Development of a sensitive and inexpensive micropush-pull technique for the continuous analysis of brain...

A new, sensitive, convenient, inexpensive and low-maintenance miniaturized triple-cannula system (Zhange et al., 1990) for the push-pull perfusion of brain tissue has been constructed, and tested for its ability to exchange substances in rats, both in vitro and in vivo. In vitro: the cannula was immersed in a vial filled with 1.5 ml of a 1 microM standard solution of norepinephrine, dopamine, and their metabolites. Artificial cerebrospinal fluid was perfused through the system. Perfusate was collected at rates of 1, 2, 5, 10 and 20 microliters/min over 10 min. The recovery rate of the biogenic amines (using high performance liquid chromatography) at 10-min intervals for 60 min was consistent (57 +/- 1.31%) in low flow rate groups (1 approximately 10 microliters/ml). In vivo: [3H]choline (3 microCi, 35 nmol) was infused (6 min) into the lateral ventricle and collected by micropush-pull at 10-min intervals for 180 min, from the cerebellomedullary cistern and hippocampus. The highest [3H]choline count was reached within 10 min after infusion. Levels returned to baseline within 20 min following infusion of the tracer. The micropush-pull cannula was also adapted for chronic brain perfusion in vivo. Recovery of perfused fast green dye (0.00025%) was comparable (83 +/- 1.75%), using the same cannula for 3 sampling periods within 1 month in a freely moving rat. A cresyl violet analysis showed minimal damage to the brain tissue, with gliosis only evident in a narrow margin along the cannula track. Thus the micropush-pull technique is highly efficient in terms of exchange of material, causes minimal damage of brain tissue, and can be used chronically in awake animals. These data have been presented in preliminary form at the 1991 ASPET meetings (Zhang et al., 1991).