Drosophila painless is a heat-activated, Ca²⁺-regulated channel

Abstract

Painless was firstly found in drosophila mutant, and appeared to play a key role in avoidance from noxious heat. Since it belongs to the TRPA1 subfamily, and both mammalian and drosophila TRPA1 are thought to be sensitive to temperature changes, Painless has also been expected to be a heat-activated ion channel. Therefore, we examined the thermal response of Painless expressed in HEK293 cells using Ca²⁺-imaging and patch-clamp methods. Painless-expressing HEK293 cells showed robust intracellular Ca²⁺ increase upon heating. Activation of Painless exhibited transient currents during heat application in a whole-cell patch-clamp mode. I-V relationship displayed dual rectification and extremely high Ca²⁺ permeability. The temperature threshold for activation with or without intracellular Ca²⁺ was around 42.6°C or 44.1°C, respectively. Painless appeared to be activated by heat with physiological intracellular Ca²⁺ concentration. Multiple heat application sensitized Painless by reducing the temperature thresholds in the presence of intra and extracellular Ca²⁺. Interestingly, Painless was insensitive to any known TRPA1 or other typical thermosensitive TRP activators. On the other hand, TRPA1 blockers such as ruthenium red and camphor inhibited the Painless-mediated heat-evoked currents. We conclude that Painless is a heat-sensitive TRP channel regulated by physiological level of intracellular Ca²⁺ ion. [J Physiol Sci. 2008;58 Suppl:S51]