The absorptive transport of many nutrients across the brush-border membrane of the small intestinal wall is mediated by cotransport through ion-coupled transport proteins. The energy which drives these movements is stored in transepithelial and transmembrane ionic gradients. The human intestinal H⁺-coupled di/tripeptide transporter hPepT1 plays essential roles in nutrient (protein in the form of small peptides) absorption and in the high bioavailability of a variety of orally active peptidomimetic drugs (e.g. antibiotics, ACE inhibitors). For optimal peptide transport though hPepT1 to occur it is essential that the driving force (the transmembrane H⁺ gradient) is maintained during absorption. Thus the ionic composition of the microenvironments bathing both intra- and extracellular surfaces of the brush-border membrane will control transport. Despite the fact that the isolated H⁺-peptide symporter hPepT1 is a Na⁺-independent transporter, dipeptide transport in intact intestinal epithelia shows a degree of Na⁺-dependence. Ganapathy & Leibach (1985) proposed a model to account for this apparent Na⁺-dependence where hPepT1 functions in a cooperative way with an apically-localised Na⁺-H⁺ exchanger (now known as NHE3). This hypothesis was tested using a range of pharmacological and physiological tools including NHE inhibitors (e.g. S1611, EIPA) and modulators of the PKA pathway (e.g. forskolin, VIP). Experiments were performed in which hPepT1 and NHE3 function were determined using intact monolayers of the human intestinal epithelial cell line (Caco-2) or hPepT1-expressing Xenopus laevis oocytes (Thwaites et al. 1999, 2002; Kennedy et al. 2002, 2005; Anderson et al. 2003). At the apical surface of Caco-2 cell monolayers, H⁺-dipeptide transport via hPepT1 led to a selective activation of NHE3 and Na⁺-dependent H⁺-efflux suggesting that activity of the two transporters is coupled functionally. Similarly modulators of NHE3 activity (e.g. S1611 or VIP) decreased dipeptide transport capacity across the apical membrane of Caco-2 cell monolayers but only under conditions in which NHE3 was active. In conclusion, the two transporters hPepT1 (a symporter) and NHE3 (an exchanger) function cooperatively to mediate dipeptide and Na⁺ absorption across the apical membrane whilst maintaining intracellular pH and the transmembrane H⁺ electrochemical gradient.