

Materials

Enzymes, substrates (ATP, phosphoenol pyruvate), and coenzyme (NADH) were obtained from Boehringer Mannheim. Ouabain was obtained from Sigma Chemicals. HEPES and Tris were obtained from Serva, and sucrose from Merck. All other chemicals used were of reagent analytical grade.

Animal Model

Ischemia was produced by bi|ateral occlusion of the common carotid arteries of Mongolian gerbils (Meriones unguiculatus) according to Crockard et al. (1980). Two groups of animals were used: group A, induced ischemia and group B, induced ischemia followed by reperfusion. Group A animals were anesthetized with pentobarbital (60 mg/kg, ip) and carotid arteries were occluded for 15 min, following which the animals were decapitated and the heads placed in liquid nitrogen for 3 s. Ischemia was induced in the same way for group B animals. Carotid arteries were released after 15 min, and animals were allowed 60 min of reperfusion prior to decapitation. Sham-operated animals were used as controls. The control animals were anesthesized with pentobarbital, 60 mg/kg ip and in a stable state of anesthesia (corneal areflection was observed), surgery was carried out. The carotid arteries were exposed, but not clamped. Animals were allowed to remain for 15 min in normothermic conditions (rectal temperature was monitored) prior to being killed by decapitation.

During ischemia and ischemia/reperfusion, animals were maintained in a thermostatically controlled, ventilated (1 L/min) holding unit. Brain



40

39

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3"1 E

F -

36

35

T o-brain t:::Z:~:i:: e

lischemia lreperfusi0n T m

i I l I

15 30 45 60 75

Time (min)

Fig. 1. Brain and rectal temperature changes during 15 rain of forebrain ischemia and following 60 min of reperfusion produced in mongolian gerbils (n -- 5) by reversible bilateral common carotid arteries occlusion under pentobarbital anesthesia.

and body temperatures were monitored with brain temperature maintained at 36.5 + 0.8~ (normothermic conditions). During reperfusion, the heating system was shut down, and the unit was allowed to respond to ambient temperature (Fig. 1). The brain temperature was monitored in 10 animals (5 from each group). Brain temperature was monitored using a semiconductor needle thermoprobe (AP-212, Tesla) introduced into the frontoparietal cortex. A second semiconductor thermoprobe (KA-206, Tesla) monitored rectal temperature.

Preparation of Synaptosomal Fraction Subcellular fractions were prepared from the whole forebrain accord-

ing to Edelman et al. (1985). All procedures were carried out at 0-4~ Brain tissue was homogenized in 0.32M sucrose buffered with 5 mM HEPES-Tris (pH 7.2) 1:10 (w:v) using a TeflonWM-glass homogenizer. Synaptosomes were obtained by both differential centrifugation of the brain tissue homogenate and centrifugation of the crude mitochondrial fraction using a discontinual four-step sucrose density gradient. The synaptosomal fraction containing plasma membrane vesicles was removed from the surface of the 1M sucrose layer, diluted to 0.45M sucrose with HEPES-Tris buffer, pH 7.2, and centrifuged at 100,000g for 30 min. The pellet was resuspended with 0.32M sucrose, and aliquots were taken for determination of protein content (Lowry et al., 1951).


Na, K.ATPase During Forebrain lschemia 71

NabK-ATPase Assay The Na,K-ATPase activity was measured spectrophotometrically with

coupled enzymatic reactions (pyruvate kinase plus lactate dehydrogenase) as described by Shonfeld et al. (1986) with some modifications. The medium for estimating Na,K-ATPase activity contained 25 mM imidazole- HCt buffer (pH 7.4), 5 mM MgC12, 10 mM KC1, 100 mM NaC1, 3 mM ATP, 5 mM NAN3, 1 mM phosphoenol pyruvate, 0.5 mM EGTA, 0.225 mM NADH, 16 IU pyruvate kinase, 16 IU lactate dehydrogenase, alamethicin (40 #g/mL), and 19.2 mM (NH4)2SO 4. The reaction was started by addition of a membrane suspension containing 20 #g protein to 0.4 mL of assay medium. Activity was measured at 37~ by decreased absorption at 340 nm on the Pharmacia LKB-Ultrospec III spectrophotometer. Na, K-ATPase activity was determined to be an ouabain-inhibited activity. For determin- ing ATP-, Na+-, and K§ the ligand concentration in the assay medium varied from 2 ~ to 5 raM, 3 to 150 mM, and 0.1 to 15 mM, respectively.

RESULTS

We studied the activity and the kinetic parameters of synaptosomal Na,K-ATPase during 15 min of forebrain ischemia and ischemia followed by 60 min of reperfusion.

The rate of ATP hydrolysis was measured within an ATP concentration range of 2 ~ to 5 mM. It was found that ATP concentration dependence of ATP hydrolysis by synaptosomal Na,K-ATPase exhibits two components; similar results have been shown for other E1/E2-ATPases (Muallem and Karlish, 1980; Garrahan and Rega, 1990; Boldyrev et al., 1991). This could be explained by the existence of two types of ATP bind~ ing sites: one high and one low affinity. The high-affinity ATP binding site should have a catalytic effect, and the low-affinity site, a regulatory character as proposed in the literature (Buxbaum and Schoner, 1991; Jorgensen, 1992; Ward and Cavieres, 1993).

Values of Kin1, Vmaxl (high-affinity site) and Kin2, Vmax2 (low-affinity site) were obtained from double-reciprocal plots (Lineweaver-Burk plot) (Fig. 2). Control values of Kin1 and 1(,,2 for ATP are 16.49 + 1.97 and 435.45 + 59.17 /~d, respectively, and control values of Vmaxl and Wrnax2 a r e 397.19 + 22.12 and 2535.69 + 232.19 nmol Pi/mg of protein/min, respectively. We observed after 15 min of ischemia and ischemia followed by 60 min of reperfusion that the low-affinity site was more affected than the high- affinity binding site. In fact, the maximal velocity of ATP hydrolysis (Vm~• decreased by 43% after 15 rain of ischemia and by 42% after 15 min of ischemia followed by 60 min of reperfusion. Km2 for ATP decreased by 52% after ischemia and by 47% after ischemia followed by 60 min of reperfusion. The high-affinity parameters are not s!gnificantly altered either


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0 I I I ~ r T I ?

0.05 0.1

1/[ATP] grnol/I

Fig. 2. Double-reciprocal plot of ATP concentration dependence on ATP hydrolysis by Na,K-ATPase. Straight lines were obtained separately by linear regression analysis from each portion of nonlinear plots. Results represent data from single experiments.

Table 1 The Effect of Ischemia and Ischemia Followed by Reperfusion

on Km (ATP) and Vmax Values of ATF Hydrolysis by Na,K-ATPase

High-affinity site Low-affinity site

Km 1 Vmaxl Kin2 Vmax2 n /~mol/L nmol Pi/mg/min #mol/L nmol Pi/mg/min

Control 7 16.5 + 1.97 397 + 22.1 435 + 59.2 2536 + 232.2 Ischemia 8 12.2 + 2.22 302 + 37.5 211 + 19.7 a 1439 + 77.8 b

15 min Reperfusion 10 19.1 _+ 1.40 c 346 + 24.2 221 _+ 34.4 a 1474 + 58.3 b

60 min

~p < 0.01. bp < 0.001. Cp < 0.05. Values are statistically significant in comparison with control (a), value is statistically

significant in comparison with ischemia (c). Data are expressed as means + SEM. n represents number of experiments performed. Kin1, Vmaxl and Km2, Vmax2 values for ATP were obtained from nonlinear Lineweaver-Burk (LWB) plots.

in t he i schemic c o n d i t i o n or in i s c h e m i c / r e p e r f u s i o n c o n d i t i o n s as com- p a r e d w i t h the cont ro l va lues (Table I a n d Fig. 3). H o w e v e r , the Kin1 va lues af ter i s c h e m i a / r e p e r f u s i o n w e r e s i g n i c a n t l y i n c r e a s e d in c o m p a r i s o n w i t h i s chemic K~I va lues , T h e s e resu l t s s h o w tha t d e c r e a s e d e n z y m e act ivi ty is a c c o m p a n i e d by a n inc reased af f in i ty of Na ,K-ATPase for ATP at t he low-af f in i ty ATP b i n d i n g site (pu ta t ive r e g u l a t o r y site) r e p r e s e n t e d by Kin2 va l ue s d u r i n g b o t h i schemic a n d i s c h e m i a / r e p e r f u s i o n cond i t i ons ,


Na, K.A TPase During Forebrain lschemia 73

60

150

120

90

30

0

Kin1

%

Vrnax 1

150

A

120

9O

% 6O

30

I

Kin2 Vmax2 Fig. 3. The effect of ischemia and ischemia/reperfusion on the Km and Vma•

of ATP hydrolysis by Na,K-ATPase in high-affinity (A) and low-affinity (B) sites. The columns represent mean values in percent + SEM, control values (open columns) equal 100, hatched columns show changes afer 15 rain of ischemia-- Kin1 = 74 + 16, V m a x l = 76 + 10, Krn2 = 48 _+ 8, Vmax2 = 57 + 6; double-hatched columns show changes after 15 rain of ischemia followed by 60 rain of reperfusion --Kin1 = 116 + 16, V m a x l = 97 • 9, K . z 2 = 53 • 11, Vmax2 = 58 • 6.

The appa ren t affinities of Na, K-ATPase for Na § and K § were de f ined f rom the ATP hydro lys i s rate as a funct ion of Na § and K § concent ra t ions . The ion concent ra t ions were var ied f rom 5 to 100 and 0.5 to 10 raM, respect ively. The half-maximal activation cons tan t Ka and Wmax values were ob ta ined f rom the linear por t ion of the double-reciprocal plots of ATP hydro lys i s d e p e n d e n c e vs Na § or K § concent ra t ions . We o b s e r v e d

Molecular and Chemical Neuropathology Vol. 29, I996


Table 2 The Effect of Ischemia and Ischemia Followed

by Reperfusion on Half-Maximal Activating Concentration of K § and Na +

n Ka (K+), mmol/L Ka (Na+), mmol/L

Control 5 1.27 + 0.16 13.84 + 0.51 Ischemia 6 2.03 + 0.23 a 12.79 + 0.94

15 min Reperfusion 4 3.12 + 0.53 b 11.58 + 0.38"

60 min

ap < 0.05. bp < 0.005. Data are expressed as means + SEM. n represents number of experiments performed.

Values of Ka were evaluated by linear regression analysis of linear part of LWB plots. Linear part of these plots correspond to the following concentration intervals: for K + - 0.5-10 mmol/L, for Na+--5-100 mmol/L.

300

250

200

150

100

50

0

K.(K +) Ka(Na*) Fig. 4. The effect of ischemia and ischemia/reperfusion on half-maximal

activating concentration of K + and Na+. The columns represent mean values of /G in percent + SEM. Control values (open columns) equal 100, hatched columns show changes afer 15 rain of ischemia--K,(K +) = 60 + 27, Ka(Na +) = 92 + 7; double-hatched columns show changes after 15 min of ischemia followed by 60 min of reperfusion/G(K +) = 246 + 52, Ka(Na +) = 84 + 4.

that fol lowing ischemia there was a 60% increase of the half-maximal acti- vat ion constant for K + that increased fur ther dur ing reperfus ion (by 146%). Converse ly , the K, for Na+ was decreased pr imari ly dur ing reper fus ion , b y 16% (Table 2 and Fig. 4). These data imply that appa ren t affinity for K § is decreased , w h e r e a s affinity for Na § is increased.


Na, K.A TPase During Forebrain lschemia 7 5

DISCUSSION

Na,K-ATPase appears to be one of the key enzymes involved in the development of neuronal ischemic damage. The results of this study show that both ischemia and ischemia/reperfusion lead to changes of synaptosomal Na,K-ATPase, resulting in decreased maximal velocity and changes in the apparent affinities for ligands. These changes are demonstrated primarily through the properties of the low-affinity ATP binding site (putative regulatory site) as well as by the altered apparent affinities to the activators Na § and K § . Thus, K,,~ of the low-affinity ATP binding site is decreased after both ischemia and ischemia followed by reperfusion, indicating an increased affinity of the enzyme for ATP. The decreased constant of half-maximal activation (KaNa +) of Na,K-ATPase indicates increased apparent affinity for Na § . On the other hand, apparent affinity of the enzyme for K § is decreased.

As a member of El/E2 type of ATPase, the enzyme alternates between those two states. In the E1 form, the enzyme binds sodium ions and ATP with high affinity. Alternatively, the K§ form (E2) binds ATP with low affinity and undergoes dephosphorylation, accompanied by ex- tracellular exchange of Na § for K § (Jorgensen, 1992). Our results imply that ischemia/reperfusion mainly affects the dephosphorylation step of the reaction cycle. The decrease of the enzyme's potassium affinity could lead to deceleration of the dephosphorylation step and, thus, to decreased maximal velocity of ATP hydrolysis. On the other hand, the increased affinity in the low-affinity ATP binding site might be a compensatory mechanism responding to the ATP loss triggered by ischemia and ischemia/ reperfusion. A similar mechanism was suggested in the case of heart sarcolemmal Na,K-ATPase under ischemic conditions (Vrbjar et al., 1991).

Several possible explanations have been derived regarding the altered enzyme affinities and maximal velocity induced by ischemia/reperfusion. However, two possibilities are highly probable. The first is structural folding in the protein molecule, and the second is environmental changes caused by degradation of membrane phospholipids surrounding enzyme molecule owing to free radical formation (Ra4ay et al., 1994) and by phos- pholipase action. Indeed it has been shown that products of phospholipid degradation--polyunsaturated free fatty acids and diacylglycerol--inhibit Na,K-ATPase (Farooqui et al., 1992). Structural folding of the protein molecule could be caused by a rapid decrease in both intra- and extra- cellular pH owing to lactate formation during ischemia as well as by free radical action. Influence of other modifiers is also possible, e.g., addi- tional phosphorylation of the protein molecule by protein kinase C (PKC) is known to alter Na, K-ATPase activity (Bertorello et al., 1991). There is evidence that ischemia produces translocation of PKC from the cytosol to the membrane-bound form (Domaflska-Janik and Zablocka, 1993). Inhibi- tion of Na, K-ATPase by PKC activators (which increase in ischemia) was


76 Matej 'ovi~ov~ et al.

found in tissue other than brain, particularly in intact renal proximal tubule cells (Bertorello et al., 1992). However , protein kinases are inhibited in early phases of ischemia. In the case of pKC, it might be con- nected with pKC translocation to the m e m b r a n e fraction as well as with s t imulat ion of Ca 2+-induced proteolysis. The relevance of these specula- tions mus t be examined in addit ional exper iments .

Neurons are very sensitive to oxygen deprivation, since that insuffi- ciency causes a lack of ATP and energy store deplet ion in the brain tissue. The metabolic rate declines rapidly and reaches about 10% of control values after 2 min of ischemia. Since ion t ransport in vivo appears to be exclusively d e p e n d e n t on ATP, it is not surpris ing that 25-75% of all ATP p roduced by neurons is c o n s u m e d by Na,K-ATPase (Whit t ingham, 1990). Thus, the effect of the ischemia on brain ion homeostasis is p redominant ly owing to lack of energy sources.

It is becoming clear that dissipation of ion homeostas is leads to the secondary ischemic changes of the neuron . Modification of the Na,K- ATPase molecule and/or its su r round ings demons t ra t ed by the decreased maximal velocity of ATP hydrolysis and altered enzyme affinities to l igands could be included in these changes.

REFERENCES

Bamberg E., Butt H.-J., Eisenrauch A., and Fendler K. (1993) Charge transport of ion pumps on lipid bilayer membranes. Quart. Rev. Biophys. 26, 1-25.

Bertoni J. M. and Sprenkle P. M. (1988) Inhibitors of cation pump enzyme equally present in normal and ischemic gerbil brain. Life Sci. 42, 1955-1962.

Bertorello A. M., Aperia A., Walaas S. I., Nairn A. C., and Greengard P. (1991) Phosphorylation of the catalytic subunit of Na +-K +-ATPase inhibits the activity of the enzyme. Proc. Natl. Acad. Sci. USA 88, 11359-11362.

Boldyrev A. A., Fedosova N. U., and Lopina O. D. (1991) The mechanism of the modifying effect of ATP on Na +-K § Biomed. Sci. 2, 450-454.

Buxbaum E. and Schoner W. (1991) Phosphate binding and ATP-binding sites co- exist in Na +/K+-transporting ATPase, as demonstrated by the inactivating MgPO4 complex analogue Co(NH3)4PO 4. Eur. J. Biochem. 195, 407-419.

Carafoli E. and Chiesi M. (1992) Calcium pumps in the plasma and intracellular membranes. Curr. Top. Cell. Reg. 32, 209-241.

Crockard A., Iannotti F., Hunstock A. T., Smith R. D., Harris R. D., and Symon L. (1980) Cerebral blood flow and edema following carotid occlusion in the gerbil. Stroke 11, 494-498.

Decollogne S., Bertrand I. B., Ascensio M., Drubaix I., and Lelievre L. G. (1993) Na+,K+-ATPase and Na+/Ca2+-exchange isoforms: Physiological and physiopathological relevance. J. Cardiovasc. Pharmacol. 22 (supl. 2), $96-$98.

Dixon I. M. C., Hata T., and Dhalla N. S. (1992) Sarcolemmal Na+-K+-ATPase activity in congestive heart failure due to myocardial infarction. Am. J. Physiol. 262, C664-C671.


Na, K.ATPase During Forebrain Ischemia 7 7

Domafiska-Janik K., and Zablocka B. (1993) Protein kinase C as an early and sensitive marker of ischemia-induced progressive neuronal damage in gerbil h ippocampus. Mol. Chem. Neuropathol. 20, 111-123.

Edelman A. M., Hunter D. D., Hendrickson A. E., and Krebs E. G. (1985) Subcel- lular distribution of calcium- and calmodulin-dependent myosin light chain phosphorylat ing activity in rat cerebral cortex. J. Neurosci. 10, 2609-2617.

Farooqui A. A., Hirashima Y., Farooqui T., and Horrocks L. A. (1992) Involve- ment of calcium, lipolytic enzymes, and free fatty acids in ischemic brain trauma, in Neurochemical Correlates of Cerebral Ischemia (Bazan N. G., Braquet P., and Ginsberg M. D., eds.), pp. 117-138, Plenum, New York.

Garrahan P. J. and Rega A. F. (1990) Plasma membrane calcium pump, in Intracel- lular Calcium Regulation (Bronner F., ed.), pp. 271-303, A. R. Liss, New York.

Goplerud J. M., Mishra O. P., and Delivoria-Papadopoulos M. (1992) Brain cell membrane disfunction following acute asphyxia in newborn piglets. Biol. Neonate 61, 33-41.

Horisberger J.-D., Lemas V., Kraehenbuhl J.-P., and Rossier B. (1991) Structure- function relationship of Na +,K § Annu. Rev. Physiol. 53, 565-584.

Jorgensen P. L. (1992) Na, K-ATPase, structure and transport mechanism, in Molecular Aspects of Transport Proteins (De Pont J. J. H. H. M., ed.), pp. 1-26, Elsevier, Amsterdam.

Lingrel J. B. (1992) Na, K-ATPase: isoform structure, function, and expression. J. Bioenerg. Biomembr. 24, 263-270.

Lowry O. H., Rosebrough N. J., Farr A. L., and Randall R. J. (1951) Protein mea- surement with the Folin phenol reagent. J. Biol. Chem. 193, 265-275.

MacMillan V. (1982) Cerebral Na+,K +-ATPase activity during exposure to and recovery from acute ischemia. J. Cereb. Blood Flow Metabol. 2, 457-465.

Marzatico F., Gaetani P., Buratti E., Messina A. L., Ferlenga P., and Rodriguez Y Baena R. (1990) Effects of high-dose methylprednisolone on Na+,K § ATPase and lipid peroxidation after experimental subarachnoid hemorrhage. Acta Neurol. Scand. 82, 263-270.

Marzatico F., Gaetani P., Buratti E., Messina A. L., Ferlenga P., and Rodriguez Y Baena R. (1991) Effect of nicardipine treatment on the Na+,K+-ATPase and lipid peroxidation after experimental subarachnoid hemorrhage. Acta Neurol. Scand. 108, 128-133.

Mishra O. P. and Delivoria-Papadopoulos M. (1988) Na § ,K +-ATPase in develop- ing guinea pig brain during normoxia and hypoxia. Neurochem. Res. 13, 761-766.

Muallem S. and Karlish S. D. J. (1980) Regulatory interaction between calmodulin and ATP on the red cell Ca 2+ pump. Biochim. Biophys. Acta 597, 631-636.

Nagafuji T., Koide T., and Takato M. (1992) Neurochemical correlates of selective neuronal loss following cerebral ischemia: role of decreased Na+,K +- ATPase activity. Brain Res. 571, 265-271.

Ra~ay P., Bezfikovfi G., Kaplfin P., Lehotsk~ J., and M6ze~ovfi V. (1994) Altera- tion in rabbit brain endoplasmic reticulum Ca 2§ transport by free oxygen radicals in vitro. Biochem. Biophys. Res. Commun. 199, 63-69.

Schonfeld W., Schonfeld R., Menke K.-H., Weiland J., and Repke K. R. H. (1986) Origin of differences of inhibitory potency of cardiac glycosides in Na § §


78 Matej'ovi~ov~ et al.

transporting ATPase from human cardiac muscle, human brain cortex and guinea-pig cardiac muscle. Biochem. Pharmacol. 35, 3221-3231.

Schwartz J. P., Mrsulja B. B., Mrsulja B. J., Passonneau J. V., and Klatzo I. (1976) Alteration of cyclic nucleotide-related enzymes and ATPase during unilateral ischemia and recirculation in gerbil cerebral cortex. J. Neurochem. 27, 101-107.

Stojanovic T. and Mrsulja B. (1988) Alternations in synaptosomal membrane Na,K-ATPase of the gerbil cortex and hippocampus following reversible brain ischemia. Metab. Brain Dis. 3, 265-272.

Vrbjar N., Slezak J., Ziegelhoffer A., and Tribulova N. (1991) Features of the (Na,K)-ATPase of cardiac sarcolemma with particular reference to myocardial ischaemia. Eur. Heart J. 12, 149-152.

Ward D. G. and Cavieres J. D. (1993) Solubilized c~B Na,K-ATPase remains pro- tomeric during turnover yet shows apparent negative cooperatively toward ATP. Proc. Natl. Acad. Sci. USA 90, 5332-5336.

Whittingham T. S. (1990) Aspects of brain energy metabolism and cerebral ischemia, in Cerebral Ischemia and Resuscitation (Schurr A. and Rigor B. M., eds.), pp. 101,102, CRC, Boca Raton.


Synaptosomal Na, K-ATPase during forebrain ischemia in Mongolian gerbils

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