A method for measuring brain partial pressure of oxygen in unanesthetized unrestrained subjects: the effect of acute and chronic hypoxia on brain tissue PO(2)

Abstract

The level of tissue oxygenation provides information related to the balance between oxygen delivery, oxygen utilization, tissue reactivity and morphology during physiological conditions. Tissue partial pressure of oxygen (PtO(2)) is influenced by the use of anesthesia or restraint. These factors may impact the absolute level of PtO(2). In this study we present a novel fiber optic method to measure brain PtO(2). This method can be used in unanesthetized, unrestrained animals, provides absolute values for PO(2), has a stable calibration, does not consume oxygen and is MRI compatible. Brain PtO(2) was studied during acute hypoxia, as well as before and after 28 days of high altitude acclimatization. A sensor was chronically implanted in the frontal cortex of eight Wistar rats. It is comprised of a fiber optic probe with a tip containing material that fluoresces with an oxygen dependent lifetime. Brain PtO(2) declines by 80% and 76% pre- and post-acclimatization, respectively, when the fraction of inspired oxygen declines from 0.21 to 0.08. In addition, a linear relationship between brain PtO(2) and inspired O(2) levels was demonstrated r(2)=0.98 and r(2)=0.99 (pre- and post-acclimatization). Hypoxia acclimatization resulted in an increase in the overall brain PtO(2) by approximately 35%. This paper demonstrates the use of a novel chronically implanted fiber optic based sensor for measuring absolute PtO(2). It shows a very strong linear relationship in awake animals between inspired O(2) and tissue O(2), and shows that there is a proportional increase in PtO(2) over a range of inspired values after exposure to chronic hypoxia.

Fig. 1

The implantable optode (a) micrograph of the fiber optic probe (10X). (b) Photograph of a chronically implantable probe supported by a transport sleeve. (c) Photograph of implanted fiber optic probe showing size, location and the supporting ring of dental cement. The picture was taken when the animal was awake.

Fig. 2

Description of implant region. (a & c) Coronal sections stained with H&E of the brain of two example subjects (4X magnification). The black square shows the location of the probe in the frontal cortex. (b & d) 10X magnification of the probe location. The tip was largely in cortical layers 4-5. There is no sign of inflammation. (e) Gradient echo MRI (4 days post-implantation) and (f) Spin echo MRI taken 30 days post implantation of the corresponding subject shown above. Black arrows show the location of the probe.

Fig. 3

Temperature effect on measurements of PO₂. Measurements were taken of the same solution with, and without temperature correction. The probe was immersed in a bath with a fixed PO₂ and the bath temperature ranged from 34-42 °C. The open circles show the PO₂ measured using automatic temperature correction. The closed circles show the PO₂ of the same bath when a fixed input temperature of 37 °C was used. Although there was a drift up, the change was not significant and the absolute difference from an automatic calibration was less than 1mmHg, well within the natural variation seen in a subject.

Fig. 4

Brain PtO₂ measured in awake unrestrained animals during acute hypoxia, before and after acclimatization to hypoxia. White circles are pre-acclimatization and the black circles are after 28 days acclimatization (375 ± 2 mmHg). These data show the degree of subject variability.

Fig. 5

The relationship between PtO₂ and inspired O₂ before and after acclimatization. (a) A linear regression is fit through the data, not including the final 2 points at low values of inspired O₂. Black circles, pre-acclimatization, r²=0.98, Open circles, post acclimatization. r²=0.99. The slopes of the curves before and after acclimatization are 0.31 and 0.37 respectively. The slopes are significantly different (p<0.001)(mean ± S.E. p≤ 0.01). Two way ANOVA with a Holm-Sidak post-hoc method was used to test for changes at each FiO₂ between pre- and post-acclimatization (* indicates significance, p < 0.05).