Abstract

ABSTRACT Paramagnetic deoxyhemoglobin in venous blood is a naturally occurring contrast agent for magnetic resonance imaging (MRI). By accentuating the effects of this agent through the use of gradient-echo techniques in high fields, we demonstrate in vivo images of brain microvasculature with image contrast reflecting the blood oxygen level. This blood oxygenation level-dependent (BOLD) contrast follows blood oxygen changes induced by anesthetics, by insulininduced hypoglycemia, and by inhaled gas mixtures that alter metabolic demand or blood flow. The results suggest that BOLD contrast can be used to provide in vivo real-time maps of blood oxygenation in the brain under normal physiological conditions. BOLD contrast adds an additional feature to magnetic resonance imaging and complements other techniques that are attempting to provide positron emission tomographylike measurements related to regional neural activity. Magnetic resonance imaging (MRI) is a widely accepted modality for providing anatomical information. Current research (1) involves extending MRI methods to provide information about biological function, in addition to the concomitant anatomical information. In addition to localized spectroscopy (2) and chemical shift imaging (3) that are applicable to many chemical species, MRI of water protons has been functionally extended to NMR angiography (4), perfusion imaging It has previously been demonstrated (8, 9) that the presence of deoxyhemoglobin in blood changes the proton signal from water molecules surrounding a blood vessel in gradientecho MRI, producing blood oxygenation level-dependent (BOLD) contrast. BOLD contrast has its origin in the fact that when normally diamagnetic oxyhemoglobin gives up its oxygen, the resulting deoxyhemoglobin is paramagnetic. The presence of paramagnetic molecules in blood produces a difference in magnetic susceptibility between the blood vessel and the surrounding tissue. This susceptibility difference is "felt" both by the water molecules in the blood and by those in the surrounding tissue, the effect extending significantly beyond the vessel wall. This increase in the number of spins affected by deoxyhemoglobin is a form of amplification. When the susceptibility-induced local field differences exist within an imaging voxel, there is a resultant distribution of shifts in water resonance frequencies. In the gradient-echo method, a phase dispersion of water proton signals is produced at the echo time. This dispersion reduces the signal intensity and the voxel appears dark in the image. These intensity losses, which at high magnetic fields (-4 T) extend significantly beyond the boundary of the blood vessel, are the source of BOLD contrast. This form of contrast is not observed in spin-echo images. Through simulations (9), we have shown that vessels as small as 50 ,um in diameter can be detected in images with a pixel size of 100 pum. We have also demonstrated that the size of the susceptibility-induced local field depends on (i) the concentration of paramagnetic deoxyhemoglobin and (ii) the orientation of the vessel relative to the main magnetic field (8, 9). Since BOLD contrast depends on the state of blood oxygenation, physiological events that change the oxy/ deoxyhemoglobin ratio should lend themselves to noninvasive detection through the accentuation of BOLD contrast in gradient-echo proton images at high magnetic fields. We report here that this is indeed the case and demonstrate changes in BOLD-contrast microimages ofbrain produced by changes in inhaled gas mixture under urethane anesthesia, by insulin-induced hypoglycemia under diazepam sedation, and by changes in the level of halothane anesthesia. The observed changes in BOLD contrast correlate with the anticipated changes in blood oxygen level produced by altered metabolic load or blood flow (10). MATERIALS AND METHODS Physiology and Anesthetics. Sprague-Dawley rats (female, 250 ± 25 g) were anesthetized with the described anesthetics and ventilated without assistance. Animal care and the administration of anesthetics were in accordance with National Institutes of Health guidelines. For MRI experiments, the animal was placed in a thermally regulated compartment within the magnet and rectal temperature and electrocardiogram were monitored. Blood was periodically sampled from the femoral artery and blood gas concentrations were measured with a Radiometer model ABL330 gas analyzer. Blood glucose was estimated using a Boehringer-Mannheim AccuCheck Ilm. To measure the electroencephalogram (EEG) during MRI experiments, small tubular salt bridges (0.8 mm i.d.) were implanted in small holes through the skull that were located inside the area encircled by the surface coil used for NMR signal detection. Care was taken not to break the dura. Magnetic Resonance Microimaging. Gradient-echo images were obtained with a 7-T horizontal magnet at 300 MHz (Spectroscopy Imaging Systems, Fremont, CA). Surface coils of 20 to 25-mm diameter were positioned on the surface of the cranium and used for both excitation and detection of signals. The maximum field gradient used to produce images was 4 G/cm (1 G = 0.1 mT). The slice width of the images shown was 500 ,4m and the in-plane pixel size was 117 ,um X Abbreviations: MRI, magnetic resonance imaging; BOLD, blood oxygenation level-dependent; EEG, electroencephalogram. 9868 The publication costs of this article were defrayed in part by page charge payment. 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