Anesthetic drugs are vasoactive and thus may alter cerebral blood volume (CBV) and thereby intracranial pressure (ICP) (1). For patients with increased ICP secondary to a space-occupying lesion, the ideal anesthetic is one that reduces CBV and ICP, while maintaining cerebral blood flow (CBF) and adequate oxygen delivery. In addition, because intraoperative hyperventilation is often used, the ideal anesthetic should also amplify, or at least not hinder, the reduction in CBV to hypocapnia. Whereas there is some information on the effect of hyperventilation on CBF in brain tumors (2–6), there are no data on the combined effects of commonly used anesthetics and hyperventilation on both CBV and CBF in tumor and peri-tumor regions of the brain. One reason is the technical difficulty in measuring CBV repeatedly in the same subject.
In the present study, our aim was to simultaneously investigate CBV and CBF responses to hyperventilation in a rabbit brain tumor model during isoflurane or propofol anesthesia. Using our previously described dynamic computed tomography (CT) method (7,8), and for each anesthetic, absolute CBF and CBV measurements were made in tumor, peri-tumor, and contralateral normal regions of the brain at normocapnia and at hypocapnia.
Brain Tumor Model
To obtain a standard-sized tumor in a select region of the brain, a measured number of VX2 carcinoma cells (approximately 5 × 105 cells) were injected through a burr hole into the right parietal lobe (2–3 mm below the dura mater) of the study animal. Ketamine/xylazine was used to sedate the animals for this procedure. When the tumor reached approximately 0.4 cm in diameter (as determined by repeated CT scans), the anesthesia study was performed.
Sixteen male New Zealand white rabbits, average weight 3.2 ± 0.3 kg, were used in experiments approved by the Animal Ethics Committee of the University of Western Ontario. Each animal with an implanted brain tumor was surgically prepared by inducing anesthesia via a mask with halothane, and one ear vein was cannulated for the administration of muscle relaxant (vecuronium bromide) and radiograph contrast material during the experiment. After a tracheotomy was performed, the animals were ventilated with an air/O2 mixture. Halothane was then gradually withdrawn as the study anesthetic was introduced: either 1 minimum alveolar concentration isoflurane (2% isoflurane) (9) or propofol (2% Diprivan®; Zeneca Pharmaceuticals) at a rate of 1.6 mg · kg−1 · min−1 for the initial 30 min followed by 1.2 mg · kg−1 · min−1 thereafter (10). Both femoral arteries were cannulated with 18-gauge catheters to allow continuous measurement of mean arterial blood pressure (MAP), as well as arterial blood sampling. Both femoral veins were also catheterized with long 18-gauge catheters to allow continuous measurement of central venous pressure from the inferior vena cava, and for fluid and drug administration. An intraventricular catheter (PE-50 cannula) was placed into the left lateral ventricle for continuous ICP measurement. All surgical wounds were infiltrated with lidocaine 1%. All pressure transducers were placed at the level of the head.
There were two experimental groups: the animals were randomly assigned to receive either isoflurane or propofol anesthesia. In both groups, CBV and CBF were measured at two different arterial carbon dioxide partial pressures (Paco2) performed in random order and corresponding to normocapnia (40 mm Hg) and hypocapnia (25 mm Hg). All sequential CBV and CBF measurement studies were separated by at least 30 min to allow for washout of contrast material as well as to establish a steady-state designated Paco2. The target Paco2 was achieved by varying the rate of mechanical ventilation.
Rectal temperature was maintained at 38.5°C with a water-heating blanket, heat lamp, or ice packs. Hematocrit was measured every half-hour and arterial blood gases were determined before and after each CBV and CBF measurement. Phenylephrine was administered IV as required to maintain MAP between 75 and 85 mm Hg.
Cine CT Scanning Protocol
The CBV and CBF measurements were acquired by using a slip-ring CT scanner (GE High Speed Advantage CT, Milwaukee, WI). With the animal positioned prone in the scanner, we selected a 3-mm coronal section through the brain at the level where the tumor was largest. All subsequent scans were taken at this same location.
Contrast-enhanced cine CT scans were then performed (i.e., the chosen slice was repeatedly and continuously scanned without any time delay between scans). The CT scanning variables were as follows: 80 kVp, 80 mA, 512 matrix size, 10-cm field of view, 3-mm slice thickness, and 1 s per scan. In the reconstruction of CT images, a back-projection filter with a cut-off frequency of 10 line pairs per centimeter was used. Cine CT scanning was initiated 5 s before a bolus of Ultravist 300 contrast (1.5 mL per kg mass) was injected IV by using an automated injector (Medrad Injector; Medrad, Indianola, PA) with an infusion rate of 0.3 mL per second. This delay in contrast injection allowed for the acquisition of nonenhanced, baseline images (i.e., background data for image analysis). Cine scanning was maintained during the bolus injection of contrast and continued for 1 min. From the raw CT projection data, images were retrospectively constructed at 0.5-s intervals; hence, each cine CT study contained 119 sequential images (60 prospective, 59 retrospective).
CT Data Analysis
The CT images were transferred to an Ultra I workstation (Sun Microsystems, Mountain view, CA) for analyses by a technician who was blinded to the experimental group. Regions of interest (ROI) in the brain were drawn in the tumor, peri-tumor, and contralateral normal tissue regions. An ROI was first drawn incorporating the entire tumor that was the most enhanced area of the right parietal lobe. The peri-tumor ROI was created by extending the tumor ROI by five to six pixels. The contralateral normal ROI was drawn incorporating the entire contralateral hemisphere (i.e., parietal and temporal lobes). Tissue ROIs were drawn such that no major intracerebral blood vessels were present within the regions. Necrotic regions visible within the tumor were delineated and excluded in the ROI analysis of the tumor. The mean CT number in the tissue ROIs was determined for each of the 119 sequential images over the 60-s interval. Because the measured change in CT number (i.e., enhancement) is linearly related to the concentration of contrast in the tissue (11), the tissue contrast concentration curve, Q(t), was obtained by subtracting the mean baseline CT number in precontrast images from the mean CT number in sequential contrast-enhanced images.
There are three types of arteries in the plane of the CT image: ear, cerebral, and radial. The artery used for determining Ca(t) was the one that was most apparent in the CT image (i.e., the largest and most distinct). This was done to minimize partial volume averaging inherent when imaging small objects (e.g., arteries) with CT scanners. In most rabbit studies, Ca(t) was obtained from one of the radial or ear arteries. The arterial contrast concentration curve, Ca(t), was determined by drawing a two-pixel-radius circular ROI in the chosen artery. Ca(t) was determined by subtracting the mean baseline CT number in the vessel ROI in precontrast scans from the mean CT number in contrast-enhanced scans. The measured Ca(t) was then corrected for partial volume averaging (7).
From the CT-measured contrast-enhancement curves, regional CBV and CBF measurements were obtained by using our previously described deconvolution-based method (7,8), and the Central Volume Principle (10,12).
Statistical analysis was performed by using the Jandel Scientific software package (SigmaPlot® and SigmaStat®; SPSS, Chicago, IL). For monitored physiologic data, a paired t-test (two-tailed) was used to determine statistically significant changes in normally distributed data. Because CBV and CBF were mea-sured for each rabbit under two conditions, anesthesia and level of Paco2, we used a 2 × 2 factorial design with repeated measures to compare intra- and interpopulation differences. A separate analysis was performed for each of the regions of the brain. Population sample means were given as mean ± sd. Statistical significance was declared at the P < 0.05 level.
Table 1 shows physiologic data for the Propofol and Isoflurane groups at normocapnia and hypocapnia. No significant differences (P > 0.10) in MAP, central venous pressure, temperature, hematocrit, or ICP were found during the sequential study periods for the same anesthetic group. Moreover, there were no intergroup differences in the above variables for both anesthetics at normocapnia and at hypocapnia. There was no difference between groups in the amount of phenylephrine used. The ages of the brain tumors (number of days postimplantation when the anesthetic study was performed) for the Propofol and the Isoflurane rabbits were 12 ± 2 and 10 ± 4 days, respectively.
Regional CBV and CBF values at normocapnia and at hypocapnia are shown in Tables 2 and 3 for isoflurane and propofol, respectively. In both anesthetic groups, absolute CBV and CBF were highest in the tumor region, and lowest in the contralateral normal tissue. For isoflurane, a significant decrease in both CBV and CBF was observed in all tissue regions with hyperventilation (P < 0.05) (Table 2). However, the percent reduction in regional CBF with hypocapnia was on average two times larger than that observed in the CBV response (P < 0.01). In contrast, there were no statistically significant changes in CBV and CBF in the Propofol group with hyperventilation for all regions (P > 0.10). The slope of CBV versus Paco2 for propofol and isoflurane for tumor and contralateral normal regions of the brain were significantly different (P < 0.02 and P < 0.002, respectively).
At normocapnia, significantly higher CBV values were found in all regions for isoflurane when compared with propofol (P < 0.02 for tumor and peri-tumor;P < 0.001 for contralateral normal). For CBF values at normocapnia, the values in the tumor and peri-tumor regions were not statistically different between the two anesthetics, except in the contralateral normal region where CBF values were greater for isoflurane (P < 0.01).
There were no significant differences between CBV values for isoflurane at hypocapnia compared with CBV values for propofol at normo- or hypocapnia (P > 0.34 and P > 0.35, respectively, in the tumor regions). There were also no significant differences between CBF for isoflurane at hypocapnia compared with CBF values for propofol at normo- or hypocapnia (P > 0.1, P > 0.4, and P > 0.06, respectively, in tumor, peri-tumor, and contralateral normal regions).
On average, tumor tissue CBV and CBF values were approximately three to four times greater than the normal contralateral tissue, whereas CBV and CBF values in the peri-tumor regions were only two to three times greater than normal contralateral tissue.
The experimental protocol took 2 h to complete. Although we did not include a time control group, previous studies using a similar experimental protocol demonstrated that CBV and CBF did not change over time (10,13).
Changes in cerebral vascular tone result in changes in CBF and CBV. Anesthetics are vasoactive and can either constrict or dilate blood vessels, which may result in potentially deleterious cerebrovascular effects in patients with space-occupying lesions. Paco2 is one of the most effective chemical mediators of cerebral vascular tone. In healthy phencyclidine-anesthetized rhesus monkeys, Grubb et al. (14) demonstrated that changes in CBV and CBF are approximately linear to changes in Paco2 in the range 20–70 mm Hg. However, the percent change in CBV was substantially less than that in CBF for each mm Hg change, thus providing useful insight on CBV and CBF responses to changing Paco2 under normal conditions. However, these effects cannot be extrapolated to other anesthetics and may differ in the presence of intracranial pathology.
We examined the simultaneous effects of hypocapnia on regional CBV and CBF in a brain tumor model during isoflurane or propofol anesthesia. In the Isoflurane group, we observed a significant decrease in both CBV and CBF in all regions of the brain with hyperventilation. These findings suggest that the cerebrovascular CO2 reactivity of tumor and peri-tumor vessels was maintained during isoflurane anesthesia. The similar relative changes of both CBF and CBV in normal and tumor tissue regions with hyperventilation show that these neoplastic vessels reacted to CO2 in a manner similar to normal vessels. We also found, with isoflurane, that for the same degree of hyperventilation there was a differential reduction in CBF compared with CBV (36% versus 15% for the tumor region). This differential reduction in CBF and CBV is similar to that shown in rabbits without an intracranial lesion during isoflurane anesthesia (13) and is also consistent with other studies (15,16). Although CBV was significantly altered by hypocapnia with isoflurane, ICP was not. This likely reflects the fact that ICP was not critically increased and elastance was therefore high. Reasons for this are not clear, but our technique measures regional CBV and CBF in the capillary bed, a compartment that may exert only a small influence on ICP.
In contrast to the Isoflurane group, the cerebrovascular CO2 reactivity was not maintained in normal or tumor vessels in the Propofol group. These results are similar to our previous study in healthy rabbits (13) and supported by another study in which CBF did not change with hyperventilation during propofol anesthesia (17). It is possible that the effect of hyperventilation in decreasing cerebral vascular tone was attenuated by the already near maximal constriction induced by propofol anesthesia. However, our findings contrast with other published studies (18,19) showing that CBF responsiveness is maintained with propofol. It is possible that the differences between our study and the above-mentioned human studies may reflect either an interspecies difference in response to propofol, or a difference in CBF measurement technique. However, these and other studies reveal that at the lower ranges of Paco2, the CBF-CO2 responsiveness to propofol is reduced (3,20). Because cerebral vessels have a limited capacity to vasoconstrict, this reduced responsiveness to hypocapnia may reflect the already-constricted state of the vessels brought on by propofol. Indeed, a study of brain tumor patients under propofol anesthesia showed that with hyperventilation, CBF decreased to levels approaching the cerebral ischemic threshold (20).
A comparison of absolute regional CBF and CBV values at normocapnia for both anesthetic studies supports the different vasoactive effects these anesthetics have on the cerebral vasculature. Mean regional CBV was 1.4 times lower in the Propofol than the Isoflurane rabbits for all tissue regions (Tables 2, 3). In the contralateral hemisphere, CBF was 2.2 times lower with propofol than with isoflurane. These results are in agreement with other comparative studies in animals with no intracranial pathology (17,21).
The measurement of regional CBF in our study reflects capillary flow or tissue perfusion, whereas regional measurements of CBV relate to the vascularity or density of blood vessels in the capillary bed. Hence, a comparison of regional CBF and CBV values in normal tissue with that in tumor and peri-tumor can characterize differences in tissue perfusion and vascular density between normal and neoplastic tissues. For both anesthetics, CBF and CBV were approximately 4.0 and 3.1 times greater in the tumor than the contralateral normal regions, respectively, indicating an increase in flow and vascularity. The high CBF and CBV values in this VX2 carcinoma are supported by a previous histological study showing increased vascular proliferation in the tumor core compared with normal tissue (22).
We used a cine CT scanning method that was previously validated both in normal and in brain tumor studies against microspheres, a CBF reference standard (7,8). The correlation between cine CT and microsphere CBF measurements was similar in both circumstances (r = 0.99 ± 0.03). In the brain tumor rabbits, the precision of our cine CT CBF and CBV for repeated measurements was respectively 13% and 7%. Compared with the tumor region of interest, we found that the signal-to-noise ratio (SNR) was much lower in the corresponding contralateral normal region of the same size. The low SNR in such small normal tissue regions would decrease the precision of our deconvolution-based CBF measurements (23,24). In the present study, the entire contralateral normal hemisphere was used in order to increase the SNR, and the precision of our measurements in normal tissue. Compared with three-dimensional positron emission tomography perfusion studies, our CT method is restricted to a single slice. Our study measures regional CBV and CBF at a single slice and thus may not provide an overall global evaluation of the cerebral vascular dynamics. However, because of the well-circumscribed tumor used in this study, CBF and CBV measurements at a representative slice (i.e., the largest cross-sectional area) may be sufficient for the evaluation of the nature and the pathophysiology of the disease.
In summary, we have compared the effects of two anesthetics and Paco2 on regional CBF and CBV in a rabbit brain tumor model using our previously validated cine CT method. Our results indicate that, during steady-state normocapnic conditions, propofol increases cerebral vascular tone in both neoplastic and normal tissue vessels compared with isoflurane. This resulted in significantly lower CBV for propofol compared with isoflurane at normocapnia. Although normocapnic CBV and CBF were significantly greater in all regions (tumor, peri-tumor, and contralateral normal tissue) with isoflurane, CBV and CBF remained responsive to hyperventilation.
We gratefully acknowledge the animal care assistance provided by Sarah Henderson.
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