Prostaglandins are involved in the modulation of various central functions (neurotransmitters and hypothalamic hormone release, thermoregulation, cerebro-vascular tone) and their levels increase in pathological situations [subarachnoid hemorrhage (SAH), stroke, convulsive disorders, etc.]. This study, using sensitive and specific antibodies, examined levels of four eicosanoids, Prostaglandins E2 and F2α(PGE2, PGF2α); and the metabolites of PGI2, 6-keto-prostaglandin F1α (6-keto-PGF1α) and of thromboxane A2, thromboxane B2 (TxB2), in the cerebrospinal fluid (CSF) obtained atraumatically from three species (human, canine, and feline). An assessment of the methodologic procedures (extraction and radioimmunoassay) was carried out. Human lumbar cerebrospinal fluid was shown to contain PGF2α (15–44 pg/ml), 6-keto-PGF1α (undetectable to 39 pg/ml), and TxB2 (un-detectable to 28 pg/ml), whereas PGE2 was undetectable (>18 pg) in all cases. In both animals species the eico-sanoid concentrations were 3-to 30-fold higher than humans for every prostaglandin examined. Although the prostaglandin profile for a given species remained constant (cat, PGE2:6-keto-PGF1α:TxB2:PGF2α; dog, TxB2:PGE2:6-keto-PGF1α:PGF2α), the absolute levels were found to be lower in the pentobarbital-anesthetized animals than in conscious cats. The correspondence of the prostaglandin profiles found in cerebrospinal fluid with the profiles reported in the literature in brain homogenates for the same species supports the hypothesis that cerebrospinal fluid levels of prostaglandins reflect the relative rates of synthesis in neural tissue.