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Published ahead of print on May 28, 2003, doi:10.1164/rccm.200211-1264OC
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American Journal of Respiratory and Critical Care Medicine Vol 168. pp. 469-475, (2003)
© 2003 American Thoracic Society

Original Article

Cyclooxygenase 2 and Intermittent Hypoxia-induced Spatial Deficits in the Rat

Richard C. Li, Barry W. Row, Evelyne Gozal, Leila Kheirandish, Qiang Fan, Kenneth R. Brittian, Shang Z. Guo, Leroy R. Sachleben, Jr. and David Gozal

Kosair Children's Hospital Research Institute, Department of Pediatrics, and Department of Pharmacology and Toxicology, University of Louisville, Louisville, Kentucky

Correspondence and requests for reprints should be addressed to David Gozal, M.D., Kosair Children's Hospital Research Institute, 570 South Preston Street, Suite 321, Department of Pediatrics, University of Louisville, Louisville, KY 40202. E-mail: david.gozal{at}louisville.edu


   ABSTRACT
TOP
 ABSTRACT
METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Intermittent hypoxia (IH) during sleep, a critical feature of sleep apnea, induces significant neurobehavioral deficits in the rat. Cyclooxygenase (COX)-2 is induced during stressful conditions such as cerebral ischemia and could play an important role in IH-induced learning deficits. We therefore examined COX-1 and COX-2 genes and COX-2 protein expression and activity (prostaglandin E2 [PGE2] tissue concentration) in cortical regions of rat brain after exposure to either IH (10% O2 alternating with 21% O2 every 90 seconds) or sustained hypoxia (10% O2). In addition, the effect of selective COX-2 inhibition with NS-398 on IH-induced neurobehavioral deficits was assessed. IH was associated with increased COX-2 protein and gene expression from Day 1 to Day 14 of exposure. No changes were found in COX-1 gene expression after exposure to hypoxia. IH-induced COX-2 upregulation was associated with increased PGE2 tissue levels, neuronal apoptosis, and neurobehavioral deficits. Administration of NS-398 abolished IH-induced apoptosis and PGE2 increases without modifying COX-2 mRNA expression. Furthermore, NS-398 treatment attenuated IH-induced deficits in the acquisition and retention of a spatial task in the water maze. We conclude that IH induces upregulation and activation of COX-2 in rat cortex and that COX-2 may play a role in IH-mediated neurobehavioral deficits.

Key Words: sleep apnea • neurocognitive deficits • inflammation • episodic hypoxia

Obstructive sleep apnea, a condition characterized by repeated episodes of upper airway obstruction during sleep, affects 2–5% of the general population and leads to substantial neurocognitive dysfunction (1). We have recently established that exposure to intermittent hypoxia (IH) during the sleep cycle of adult rats is associated with significant spatial learning deficits as well as with increased neuronal apoptosis within susceptible brain regions such the hippocampus and cortex (2). However, the molecular mechanisms underlying IH-associated neurobehavioral deficits are currently unknown.

Cyclooxygenase (COX)-2 is constitutively present in selected brain neurons (3–5), and its expression is upregulated in several neurologic diseases, including stroke (6), Alzheimer's disease (7, 8), and seizures (9). Furthermore, mounting evidence points to COX-2 as being involved in mechanisms of ischemic brain injury (9–15), as evidenced by the significant reduction in brain injury in COX-2–deficient mice after occlusion of the middle cerebral artery (15). In further support of this notion, administration of selective COX-2 inhibitors attenuates ischemic brain injury (15–18). Taken together, these studies suggest that COX-2 is involved in a wide range of degenerative brain injury and that COX-2 could be involved in hypoxia-induced neuronal loss because the induction of COX-2, but not COX-1 gene expression, was linked to apoptotic neuron death (19). The increases in apoptosis found in our rodent model could therefore be mediated by COX-2 induction.

Based on previously mentioned considerations, we hypothesized that COX-2 expression and activity may be upregulated by hypoxia and that inhibition of COX2 activity may palliate hypoxia-induced neuronal injury and spatial memory deficits in the rat.


   METHODS
TOP
 ABSTRACT
 METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Hypoxic Exposures
Male Sprague Dawley rats (172–225 g) were randomly assigned to three experimental groups consisting of (1) chronic hypoxia (CH), 10% O2 for 12 hours during daylight phase; (2) IH, alternating 10% O2 and 10% O2 every 90 seconds for 12 hours during daylight; and (3) normoxic exposures throughout as the control (RA) (see the online supplement). For each exposure time point, four rats were assigned to studies of COX-1 and COX-2 mRNA, COX-2 protein, and prostaglandin E2 (PGE2) concentration, whereas eight rats were assigned to the NS-398 study.

NS-398 Administration
The IC50 of NS-398 for COX-2 is 168 times smaller than that for COX-1 (20). NS-398 effectively decreases PGE2 accumulation in postischemic brain (15–18). It has been also reported that NS-398 inhibits not only the synthesis of PGE2 but also other prostaglandins, such as PGI2 (21). NS-398 (15 mg/kg/day, intraperitoneally; Cayman Chemical, Ann Arbor, MI) or vehicle were administered daily in rats exposed to IH or RA for 7 days. Cortical samples were harvested 2 hours after the last dose of NS398 and were kept at -80°C for PGE2 and COX-2 mRNA measurements.

Morris Water Maze
The Morris water maze was used for all behavioral testing (see the online supplement). After acclimatization procedures, animals were given three daily training sessions consisting of three blocks of four training trials, followed by probe trials and cued trials. The swim distances (path lengths) derived from the reference memory task (averaged over blocks of four trials) were analyzed using repeated-measures analyses of variance. The relative average proximity of the rat to the target platform location during the probe trials was analyzed using one-way analysis of variance.

Western Blotting Cortical Tissues were Homogenized by Standard Procedures
Protein concentration was determined by the Lowry protein assay. Homogenate proteins (50 µg) were heated for 10 minutes at 90°C and then loaded onto gradient (4–12%) sodium dodecyl sulfate gels. The protein was transferred electrophoretically onto nitrocellulose membranes. The membranes were blocked with 5% nonfat dry milk and subsequently incubated overnight at 4°C with primary anti-COX-2 antibody (1:1,000; Exalpha Biological, Boston, MA). Specific proteins was detected with horseradish peroxidase-conjugated anti-mouse secondary antibodies (1:5,000; Santa Cruz Biotechnology, Santa Cruz, CA) and visualized by enhanced chemiluminescence reagents (Amersham, Piscataway, NJ). Prestained molecular weight (MW) standards were used as markers. For loading control across lanes, ß-actin immunoreactivity was assessed by reprobing all membranes. Densitometric analysis was performed with a gel scanning densitometer (Molecular Dynamics, Sunnyvale, CA).

Immunohistochemistry
Four groups of rats (RA + vehicle, RA + NS398, IH + vehicle, and IH + NS398, n = 8 per group) were anesthetized with pentobarbital (50 mg/kg intraperitoneally) and perfused transcardially with 200 ml of phosphate-buffered saline (PBS) at ambient temperature and then with 2.5% paraformaldehyde in cold PBS containing 5% sucrose, pH 7.4. The brain was removed immediately from the skull after perfusion and placed overnight in a fixative containing 1% paraformaldehyde in PBS and 30% sucrose at 4°C. Coronal sections (40 µm) were cut on a freezing microtome, washed extensively in PBS, incubated in 0.4% triton X-100 in PBS containing 1.5% normal goat serum for 1 hour, and incubated with cleaved caspase 3 antibody (1:2,500; Cell Signaling Technology, Beverly, CA). Once the primary antibody reaction was completed, the sections were then washed extensively in PBS, incubated in biotinylated anti-rabbit IgG (Vector Laboratories, Burlingame, CA) diluted in 0.4% triton X-100 in PBS for 1 hour, washed three times in PBS, incubated for 1 hour in avidin-biotinylated horseradish peroxidase (Vectastain Elite kit; Vector) diluted in 0.4% triton X-100 in PBS, rinsed 3 times in Tris (pH 7.6), and incubated in 0.05% diaminobenzidine tetrahydrochloride and 0.005% H2O2 diluted in Tris (pH 7.6) for 1–3 minutes. The reaction was stopped in PBS, and the sections were mounted onto slides. Sections were assessed using a Nikon Ellipse E800 microscope (Nikon USA, Melville, NY), and images were taken by SPOT digital camera (Diagnostic Instruments, Inc., Sterling Heights, MI). Five adjacent sections were assessed for every animal, and the mean number of cleaved caspase 3 positively labeled cells per section was then calculated for the group (n = 8).

Quantitative Polymerase Chain Reaction
cDNA equivalent to 20 ng of total RNA were subjected to real-time polymerase chain reaction analysis (MX4000; Stratagene, La Jolla, CA) with the following primers and probes: COX2: forward primer, 5'-ctgttacaaggagggaaatg-3', reverse primer, 5'-tcggttgctcctaacatagt-3'; Taqman probe, 5'-(FAM)-tgcattgttggttatgactgtgtca-(BHQ-1)-3'; COX-1: forward primer, 5'-gagtctctcgctccagtttcc-3'; reverse primer, 5'-agggaatgactggtgagggta-3'; Taqman probe, 5'-(FAM)-tgctgctgctcctgctgctgct-(BHQ-1)-3'; and ß-actin: forward primer, 5'-acccagatcatgtttgagac-3'; reverse primer 5'-gcatacagggacaacacag-3'; Taqman probe, 5'-(FAM)-agccatgtacgtagccatccag-(BHQ-1)-3'. Standard curves for COX-1, COX-2, and ß-actin were included in each reaction (see the online supplement). Real-time polymerase chain reaction results were analyzed using MX4000 software (Stratagene).

PGE2 Enzyme Immunoassay
Cortical tissue concentrations of PGE2 were determined using a commercially available enzyme immunoassay kit (Oxford Biomedical Research, Oxford, MI; see online supplement).

Data Analysis
Data in text and figures are expressed as mean ± SE. Two group comparisons were evaluated by paired or unpaired t tests, as appropriate. Multiple comparisons were analyzed by analysis of variance and Tukey's or Newman Keuls post hoc tests. Differences were considered statistically significant for p < 0.05.


   RESULTS
TOP
 ABSTRACT
 METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
IH Induces a Significant Increased Expression of COX 2 Protein and mRNA
To establish whether alteration of COX-2 expression in cortical tissue occurs after exposure to CH or IH, COX-2 protein expression was assessed by Western blotting. Cortical COX-2 protein expression was unchanged after exposure to CH (p > 0.05; Figure 1A) . In contrast, IH induced marked increases in COX-2 protein expression at Day 1 (p < 0.01), which were sustained throughout IH exposures (Figure 1A). COX-2 mRNA expression after exposure to IH or CH was measured by quantitative polymerase chain reaction. COX-2 mRNA was slightly increased at 1 and 3 days of CH exposure (p < 0.05) but not thereafter. However, COX-2 mRNA was significantly elevated in the cortical tissue after IH exposure, peaking at Day 1 and remaining elevated until Day 14 (p < 0.01; Figure 1B). No significant changes was detected in COX-1 mRNA after IH exposure (p > 0.05; Figure 1C).



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  		Figure 1. (A) Effect of chronic hypoxia (CH) or intermittent hypoxia (IH) exposure on cyclooxygenase (COX)-2 protein expression. After exposure to CH or IH for 1, 3, 7, and 14 days, cortical tissues were collected and COX-2 protein expression was assessed by Western blotting (n = 4 per group). The upper panel represents the Western blots of COX-2 and ß-actin after CH or IH exposure. The lower panel represents the densitometry analysis of COX-2 Western blots. Data were expressed as densitometry units normalized by ß-actin (mean ± SE). Densitometry analysis revealed significant increases in COX-2 protein expression after IH exposure (*p < 0.01 vs. normoxia [RA]) but not after CH exposure (p > 0.05). (B) Time course of COX-2 mRNA expression in rat cortical tissue. After exposure to IH for 1, 3, 7, and 14 days, cortical tissues were collected and the expression of COX-2 mRNA was assessed by quantitative real-time polymerase chain reaction (n = 4). Data were expressed as cDNA copy number normalized by ß-actin (mean ± SE). Data analysis showed that CH induced a slight increase of COX-2 mRNA only on Days 1 and 3 (*p < 0.05 vs. RA). In contrast, IH induced marked increases of COX-2 mRNA in cortical tissue over time (*p < 0.01 vs. RA). (C) Time course of COX-1 mRNA expression in rat cortical tissue. After exposure to CH or IH for 1, 3, 7, and 14 days, cortical tissues were collected and COX-1 mRNA was measured by quantitative real-time polymerase chain reaction (n = 4). Data were expressed as cDNA copy number and normalized for ß-actin (mean ± SE). No significant changes occurred in COX-1 mRNA after exposure to either CH or IH (p > 0.05 vs. RA).

 
IH Increases PGE2 Induction in Cortical Tissue
To determine whether the increased expression of COX-2 protein and mRNA is associated with induction in COX-2 enzymatic activity, the concentration of PGE2 was measured. No significant changes in PGE2 concentration occurred after exposure to CH at any given time point (Figure 2A) . In contrast, PGE2 concentrations were markedly increased after exposure to IH form at Day 1 (p < 0.01) and remained elevated until Day 14 (p < 0.01; Figure 2A).



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  		Figure 2. (A) Time course of prostaglandin E2 (PGE2) induction in rat cortical tissue. After exposure to either CH or IH for 1, 3, 7, and 14 days, cortical tissues were collected and levels of PGE2 were measured by immunoassay (n = 4). Data were expressed as ng/mg protein (mean ± SE). The production of PGE2 was significantly increased by IH exposure (*p < 0.01 vs. RA) but remained unaltered after CH exposure (p > 0.05 vs. RA). (B) Effect of NS-398 on IH-induced elevation of PGE2. During a 7-day exposure to RA or IH, rats were administered daily with either NS-398 or vehicle. After exposure, cortical tissues were collected, and PGE2 tissue concentrations were measured by immunoassay (n = 8). Data were expressed as ng/mg protein (mean ± SE). There was a significant increase in PGE2 concentrations after IH exposure (*p < 0.01 vs. RA), and NS-398 treatment markedly reduced PGE2 concentrations (#p < 0.01 vs. IH + vehicle [V]). (C) Effect of NS-398 on IH-induced expression of COX-2 mRNA. During a 7-day exposure to RA or IH, rats were administered daily with either NS-398 or vehicle. After exposure, cortical tissues were collected, and expression of COX-2 mRNA was assessed by quantitative polymerase chain reaction (n = 8). Data were expressed as cDNA copy number after normalization with ß-actin. There was a significant increase of COX-2 mRNA after exposure to IH (*p < 0.01 vs. RA). NS-398 did not significantly alter IH-induced expression of COX-2 mRNA (p > 0.05 vs. IH + V).

 
NS-398 Blocks IH-induced PGE2 without Altering COX2 mRNA
To ascertain that COX-2 inhibition with NS398 would effectively suppress IH-induced increase of PGE2 and COX-2 mRNA, the induction of PGE2 and expression of COX-2 mRNA in cortical tissue were assessed after 7 days of treatment with NS-398 or vehicle. The treatment of NS-398 suppressed PGE2 synthesis in both RA and IH treatment groups (p < 0.01; Figure 2B). However, there was no significant modification on IH-induced expression of COX2 mRNA after treatment with NS-398 (p > 0.05 vs. IH + V; Figure 2C).

NS-398 Decreases IH-induced Neuronal Apoptosis
To examine whether administration of NS398 would decrease IH-induced neuronal apoptosis, cleaved (i.e., activated) caspase 3 expression was determined after exposure to either RA or IH treated with NS-398 or vehicle. Cleaved caspase 3 expression was significantly increased after IH exposure in animals treated with vehicle (Figure 3) , suggesting that IH exposure induced robust neuronal apoptosis in the cortical tissue. In addition, IH-induced increases of activated caspase 3 expression were markedly attenuated by treatment with NS-398 (Figure 3).



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  		Figure 3. (A) Effect of NS-398 on cleaved caspase 3 expression after exposure to IH or RA for 2 days, in which rats were treated daily with either NS-398 or V. After exposure, brains sections were assessed by immunohistochemistry (n = 8). Positively labeled cells for cleaved caspase 3 were significantly increased in the IH + V sections (left panel of A; scale bar, 50 µm for low magnification; scale bar = 5 µm for high magnification). In contrast, the number of cleaved caspase 3 positively labeled cells was markedly decreased with NS-398 treatment (right panel of A; scale bar= 50 µm). (B) Effect of NS-398 or vehicle on the number of cleaved caspase 3 positively stained cells after a 2-day exposure to RA or IH (n = 8). Data are expressed as mean number of positively stained cells of caspase 3 per section per animal. Cleaved caspase 3 positively labeled cells were significantly increased in IH + V (*p < 0.01 vs. RA + V), and NS-398 treatment was associated with normalization of cleaved caspase 3 expression (#p < 0.01 vs. IH + V).

 
NS-398 Attenuates IH-induced Neurobehavioral Deficits
During training trials, vehicle-treated rats exposed to IH showed significantly prolonged latencies and path lengths compared with vehicle-treated RA, NS-398–treated RA, and NS-398–treated IH animals during the third block of trials, that is, when spatial acquisition of the task should have occurred (p < 0.002, analysis of variance; Figure 4A) . Similarly, during probe trials, IH-exposed animals treated with vehicle displayed significantly lower spatial bias (reduced proximity to the platform) compared with any of the three other experimental groups (p < 0.01; Figure 4B). Post hoc analyses further confirmed that IH-exposed animals treated with NS-398 were not significantly different from control subjects, suggesting that NS-398 attenuated IH-induced deficits in spatial learning and retention. No group differences were observed on the cued task, indicating that the effects of either IH or NS-398 were not due to sensorimotor impairments.



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  		Figure 4. (A) Mean swim distances (cm) to locate the target platform during place training in rats exposed to 14 days of IH during sleep (open symbols) or room air (RA; filled symbols), receiving either vehicle (V; square symbols), or NS-398 treatments (circle symbols) (n = 8 per group; *p < 0.001, analysis of variance). (B) Proximity to platform location in rats exposed to either RA or to 7-days IH after administration of either vehicle or NS-398 to determine spatial bias after training in the Morris water maze. Data are expressed in centimeters (mean ± SE; n = 8; *p < 0.01 vs. RA control subjects, #p < 0.01 vs. IH + V).

 

   DISCUSSION
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
In this study, we found that COX2 mRNA was induced by IH and CH, albeit to a greater magnitude and duration in the earlier compared with the latter. In contrast, COX-2 protein expression was only increased after intermittent hypoxic exposures of freely behaving rats during their sleep circadian phase. These changes were associated with corresponding increases in PGE2 concentrations in cortical tissue. Furthermore, administration of the selective COX2 inhibitor NS-398 not only abolished the enhanced generation of PGE2 but also attenuated IH-induced apoptosis and neurobehavioral deficits, suggesting that the latter are, at least in part, due to increased expression of COX-2 in the brain.

The original assumption, based on work with inflammation cells, was that COX-1 was necessary to produce low basal amounts of prostaglandins for physiologic purposes, whereas the inducible COX-2 generated prostanglandins in larger amounts and contributed to tissue injury. This view appears to have been substantially challenged in recent years, as mice lacking COX-2 display significant renal abnormalities (22) and COX-2–derived products are produced by the normal vasculature (23) and are required for successful reproduction (24). Furthermore, COX-2 is constitutively expressed in neurons (3) and is dynamically regulated by physiologic synaptic activity (4, 5). These observations together with additional evidence supporting a role for COX-2 in spatial memory consolidation (25) suggest that COX-2 is an important modulator of neuronal function during physiologic conditions. However, in conditions leading to COX-2 induction, such as repeated spreading depression (26, 27), stroke (6, 12, 13), or excitotoxicity induced by glutamatergic agonists (11, 28–31), COX-2 activity appears to play a critical role in neuronal cell injury and death. Because hypoxia leads to excessive release of glutamate (32, 33) and activation of glutamatergic receptors in the brain that in turn can induce COX-2 (30) and because the opposite is also possible (i.e., COX-2 activation by hypoxia can lead to release of glutamate) (34), we focused our attention on the changes in COX-2 expression and activity after exposure during the lights-on period to our recently developed IH model of sleep-disordered breathing. Of note, this model employs a relatively mild hypoxic stimulus that is not associated with either behavioral or metabolic evidence of cellular energy limitation (2). However, to establish further whether mild hypoxia or whether the mode of presentation of the mild hypoxic stimulus were of importance in the process of inducing COX-2 in rat brain cortex, we incorporated CH exposures of similar magnitude to the experimental design. Our findings indicate that IH leads to an induction of COX-2 mRNA and upregulation of COX-2 protein, resulting in substantial increases in PGE2 production. Thus, the cyclical nature of hypoxia during sleep, rather than hypoxia per se, may lead to increased oxidative stress (35), and the latter may underlie the signaling pathways mediating COX-2 induction. One possible explanation for the discrepancy between IH and CH could relate to either difference in the oxidative stress each of these hypoxic stimuli generates, or alternatively, the differences in COX-2 induction between IH and CH could represent post-translational modifications associated with adaptation processes. In addition, IH could be associated with upregulation of proinflammatory cytokines, which would in turn lead to increased COX-2 expression (36). In support of the latter contention, we found that IH, but not CH, elicit increased mRNA expression of interleukin-1 in cortical tissue (data not shown).

There is increasing evidence that COX-2 is involved in the pathogenesis of several neurologic disorders including cerebral ischemia (10, 11, 13–15, 17), traumatic brain injury (37–41), and Alzheimer disease (7, 8). Cerebral ischemia is associated with prominent upregulation of COX-2 in neurons, inflammatory cells, glial cells, and the vasculature (3, 10, 14, 42) and administration of the selective COX-2 inhibitor, NS-398, or genetic deletion of COX-2 in mice will substantially attenuate cerebral ischemic injury (11, 12, 15–18), suggesting that COX-2 is mechanistically involved in the mechanisms leading to cerebral ischemic injury.

In preliminary studies, we found that the nadir of IH-induced spatial learning deficits occurred at 7 days of exposure, and therefore, we selected this time point for the COX-2 inhibitor experiments. To further establish relationships between IH-induced COX-2 and IH-induced neurobehavioral deficits, animals were treated with NS-398, a selective COX-2 inhibitor with 168 times higher affinity for COX-2 compared with COX-1 (20). NS398 attenuated COX-2–mediated neurotoxicity in vitro by blocking lipopolysaccharide-induced elevation of PGE2 (43, 44) as well as NMDA excitotoxicity (27, 31, 45, 46). Similarly, in vivo treatment with NS398 reduced the infarct size after occlusion of middle cerebral artery in mice (12). In this study, NS-398 attenuated induction of caspase 3 during IH, blocked IH-induced PGE2 elevation, and attenuated the neurobehavioral deficits, strongly suggesting that COX-2 expression and activity are critical contributors to the adverse functional outcomes associated with episodic hypoxia during sleep. Of note, other COX-2 inhibitors, such as celecoxib and rofecoxib, are now routinely employed in clinical practice. In this study, we have established that NS398 crosses the blood–brain barrier, whereas it remains unclear whether other clinically used COX-2 inhibitors will do so. The mechanisms whereby COX-2 and its reaction product PGE2 contribute to IH-induced apoptosis remain to be defined (2, 45, 46). COX2 activity and downstream generation of prostanoids and superoxide radicals are likely to be involved. In a recent study, Kondo and colleagues (47) suggested that prostaglandin D2 undergoes dehydration to yield bioactive cyclopentenone-type prostaglandins of the J2 series, such as 15-deoxy-Delta(12,14)-prostaglandin J2. They further showed that this compound induced apoptotic cell death in a neuroblastoma cell line through accumulation of p53 and activation of a death-inducing caspase cascade involving Fas and the Fas ligand (47). Thus, several prostanoids generated through COX-2 activation could directly induce neuronal cell death mechanisms (43). Alternatively, PGE2 could also enhance the induction of cytokines and thereby contribute to neurotoxicity (36, 48, 49) or could exacerbate neurotoxicity by facilitating astrocytic glutamate release (49). In fact, COX-2 inhibition by NS-398 protects neuronal cultures from glutamate neurotoxicity (31) and the damage produced by N-methyl-D-aspartate microinjection is attenuated in COX-2–null mice (11). It should be also stressed that COX-2 inhibition per se could lead to apoptosis in selected biologic substrates, possibly through alteration in mitochondrial calcium homeostasis (50). Although we did not find evidence for such occurrence in our rodent model, it is unclear whether long-term treatment with COX-2 inhibitors may lead to increased apoptosis in neural tissue.

In summary, we have shown that COX-2 plays a prominent role in the pathophysiologic mechanisms mediating spatial learning deficits associated with the episodic hypoxia of sleep-disordered breathing. These findings are consistent with the concept whereby IH triggers the initiation of an inflammatory cascade in vulnerable brain regions involving COX-2 increased expression and activity. It is therefore possible that COX-2 inhibitors may have potential therapeutic applications for the prevention of obstructive sleep apnea–associated neurobehavioral deficits.


   FOOTNOTES
 
Supported by National Institutes of Health grants HL69932, HL63912, and HL66385; American Heart Association grant AHA-0050442N; the Commonwealth of Kentucky Research Challenge Trust Fund; and F32 HD42396 from the National Institutes of Health (B.W.R.).

This article has an online supplement, which is accessible from this issue's table of contents online at www.atsjournals.org

Received in original form November 2, 2002; accepted in final form May 21, 2003


   REFERENCES
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

  1. Beebe DW, Gozal D. Obstructive sleep apnea and the prefrontal cortex: towards a comprehensive model linking nocturnal upper airway obstruction to daytime cognitive and behavioral deficits. J Sleep Res 2002;11:1–16.[Medline]
  2. Gozal D, Daniel JM, Dohanich GP. Behavioral and anatomical correlates of chronic episodic hypoxia during sleep in the rat. J Neurosci 2001;21:2442–2450.[Abstract/Free Full Text]
  3. Breder CD, Dewitt D, Kraig RP. Characterization of inducible cyclooxygenase in rat brain. J Comp Neurol 1995;355:296–315.[CrossRef][Medline]
  4. Kaufmann WE, Worley PF, Pegg J, Bremer M, Isakson P. COX-2, a synaptically induced enzyme, is expressed by excitatory neurons at post-synaptic sites in rat cerebral cortex. Proc Natl Acad Sci USA 1996;93:2317–2321.[Abstract/Free Full Text]
  5. Yamagata K, Andreasson KI, Kaufmann WE, Barnes C, Worley PF. Expression of a mitogen-inducible cyclooxygenase in brain neurons: regulation by synaptic activity and glucocorticoids. Neuron 1993;11:371–386.[CrossRef][Medline]
  6. del Zoppo G, Ginis I, Hallenbeck JM, Iadecola C, Wang X, Feuerstein GZ. Inflammation and stroke: putative role for cytokines, adhesion molecules and iNOS in brain response to ischemia. Brain Pathol 2000;10:95–112.[Medline]
  7. McGeer PL, McGeer EG. Inflammation, autotoxicity and Alzheimer disease. Neurobiol Aging 2001;22:799–809.[CrossRef][Medline]
  8. Xiang Z, Ho L, Valdellon J, Borchelt D, Kelley K, Spielman L, Aisen PS, Pasinetti GM. Cyclooxygenase (COX)-2 and cell cycle activity in a transgenic mouse model of Alzheimer's disease neuropathology. Neurobiol Aging 2002;23:327–334.[CrossRef][Medline]
  9. McCown TJ, Knapp DJ, Crews FT. Inferior collicular seizure generalization produces site-selective cortical induction of cyclooxygenase 2 (COX-2). Brain Res 1997;767:370–374.[CrossRef][Medline]
  10. Iadecola C, Forster C, Nogawa S, Clark HB, Ross ME. Cyclooxygenase-2 immunoreactivity in the human brain following cerebral ischemia. Acta Neuropathol (Berl) 1999;98:9–14.[CrossRef][Medline]
  11. Iadecola C, Niwa K, Nogawa S, Zhao X, Nagayama M, Araki E, Morham S, Ross ME. Reduced susceptibility to ischemic brain injury and N-methyl-D-aspartate-mediated neurotoxicity in cyclooxygenase-2-deficient mice. Proc Natl Acad Sci USA 2001;98:1294–1299.[Abstract/Free Full Text]
  12. Nogawa S, Zhang F, Ross ME, Iadecola C. Cyclo-oxygenase-2 gene expression in neurons contributes to ischemic brain damage. J Neurosci 1997;17:2746–2755.[Abstract/Free Full Text]
  13. Sairanen T, Ristimaki A, Karjalainen-Lindsberg ML, Paetau A, Kaste M, Lindsberg PJ. Cyclooxygenase-2 is induced globally in infarcted human brain. Ann Neurol 1998;43:738–747.[CrossRef][Medline]
  14. Toti P, De Felice C, Schurfeld K, Stumpo M, Bartolommei S, Lombardi A, Petraglia E, Buonocore G. Cyclooxygenase-2 immunoreactivity in the ischemic neonatal human brain: an autopsy study. J Submicrosc Cytol Pathol 2001;33:245–249.[Medline]
  15. Nagayama M, Niwa K, Nagayama T, Ross ME, Iadecola C. The cyclooxygenase-2 inhibitor NS-398 ameliorates ischemic brain injury in wild-type mice but not in mice with deletion of the inducible nitric oxide synthase gene. J Cereb Blood Flow Metab 1999;19:1213–1219.[CrossRef][Medline]
  16. Domoki F, Perciaccante JV, Puskar M, Bari F, Busija DW. Cyclooxygenase-2 inhibitor NS398 preserves neuronal function after hypoxia/ischemia in piglets. Neuroreport 2001;12:4065–4068.[CrossRef][Medline]
  17. Hara K, Kong DL, Sharp FR, Weinstein PR. Effect of selective inhibition of cyclooxygenase 2 on temporary focal cerebral ischemia in rats. Neurosci Lett 1998;256:53–56.[CrossRef][Medline]
  18. Hurley SD, Olschowka JA, O'Banion MK. Cyclooxygenase inhibition as a strategy to ameliorate brain injury. J Neurotrauma 2002;19:1–15.[CrossRef][Medline]
  19. Ho L, Osaka H, Aisen PS, Pasinetti GM. Induction of cyclooxygenase (COX)-2 but not COX-1 gene expression in apoptotic cell death. J Neuroimmunol 1998;89:142–149.[CrossRef][Medline]
  20. Vane JR, Botting RM. The future of NSAID therapy: selective COX-2 inhibitors. Int J Clin Pract 2000;54:7–9.[Medline]
  21. Futaki N, Takahashi S, Kitagawa T, Yamakawa Y, Tanaka M, Higuchi S. Selective inhibition of cyclooxygenase-2 by NS-398 in endotoxin shock rats in vivo. Inflamm Res 1997;46:496–502.[CrossRef][Medline]
  22. Dinchuk JE, Car BD, Focht RJ, Johnston JJ, Jaffee BD, Covington MB, Contel NR, Eng VM, Collins RJ, Czerniak PM. Renal abnormalities and an altered inflammatory response in mice lacking cyclooxygenase II. Nature 1995;378:406–409.[CrossRef][Medline]
  23. McAdam BF, Catella-Lawson F, Mardini IA, Kapoor S, Lawson JA, FitzGerald GA. Systemic biosynthesis of prostacyclin by cyclooxygenase (COX)-2: the human pharmacology of a selective inhibitor of COX-2. Proc Natl Acad Sci USA 1999;96:272–277.[Abstract/Free Full Text]
  24. Reese J, Zhao X, Ma WG, Brown N, Maziasz TJ, Dey SK. Comparative analysis of pharmacologic and/or genetic disruption of cyclooxygenase-1 and cyclooxygenase-2 function in female reproduction in mice. Endocrinology 2001;142:3198–3206.[Abstract/Free Full Text]
  25. Teather LA, Packard MG, Bazan NG. Post-training cyclooxygenase-2 (COX-2) inhibition impairs memory consolidation. Learn Mem 2002;9:41–47.[Abstract/Free Full Text]
  26. Koistinaho J, Chan PH. Spreading depression-induced cyclooxygenase-2 expression in the cortex. Neurochem Res 2000;25:645–651.[CrossRef][Medline]
  27. Miettinen S, Fusco FR, Yrjanheikki J, Keinanen R, Hirvonen T, Roivainen R, Narhi M, Hokfelt T, Koistinaho J. Spreading depression and focal brain ischemia induce cyclooxygenase-2 in cortical neurons through N-methyl-D-aspartic acid-receptors and phospholipase A2. Proc Natl Acad Sci USA 1997;94:6500–6505.[Abstract/Free Full Text]
  28. Baik EJ, Kim EJ, Lee SH, Moon C. Cyclooxygenase-2 selective inhibitors aggravate kainic acid induced seizure and neuronal cell death in the hippocampus. Brain Res 1999;843:118–129.[CrossRef][Medline]
  29. Chen J, Marsh T, Zhang JS, Graham SH. Expression of cyclo-oxygenase 2 in rat brain following kainate treatment. Neuroreport 1995;6:245–248.[Medline]
  30. Domoki F, Thrikawala N, Robins GS, Bari F, Busija DW. Kainic acid rapidly induces cyclooxygenase (COX)-2 in piglet cerebral cortex. Neuroreport 2000;11:3435–3438.[Medline]
  31. Hewett SJ, Uliasz TF, Vidwans AS, Hewett JA. Cyclooxygenase-2 contributes to N-methyl-D-aspartate-mediated neuronal cell death in primary cortical cell culture. J Pharmacol Exp Ther 2000;293:417–425.[Abstract/Free Full Text]
  32. Phillis JW, Walter GA. Hypoxia/hypotension evoked release of glutamate and aspartate from the rat cerebral cortex. Neurosci Lett 1989;106:147–151.[CrossRef][Medline]
  33. Mitani A, Kadoya F, Nakamura Y, Kataoka K. Visualization of hypoxia-induced glutamate release in gerbil hippocampal slice. Neurosci Lett 1991;122:167–170.[CrossRef][Medline]
  34. Bezzi P, Carmignoto G, Pasti L, Vesce S, Rossi D, Rizzini BL, Pozzan T, Volterra A. Prostaglandins stimulate calcium-dependent glutamate release in astrocytes. Nature 1998;391:281–285.[CrossRef][Medline]
  35. Figueiredo-Pereira ME, Li Z, Jansen M, Rockwell P. N-acetylcysteine and celecoxib lessen cadmium cytotoxicity which is associated with cyclooxygenase-2 up-regulation in mouse neuronal cells. J Biol Chem 2002;277:25283–25289.[Abstract/Free Full Text]
  36. Hoozemans JJ, Veerhuis R, Janssen I, Rozemuller AJ, Eikelenboom P. Interleukin-1beta induced cyclooxygenase 2 expression and prostaglandin E2 secretion by human neuroblastoma cells: implications for Alzheimer's disease. Exp Gerontol 2001;36:559–570.[CrossRef][Medline]
  37. Bidmon HJ, Oermann E, Schiene K, Schmitt M, Kato K, Asayama K, Witte OW, Zilles K. Unilateral upregulation of cyclooxygenase-2 following cerebral, cortical photothrombosis in the rat: suppression by MK-801 and co-distribution with enzymes involved in the oxidative stress cascade. J Chem Neuroanat 2000;20:163–176.[CrossRef][Medline]
  38. Dash PK, Mach SA, Moore AN. Regional expression and role of cyclooxygenase-2 following experimental traumatic brain injury. J Neurotrauma 2000;17:69–81.[Medline]
  39. Cernak I, O'Connor C, Vink R. Activation of cyclo-oxygenase-2 contributes to motor and cognitive dysfunction following diffuse traumatic brain injury in rats. Clin Exp Pharmacol Physiol 2001;28:922–925.[CrossRef][Medline]
  40. Schwab JM, Seid K, Schluesener HJ. Traumatic brain injury induces prolonged accumulation of cyclooxygenase-1 expressing microglia/brain macrophages in rats. J Neurotrauma 2001;18:881–890.[CrossRef][Medline]
  41. Strauss KI, Barbe MF, Marshall RM, Raghupathi R, Mehta S, Narayan RK. Prolonged cyclooxygenase-2 induction in neurons and glia following traumatic brain injury in the rat. J Neurotrauma 2000;17:695–711.[CrossRef][Medline]
  42. Kelley KA, Ho L, Winger D, Freire-Moar J, Borelli CB, Aisen PS, Pasinetti GM. Potentiation of excitotoxicity in transgenic mice overexpressing neuronal cyclooxygenase-2. Am J Pathol 1999;155:995–1004.[Abstract/Free Full Text]
  43. Takadera T, Yumoto H, Tozuka Y, Ohyashiki T. Prostaglandin E (2) induces caspase-dependent apoptosis in rat cortical cells. Neurosci Lett 2002;317:61–64.[CrossRef][Medline]
  44. Araki E, Forster C, Dubinsky JM, Ross ME, Iadecola C. Cyclooxygenase-2 inhibitor ns-398 protects neuronal cultures from lipopolysaccharide-induced neurotoxicity. Stroke 2001;32:2370–2375.[Abstract/Free Full Text]
  45. Kim EJ, Lee JE, Kwon KJ, Lee SH, Moon CH, Baik EJ. Differential roles of cyclooxygenase isoforms after kainic acid-induced prostaglandin E(2) production and neurodegeneration in cortical and hippocampal cell cultures. Brain Res 2001;908:1–9.[CrossRef][Medline]
  46. Mirjany M, Ho L, Pasinetti GM. Role of cyclooxygenase-2 in neuronal cell cycle activity and glutamate-mediated excitotoxicity. J Pharmacol Exp Ther 2002;301:494–500.[Abstract/Free Full Text]
  47. Kondo M, Shibata T, Kumagai T, Osawa T, Shibata N, Kobayashi M, Sasaki S, Iwata M, Noguchi N, Uchida K. 15-Deoxy-Delta(12,14)-prostaglandin J(2): the endogenous electrophile that induces neuronal apoptosis. Proc Natl Acad Sci USA 2002;99:7367–7372.[Abstract/Free Full Text]
  48. Blom MA, van Twillert MG, de Vries SC, Engels F, Finch CE, Veerhuis R, Eikelenboom P. NSAIDS inhibit the IL-1 beta-induced IL-6 release from human post- mortem astrocytes: the involvement of prostaglandin E2. Brain Res 1997;777:210–218.[CrossRef][Medline]
  49. Fiebich BL, Schleicher S, Spleiss O, Czygan M, Hull M. Mechanisms of prostaglandin E2-induced interleukin-6 release in astrocytes: possible involvement of EP4-like receptors, p38 mitogen- activated protein kinase and protein kinase C. J Neurochem 2001;79:950–958.[CrossRef][Medline]
  50. Johnson AJ, Hsu AL, Lin HP, Song X, Chen CS. The cyclo-oxygenase-2 inhibitor celecoxib perturbs intracellular calcium by inhibiting endoplasmic reticulum Ca2+-ATPases: a plausible link with its anti-tumour effect and cardiovascular risks. Biochem J 2002;366:831–837.[Medline]



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