Goran Bacic

Assessment of cerebral pO2 by EPR oximetry in rodents: effects of anesthesia, ischemia, and breathing gas

This report describes experiments designed to assess and illustrate the effectiveness of a new method for the measurement of cerebral interstitial pO2 in conscious rodents. It is based on the use of low frequency electron paramagnetic resonance (EPR) spectroscopy with lithium phthalocyanine as the oxygen sensitive probe. Magnetic resonance imaging was used to document placement of the probe in the brain, and to assess potential cerebral changes associated with the placement. The technique provided accurate and reproducible measurements of localized pO2 in the brains of conscious rodents under a variety of physiological conditions and for time periods of at least 2 weeks. Using this approach we quantitated the depressing effects on cerebral pO2 of three representative anesthetics, isoflurane, ketamine/xylazine, and sodium pentobarbital. The effects of changing the content of oxygen in the breathing gas was investigated and found to change the cerebral pO2. In experiments with gerbils, crystals of lithium phthalocyanine were implanted in each side of the brain and using a one-dimensional magnetic field gradient, simultaneous measurement of pO2 values from normal and ischemic (ischemia induced by unilateral ligation of a carotid artery) hemispheres of the brain were obtained. These results demonstrate that EPR oximetry with lithium phthalocyanine is a versatile and useful method in the measurement of cerebral pO2 under various physiological and pathophysiological conditions.

Endotoxin-induced changes in intrarenal pO2, measured by in vivo electron paramagnetic resonance oximetry and magnetic resonance imaging.

Electron Paramagnetic Resonance (EPR) oximetry was used to measure tissue oxygen tension (pO2-partial pressure of oxygen) simultaneously in the kidney cortex and outer medulla in vivo in mice. pO2 in the cortex region was higher compared to that in the outer medulla. An intravenous injection of endotoxin resulted in a sharp drop in pO2 in the cortex and an increase in the medulla region, resulting in a transient period of equal pO2 in both regions. In control kidneys, functional Magnetic Resonance (MR) images showed the cortex region to have high signal intensity (T2*-weighted images), indicating that this region was well supplied with oxygenated hemoglobin, whereas the outer medulla showed low signal intensity. After administration of endotoxin, we observed an immediate increase in signal intensity in the outer medulla region, reflecting an increased level of oxygenated blood in this region. Pretreatment of mice with NG-monomethyl-L-arginine prevented both the changes in tissue pO2 and distribution of oxygenated hemoglobin, suggesting that localized production of nitric oxide has a critical role to play in renal medullary hemodynamics. In combining in vivo EPR with MR images of kidneys, we demonstrate the usefulness of these techniques for monitoring renal pO2 and changes in the distribution of oxygen.

Measurements of pO2 in Vivo, Including Human Subjects, by Electron Paramagnetic Resonance

The purpose of this paper is to provide an illustrative description of the current state of development of the use of electron paramagnetic resonance (EPR, or completely equivalently, electron spin resonance or ESR) to measure the partial pressure of oxygen (pO2) in tissues in vivo under physiological conditions. This summary is based on published and unpublished results from our laboratory (1–7) and does not attempt to describe the results of other laboratories which also are working along related lines (8–10). The pertinent features of our technique are illustrated. We also consider the current limitations of the technique and likely developments in the near future. Our evaluation is that: this technique now is suitable for immediate use in small animals; within a short period of time instruments will be available facilitating its use in larger animals; and preliminary studies are imminent in human subjects (7).

Histological Assessment of Rodent CNS Tissues to EPR Oximetry Probe Material

The effects of the paramagnetic oxygen sensing material, lithium phthalocyanine (LiPc) and fusinite were assessed in the brain of Mongolian gerbils and the spinal columns of rats respectively, to determine if there are histologically discernible changes in the tissue surrounding the probe material. This information is essential for the evaluation of the role of EPR oximetry in the measurements of pO2 in the CNS; the technique has great potential value for such measurements because it reports on the pO2 accurately and sensitively and, after the initial placement, measurements can be made repeatedly without invasive procedures or anesthesia. Histologic assessments demonstrated the inert nature of both the fusinite and LiPc EPR probes in rodent CNS tissue over relatively long (2 month) time periods. The fusinite suspensions and LiPc crystals (size range of approximately 100-200 microns) remained well localized to the point of injection and created mild acute tissue reaction on implantation (which appeared to resolve quickly) and virtually no tissue reaction at later times. The majority of the implanted fusinite and LiPc material was present extracellularly in the brain and spinal cord. MRI provided an accurate, noninvasive assessment of probe placement and was able to investigate pathologic effects (hemorrhage, edema, necrosis) associated with the probe placement and treatment effects.

In vivo EPR spectroscopy

This chapter is intended to provide a brief overview of the principles of electron paramagnetic resonance (EPR, or completely equivalently electron spin resonance, ESR) spectroscopy applied to living animals. It attempts to indicate especially those areas in which this approach is likely to be of value because it can provide useful information that cannot be provided as well by other approaches. As a matter of convenience the descriptions are drawn principally from the authors’ laboratory but it should be noted that there are a number of laboratories around the world, especially in Japan, which are also actively pursuing these developments. Because of the need for brevity in this volume, the coverage is illustrative rather than comprehensive but this fits well with the aim of the book which is to provide a review that will be useful for the longer term rather than only a review of the current state of development.