In ischemic and in inflamed tissues, pH levels down to 5.4 have been measured, and this local acidosis may contribute to pain and hyperalgesia in disease states. To evaluate the role of acid pH in nociception, we have studied identified primary afferents in a rat skin- saphenous nerve preparation in vitro where the receptive fields can be superfused at the highly permeable corium side with controlled solutions. The nerve endings were exposed to CO2-saturated synthetic interstitial fluid (SIF;pH 6.1) and to carbogen-gassed SIF phosphate buffered to different acid pH levels (5 min duration, 10 min intervals). Mechanical thresholds were repeatedly tested in a “blind” fashion by von Frey hair stimulation. Low-threshold mechanosensitive A beta- (n = 12) and A delta-fibers (n = 11) were not excited or sensitized by acid pH levels. In 24 of 96 nociceptor type C- and A delta-fibers, irregular low-frequency discharge with poor response characteristics was induced. However, a distinct subpopulation of mechanoheat sensitive, “polymodal” C-units (n = 25; 38%) showed stimulus-related responses increasing with proton concentration and encoding the time course of the pH change. Threshold levels were found to range from pH 6.9 to 6.1; mean maximum discharge was at pH 5.2. All such fibers responded to CO2 as well as to phosphate-buffered solution at the same pH 6.1. The CO2 responses, however, displayed significantly shorter latencies and more pronounced dynamic phases. The carboanhydrase blocker acetazolamide markedly delayed and reduced the CO2 responses. Prolonged application of acid pH (30 min) evoked nonadapting activity irrespective of oxygen supply. Many, but certainly not all, fibers sensitive to protons were also driven by capsaicin (10(- 6) M, 10(-5) M) and vice versa. Repeated or prolonged treatment with low pH induced a significant and lasting decrease of the mechanical (von Frey) thresholds in almost all C-fibers tested (from 35 to 16 mN, on average), and this occurred whether or not a fiber was excited by protons. The sensitizing effect was more pronounced the higher the initial von Frey thresholds (0.75 rank correlation). This sensitization to mechanical stimulation was in contrast to the combined action of other inflammatory mediators, bradykinin, 5-HT, histamine and prostaglandin E2. In conclusion, we suggest that pH sensitivity of nociceptors may be an important source of pain and hyperalgesia.