MICRODIALYSIS PROBES CALIBRATION - GRADIENT AND TISSUE-DEPENDENT CHANGES IN NO NET FLUX AND REVERSE DIALYSIS METHODS

Probe calibrations are required for accurate estimations of extracellu lar concentrations in microdialysis experiments. Several methods have been developed and validated for in vivo determination of dialysis mem brane recovery such as the perfusion rate method and the No Net Flux m ethod. In this study, the No Net Flux and the reverse dialysis methods were investigated. Both measure the net transport of drug across the dialysis membrane. The recovery was defined as R = (Cin - Cout)/Cin, w here Cin and Cout were the concentrations of a compound in the perfusa te and in the dialysate, respectively. First, the accuracy of the No N et Flux method to estimate in vivo recovery was compared in two situat ions: diffusion from the probe into the dialysis medium and diffusion from the outer medium into the probe. The point of no net transport wa s used to estimate the concentration surrounding the probe. Neither di fference between extracellular concentrations (intercept values) nor d ifference between recoveries were observed. Then the reverse dialysis method was tested to estimate the relative loss of drug from the perfu sate when the probe was placed in a drug-free medium. Finally comparis ons of the behavior of the drug diffusion across the membrane under in creasing gradient conditions have shown an asymptotic profile, specifi c of the tissue: blood, muscle, and adipose tissue. The faster a drug was removed by microvascular transport (blood>muscle>adipocytes), the higher was the recovery, until the perfusate concentration reached a t hreshold value where the transport process became gradient limited and no more tissue limited. The usefulness of the reverse dialysis method has allowed us (a) to estimate the recovery of a dialysis probe in vi vo without a systemic administration of a drug and (b) to characterize the diffusion process as a function of concentration gradient and tis sue.