1. Three human adenocarcinoma cell lines, Colony-24 (Col-24), Col-6 and Col-1 have been studied as confluent epithelial layers able to transport ions vectorially in response to basolateral vasoactive intestinal polypeptide (VIP) and pancreatic polypeptides (PP). 2. Different species PP stimulated responses in Col-24 with Y(4)-like pharmacology. Bovine (b)PP, human (h)PP and porcine (p)PP were equipotent (EC(50) values 3.0--5.0 nM) while rat (r)PP, avian (a)PP and [Leu(31), Pro(34)]PYY (Pro(34)PYY) were significantly less potent. PYY was inactive. The PP pharmacology in Col-1 was comparable with Col-24. However, Col-6 cells were different; pPP had an EC(50) intermediate (22.0 nM) between that of bPP (3.0 nM) and hPP (173.2 nM), with aPP and rPP being at least a further fold less potent. 3. Deamidation of Tyr(36) in bPP (by O-methylation or hydroxylation) or removal of the residue resulted in significant loss of activity in Col-24. 4. GR231118 (1 microM) had no PP-like effects. In Col-24 and Col-1, GR231118 significantly attenuated bPP (30 nM) or hPP (100 nM) responses, but it did not alter bPP responses in Col-6. BIBP3226 and GR231118 both inhibited Y(1)-mediated responses which were only present in Col-6. 5. RT--PCR analysis confirmed the presence of hY(4) receptor mRNA in Col-24 and Col-1 epithelia but a barely visible hY(4) product was observed in Col-6 and we suggest that an atypical Y(4) receptor is expressed in this cell line.