title: Inhibition of intestinal dipeptide transport by the neuropeptide VIP is an anti-absorptive effect via the VPAC1 receptor in a human enterocyte-like cell line (Caco-2).
abstract: 1. Optimal dipeptide and peptidomimetic drug transport across the intestinal mucosal surface is dependent upon the co-operative functional activity of the di/tripeptide transporter hPepT1 and the Na(+)/H(+) exchanger NHE3. The ability of the anti-absorptive enteric neuropeptide VIP (vasoactive intestinal peptide) to modulate dipeptide uptake was determined using human intestinal (Caco-2) epithelial cell monolayers. 2. Uptake of glycylsarcosine (Gly-Sar) across the apical membrane of Caco-2 cell monolayers is inhibited by basolateral exposure to either VIP, pituitary adenylate cyclase-activating polypeptide (PACAP), or the VPAC(1) receptor agonist [(11,22,28)Ala]-VIP. Inhibition of Gly-Sar uptake is observed only in the presence of extracellular Na(+). Reverse-transcription polymerase chain reaction (RT-PCR) demonstrates that VPAC(1) mRNA is expressed in Caco-2 cells whereas VPAC(2) mRNA is not detected. 3. The VIP-induced inhibition of Gly-Sar uptake is abolished in the presence of the protein kinase A (PKA) inhibitor H-89 (N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide.2HCl). 4. (22)Na(+) uptake across the apical membrane is inhibited by the selective NHE3 inhibitor S1611. Experiments with BCECF [2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein]-loaded Caco-2 cells demonstrate that VIP reduces the NHE3-dependent recovery of intracellular pH (pH(i)) after dipeptide-induced acidification. Western blot of Caco-2 cell protein demonstrates expression of the NHE regulatory factor NHERF1 (expression of which is thought to be required for PKA-mediated inhibition of NHE3). 5. VIP has no effect on Gly-Sar uptake in the presence of S1611 suggesting that VIP and S1611 both modulate dipeptide uptake via the same mechanism. 6. These observations demonstrate that VIP (and PACAP) modulate activity of the H(+)/dipeptide transporter hPepT1 in a Na(+)-dependent manner consistent with the modulation being indirect through inhibition of NHE3.
authors.affiliation: School of Cell & Molecular Biosciences, The Medical School, University of Newcastle upon Tyne, Newcastle upon Tyne NE2 4HH.
authors.forename: Catriona M H
authors.initials: CM
authors.lastname: Anderson
authors.forename: Maria E
authors.initials: ME
authors.lastname: Mendoza
authors.forename: David J
authors.initials: DJ
authors.lastname: Kennedy
authors.forename: Demetrio
authors.initials: D
authors.lastname: Raldua
authors.forename: David T
authors.initials: DT
authors.lastname: Thwaites
mesh_terms: D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D018938:Caco-2 Cells / Q000187:drug effects* / Q000378:metabolism; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007408:Intestinal Absorption / Q000187:drug effects* / Q000502:physiology; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000166:cytology / Q000187:drug effects; D000074011:Peptide Transporter 1; D018005:Receptors, Vasoactive Intestinal Peptide / Q000037:antagonists & inhibitors* / Q000378:metabolism; D051238:Receptors, Vasoactive Intestinal Polypeptide, Type I; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000378:metabolism; D027981:Symporters; D014660:Vasoactive Intestinal Peptide / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D002352:Carrier Proteins; D000074011:Peptide Transporter 1; D018005:Receptors, Vasoactive Intestinal Peptide; D051238:Receptors, Vasoactive Intestinal Polypeptide, Type I; C082186:SLC15A1 protein, human; D017923:Sodium-Hydrogen Exchangers; D027981:Symporters; D014660:Vasoactive Intestinal Peptide