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Probe calibrations are required for accurate estimations of extracellular concentrations in microdialysis experiments. Several methods have been developed and validated for in vivo determination of dialysis membrane recovery such as the perfusion rate method and the No Net Flux method. 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, where Cin and Cout were the concentrations of a compound in the perfusate and in the dialysate, respectively. First, the accuracy of the No Net Flux method to estimate in vivo recovery was compared in two situations: diffusion from the probe into the dialysis medium and diffusion from the outer medium into the probe. The point of no net transport was used to estimate the concentration surrounding the probe. Neither difference between extracellular concentrations (intercept values) nor difference between recoveries were observed. Then the reverse dialysis method was tested to estimate the relative loss of drug from the perfusate when the probe was placed in a drug-free medium. Finally comparisons of the behavior of the drug diffusion across the membrane under increasing gradient conditions have shown an asymptotic profile, specific 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 threshold 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 vivo without a systemic administration of a drug and (b) to characterize the diffusion process as a function of concentration gradient and tissue. |
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