Department of Anatomy and Cell Biology, Queen's University, Kingston, Ontario, Canada K7L 3N6
![]() |
Abstract |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
An important process associated with ischemia is anoxic depolarization (AD) which was described originally as a propagating electrical silence following interruption of the cerebral circulation (Leao, 1947). It was described as spreading depression-like, a sudden and profound depolarization to the point where neurons can no longer discharge. Spreading depression (SD) involves loss of ion homeostasis and water influx under normoxic conditions (Somjen et al., 1992
) that arises focally and then engulfs neurons and glia at a rate of 25 mm/min. The cerebral cortex, where it was first measured as a wave of electrical silence lasting several minutes (Leao, 1944
), is particularly susceptible. The cause of SD (and thus its prevention) is not understood and its characteristics depend on the tissue's metabolic status. SD without metabolic compromise (as occurs in migraine aura) causes no discernible damage to intact neocortex (Lauritzen, 1987
; Nedergaard and Hansen, 1988
) or to neocortical slices (Footit and Newberry, 1998
; Anderson et al., 1999
). In vivo this normoxic version of SD is blocked by N-methyl-D-aspartatic acid (NMDA) receptor antagonists but the AD is not blocked by NMDA or non-NMDA receptor antagonists (Hernandez-Caceres et al., 1987
; Marranes et al., 1988; Nedergaard and Hansen, 1988
; Lauritzen and Hansen, 1992
; Nellgard and Wieloch, 1992
). For 34 h following focal ischemia onset, recurring SD-like events expand the ischemic core and increase the number of at risk neurons in the penumbra (Hossman, 1994, 1996; Nedergaard, 1996
; Takano et al., 1996
; Irwin and Walz, 1999
). Preventing penumbral infarction reduces neurological impairment clinically (Furlan et al., 1996
), so inhibiting the initiation and propagation of recurrent SD during ischemia should reduce infarct volume.
Cell swelling during ischemia-like conditions contributes a major component of the measured intrinsic optical signal (IOS) which is associated with changes in light transmittance in brain slices under ischemia-like conditions (Basarsky et al., 1998; Obeidat and Andrew, 1998
; Aitken et al., 1999
; Andrew et al., 1999
; Kreisman et al., 2000
). In submerged slices, an initial LT increase represents the propagating depolarizing front, whereas a subsequent LT reduction is attributed to dendritic damage (Andrew et al., 1999
; Jarvis et al., 1999
; Obeidat et al., 2000
). Thus the imaging technique permits an assessment of both AD initiation and the acute neuronal damage left in its wake.
In this study, we optically map the effects of simulated global ischemia by depriving the neocortical slice preparation of O2 and lowering glucose to 1 mM. Our results indicate that the so-called anoxic depolarization that arises during global ischemia initially appears as a multi-focal event, each focus spreading out concentrically over the cortex and exacerbating neuronal damage in gray matter. Preliminary data have been presented by Jarvis and Andrew (Jarvis and Andrew, 1998).
![]() |
Materials and Methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Male SpragueDawley rats, 2130 days old (Charles River, St Constant, Quebec, Canada), were housed in a controlled environment (25°C, 12 h light/dark cycle) and fed Purina lab chow and water ad libitum. A rat was placed in a rodent restrainer and guillotined. The brain was excised and placed in ice cold oxygenated (95% O2/5% CO2) artificial cerebrospinal fluid (aCSF). Coronal slices (350400 µm) of neocortex and underlying striatum or hippocampus were cut using a vibrating microtome (Leica VT100S). Slices were incubated (26 h) at 30°C in aCSF before transfer to the recording chamber. A slice was submerged in oxygenated aCSF flowing at a rate of 12 ml/min at 37.5°C.
Solutions
The aCSF contained (in mM): NaCl 120, KCl 3.3, NaHCO3 26, MgSO4 1.3, NaH2PO4 1.2, D-glucose 11, CaCl2 1.8. The pH was 7.37.4 and the osmolarity was 292295 mOsm. O2/glucose deprivation (OGD) which simulates ischemia in vitro, was induced by reducing aCSF glucose from 11 to 1 mM and gassing the aCSF with 95% N2/5% CO2. NMDA (100 µM) or ouabain (100 µM) were added to the aCSF as required. Furosemide (5 mM), kynurenic acid (2 mM), DL-2-amino-5-phosphonovaleric acid (AP-5, 50 µM) or (6-cyano-7-nitroquinozaline-2,3-(1H,4H)-dione (CNQX, 10 µM), were added to the aCSF as required. The 50:50 racemic stock of AP-5 made up to 50 µM contained 25 µM of the active D-isomer. All chemicals were purchased from the Sigma Chemical Co.
Imaging Intrinsic Optical Signals (IOSs)
IOSs are generated by changes in light scattering or absorbance within living tissue. In terms of optics, the simplest paradigm is to image change in light transmitted by a submerged brain slice, thereby avoiding complexities associated with measuring reflectance and the tissue/air interface [reviewed by Jarvis et al. (Jarvis et al., 1999)]. A neocortical slice was placed in an imaging chamber with a coverslip as the base. The slice was superfused with flowing aCSF, transilluminated with a broadband halogen light source, and viewed with a 1.25 objective on an inverted microscope (Zeiss Axiovert TV 100). Video images were collected using a charge coupled device (CCD, Cohu) connected to an image processing board (DT 3155, Translation) in a PC controlled by Axon Imaging Workbench software (Axon Instruments). The CCD was set at maximum gain and low black level. The gamma level was set to 1.0 so that CCD output was linear with respect to changes in light intensity. With appropriate filters, the IOS signal comprised the far red to near infrared spectrum (6901000 nm). Video frames were acquired at 1.38 s intervals with 32256 frames averaged per single image. The transmittance value (T) of the first image (Tcont) was subtracted from each subsequent image (Texpt) of the series, so the difference image (TexpTcont) revealed areas where LT changed over time. To visualize these areas better, light transmittance changes (
LT) were pseudocolored. Data were also quantified and graphically displayed by averaging the digital intensities from selected zones of interest (ZOI). Since various regions differed in Tcont, the data were normalized as follows. The change in light transmittance =
![]() |
There are several biophysical changes when cells take up water and swell which act to reduce light scattering (thereby elevating LT). However, when dendritic beading accompanies swelling (as during excitotoxicity or O2/glucose deprivation) the beading becomes the most important biophysical factor, scattering light even as the tissue continues to swell (Jarvis et al., 1999).
Electrophysiology
Extracellular recording of evoked field potentials served as an indicator of synaptic function and slice viability. Simultaneous extracellular recording and IOS imaging during OGD was performed to correlate the onset of the negative shift and LT changes resulting from OGD. The recording micropipette was placed in layer II/III of the neocortical slice and a concentric bipolar stimulating electrode was placed in the immediately underlying V/VI layers. A current pulse (0.1 ms duration; 0.25 Hz) was applied to produce a near-maximal amplitude population spike. Digitized data were plotted using pCLAMP software (Axon Instruments). The recording pipette also served to measure a negative DC shift in the extracellular potential induced by OGD. The DC shift is the electrical signature of the AD and SD.
Histology
For histological analyses, control slices were maintained in the imaging chamber for 15 min at 37.5°C then fixed in Bouin's fluid. Experimental slices were exposed to OGD for 10 min at 37.5°C which evoked an AD episode. After 15 min they were placed in fixative. Following fixation for 24 h in Bouin's fluid, slices were stored in 70% ethanol and processed for paraffin embedding. Sections (7 µm) were stained with hematoxylin/ eosin and photographed using a 40x objective.
![]() |
Results |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Within 10 min, OGD at 37.5°C induced AD in all neocortical slices tested with a mean latency of 4:26 ± 1:24 (min:s ± SD, n = 32). AD was first detected as one or two focal increases in LT in layers II/III. Each focus usually first appeared as a sphere ~0.5 mm in diameter that migrated out along adjacent cortex of all six layers. The ignition site could be in lateral, central or midline neocortex. Changes in LT were plotted from several zones of interest across the neocortex. AD propagated as a front of elevated LT in gray but not white matter at a rate of 1.50 ± 0.83 mm/min (n = 32). A dramatic decrease in LT then followed in the wake of the elevated LT front. Figure 1A shows migration of the AD front (initiated out of frame) first in one hemisphere and then as an independent event contralaterally. The elevated LT front propagates across the gray matter (Fig. 2A
), activating all neocortical layers of a single column concurrently (Fig. 2B
).
|
|
Simultaneous IOS imaging (Fig. 3A) and extracellular recording (Fig. 3B
) were performed to correlate changes in extracellular potential with changes in LT during OGD. A recording electrode in layer II/III monitored the extracellular potential (Fig. 3B
) as well as the population spike (Fig. 3D
) evoked by a stimulating electrode positioned in underlying layers V/VI. A sudden negative shift of 510 mV (Fig. 3B
) correlated temporally and spatially with the passage of the elevated LT front past the electrode tip (Fig. 3A
,C; n = 5). The negative shift started to return to baseline during OGD exposure, usually reaching baseline within 23 min but often not returning completely. Significantly, the evoked field potential recorded in layers II/III prior to OGD was abolished following AD and showed no recovery over the following 30 min (Fig. 3D
).
|
Histology/Temperature/Furosemide
Histological sections of control tissue (n = 4) maintained at 37.5°C for 15 min (no AD) were compared with experimental slices (n = 4) that supported AD in response to OGD for 10 min at 37.5°C. In contrast to control tissue (Fig. 4A), neocortical neurons that generated the AD displayed nuclear and cytoplasmic swelling (Fig. 4B
). In addition, the large primary dendrites that could be discerned running perpendicular to the cortical layers in control slices (Fig. 4A
, arrowheads) were not apparent in experimental slices where the neuropil instead appeared mottled (Fig. 4B
), suggesting altered structure of these cortical neurons, which were also unable to generate an evoked field potential.
|
|
Glutamate Receptors and SD Induced by OGD
To test the potential role of glutamate receptors in the OGD response, the non-specific glutamate antagonist kynurenate (2 mM) was applied 1540 min before, during and after OGD at 37.5°C. Treatment with kynurenate did not prevent the initiation and propagation of AD induced by 10 min of OGD in nine of nine neocortical slices tested (Fig. 5A). Moreover, kynurenate did not significantly alter AD onset time (Fig. 6A
) nor propagation rate (Fig. 6B
). The negative shift recorded extracellularly in layers II/III was similar in waveform to slices without kynurenate (n = 6, not shown). In support of these findings, treatment of slices with a combination of an NMDA and a non-NMDA receptor antagonist, AP5 (D-isomer, 25 µM) and CNQX (10 µM) respectively, did not prevent OGD-induced AD in five of five slices tested (Fig. 5
). Neither the time to onset (Fig. 5A
) nor the propagation rate (Fig. 5B
) were altered by AP-5/CNQX treatment. Likewise, treatment with 50 µM of the D-isomer of AP-5 alone was ineffective in five of five slices tested (not shown).
|
If extracellular glutamate accumulation has a role in OGD-induced AD, then glutamate receptor agonists should elicit AD in a pattern similar to OGD. However, the application of 100 µM NMDA at 37.5°C (n = 8, Fig. 7A, B) first produced a generalized, not focal, elevation in LT which developed more slowly than AD onset induced by OGD (Figs 1A
, 2A
). However, there then followed a sudden spreading optical signal reduction (Fig. 7A, B
) coincident with a negative shift in layers II/III (Fig. 7C
). A generalized elevation in LT was also observed in the hippocampus, but no spreading event was observed in eight of eight hippocampal slices, as previously reported (Jarvis et al., 1999
).
|
![]() |
Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
In animal models of ischemia, the extent of necrotic brain injury is quantified by determining infarct size or neuronal loss sampled histologically hours or days post-insult (Corbett and Nurse, 1998). Recurrent SD, also termed peri-infarct depolarization (PID), contributes to this damage in the penumbra (Nedergaard, 1996
; Hossman 1996; Strong et al., 1999
). Mapping intrinsic optical signals (IOSs) reveals real-time responses to glutamate agonists or to simulated ischemia in brain slices (Andrew et al., 1996
; Obeidat and Andrew, 1998
; Polischuk et al., 1998
; Aitken et al., 1999
; Jarvis et al., 1999
). IOS imaging detects cell swelling, clearly demarcating the ignition site and migration front of AD across live submerged slices of hippo-campus or neocortex (Basarsky et al., 1998
; Obeidat and Andrew, 1998
). The present study suggests that during global ischemia, the AD initiates at multiple sites and propagates outwards like the ripples from several stones thrown in a pond. In their wake the ripples leave further energetically compromised gray matter.
SD involves a sudden and massive increase in membrane permeability causing influx of Na+, Ca2+, Cl and water and efflux of K+ (Somjen et al., 1992). The resultant cell swelling decreases extracellular space (Nicholson and Kraig, 1981
; Hansen, 1985
). Under normoxic conditions, ion redistribution returns to normal within minutes with no neuronal or glial damage. However, the AD generated during ischemic-like conditions (Leao, 1947
) exacerbates neuronal damage (Nedergaard and Hansen, 1993
; Mies et al., 1994
; Back et al., 1996
), presumably by further elevating the metabolic load in the vulnerable penumbra (Obrenovitch, 1995
).
In the present study, submerged neocortical slices were imaged at low magnification in response to metabolic compromise by O2/glucose deprivation (OGD) or by ouabain (Balestrino, 1995; Balestrino et al., 1999
), both simulating a global ischemia. Either treatment evokes an increase in LT that arises focally at one or several sites and propagates into adjacent cortical tissue. The AD propagation rate of 1.52.0 mm/min is in the lower range of those reported for SD in intact cortex (Nedergaard, 1996
).
In the hippocampal slice (Obeidat et al., 2000), OGD induced a propagating increase in LT (cell swelling) followed by a rapid decrease in LT over several minutes. This decrease might represent cellular shrinkage because decreased LT is reversibly evoked by hyperosmotic saline (Andrew and MacVicar, 1994
; Andrew et al., 1997
). However, the more likely cause is dendritic beading which develops even as the tissue continues to swell and is indicative of neuronal damage. The necklace-like conformation of hundreds of dendritic processes is highly efficient at scattering light, such that bead formation over several minutes dramatically reduces light transmittance (Polischuk et al., 1998
; Jarvis et al., 1999
; Obeidat et al., 2000
). Where dendrites are lacking (as in hippocampal cell body layers), LT continues to increase. In contrast, the high density of dendrites in all neocortical layers leads to a decreased LT across gray matter once enough beading overwhelms the initial increase in LT generated by cell swelling. Dendritic beading has been observed in cultured neurons following O2/glucose deprivation (Park et al., 1996
) and in vivo following ischemia (Hori and Carpenter, 1994
). The optical sequence evoked by ouabain is indistinguishable from that evoked by OGD, consistent with previous observations in hippocampus (Obeidat and Andrew, 1998
). In both cases there is loss of Na+/K+ pump function causing increases in [K+]o, which appears to be a critical step in the induction of the AD.
IOS imaging and simultaneous recording of the extracellular potential in layers II/III reveal a temporal and spacial correlation between signals. The negative voltage shift recorded extracellularly, which is the electrophysiological signature of the AD, arises as the LT front passes the site of the recording electrode. The negative shift may return to near baseline, but this does not indicate physiological recovery because the evoked field potential is permanently lost following the AD. Moreover, an irreversible decrease in LT develops following the AD. In addition, histological analysis reveals swelling of neuronal nuclei and cell bodies in all cortical layers. The neuropil displays a mottled appearance and indistinct primary dendrites. None of these indicators of neuronal damage are observed in OGD-exposed tissue unless the AD is generated. In contrast, SD can be generated repetitively in our slices each time extracellular K+ is briefly increased and there are no signs of tissue damage (Anderson et al., 1999), so SD without OGD appears innocuous.
AD Resists Pharmacological Blockade
Bath application of the glutamate receptor agonist NMDA produced a general LT increase in neocortex and hippocampus (particulary in the CA1 region). Unlike the AD, the signal developed slowly and uniformly over the slice, did not propagate and was blocked by AP-5. An SD-like negative shift developed just at the time that the LT increase peaked and abruptly reversed, which was the point at which we conjecture light scattering by beaded dendrites began to overwhelm the swelling signal. Thus the NMDA-evoked signal sequence had some elements of the AD, but it appeared to engage the entire slice almost simultaneously.
NMDA receptor antagonists block SD under normoxic conditions in vivo (Hernandez-Caceres et al., 1987; Marranes et al., 1988; Nedergaard and Hansen, 1988
; Lauritzen and Hansen, 1992
; Nellgard and Wieloch, 1992
). These same studies show that glutamate receptors do not play a major role in the AD induced by ischemia. A recent study (Rossi et al., 1999
) showed that the AD could be blocked by a combination of NMDAR and non-NMDAR antagonists, but used slices from 12-day-old rats where NMDAR density is higher than adults. MK801 is ~10-fold more potent as a neuroprotectant against NMDA- and OGD- mediated neuronal injury in immature rodents than in adults (McDonald and Johnston, 1990
). Our study supports in vivo work cited above by showing that NMDA receptor antagonism does not block the AD in slices or offer protection from acute post-AD damage. This is unlike normoxic SD in sister brain slices (Anderson et al., 1999
) where NMDA receptor antagonists are effective blockers. This indicates a contribution by these receptors in generating the milder SD. However, the fact that glutamate begins to accumulate only after the anoxic depolarization (Obrenovitch and Urenjak, 1997a
; Obrenovitch, 1999
) further argues against a role for glutamate in AD initiation and propagation under metabolically compromised conditions.
While both neurons and glia depolarize during AD, their relative contribution to the optical signal is still an open question. A previous study using furosemide (to interfere with the Na+/K+/2Clco-transporter) blocked weak IOSs generated by synaptic stimulation (MacVicar and Hochman, 1991). Swelling by cultured glial cells can be blocked by furosemide (Walz and Hertz, 1984
; Kempski et al., 1991
). Furosemide has been reported to reduce the duration of normoxic SD induced by KCl application in cat neocortex in vivo (Read et al., 1997
). However, in the present study the initiation, propagation and post-AD damage were each furosemide-insensitive, so a glial contribution to the swelling signal was not demonstrated.
Slight reductions in temperature can lessen neuronal damage associated with ischemia (Chen et al., 1993; Dietrich et al., 1997
). In the present study, OGD-induced AD persisted in neocortical slices maintained at a lowered temperature of 32°C but not at 22°C, probably because metabolic demand was reduced enough to avoid AD induction. We suggest that reduced temperature can be neuroprotective because recurrent SD in the penumbra is suppressed. Hypothermia reduces the propensity of cortical tissue to propagate SD in the rat (Takaoka et al., 1996
) and in rat hippocampal slices (Obeidat et al., 2000
). Lowered temperature suppresses AD and reduces infarct size following middle cerebral artery occlusion (Chen et al., 1993
; Colbourne et al., 1997
; Corbett et al., 2000
).
In the present study TTX did not block AD, as also found by Taylor et al. (Taylor et al., 1999) in half of their slices exposed to OGD. In the other half, it was delayed. We found that treatment with glutamate antagonists (kynurenate or AP-5/CNQX) had no effect. Combining TTX and kynurenate delayed AD onset by 8 min, so blockade of both glutamate receptors and voltage-sensitive Na+ channels can delay, but not prevent, the onset of AD. This supports findings by Aitken et al. (Aitken et al., 1988
). Other slice studies show that NMDA receptor antagonists are ineffective in blocking hypoxic SD (Jing et al., 1993
). Clearly, an important reason why acute stroke damage is difficult to prevent (other than by restoring blood supply or lowering temperature) is because the AD and recurrent SD are so resistant to pharmacological blockade. However, we have recently found that sigma receptor ligands such as dextromethorphan block ischemic SD (i.e. AD) or normoxic SD in neocortical slice preparations (Anderson et al., 2000
). The findings presented here suggest that an ideal stroke treatment would uncouple the AD from ischemia yet be clinically tolerable. Sigma receptor ligands may prove useful in this regard.
To conclude, this study indicates that AD is an important contributor to neuronal damage immediately following the onset of ischemia. Glutamate receptor antagonists appear to be of little benefit in vivo or in cortical slices during and following the initial ischemic period. The inhibition of AD or recurrent SD in the penumbra could prove to be an effective therapeutic strategy in the treatment of stroke if clinically tolerable drugs could be taken prophylactically or introduced during the 3 h period following stroke when peri-infarct depolarizations recur.
![]() |
Notes |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Address correspondence to R. David Andrew, Department of Anatomy and Cell Biology, Queen's University, Kingston, Ontario, Canada K7L 3N6. Email: andrewd{at}post.queensu.ca.
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Aitken PG, Tombaugh GC, Turner DA, Somjen GG (1998) Similar propagation of SD and hypoxic SD-like depolarization in rat hippocampus recorded optically and electrically. J Neurophysiol 80:15141521.
Aitken PG, Fayuk D, Somjen GG, Turner DA (1999) Use of intrinsic optical signals to monitor physiological changes in brain tissue slices. Methods: A Companion to Methods in Enzymology 18:91103.[ISI]
Anderson, TA, Jarvis CR, Andrew RD (1999) Imaging repetitive spreading depression in submerged neocortical slices. Soc Neurosci Abstr 25: 740.
Anderson TA, Biedermann AJ, Andrew RD (2000) Sigma receptor ligands block spreading depression in rat neocortical slices. Soc Neurosci Abstr 26:282.15.
Andrew RD, MacVicar BA (1994) Imaging cell volume changes and neuronal excitation in the hippocampal slice. Neuroscience 62: 371383.[ISI][Medline]
Andrew RD, Adams JR, Polischuk TM (1996) Imaging NMDA- and kainate- induced intrinsic optical signals from hippocampal slice. J Neurophysiol 76: 27072717.
Andrew RD, Lobinowich ME, Osehobo EP (1997) Evidence against volume regulation by cortical brain cells during acute osmotic stress. Exp Neurol 142:300312.
Andrew RD, Jarvis CR, Obeidat AS (1999) Potential sources of intrinsic optical signals imaged in live brain slices. Methods: A Companion to Methods in Enzymology 18:185196.[ISI]
Back T, Ginsberg MD, Dietrich WD, Watson BD (1996) Induction of spreading depression in the ischemic hemisphere following experimental middle cerebral artery occlusion: effect on infarct morphology. J Cereb Blood Flow Metab 16:202213.[ISI][Medline]
Balestrino M (1995) Studies on anoxic depolarization. In: Brain slices in basic and clinical research (Shurr A, Rigor BM, eds), pp. 273293. London: CRC Press.
Balestrino M, Young J, Aitken P (1999) Block of (Na+, K+) ATPase with ouabain induces spreading depression-like depolarization in hippocampal slices. Brain Res 838:3744.[ISI][Medline]
Basarsky TA, Duffy SN, Andrew RD, MacVicar BA (1998) Imaging spreading depression and associated intracellular calcium waves in brain slices. J Neurosci 18:71897199.[Abstract]
Chen Q, Chopp M, Bodzin G, Chen H (1993) Temperature modulation of cerebral depolarization during focal cerebral ischemia in rats: correlation with ischemic injury. J Cereb Blood Flow Metab 13: 389394.[ISI][Medline]
Colbourne F, Sutherland G, Corbett D (1997) Postischemic hypothermia: a critical appraisal with implications for clinical treatment. Mol Neurobiol 14:171201.[ISI][Medline]
Corbett D, Nurse S (1998) The problem of assessing effective neuroprotection in experimental cerebral ischemia. Prog Neurobiol 54:531548.[ISI][Medline]
Corbett D, Hamilton M, Colbourne F (2000) Persistent neuroprotection with prolonged postischemic hypothermia in adult rats subjected to transient middle cerebral artery acclusion. Exp Neurol 163:200206.[ISI][Medline]
Dietrich WD, Busto R, Globus MYT, Ginsberg, MD (1997) Brain damage and temperature: cellular and molecular mechanisms. Adv Neurol 71: 177197.[ISI]
Footit DR, Newberry NR (1998) Cortical spreading depression induces an LTP-like effect in rat neocortex in vivo. Brain Res 781:339342.[ISI][Medline]
Furlan M, Marchal G, Viader F, Derlon JM, Baron JC (1996) Spontaneous neurological recovery after stroke and the fate of the ischemic penumbra. Ann Neurol 40:216226.[ISI][Medline]
Hansen AJ (1985) Effect of anoxia on ion distribution in the brain. Physiol Rev 65:101148.
Hernandez-Caceres J, Maclas-Gonzales K, Brozek G, Bures J (1987) Systemic ketamine blocks cortical spreading depression but does not delay the onset of terminal anoxic depression. Brain Res 437:360364.[ISI][Medline]
Hori N, Carpenter DO (1994) Functional and morphological changes induced by transient in vivo ischemia. Exp Neurol 129:279289.[ISI][Medline]
Hossmann KA (1994) Glutamate-mediated injury in focal cerebral ischemia: the excitotoxic hypothesis revised. Brain Pathol 4:2326.[ISI][Medline]
Hossmann KA (1996) Periinfarct depolarizations. Cerebrovasc Brain Metab Rev 8:195208.[ISI][Medline]
Irwin A, Walz W (1999) Spreading depression waves as mediators of secondary Injury and protective mechanisms. In Cerebral ischemia: molecular and cellular pathophysiology (Walz W, ed.). Totow, NJ: Humana Press.
Jarvis CR, Andrew RD (1998) Spreading depression mediates acute neuronal damage induced by oxygen/glucose deprivation in neocortical slices. Soc Neurosci Abstr 24:982.
Jarvis CR, Lilge L, Vipond GJ, Andrew RD (1999) Interpretation of intrinsic optical signals and calcein fluorescence during acute excitotoxic insult in the hippocampal slice. NeuroImage 10:357372.[ISI][Medline]
Jing J, Aitken PG, Somjen GG (1993) Role of calcium channels in spreading depression in rat hippocampal slices. Brain Res 604: 251259.[ISI][Medline]
Kempski O, von Rosen S, Weigt H, Staub F, Peters J, Baethman A (1991) Glial ion transport and volume control. Ann NY Acad Sci 633:306317.[Abstract]
Kreisman NR, Soliman S, Gozal D (2000) Regional differences in hypoxic depolarization and swelling in hippocampal slices. J Neurophysiol 83: 10311038.
Lauritzen M (1987) Cerebral blood flow in migraine and cortical spreading depression. Acta Neurol Scand Suppl 76:940.
Lauritzen M, Hansen AJ (1992) The effect of glutamate receptor blockade on anoxic depolarisation and cortical spreading depression. J Cereb Blood Flow Metab 12:223229.[ISI][Medline]
Leao APP (1944) Spreading depression of activity in the cerebral cortex. J Neurophysiol 7:359390.
Leao APP (1947) Further observations on the spreading depression of activity in the cerebral cortex. J Neurophysiol 10:409414.
MacVicar BA, Hochman D (1991) Imaging of synaptically evoked intrinsic optical signals in hippocampal slices. J Neurosci 11:14581469.[Abstract]
Marrannes R, Willems R, DePrins E (1988) Evidence for a role of the NMDA receptor in cortical spreading depression in the rat. Brain Res 457:226240.[ISI][Medline]
McDonald JR, Johnston MV (1990) Physiological and pathophysiological roles of excitatory amino acids during central neurons system development. Brain Res Rev 15:4170.[ISI][Medline]
Mies G, Kohno K, Hossmann KA (1994) Prevention of periinfarct direct current shifts with glutamate antagonist NBQX following occlusion of the middle cerebral artery in the rat. J Cereb Blood Flow Metab 14:802807.[ISI][Medline]
Nedergaard M (1996) Spreading depression as a contributor to ischemic brain damage. Adv Neurobiol 71:7584.
Nedergaard M, Hansen AJ (1988) Spreading depression is not associated with neuronal injury in the normal brain. Brain Res 449:395398.[ISI][Medline]
Nedergaard M, Hansen AJ (1993) Characterization of cortical depolarizations evoked in focal cerebral ischemia. J Cereb Blood Flow Metab 13:568574.[ISI][Medline]
Nellgard B, Wieloch T (1992) NMDA-receptor blockers but not NBQX, an AMPA-receptor antagonist, inhibit spreading depression in the rat brain. Acta Physiol Scand 146:497503.[ISI][Medline]
Nicholson C, Kraig RP (1981) The behavior of extracellular ions during spreading depression. In: The application of ion-selective electrodes (Zeuthen T, ed.). Amsterdam: Elsevier.
Obeidat, A, Andrew RD (1998) Spreading depression determines acute cellular damage in the hippocampal slice during oxygen/glucose deprivation. Eur J Neurosci 10:34513461.[ISI][Medline]
Obeidat A, Jarvis CR, Andrew RD (2000) Glutamate does not mediate acute neuronal damage following spreading depression induced by O2/glucose deprivation in the hippocampal slice. J Cereb Blood Flow Metab 20:412422.[ISI][Medline]
Obrenovitch TP (1995) The ischemic penumbra: twenty years on. Cerebrovasc Brain Metab Rev 7:297323.[ISI][Medline]
Obrenovitch TP (1999) High extracellular glutamate and neuronal death in neurological disorders: cause, contribution, or consequence. Ann NY Acad Sci 890:273286.
Obrenovitch TP, Urenjak J (1997a) Altered glutamatergic transmission in neurological disorders: from high extracellular glutamate to excessive synaptic efficacy. Prog Neurobiol 51:3987.[ISI][Medline]
Obrenovitch TP, Urenjak J (1997b) Is high extracellular glutamate the key to excitotoxicity in traumatic brain injury? J Neurotrauma 14: 677697.[ISI][Medline]
Park JS, Bateman MC, Goldberg MP (1996) Rapid alterations in dendrite morphology during sublethal hypoxia or glutamate receptor activation. Neurobiol Disease 3:215227.[ISI][Medline]
Polischuk TM, Jarvis CR, Andrew RD (1998) Intrinsic optical signaling denoting neuronal damage in response to acute excitotoxic insult in the hippocampal slice. Neurobiol Disease 4:423437.[ISI][Medline]
Read SJ, Smith MI, Benham CD, Hunter AJ, Parsons AA (1997) Furosemide inhibits regenerative cortical spreading depression in anaesthetized cats. Cephalalgia 17:826832.[ISI][Medline]
Rossi DJ, Oshima T, Attwell D (1999) Glutamate release in severe brain ischemia is mainly by reversed uptake. Nature 403:316321.[ISI]
Strong AJ, Smith SE, Whittington DJ, Meldrum, BS, Parsons AA, Krupinski J, Hunter AJ, Patel S (1999) Factors influencing the frequency of fluorescence transients as markers of peri-infarct depolarizations in focal cerebral ischemia. Stroke 31:214221.
Somjen GG, Aitken PG, Czeh GL, Herreras O, Jing J, Young JN (1992) Mechanisms of spreading depression: a review of recent findings and a hypothesis. Can J Physiol Pharmacol 70:S248S254.[ISI][Medline]
Takano K, Latour LL, Formato JE, Carano RA, Helmer KG, Hasegawa Y, Sotak CH, Fisher M (1996) The role of spreading depression in focal ischemia evaluated by diffusion mapping. Ann Neurol 39:308318.[ISI][Medline]
Takaoka S, Pearlstein RD, Warner DS (1996) Hypothermia reduces the propensity of cortical tissue to propagate direct current depolarization in the rat. Neurosci Lett 218:2528.[ISI][Medline]
Taylor CP, Weber ML, Gaughan CL, Lehning EJ, LoPachin RM (1999) Oxygen/glucose deprivation in hippocampal slices: altered intraneuronal elemental composition predicts structural and functional damage. J Neurosci 19:619629.
Verity MA (1991) Use and abuse of tissue cultures in neurotoxicity studies. Neurotoxicology 12:457459.[ISI][Medline]
Walz W (1987) Swelling and K+ uptake in cultured astrocytes. Can J Physiol Pharmacol 65:10511057.[ISI][Medline]
Walz W, Hertz L (1984) Intense furosemide-sensitive potassium accumulation in astrocytes in the presence of pathologically high extracellular potassium levels. J Cereb Blood Flow Metab 4:301304.[ISI][Medline]
Weber ML, Taylor CP (1994) Damage from oxygen and glucose deprivation in hippocampal slices is prevented by tetrodotoxin, lidocaine and phenytoin without blockade of action potentials. Brain Res 664:167177.[ISI][Medline]