A sample pretreatment scheme involving PVC–metal ion interactions was optimized. A monitoring method for rat brain extracellular Fe(II)/Fe(III) was developed. The DLs were in the range 0.9–6.9 μg Fe L−1 based on a 12-μL microdialysate. The basal rat brain extracellular Fe(II)/Fe(III) concentrations were determined. The dynamic changes in response to neural depolarization were in vivo profiled.