Electron paramagnetic resonance imaging (EPRI) is a powerful technique that enables spatial mapping of free radicals or other paramagnetic compounds; however, it does not in itself provide anatomic visualization of the body. Proton magnetic resonance imaging (MRI) is well suited to provide anatomical visualization. A hybrid EPR/NMR coimaging instrument was constructed that utilizes the complementary capabilities of both techniques, superimposing EPR and proton-MR images to provide the distribution of paramagnetic species in the body. A common magnet and field gradient system is utilized along with a dual EPR and proton-NMR resonator assembly, enabling coimaging without the need to move the sample. EPRI is performed at ϳ1.2 GHz/ ϳ40 mT and proton MRI is performed at 16.18 MHz/ϳ380 mT; hence the method is suitable for whole-body coimaging of living mice. The gradient system used is calibrated and controlled in such a manner that the spatial geometry of the two acquired images is matched, enabling their superposition without additional postprocessing or marker registration. The performance of the system was tested in a series of phantoms and in vivo applications by mapping the location of a paramagnetic probe in the gastrointestinal ( Key words: proton MRI; EPR imaging; free radicals, oxygen; image coregistration; in vivo NMR; in vivo EPRThe techniques of electron paramagnetic resonance (EPR) spectroscopy and imaging (EPRI) have been widely used to measure and map the distribution of paramagnetic materials and free radicals in biological systems, including ex vivo tissues and in vivo living animals (1-8). These techniques provide unique information that enables measurement and imaging of the processes of free radical metabolism, tissue oxygenation, and nitric oxide generation in normal physiology and disease (9 -17). While EPRI provides specific mapping of the location of a given paramagnetic species, this is often not sufficient in itself to enable anatomic mapping of the precise organ-specific location of the paramagnetic probe in the body of a living animal. On the other hand, proton magnetic resonance imaging (MRI) has been established as a powerful diagnostic imaging technique capable of obtaining high-resolution images of the anatomical structure of humans and animals (18 -22). While proton MRI is a powerful technique for imaging biological systems based on their high content of water, it can not directly localize low, submillimolar levels of free radicals.Several noninvasive MRI-based methods for detection and mapping of paramagnetic substances in vivo are under development. Both contrast-enhanced MRI (15,16) and proton electron double resonance imaging (PEDRI) (23,24) utilize proton MRI-based detection and therefore automatically provide coregistration of free radical distribution with anatomical structure. The availability of anatomic information, as well as superior spatial and temporal resolution, make MRI-based techniques more efficient in these aspects compared to direct EPRI methods. However, since t...

A new concept of Variable Field Proton-Electron Double-Resonance Imaging (VF PEDRI) is proposed. This allows for functional mapping using specifically designed paramagnetic probes (e.g. oxygen or pH mapping) with MRI high quality spatial resolution and short acquisition time. Studies performed at 200 G field MRI with phantoms show that a pH map of the sample can be extracted using only two PEDRI images acquired in 140 s at pre-selected EPR excitation fields providing pH resolution of 0.1 pH units and a spatial resolution of 1.25 mm. Note that while concept of functional VF PEDRI was demonstrated using the pH probe, it can be applied for studies of other biologically relevant parameters of the medium such as redox state, concentrations of oxygen or glutathione using specifically designed EPR probes.

Proton-Electron Double-Resonance Imaging (PEDRI) offers rapid image data collection and high resolution for spatial distribution of paramagnetic probes. Recently we developed the concept of Variable Field (VF) PEDRI which enables extracting a functional map from a limited number of images acquired at pre-selected EPR excitation fields using specific paramagnetic probes (Khramtsov et al. 2010, JMR, 202, 267-273). In this work, we propose and evaluate a new modality of PEDRI-based functional imaging with enhanced temporal resolution which we term Variable Radio Frequency (VRF) PEDRI. The approach allows for functional mapping (e.g., pH mapping) using specifically designed paramagnetic probes with high quality spatial resolution and short acquisition times. This approach uses a stationary magnetic field but different EPR RFs. The ratio of Overhauser enhancements measured at each pixel at two different excitation frequencies corresponding to the resonances of protonated and deprotonated forms of a pH-sensitive nitroxide is converted to a pH map using a corresponding calibration curve. Elimination of field cycling decreased the acquisition time by exclusion periods of ramping and stabilization of the magnetic field. Improved magnetic field homogeneity and stability allowed for the fast MRI acquisition modalities such as fast spin echo. In total, about 30-fold decrease in EPR irradiation time was achieved for VRF PEDRI (2.4 s) compared with VF PEDRI (70 s). This is particularly important for in vivo applications enabling one to overcome the limiting stability of paramagnetic probes and sample overheating by reducing RF power deposition.

In vivo mapping of alterations in redox status is important for understanding organ specific pathology and disease. While electron paramagnetic resonance imaging (EPRI) enables spatial mapping of free radicals, it does not provide anatomic visualization of the body. Proton MRI is well suited to provide anatomical visualization. We applied EPR/NMR co-imaging instrumentation to map and monitor the redox state of living mice under normal or oxidative stress conditions induced by secondhand cigarette smoke (SHS) exposure. A hybrid co-imaging instrument, EPRI (1.2 GHz) / proton MRI (16.18 MHz), suitable for whole-body co-imaging of mice was utilized with common magnet and gradients along with dual EPR/NMR resonators that enable co-imaging without sample movement. The metabolism of the nitroxide probe, 3–carbamoyl–proxyl (3-CP), was used to map the redox state of control and SHS-exposed mice. Co-imaging allowed precise 3D mapping of radical distribution and reduction in major organs such as the heart, lungs, liver, bladder and kidneys. Reductive metabolism was markedly decreased in SHS-exposed mice and EPR/NMR co-imaging allowed quantitative assessment of this throughout the body. Thus, in vivo EPR/NMR co-imaging enables in vivo organ specific mapping of free radical metabolism and redox stress and the alterations that occur in the pathogenesis of disease.