IOVS
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

(Investigative Ophthalmology and Visual Science. 2005;46:4139-4146.)
© 2005 by The Association for Research in Vision and Ophthalmology, Inc.
DOI:  10.1167/iovs.05-0258
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via ISI Web of Science (2)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Petropoulos, I. K.
Right arrow Articles by Pournaras, C. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Petropoulos, I. K.
Right arrow Articles by Pournaras, C. J.

Effect of Carbogen Breathing and Acetazolamide on Optic Disc PO2

Ioannis K. Petropoulos,1 Jean-Antoine C. Pournaras,2 Jean-Luc Munoz,1 and Constantin J. Pournaras1

1From the Department of Ophthalmology, University Hospitals of Geneva, Geneva, Switzerland; and the 2Jules Gonin Eye Hospital, University of Lausanne, Lausanne, Switzerland.


   Abstract
Top
 Abstract
Materials and Methods
 Results
 Discussion
 References
 
PURPOSE. Acetazolamide was previously shown to increase optic disc partial pressure of oxygen (PO2). The study was conducted to evaluate optic disc PO2 variations during normoxia, hyperoxia (100% O2), and carbogen breathing (95% O2, 5% CO2), before and after intravenous administration of acetazolamide.

METHODS. PO2 measurements were obtained at intervascular areas of the optic disc in nine anesthetized minipigs using oxygen-sensitive microelectrodes (10-µm tip diameter) placed at <50 µm from the optic disc. PO2 was measured continuously during 10 minutes under normoxia, hyperoxia, or carbogen breathing. Oxygen measurements were repeated under these conditions after intravenous injection of acetazolamide (500-mg bolus).

RESULTS. In hyperoxia, optic disc PO2 increased moderately ({Delta}PO2 = 4.81 ± 1.16 mm Hg (mean ± SD; 24%; P < 0.001) after a much larger increase in systemic PaO2. Carbogen breathing induced a significant increase in both systemic PaO2 and PaCO2, which resulted in a large increase in optic disc PO2 ({Delta}PO2 = 13.17 ± 2.18 mm Hg; 67%; P < 0.001). Acetazolamide induced a slow and progressive increase in both systemic PaCO2 and optic disc PO2 (30 minutes {Delta}PO2 = 4.24 ± 2.45 mm Hg; 24%; P < 0.04). However, it was when carbogen was simultaneously administered that optic disc PO2 increased most substantially ({Delta}PO2 = 18.91 ± 5.23 mm Hg; 90%; P < 0.002).

CONCLUSIONS. Carbogen breathing increases optic disc PO2 significantly in minipigs, more than hyperoxia. The association of acetazolamide injection with carbogen breathing could induce an additional increase in optic disc PO2 through the effect of higher systemic PaCO2.

Blood flow modifications and perturbations of the autoregulatory mechanisms of the optic nerve head (ONH) circulation have been incriminated in several ocular disorders, including glaucomatous and ischemic optic neuropathy.1 2 Knowledge of the parameters that can influence the autoregulatory responses of the ONH might improve our understanding of the pathogenesis of those diseases and might lead to new therapeutic modalities.

Retinal and ONH blood flow is closely dependent on systemic partial pressure of oxygen (PaO2) and systemic partial pressure of carbon dioxide (PaCO2). High PaO2 (hyperoxia) causes vasoconstriction of cerebral arteries3 and retinal arterioles and a decrease in retinal blood flow.4 5 However, hyperoxia does not affect the tissue oxygen supply, as tissue partial pressure of oxygen (PO2) remains relatively stable in the inner retina6 7 or at intervascular areas of the ONH.8 9 10

In contrast, carbon dioxide is a well-known dilator of arterial circulation, and high PaCO2 has been shown to dilate retinal arterioles and to increase retinal blood flow in monkeys11 and in humans.12 Furthermore, carbogen breathing (95% O2, 5% CO2), which has been shown to increase brain tumor oxygenation,13 has also been reported to increase preretinal PO2 in cats,14 rats,15 and minipigs.16 Carbogen may also be effective in increasing oxygen delivery in the ONH, though it has not yet been shown to increase optic disc PO2 in an experimental setting.

Blood flow can be influenced by several other chemical substances or drugs. Intravenous administration of acetazolamide has been shown to increase cerebral blood flow17 18 19 and has been used as a provocation test to assess cerebrovascular reserve capacity.20 21 It has also been reported to increase retinal22 and choroidal blood flow,23 as well as optic disc PO2.10 24 High PaCO2 leading to arterial vasodilation25 26 is thought to be the mechanism of increased tissue blood flow by acetazolamide. CO2-associated vasodilation is apparently mediated in the extravascular tissue through acidification of the interstitial space of the inner retina,25 a mechanism similar to that occurring at the cerebral cortex.27

Provided there is no change in tissue oxygen consumption, tissue oxygen tension reflects corresponding blood flow variations28 and enables an accurate estimation of tissue oxygen supply in the physiological state or ischemic disease. Optic disc PO2 was first measured in vivo with microelectrodes in cats29 and in monkeys.30 Thereafter, PO2 measurements were obtained within the ONH tissue at various depths in minipigs8 9 and in cats.31 A dose-dependent significant increase in optic disc PO2 in pigs after intravenous administration of acetazolamide or dorzolamide using polarographic oxygen electrodes placed in front of the optic disc was recently described.24 However, the effect of carbogen inhalation on the optic disc oxygenation has not been tested. It would therefore be reasonable to see whether carbogen is sufficient to increase optic disc PO2, as well as to test the effect of the combined use of acetazolamide and carbogen on optic disc PO2. This may be useful in defining new therapeutic modalities for ischemic diseases of the optic nerve.

Based on these considerations, we sought in the present study to evaluate the variations of optic disc PO2 during normoxia, systemic hyperoxia (breathing of 100% O2), and carbogen breathing (95% O2, 5% CO2), before and after intravenous administration of acetazolamide.


   Materials and Methods
Top
 Abstract
 Materials and Methods
Results
 Discussion
 References
 
Experiments were conducted in one eye of 9 minipigs (Arare Animal Facility, Geneva, Switzerland) weighing 10 to 12 kg. The advantage of using the minipig is the anatomic similarity of its optic nerve to the optic nerve of the primates,32 except that the retinal arteries arise from the ciliary circulation.32 All the experiments were performed in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research.

Animal Preparation
Minipigs were prepared for the experiments, as previously described.7 In brief, after the minipigs received intramuscular injection of 2 mL (10 mg) of the tranquilizer midazolam maleate (Dormicum; Roche Pharma, Reinach, Switzerland), 3 mL (120 mg) of the tranquilizer azaperon (Stresnil; Janssen Pharmaceutica, Beerse, Belgium), and 1 mL (0.5 mg) of atropine, anesthesia was induced with 2 to 3 mg thiopental sodium (Pentothal; Abbott AG, Baar, Switzerland) injected into the ear vein. Curarization was performed with 2 mL (4 mg) of pancuronium bromide (Pavulon; Organon SA, Pfäffikon, Switzerland). The animals were intubated and artificially ventilated. After arterial, venous, and bladder catheterization, anesthesia and myorelaxation were maintained throughout the experiment by continuous perfusion of thiopental and pancuronium, respectively.

Each animal was ventilated at approximately 18 strokes/min, with a continuous flow of 20% O2 and 80% N2O, through a variable-volume respirator. Systolic, diastolic, and mean arterial pressures were monitored through the femoral artery with a transducer. Temperature was maintained between 36°C and 37°C with a thermoblanket. PaO2, PaCO2, and pH were measured intermittently from the same artery with a blood gas analyzer (Labor-system; Flukiger AG, Menziken, Switzerland) and kept under control by adjusting ventilatory rate, stroke volume, and composition of the inhaled gas.

A head-holder was used to avoid movements due to respiration. Upper and lower eyelids were removed, as well as a rectangular area of skin surrounding the eye; the bulbar conjunctiva was detached; the sclera was carefully cleaned to 5 mm from the limbus; the superficial scleral vessels were thermocauterized; the globe was fixed with a metal ring sutured around the limbus; and a sclerotomy at the pars plana was performed. The pupil was dilated with 1% atropine eye drops and the fundus was observed using an operating microscope (Carl Zeiss Surgical GmbH, Oberkochen, Germany).

PO2 Measurements
Optic disc PO2 measurements were obtained using double-barreled, recessed oxygen-sensitive microelectrodes with a 10-µm tip diameter, as previously described.7 The microelectrodes were prepared in our laboratory according to a technique described in detail in previous papers.33 34 They were calibrated before insertion into the eye and again after withdrawal in a buffered saline solution at 35°C, which was equilibrated with air. The microelectrodes were inserted into the vitreous cavity through a sclerotomy placed 2 to 3 mm posterior to the limbus, aided by an electronic micromanipulator.35 The microelectrode tip was positioned at a distance of less than 50 µm from the optic disc, over intervascular areas, as follows: When the microelectrode tip touched the optic disc surface, the reference barrel recorded a sudden negative DC potential shift of several millivolts. At the moment of contact, the position display was zeroed, and the microelectrode was then withdrawn less than 50 µm from the optic disc surface.

The timeline of PO2 measurements was as follows: Baseline measurements under normoxia and a stable continuous recording for at least 10 minutes preceded inhalation of 100% O2 (hyperoxia) for 10 minutes. Then normoxia was again induced, with the purpose of obtaining a stable recording for at least 10 minutes. This recording was considered the baseline before inhalation of carbogen for 10 minutes. Each condition was confirmed by corresponding PaO2, PaCO2, and pH measurements. PaO2 had to be high in hyperoxia, whereas both PaO2 and PaCO2 had to be high during carbogen breathing. The timeline of measurements was performed before and after intravenous injection of acetazolamide (500 mg bolus; Diamox; Vifor SA, Fribourg, Switzerland). Though higher than that used in clinical practice, the dose of 500 mg was chosen in accordance with previous results,10 24 of studies in which various doses had been tested and the dose of 500 mg had been the lowest one to produce maximum effects on optic disc PO2.

Nitrous oxide (N2O) was present in the breathing gas during normoxia, whereas it was absent during hyperoxia or carbogen breathing. However, this was not considered to affect the results, as we have previously tested the effect of the presence or absence of N2O in the breathing gas during normoxia and we have not found any differences in pH, PaCO2, PaO2, or optic disc PO2 (data not shown).

The mean ± SD of optic disc PO2, PaO2, PaCO2, and pH, were calculated at baseline and 7 minutes after initiation of hyperoxia or carbogen breathing in each case.

Statistics
All PO2 levels, as well as their differences from baseline ({Delta}PO2) are expressed as the mean ± SD. For each result presented, n represents the number of optic disc territories where measurements were obtained. An n greater than the number of minipigs means that more than one optic disc territory was analyzed in the same eye. The Wilcoxon signed-rank test was used to detect differences in mean PO2 under each condition. In addition, the Friedman test was performed to compare the respective effect of hyperoxia and carbogen breathing in the same territory at four predetermined time points (2, 5, 7, and 10 minutes). A box-plot representation was used to provide a visual summary of the median values and of the 5%, 25%, 75%, and 95% percentiles. Finally, the Wilcoxon signed-rank test was performed to compare the respective effect of hyperoxia and carbogen breathing before and after intravenous injection of acetazolamide. In all comparisons, P < 0.05 defined statistically significant differences.


   Results
Top
 Abstract
 Materials and Methods
 Results
Discussion
 References
 
Measurements before Administration of Acetazolamide
Under normoxia (PaO2 = 104.09 ± 6.06 mm Hg; PaCO2 = 35.45 ± 2.41 mm Hg; pH = 7.45 ± 0.06; n = 28), mean optic disc PO2 recorded in intervascular areas of nine eyes was 20.11 ± 2.47 mm Hg (n = 28), a level similar to those previously described.8 9 10

The inhalation of 100% O2 induced a mean increase in optic disc PO2 of {Delta}PO2 = 4.81 ± 1.16 mm Hg or 24% (nine eyes; n = 13; Fig. 1 ) after 7 minutes. Under systemic hyperoxia, mean optic disc PO2 increased from 20.36 ± 2.88 to 25.18 ± 3.75 mm Hg and that difference, although moderate, was statistically significant (P < 0.001), yet disproportional to a substantial increase in PaO2 ({Delta}PaO2 = 334.40 ± 39.36 mm Hg; {Delta}PaCO2 = –4.66 ± 2.27 mm Hg; {Delta}pH = 0.03 ± 0.01; n = 13). The effect was fully reversible after return to normoxia.



View larger version (25K):
[in this window]
[in a new window]
  		FIGURE 1. Continuous typical recording of optic disc PO2 under hyperoxia or carbogen breathing before the injection of acetazolamide. The corresponding table shows the mean ± SD of the blood gas levels and the optic disc PO2 variation in each condition. Carbogen breathing induced a greater increase in optic disc PO2 than did hyperoxia. Both hyperoxia and carbogen induced an increase in PaO2, whereas carbogen also induced an increase in PaCO2 and a concomitant respiratory acidosis.

 
The inhalation of carbogen induced a mean increase in optic disc PO2 of {Delta}PO2 = 13.17 ± 2.18 mm Hg or 67% (nine eyes; n = 15; Fig. 1 ) after 7 minutes. Mean optic disc PO2 increased significantly from 19.89 ± 2.13 to 33.05 ± 3.45 mm Hg (P < 0.001). Under the effect of carbogen, both PaO2 and PaCO2 increased significantly ({Delta}PaO2 = 371.72 ± 95.19 mm Hg; {Delta}PaCO2 = 10.31 ± 3.71 mm Hg; n = 15). The CO2 increase induced a respiratory acidosis from a mean pH of 7.45 ± 0.06 to 7.38 ± 0.07 ({Delta}pH = –0.07 ± 0.02; n = 15). The effect was also fully reversible after return to normoxia.

Considering 12 optic disc territories of nine minipigs submitted to the same conditions at four different time points (2, 5, 7, and 10 minutes), the Friedman test revealed a statistically more significant effect of carbogen inhalation on the optic disc PO2 variations with time (P < 0.0001) than of hyperoxia (P < 0.02; Fig. 2 ). Furthermore, at each of the four analyzed times, the Wilcoxon signed-rank test showed a significantly greater effect of carbogen inhalation on the optic disc PO2 variations than that of hyperoxia (P < 0.02 at 2 minutes; P < 0.003 at 5, 7, and 10 minutes; Fig. 2 ).



View larger version (22K):
[in this window]
[in a new window]
  		FIGURE 2. Box plots displaying the optic disc {Delta}PO2 obtained in the same 12 optic disc territories of nine eyes at four different time points (2, 5, 7, and 10 minutes) under hyperoxia (left) or carbogen breathing (right) before the injection of acetazolamide. Error bars: 5th and 95th percentiles of the {Delta}PO2 level. (•) Levels higher or lower than those percentiles. The Friedman test revealed a statistically more significant effect of carbogen breathing on optic disc {Delta}PO2 with time (P < 0.0001; n = 12) compared with hyperoxia (P < 0.02; n = 12). This was further supported by the Wilcoxon signed-rank test.

 
Measurements after Intravenous Administration of Acetazolamide
After inducing a transient decrease in optic disc PO2 due to a transient systemic hypotension (confirmed by our arterial pressure monitoring) related to the injection, intravenous administration of 500 mg of acetazolamide led to a slow and progressive increase in optic disc PO2 (Fig. 3A) in parallel with a slow and progressive increase in PaCO2 (Fig. 3B) , as previously described.10 24 In six intervascular optic disc territories of six eyes, after the injection of acetazolamide and during normoxia, the optic disc PO2 measured 30 minutes later revealed an increase ({Delta}PO2 = 4.24 ± 2.45 mm Hg, or 24%). More specifically, acetazolamide had increased optic disc PO2 from a mean of 18.00 ± 3.08 to 22.13 ± 4.04 mm Hg and that difference, though moderate, was statistically significant (P < 0.04). PaCO2 had simultaneously increased by 31% in average ({Delta}PaCO2 = 10.11 ± 4.42 mm Hg; {Delta}PaO2 = 1.27 ± 6.51 mm Hg; n = 6). The CO2 increase induced a metabolic acidosis from a mean pH of 7.47 ± 0.05 to 7.39 ± 0.04 ({Delta}pH = –0.08 ± 0.03; n = 6).



View larger version (26K):
[in this window]
[in a new window]
  		FIGURE 3. (A) Continuous typical recording of optic disc PO2 during 30 minutes of normoxia after intravenous injection of acetazolamide. Optic disc PO2 increased progressively in a linear pattern under the effect of acetazolamide. (B) Optic disc PO2 mean variation and PaCO2 mean variation during 90 minutes of normoxia after intravenous injection of acetazolamide. Optic disc PO2 increased progressively, in parallel with an increase in PaCO2, in a linear pattern, after treatment with acetazolamide.

 
The inhalation of 100% O2 under the effect of acetazolamide led to a significant increase in optic disc PO2 ({Delta}PO2 = 12.86 ± 4.08 mm Hg, or 58%; six eyes; n = 10; Fig. 4 ) after 7 minutes, from a mean level of 22.97 ± 4.23 to 35.83 ± 5.52 mm Hg (P < 0.002). In hyperoxic conditions, PaO2 increased significantly ({Delta}PaO2 = 374.42 ± 43.77 mm Hg), whereas PaCO2 and pH remained practically stable ({Delta}PaCO2 = –1.42 ± 3.27 mm Hg; {Delta}pH = –0.001 ± 0.03; n = 10).



View larger version (25K):
[in this window]
[in a new window]
  		FIGURE 4. Continuous typical recording of optic disc PO2 under hyperoxia or carbogen breathing after intravenous injection of acetazolamide. The corresponding table shows the mean ± SD of the blood gas levels and the optic disc PO2 variation in each condition. Both hyperoxia and carbogen breathing led to a significant increase of optic disc PO2 under the effect of acetazolamide, but carbogen breathing induced a greater increase in optic disc PO2 than did hyperoxia. Both hyperoxia and carbogen induced an increase in PaO2, whereas carbogen also induced a more marked increase in PaCO2 and a deeper systemic acidosis.

 
During carbogen inhalation under the effect of acetazolamide, the greatest increase in optic disc PO2 was recorded ({Delta}PO2 = 18.91 ± 5.23 mm Hg, or 90%; six eyes; n = 10; Fig. 4 ) after 7 minutes, from a mean of 21.90 ± 5.03 to 40.81 ± 7.70 mm Hg (P < 0.002). Carbogen breathing induced a significant increase in both PaO2 and PaCO2 ({Delta}PaO2 = 404.14 ± 36.82 mm Hg; {Delta}PaCO2 = 10.42 ± 4.05 mm Hg; n = 10), leading to a deeper systemic acidosis from pH = 7.37 ± 0.06 to 7.30 ± 0.06 ({Delta}pH = –0.07 ± 0.02; n = 10).

With the Friedman test, the effect of hyperoxia and carbogen breathing was analyzed at four different time points (2, 5, 7, and 10 minutes) and in nine optic disc territories of six minipigs placed in the same conditions after acetazolamide injection (Fig. 5) . This test revealed the statistically significant effect of both hyperoxia or carbogen inhalation on the optic disc PO2 variations with time at all four analyzed time points (P < 0.0002). Furthermore, the Wilcoxon signed-rank test demonstrated a significantly greater effect of carbogen inhalation on the optic disc PO2 variations compared with hyperoxia (Fig. 5) , at all time points (P < 0.02) except 2 minutes (P < 0.07).



View larger version (27K):
[in this window]
[in a new window]
  		FIGURE 5. Box plots displaying the optic disc {Delta}PO2 obtained in the same nine optic disc territories of six eyes at four different time points (2, 5, 7, and 10 minutes) under hyperoxia or carbogen breathing, after intravenous injection of acetazolamide. Error bars: 5th and 95th percentiles of the {Delta}PO2 level. (•) Levels higher or lower than those percentiles. A Wilcoxon signed-rank test revealed a more significant effect of carbogen breathing on optic disc {Delta}PO2 with time than of hyperoxia, at all time points (P < 0.02; six eyes; n = 9), except 2 minutes (P < 0.07; six eyes; n = 9).

 
In addition, with the Friedman test, the effect of hyperoxia was analyzed at four different time points (2, 5, 7, and 10 minutes) and in the same eight optic disc territories of six minipigs tested before and after acetazolamide injection (Fig. 6) . This test revealed a significantly greater effect of hyperoxia after acetazolamide injection on optic disc PO2 variation with time (P < 0.0005), compared with hyperoxia before acetazolamide injection (P < 0.04). That effect of hyperoxia after acetazolamide injection was confirmed by the Wilcoxon signed-rank test, at all four time points (P < 0.02).



View larger version (23K):
[in this window]
[in a new window]
  		FIGURE 6. Box plots displaying the optic disc {Delta}PO2 obtained in the same eight optic disc territories of six eyes at four different time points (2, 5, 7, and 10 minutes) under hyperoxia before and after intravenous injection of acetazolamide. Error bars: 5th and 95th percentiles of the {Delta}PO2 levels. (•) Levels higher or lower than those percentiles. A Friedman test revealed a statistically more significant effect of hyperoxia on optic disc {Delta}PO2 after acetazolamide injection (P < 0.0005; n = 8) than of hyperoxia before acetazolamide injection (P < 0.04; n = 8) with time. This was further supported by the Wilcoxon signed-rank test.

 
Finally, with the Friedman test, the effect of carbogen inhalation was analyzed at four different time points (2, 5, 7, and 10 minutes) and in the same seven optic disc territories of six minipigs tested before and after acetazolamide injection (Fig. 7) . This test revealed a statistically significant effect of carbogen before and after acetazolamide injection on optic disc PO2 variation with time (P < 0.002). Furthermore, the Wilcoxon signed-rank test demonstrated a significantly greater effect on optic disc PO2 variation with time of carbogen breathing after acetazolamide injection than of carbogen breathing before acetazolamide injection, at all four time points (P < 0.03).



View larger version (30K):
[in this window]
[in a new window]
  		FIGURE 7. Box plots displaying the optic disc {Delta}PO2 levels obtained in the same seven optic disc territories of six eyes at four different time points (2, 5, 7, and 10 minutes) under carbogen breathing before and after intravenous injection of acetazolamide. Error bars: 5th and 95th percentiles of the {Delta}PO2 levels. A Wilcoxon signed-rank test revealed a statistically more significant effect on optic disc {Delta}PO2 of carbogen breathing after acetazolamide injection than of carbogen breathing before acetazolamide injection, with time and at all time points (P < 0.03; n = 9).

 

   Discussion
Top
 Abstract
 Materials and Methods
 Results
 Discussion
References
 
Nutrition and oxygenation of the ONH depends on its blood flow, which is proportional to the perfusion pressure in the ONH and inversely proportional to the resistance to flow. Resistance is closely dependent on PaO2 and PaCO2. As a result, elevated PaO2 decreases ONH blood flow,36 37 whereas elevated PaCO2 increases ONH blood flow,37 to ensure a constant supply of the ONH in oxygen.

Previous findings in minipigs indicate regulation of retinal blood flow during systemic hyperoxia, maintaining preretinal PO2 at a constant level despite the elevation of PaO2.6 7 In addition, previously published data demonstrated that, although juxta-arteriolar tissue PO2 of the ONH increases (due to direct O2 diffusion from the arteriolar wall), intervascular tissue PO2 within the ONH and in front of the optic disc remains practically unchanged.8 9 In our experiments, systemic hyperoxia significantly increased PaO2. This high PaO2 induced a slight increase in optic disc PO2, probably due to O2 diffusion from the arterioles (Fig. 1) .

The slight increase in optic disc PO2 recorded in our experiments during systemic hyperoxia may be related in part to the tip diameter of the microelectrode used. Juxta-arteriolar and intervascular PO2 in the vitreous are significantly different at 50 µm from the vitreoretinal interface.38 39 Given the microelectrode’s properties,33 a small tip diameter ensures an intervascular PO2 measurement at that distance. Investigations involving a tip diameter of 3 to 6 µm8 9 found no increase in optic disc PO2 over intervascular areas during systemic hyperoxia. Because our electrode had a 10-µm tip diameter, it is likely that we also recorded a small part of the O2 diffusing directly from the ONH arterioles; however, after the initial PO2 increase, the PO2 remained constant throughout measurements under hyperoxia.

Carbogen breathing induces a simultaneous increase in PaO2 and PaCO2, affecting the retinal and ONH arteriolar reactivity. Elevated PaO2 induces arteriolar vasoconstriction, whereas elevated PaCO2 induces a vasodilation11 similar to that observed in cerebral arterioles.40 In retinal circulation, due to the vasodilatory effect of elevated PaCO2, carbogen breathing leads to less reduction of retinal blood flow compared with systemic hyperoxia.41

Assuming similar reactivity of the ONH arterioles, carbogen breathing should lead to a less important arteriolar vasoconstriction at the level of the ONH compared with hyperoxia, as in the retina.41 Indeed, in our experiments, optic disc PO2 increased significantly during carbogen breathing (Fig. 1) , more than after systemic hyperoxia (67% vs. 24%), as also confirmed by statistical comparison. Our results are in agreement with those observed in normal retinas of newborn and adult rats15 and in normal and ischemic intervascular retinal territories of minipigs.16

In addition to blood flow changes induced by carbogen, the blood CO2 increase and the resultant pH decrease should affect the ability of hemoglobin to bind oxygen. Elevated PaCO2 induces a rightward shift of the oxyhemoglobin dissociation curve,42 which reflects decreased affinity of hemoglobin for oxygen, meaning that oxygen is released from hemoglobin more readily, increasing oxygen availability in the tissue. Thus, the optic disc PO2 increase measured in our experiments during carbogen breathing reflects both the effect of carbogen on the ONH circulation and the increased oxygen delivery to the ONH tissue due to a rightward shift of the oxyhemoglobin dissociation curve.

Intravenous administration of acetazolamide increases cerebral,17 18 19 retinal,22 and choroidal23 blood flow, with simultaneous increase in PaCO2.26 This PaCO2 increase is due to significant bicarbonate losses in the renal tubules, resulting in hyperchloremic metabolic acidosis. The CO2 produced by cells cannot be eliminated by carbonic anhydrase and so increases PaCO2 by diffusing in the blood through the basement membrane.43

In our experiments, acetazolamide injection induced a slow and progressive increase in optic disc PO2 in association with a slowly and progressively increasing PaCO2, as previously described.10 This probably reflects, as in the case of carbogen, a vasodilatory effect of acetazolamide on the ONH arterioles and an increased dissociation of oxygen from hemoglobin, both due to the elevated PaCO2. Rightward shift of the oxyhemoglobin dissociation curve with increased oxygen availability under the effect of acetazolamide44 has also been described in the treatment of myocardial ischemia.45 Moreover, our results indicated that concomitant administration of acetazolamide and carbogen can increase optic disc PO2 more significantly than the association of the former with hyperoxia (90% vs. 58%). This effect is apparently due to the additive action of acetazolamide and carbogen on the elevation of PaCO2. Indeed, PaCO2 was higher when acetazolamide was associated with carbogen than with hyperoxia.

To our knowledge, this is the first time that concomitant administration of acetazolamide and carbogen has been tested for the ONH. This is important, as one can extrapolate that carbogen inhalation, especially in association with intravenous injection of acetazolamide, may improve oxygen delivery to a hypoxic ONH. In disturbed perfusion through the ONH capillaries, this association improves the oxygenation of the ONH by increasing O2 diffusion from the ONH arterioles. In that case, the association would be beneficial as a treatment modality for disorders related to ONH ischemia or dysregulation of the ONH blood flow, such as glaucoma and nonarteritic anterior ischemic optic neuropathy (NAION).

Regarding glaucoma, acetazolamide is being used for its intraocular pressure (IOP)-lowering action, which increases the perfusion pressure in the ONH, but the double effect of elevated PaCO2 described herein is also possible. Flammer and Drance46 gave oral acetazolamide in patients with glaucoma and observed visual field improvement without correlation with the IOP reduction and without change in perfusion pressure, suspecting improved ONH blood flow from decreased vascular resistance. In addition, the presence of relative vasoconstriction has been shown in patients with glaucoma, at least partially reversed by hypercapnia.47 Finally, carbogen breathing was reported to improve visual fields of patients with normal-tension glaucoma, postulated to reverse an initial vasospasm.48

Regarding NAION, no established treatment exists.49 Intravenous acetazolamide was administered by Hayreh50 in patients with NAION and progressive loss of vision or visual fields, pretreatment IOP < 20 mm Hg, and (in some of them) angiographic evidence of poor perfusion in posterior ciliary artery system; further deterioration was prevented. Hayreh actually wanted to lower IOP to improve the perfusion pressure in the ONH, whereas, retrospectively, a beneficial effect through elevated PaCO2 also seems possible.

In clinical practice, if acetazolamide alone, or associated with carbogen breathing, can exert a beneficial functional effect through increased oxygenation, this should also be reflected in the results of functional tests. However, acetazolamide attenuates the amplitudes of ERG responses.51 Moreover, under hypercapnic conditions, the b-wave of ERG is depressed,52 53 as a result of reduction in extracellular K+.54 In addition, recent studies have shown either no effect55 or decreased contrast sensitivity56 57 by CO2. Thus, a blood flow increase by acetazolamide and/or carbogen breathing does not necessarily mean a functional benefit resulting from increased oxygenation of the ONH. The beneficial effect of increased ONH oxygenation by carbogen inhalation with concomitant intravenous acetazolamide injection remains to be demonstrated by clinical findings.

In conclusion, the present study shows that carbogen breathing increases optic disc PO2 significantly in minipigs, more than systemic hyperoxia. Moreover, it shows that a much more important increase in optic disc PO2 can be obtained by concomitant use of carbogen breathing and intravenous injection of acetazolamide, through the effect of elevated PaCO2, most probably inducing ONH arteriolar vasodilation and increased dissociation of O2 from hemoglobin. The association of intravenous injection of acetazolamide with carbogen breathing can improve the oxygenation of the ONH substantially and could be a good therapeutic modality in cases of ischemia or dysregulation. This would be of particular interest, as acetazolamide is already being used in the treatment of glaucoma. Clinical studies are needed, however, to confirm these speculations.


   Acknowledgements
 
The authors thank Andrew R. Whatham, DPhil, for a critical reading of the manuscript and useful suggestions.


   Footnotes
 
Presented in part at the annual meeting of the Association for Research in Vision and Ophthalmology, Fort Lauderdale, Florida, May 2003.

Supported by Grant 32-61685.00 (CJP) from the Swiss National Science Foundation.

Submitted for publication February 24, 2005; revised May 13, 2005; accepted September 6, 2005.

Disclosure: I.K. Petropoulos, None; J.-A.C. Pournaras, None; J.-L. Munoz, None; C.J. Pournaras, None

The publication costs of this article were defrayed in part by page charge payment. This article must therefore be marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Corresponding author: Constantin J. Pournaras, Department of Ophthalmology, University Hospitals of Geneva, 22 Alcide-Jentzer Street, CH-1211 Geneva 14, Switzerland; constantin.pournaras{at}hcuge.ch.


   References
Top
 Abstract
 Materials and Methods
 Results
 Discussion
 References
 

  1. Flammer J, Orgül S. Optic nerve blood-flow abnormalities in glaucoma. Prog Retin Eye Res. 1998;17:267–289.[CrossRef][ISI][Medline][Order article via Infotrieve]
  2. Hayreh SS, Zimmerman MB, Podhajsky P, Alward WL. The role of nocturnal hypotension in ocular and optic nerve ischemic disorders. Invest Ophthalmol Vis Sci. 1993;34:994–999.
  3. Demchenko IT, Oury TD, Crapo JD, Piantadosi CA. Regulation of the brain’s vascular responses to oxygen. Circ Res. 2002;91:1031–1037.[Abstract/Free Full Text]
  4. Eperon G, Johnson M, David NJ. The effect of arterial PO2 on relative retinal blood flow in monkeys. Invest Ophthalmol. 1975;14:342–352.[Abstract/Free Full Text]
  5. Luksch A, Garhofer G, Imhof A, et al. Effect of inhalation of different mixtures of O2 and CO2 on retinal blood flow. Br J Ophthalmol. 2002;86:1143–1147.[Abstract/Free Full Text]
  6. Riva CE, Pournaras CJ, Tsacopoulos M. Regulation of local oxygen tension and blood flow in the inner retina during hyperoxia. J Appl Physiol. 1986;61:592–598.[Abstract/Free Full Text]
  7. Pournaras CJ, Riva CE, Tsacopoulos M, Strommer K. Diffusion of O2 in the retina of anesthetized miniature pigs in normoxia and hyperoxia. Exp Eye Res. 1989;49:347–360.[CrossRef][ISI][Medline][Order article via Infotrieve]
  8. Pournaras CJ, Munoz JL, Abdesselem R. Régulation de la PO2 au niveau de la papille du porc miniature en hypoxie. Klin Monatsbl Augenheilkd. 1991;198:404–405.[Medline][Order article via Infotrieve]
  9. Bouzas EA, Donati G, Pournaras CJ. Distribution and regulation of the optic nerve head tissue PO2. Surv Ophthalmol. 1997;42(suppl 1)S27–S34.
  10. Petropoulos IK, Pournaras JA, Munoz JL, Pournaras CJ. Effect of acetazolamide on the optic disc oxygenation in miniature pigs. Klin Monatsbl Augenheilkd. 2004;221:367–370.[CrossRef][Medline][Order article via Infotrieve]
  11. Tsacopoulos M, David NJ. The effect of arterial PCO2 on relative retinal blood flow in monkeys. Invest Ophthalmol. 1973;12:335–347.[Abstract/Free Full Text]
  12. Dorner GT, Garhoefer G, Zawinka C, Kiss B, Schmetterer L. Response of retinal blood flow to CO2-breathing in humans. Eur J Ophthalmol. 2002;12:459–466.[ISI][Medline][Order article via Infotrieve]
  13. Dunn JF, O’Hara JA, Zaim-Wadghiri Y, et al. Changes in oxygenation of intracranial tumors with carbogen: a BOLD MRI and EPR oximetry study. J Magn Reson Imag. 2002;16:511–521.[CrossRef][ISI][Medline][Order article via Infotrieve]
  14. Alm A, Bill A. The oxygen supply to the retina. I. Effects of changes in intraocular and arterial blood pressures, and in arterial PO2 and PCO2 on the oxygen tension in the vitreous body of the cat. Acta Physiol Scand. 1972;84:261–274.[ISI][Medline][Order article via Infotrieve]
  15. Berkowitz BA. Adult and newborn rat inner retinal oxygenation during carbogen and 100% oxygen breathing: comparison using magnetic resonance imaging delta PO2 mapping. Invest Ophthalmol Vis Sci. 1996;37:2089–2098.[Abstract/Free Full Text]
  16. Pournaras JA, Petropoulos IK, Munoz JL, Pournaras CJ. Experimental retinal vein occlusion: effect of acetazolamide and carbogen (95% O2/5% CO2) on preretinal PO2. Invest Ophthalmol Vis Sci. 2004;45:3669–3677.[Abstract/Free Full Text]
  17. Vorstrup S, Henriksen L, Paulson OB. Effect of acetazolamide on cerebral blood flow and cerebral metabolic rate for oxygen. J Clin Invest. 1984;74:1634–1639.
  18. Ringelstein EB, Van Eyck S, Mertens I. Evaluation of cerebral vasomotor reactivity by various vasodilating stimuli: comparison of CO2 to acetazolamide. J Cereb Blood Flow Metab. 1992;12:162–168.[ISI][Medline][Order article via Infotrieve]
  19. Dahl A, Russell D, Nyberg-Hansen R, Rootwelt K, Bakke SJ. Cerebral vasoreactivity in unilateral carotid artery disease: a comparison of blood flow velocity and regional cerebral blood flow measurements. Stroke. 1994;25:621–626.[Abstract]
  20. Piepgras A, Schmiedek P, Leinsinger G, Haberl RL, Kirsch CM, Einhaupl KM. A simple test to assess cerebrovascular reserve capacity using transcranial Doppler sonography and acetazolamide. Stroke. 1990;21:1306–1311.[Abstract/Free Full Text]
  21. Müller M, Voges M, Piepgras U, Schimrigk K. Assessment of cerebral vasomotor reactivity by transcranial Doppler ultrasound and breath-holding: a comparison with acetazolamide as vasodilatory stimulus. Stroke. 1995;26:96–100.[Abstract/Free Full Text]
  22. Rassam SM, Patel V, Kohner EM. The effect of acetazolamide on the retinal circulation. Eye. 1993;7:697–702.
  23. Dallinger S, Bobr B, Findl O, Eichler HG, Schmetterer L. Effects of acetazolamide on choroidal blood flow. Stroke. 1998;29:997–1001.[Abstract/Free Full Text]
  24. Stefansson E, Jensen PK, Eysteinsson T, et al. Optic nerve oxygen tension in pigs and the effect of carbonic anhydrase inhibitors. Invest Ophthalmol Vis Sci. 1999;40:2756–2761.[Abstract/Free Full Text]
  25. Tsacopoulos M, Levy S. Intraretinal acid-base studies using pH glass microelectrodes: effect of respiratory and metabolic acidosis and alkalosis on inner-retinal pH. Exp Eye Res. 1976;23:495–504.[CrossRef][ISI][Medline][Order article via Infotrieve]
  26. Taki K, Kato H, Endo S, Inada K, Totsuka K. Cascade of acetazolamide-induced vasodilatation. Res Commun Mol Pathol Pharmacol. 1999;103:240–248.[ISI][Medline][Order article via Infotrieve]
  27. Harper AM. Physiology of cerebral bloodflow. Br J Anaesth. 1965;37:225–235.[Free Full Text]
  28. Tsacopoulos M. Quelques aspects des relations existant entre le fonctionnement neuronal rétinien, le métabolisme oxydatif et l’apport constant d’O2 par la microcirculation. Klin Monatsbl Augenheilkd. 1985;186:477–479.[Medline][Order article via Infotrieve]
  29. Ernest JT. In vivo measurement of optic disk oxygen tension. Invest Ophthalmol. 1973;12:927–931.[Free Full Text]
  30. Ernest JT. Optic disk oxygen tension. Exp Eye Res. 1977;24:271–278.[CrossRef][ISI][Medline][Order article via Infotrieve]
  31. Ahmed J, Linsenmeier RA, Dunn R, Jr. The oxygen distribution in the prelaminar optic nerve head of the cat. Exp Eye Res. 1994;59:457–465.[CrossRef][ISI][Medline][Order article via Infotrieve]
  32. Rootman J. Vascular system of the optic nerve head and retina in the pig. Br J Ophthalmol. 1971;55:808–819.[Free Full Text]
  33. Tsacopoulos M, Lehmenkühler A. A double-barreled Pt-microelectrode for simultaneous measurement of PO2 and bioelectrical activity in excitable tissues. Experientia. 1977;33:1337–1338.[CrossRef][ISI][Medline][Order article via Infotrieve]
  34. Tsacopoulos M, Poitry S, Borsellino A. Diffusion and consumption of oxygen in the superfused retina of the drone (Apis mellifera) in darkness. J Gen Physiol. 1981;77:601–628.[Abstract/Free Full Text]
  35. Pournaras CJ, Shonat RD, Munoz JL, Petrig BL. New ocular micromanipulator for measurements of retinal and vitreous physiologic parameters in the mammalian eye. Exp Eye Res. 1991;53:723–727.[CrossRef][ISI][Medline][Order article via Infotrieve]
  36. Riva CE, Harino S, Petrig BL, Shonat RD. Laser Doppler flowmetry in the optic nerve. Exp Eye Res. 1992;55:499–506.[CrossRef][ISI][Medline][Order article via Infotrieve]
  37. Harris A, Anderson DR, Pillunat L, et al. Laser Doppler flowmetry measurement of changes in human optic nerve head blood flow in response to blood gas perturbations. J Glaucoma. 1996;5:258–265.[ISI][Medline][Order article via Infotrieve]
  38. Molnar I, Poitry S, Tsacopoulos M, Gilodi N, Leuenberger PM. Effect of laser photocoagulation on oxygenation of the retina in miniature pigs. Invest Ophthalmol Vis Sci. 1985;26:1410–1414.[Abstract/Free Full Text]
  39. Pournaras CJ, Miller JW, Gragoudas ES, et al. Systemic hyperoxia decreases vascular endothelial growth factor gene expression in ischemic primate retina. Arch Ophthalmol. 1997;115:1553–1558.[Abstract]
  40. Reavich M. Arterial PaCO2 and cerebral hemodynamics. Am J Physiol. 1964;206:25–35.[Abstract/Free Full Text]
  41. Pakola SJ, Grunwald JE. Effects of oxygen and carbon dioxide on human retinal circulation. Invest Ophthalmol Vis Sci. 1993;34:2866–2870.[Abstract/Free Full Text]
  42. Benz EJ, Jr. Hemoglobinopathies. Braunwald E Fauci AS Kasper DL Hauser SL Longo DL Jamson JL eds. Harrison’s Principles of Internal Medicine. 2001; 15th ed. 666–667. McGraw-Hill New York.
  43. Ives HE. Diuretic agents. Katzung BG eds. Basic and Clinical Pharmacology. 2001; 8th ed. 249–252. McGraw-Hill/Appleton and Lange New York/San Francisco.
  44. Gai X, Taki K, Kato H, Nagaishi H. Regulation of hemoglobin affinity for oxygen by carbonic anhydrase. J Lab Clin Med. 2003;142:414–420.[CrossRef][ISI][Medline][Order article via Infotrieve]
  45. Weiss RG, Marco M. Preservation of canine myocardial high energy phosphates during low-flow ischemia with modification of hemoglobin oxygen. J Clin Invest. 1999;103:739–746.[ISI][Medline][Order article via Infotrieve]
  46. Flammer J, Drance SM. Effect of acetazolamide on the differential threshold. Arch Ophthalmol. 1983;101:1378–1380.[Abstract]
  47. Hosking SL, Harris A, Chung HS, et al. Ocular haemodynamic responses to induced hypercapnia and hyperoxia in glaucoma. Br J Ophthalmol. 2004;88:406–411.[Abstract/Free Full Text]
  48. Pillunat LE, Lang GK, Harris A. The visual response to increased ocular blood flow in normal pressure glaucoma. Surv Ophthalmol. 1994;38(suppl)S139–S148.
  49. Rucker JC, Biousse V, Newman NJ. Ischemic optic neuropathies. Curr Opin Neurol. 2004;17:27–35.[CrossRef][ISI][Medline][Order article via Infotrieve]
  50. Hayreh SS. Anterior ischemic optic neuropathy. III. Treatment, prophylaxis, and differential diagnosis. Br J Ophthalmol. 1974;58:981–989.[Free Full Text]
  51. Findl O, Hansen RM, Fulton AB. The effects of acetazolamide on the electroretinographic responses in rats. Invest Ophthalmol Vis Sci. 1995;36:1019–1026.[Abstract/Free Full Text]
  52. Tsacopoulos M, Baker R, Hamasaki D, David NJ. Proceedings: studies on retinal blood flow regulation: the effect of PaCO2 on blood flow, oxygen availability, oxygen consumption rate and ERG in monkeys. Exp Eye Res. 1973;17:391.
  53. Pournaras C, Tsacopoulos M. L’effet du CO2 sur l’ERG local enregistré au moyen d’une micro-électrode dans les couches synaptiques de la rétine de la grenouille (Rana ridibunda). Klin Monatsbl Augenheilkd. 1980;176:524–528.[Medline][Order article via Infotrieve]
  54. Pournaras KI, Karwoski CJ, Tsacopoulos M. L’effet du CO2 sur l’activité du K+ extracellulaire au niveau de la couche plexiforme interne de la rétine isolée de la grenouille (Rana ridibunda). Klin Monatsbl Augenheilkd. 1982;180:339–340.
  55. Harris A, Arend O, Wolf S, Cantor LB, Martin BJ. CO2 dependence of retinal arterial and capillary blood velocity. Acta Ophthalmol Scand. 1995;73:421–424.[ISI][Medline][Order article via Infotrieve]
  56. Sponsel WE, Harrison J, Elliott WR, Trigo Y, Kavanagh J, Harris A. Dorzolamide hydrochloride and visual function in normal eyes. Am J Ophthalmol. 1997;123:759–766.[ISI][Medline][Order article via Infotrieve]
  57. Hosking SL, Evans DW, Embleton SJ, Houde B, Amos JF, Bartlett JD. Hypercapnia invokes an acute loss of contrast sensitivity in untreated glaucoma patients. Br J Ophthalmol. 2001;85:1352–1356.[Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 IOVSHome page
M. H. Noergaard, D. Bach-Holm, E. Scherfig, K. Bang, P. K. Jensen, J. F. Kiilgaard, E. Stefansson, and M. la Cour
Dorzolamide Increases Retinal Oxygen Tension after Branch Retinal Vein Occlusion
Invest. Ophthalmol. Vis. Sci., March 1, 2008; 49(3): 1136 - 1141.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via ISI Web of Science (2)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Petropoulos, I. K.
Right arrow Articles by Pournaras, C. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Petropoulos, I. K.
Right arrow Articles by Pournaras, C. J.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS