Glass capillary gas chromatography in combination with thermionic detection has been developed for the measurement of trace amounts of glycine, β-alanine, aspartic acid, γ-aminobutyric acid and glutamic acid from brain perfusates collected in vivo by push-pull cannula techniques. Amino acids extracted from the dried perfusate residues are converted to the corresponding N-pentafluoropropionyl hexafluoroisopropyl esters (PFP-HFIP derivatives) and separated by gas chromatography on a glass capillary column coated with methylsiloxane. The detection limit of the amino acids (signal-to-noise ratio 3 : 1) ranges between 200 fmol and 2.0 pmol. The assay was applied to the measurement of amino acids released in vivo in the pigeon optic tectum into perfusates collected by push-pull techniques.