Pain is a physiological state promoting protective responses to harmful episodes. However, pain can become pathophysiological and become a chronic disruptive condition, damaging quality of life. The mammalian K(2P)2.1 (KCNK2, TREK-1) channel, expressed in sensory neurons of the dorsal root ganglia, was previously identified as a polymodal molecular sensor involved in pain perception. Here, we report that two pain-associated signals, external acidosis and lysophosphatidic acid (LPA), known to rise during injury, inflammation and cancer, profoundly down-modulate human K(2P)2.1 activity. The pH regulatory effect was mediated by activation of proton-sensitive G-protein coupled receptors and phospholipase C. Physiological concentrations of LPA overcame the effects of known K(2P)2.1 activators, such as arachidonic acid, lysophosphatidylcholine and temperature, by activating cell-surface receptors stimulating the G(q) pathway. Furthermore, we identified three K(2P)2.1 carboxy-terminal residues that mediate both pH and LPA regulatory effects. Our results highlight the important role of K(2P)2.1 channels as receptors for mediators known to cause nociception.