Several studies have shown that experimentally added guanine nucleotides (GN) may extracellularly modulate the glutamatergic system. However, there is no previous report of the extracellular occurrence of GN in the CNS. This study used HPLC to investigate the presence of GN in cerebrospinal fluid (CSF) samples of 26 patients. We demonstrated the extracellular presence of GN in the CNS. In human CSF, GMP was detected in a remarkably high concentration (236.20 microM). This evidence stimulates further investigation of extracellular GN modulation of neurotransmission, physiological mechanisms of action(s), and therapeutic potential(s) in the CNS.