Daily variation in the levels of cyclic nucleotides and GABA was examined in seven brain regions of male Sprague-Dawley rats. Significant daily rhythm of cyclic AMP levels was found in the cerebellum and pons medulla oblongata. Circadian variation of cyclic GMP levels was found in the cerebellum, cerebral cortex, striatum, and hypothalamus. Daily variation of GABA levels was found in the pons medulla oblongata and striatum. Cyclic GMP in the pons medulla oblongata and GABA in the hypothalamus were found to exhibit ultradian variation of levels. These observed daily fluctuations of baseline levels should be considered when examining the duration of action of various drugs upon these substances.