The cystic fibrosis transmembrane conductance regulator (CFTR) is a cAMP-regulated chloride channel localized primarily at the apical or luminal surfaces of epithelial cells that line the airway, gut, and exocrine glands; it is well established that CFTR plays a pivotal role in cholera toxin (CTX)-induced secretory diarrhea. Lysophosphatidic acid (LPA), a naturally occurring phospholipid present in blood and foods, has been reported to play a vital role in a variety of conditions involving gastrointestinal wound repair, apoptosis, inflammatory bowel disease, and diarrhea. Here we show, for the first time, that type 2 LPA receptors (LPA₂) are expressed at the apical surface of intestinal epithelial cells, where they form a macromolecular complex with Na⁺/H⁺ exchanger regulatory factor–2 and CFTR through a PSD95/Dlg/ZO-1–based interaction. LPA inhibited CFTR-dependent iodide efflux through LPA₂-mediated G_i pathway, and LPA inhibited CFTR-mediated short-circuit currents in a compartmentalized fashion. CFTR-dependent intestinal fluid secretion induced by CTX in mice was reduced substantially by LPA administration; disruption of this complex using a cell-permeant LPA₂-specific peptide reversed LPA₂-mediated inhibition. Thus, LPA-rich foods may represent an alternative method of treating certain forms of diarrhea.