Quantitative microdialysis using modified ultraslow microdialysis: direct rapid and reliable determination of free...

The only method to quantify free extracellular levels of drugs in the brain of living animals is microdialysis. However, quantitative microdialysis has been hampered by methodological issues for decades. The problems arise from the need to establish the in vivo recovery for appropriate quantitation. In dealing with these issues the "dynamic no-net-flux" (DNNF) method seemed to be the experimental method of choice. Major disadvantages were, however, the need for a very high degree of bioanalytical precision and accuracy and the need for a large number of animals. Moreover, today we know that the experimental data are not always straightforward. To improve robustness and practicality of quantitative microdialysis sampling we modified the ultraslow microdialysis approach. Ultraslow microdialysis uses very low microdialysis flow rates (<200 nl/min) which increase recovery (both in vivo and in vitro) to over 90%. However, new practical issues arise when attempting to work with these flow rates. The resulting very low volumes and long lag times make this method very impractical for general application. In the modified version, addition of a carrier flow after the dialysis process has been completed, which negates the problems of long lag times and low volumes. The resulting dilution of the dialysis sample concentration can simply be mathematically corrected. In the current study we measured the free brain levels of two CNS compounds using the classic DNNF and the new modified ultraslow dialysis method. Modified ultraslow microdialysis was shown to generate robust data with the use of only small numbers of rats. The method is a promising tool for common straightforward screening of blood-brain barrier penetration of compounds into the brain.