A new method was developed to carry out in vivo microdialysis experiments repetitively and routinely within an individual monkey. We designed and built a dialysis probe guide and holding device ('guide holder') which permits accurate placement of dialysis probes into cortical (e.g., prefrontal, hippocampus, and parietal cortices) and subcortical target areas (e.g., caudate nucleus, amygdala, and nucleus accumbens) of the rhesus monkey brain without extensive and repetitive surgery needed to expose a desired brain region. The guide holder is positioned, using MRI-guided coordinates, and fixed to the skull over an intended targeted region. This design provides an opportunity to conduct several experiments in a single monkey over an extended period and permits placement of several probes accurately into 'fresh' or 'experienced' tissue during repeated microdialysis experiments. In addition, during repeated dialysis experiments tissue trauma is minimized because no surgical procedure is necessary on the day of dialysate collection. This procedure can be readily adapted for use with an awake monkey.