TRPV1 is a member of the transient receptor potential (TRP) family of cation channels. It is expressed in sensory neurons of the dorsal root and trigeminal ganglia as well as in a wide range of non-neuronal tissues. The channel proteins serve as polymodal receptors for various potentially harmful stimuli to prevent tissue damage by mediating unpleasant or painful sensations. Using Ca imaging and voltage-clamp recordings, we found that low millimolar doses of Ni2+ (NiSO4) are able to induce non-specific cation currents in a capsaicin-sensitive population of cultured mouse trigeminal ganglion neurons. In addition, we show that NiSO4 elicits intracellular Ca2+ transients and membrane currents in HEK293 and CHO cells heterologously expressing rat TRPV1. The use of voltage ramps from -100 to +100 mV revealed a strong outward rectification of these currents. Application of NiSO4 to the cytoplasmic face of inside-out membrane patches did not induce any currents. However, delivering NiSO4 to the extracellular face during outside-out recordings, we observed a significant increase in open probability paralleled by a decrease in channel conductance. When combined with other TRPV1 agonists, NiSO4 produces a bimodal effect on TRPV1 activity, depending on the strength and concentration of the second stimulus. Outwardly directed currents induced by low doses of capsaicin and nearly neutral pH values ( approximately pH = 7.0-6.5) were augmented by low doses of NiSO4. In contrast, responses to stronger stimuli were reduced by NiSO4. Moreover, we were able to identify amino acids involved in the effect of NiSO4 on TRPV1.