Glutamate is the principal excitatory neurotransmitter in the brain, and its excessive release plays a key role in neuronal death associated with a wide range of neural disorders. Real-time monitoring of extracellular glutamate levels would be very helpful in understanding the excitotoxic process of neurotransmitters on brain injury. Toward the detection of L: -glutamate, we describe in this chapter the preparation of carbon nanotube (CNT) network probes with immobilized L: -glutamate oxidase (GLOD) by using a non-covalent functionalized method. Such GOLD-CNT network probes are evaluated with real-time electronic responses corresponding to standard glutamate solutions in vitro and a 11-vessel occlusion (11 VO) rat model in vivo. The ultrahigh sensitivity, selectivity, and fast response time of GLOD-CNT network probes are greatly promising for the real-time electronic detection of extracellular glutamate levels in brain.