The ability to monitor the chemical composition of brain interstitial fluid remains an important challenge in the field of bioanalytical chemistry. In particular, microelectrode biosensors are a promising resource for the detection of neurochemicals in interstitial fluid in both animals and humans. These biosensors can provide second-by-second temporal resolution and enzymatic recognition of virtually any redox or nonredox molecule. However, despite miniaturization of these sensors to 50–250 μm in diameter to avoid vascular and cellular injury, inflammation and foreign-body reactions still occur following their implantation. Here, we fabricated microelectrodes with platinized carbon fibers to create biosensors that have an external diameter that is less than 15 μm. Platinization was achieved with physical vapor deposition, and increased sensitivity to hydrogen peroxide and improved enzymatic detection were observed for these carbon fiber microelectrodes. When these devices were implanted in the brains of rats, no injuries to the parenchyma or brain blood vessels were detected. In addition, these microelectrodes provided different estimates of basal glucose, lactate, and oxygen concentrations compared to conventional biosensors. Induction of spreading depolarization in the cerebral cortex further demonstrated the greater sensitivity of our microelectrodes to dynamic neurochemical changes. Thus, these minimally invasive devices represent a major advance in our ability to analyze brain interstitial fluid.