Comparing the Effects of Isoflurane and Alpha Chloralose upon Mouse Physiology

Abstract

Functional magnetic resonance imaging of mice requires that the physiology of the mouse (body temperature, respiration and heart rates, blood pH level) be maintained in order to prevent changes affecting the outcomes of functional scanning, namely blood oxygenation level dependent (BOLD) measures and cerebral blood flow (CBF). The anesthetic used to sedate mice for scanning can have major effects on physiology. While alpha chloralose has been commonly used for functional imaging of rats, its effects on physiology are not well characterized in the literature for any species. In this study, we anesthetized or sedated mice with isoflurane or alpha chloralose for up to two hours, and monitored physiological parameters and arterial blood gasses. We found that, when normal body temperature is maintained, breathing rates for both drugs decrease over the course of two hours. In addition, alpha chloralose causes a substantial drop in heart rate and blood pH with severe hypercapnia (elevated blood CO2) that is not seen in isoflurane-treated animals. We suggest that alpha chloralose does not maintain normal mouse physiology adequately for functional brain imaging outcome measures.

Fig 1. Breathing Rate decreases over time during prolonged anesthesia.

There were no group differences between isoflurane and alpha chloralose-treated mice at any time (F_(1,386) = 0.001, p = 0.97), but there was a significant overall drop in breathing rates for both groups (F_(11,386) = 5.68, p<0.0001, n = 7–24 per time point).

Fig 2. Heart Rate drops under alpha chloralose sedation.

Mice retain a steady heart rate under isoflurane anesthesia, ranging from 463±20 beats/min to 489±20 beats/min. However, under alpha chloralose sedation, heart rate rapidly drops to below 350 beats/min, leading to a significant difference in heart rate between the treatment groups (F_(1,384) = 516.2,p<0.0001, n = 9–23 per time point). The shaded region indicates the normal heart rate for an awake mouse at rest (450–500 beats/min).

Fig 3. Blood oxygenation levels do not differ between isoflurane and alpha chloralose.

A) Over the course of 120 minutes of sedation/anesthesia, there were no group differences in blood oxygenation levels (F_(1,81) = 0.6, p = 0.44, n = 4–19 per time point), but there was a significant effect of time in the isoflurane group, where blood oxygenation levels spiked between 15 and 30 minutes (F_(5,82) = 4.06, p = 0.002), then returned to baseline levels. B) Blood oxygenation is not different between isoflurane and alpha chloralose after 120 minutes of anesthesia/sedation (p = 0.67).

Fig 4. Blood carbon dioxide levels increase significantly under alpha chloralose sedation over the course of 120 minutes.

A) Carbon dioxide levels in arterial blood were significantly higher under alpha chloralose sedation (F_(1,82) = 38.06, p<0.0001, n = 4–19 per time point). Although there was an overall effect of time (F_(5,82) = 4.06, p = 0.002), reflecting increasing CO₂ levels over time, these levels did not differ significantly from baseline over the course of 120 minutes under isoflurane anesthesia (p>0.05). B) After 120 minutes, carbon dioxide levels are significantly higher in alpha chloralose-sedated animals (p = 0.002).

Fig 5. Alpha chloralose sedation causes clear blood acidosis.

A) While pH drops under both types of anesthesia (F_(5,86) = 8.97, p<0.0001, n = 3–19 per time point), the drop is much more pronounced under alpha chloralose sedation (F_(1,86) = 35.72, p<0.0001), dropping from a normal reading of pH 7.39±0.05 during the first 15 minutes, to 7.13±0.07 by 120 minutes. In contrast, blood pH under isoflurane drops from 7.34±0.04 to 7.27±0.06 after two hours, which is still within a normal physiological range for the mouse. The shaded region indicates non-acidotic blood pH for the mouse (15). B) Blood is acidotic after 120 minutes of alpha chloralose sedation (p = 0.0006).