The possibility that pituitary adenylyl cyclase-activating peptide (PACAP) is an inhibitory neurotransmitter has been investigated in the taenia of the guinea-pig caecum. The action of PACAP on muscle contractility and its ability to alter levels of adenosine-3':5'-cyclic monophosphate (cyclic AMP) and guanosine-3':5'-cyclic monophosphate (cyclic GMP) were investigated. PACAP-1-27 was an effective agonist, giving relaxations comparable in magnitude to isoproterenol; its EC50 was 3.4 x 10(-7) M. PACAP (10(-6) M) caused an almost two-fold increase in cyclic AMP levels; but the level of cyclic GMP was not affected. The relaxation caused by PACAP was slow in onset, with a latency of 5.8 +/- 0.8 s and reached a maximum at 9.1 +/- 1.1 s after onset. The relaxation was significantly reduced by apamin (10(-6) M) and suramin (10(-4) M) but was not reduced by tetrodotoxin (10(-7) M). Relaxation of the taenia coli caused by electrical stimulation of the inhibitory nerves was greatly reduced by apamin but only slightly reduced by suramin. PACAP-like immunoreactivity (-IR) was localised immunohistochemically in varicose nerve fibres within the taenia coli and in the underlying myenteric plexus and circular muscle. Approx. 50% of vasoactive intestinal peptide (VIP)-IR nerve fibres in the taenia also had immunoreactivity for PACAP; conversely, almost all PACAP-IR fibres were immunoreactive for VIP. PACAP-IR and substance P (SP)-IR were generally in separate fibres; only about 5% of SP-IR fibres were PACAP-IR. Radioimmunoassay revealed tissue concentrations of PACAP-1-27 and PACAP-1-38 of 1.0 +/- 0.1 and 2.1 +/- 0.3 (SEM) pmol/g wet weight of tissue, respectively. Material with PACAP-1-27 immunoreactivity co-eluted with authentic PACAP-1-27 on gel filtration chromatography, and PACAP-1-38 immunoreactivity also co-eluted with the authentic peptide. This study provides structural, chemical and pharmacological evidence that PACAP could be involved in inhibitory neurotransmission to the taenia coli of the guinea-pig caecum.