Fig. 4.

Co²⁺ histochemistry on the HaCaT cell line expressing TRPV1 ectopically. Cells were incubated for 10 min in buffer A containing a 20 μM CAPS + 5 mM Co²⁺; b 5 mM Co²⁺ without CAPS; c 20 μM CAPS + 5 mM Co²⁺ + 300 nM CapZ; d 20 μM CAPS + 5 mM Co²⁺ + 5 μM CapZ; e 20 μM CAPS + 5 mM Co²⁺ + 100 μM CapZ; f 20 μM CAPS + 5 mM Co²⁺ + 500 nM RuRed; g 20 μM CAPS + 5 mM Co²⁺ + 7 μM RuRed; h 20 μM CAPS + 5 mM Co²⁺ + 100 μM RuRed. The dark precipitates indicate the presence of intracellular CoS that is blockable with RuRed, a channel blocker of heat and vanilloid pain signalling. Co-application of CapZ, a competitive antagonist of pungent vanilloids, also inhibited the cellular entry of Co²⁺ in a dose-dependent manner: this is a well-characterized evidence-based method of localization of intracellular Co²⁺. I: Gray values of the HaCaT cell line expressing TRPV1 measured by means of ImageJ software