Transient Receptor Potential (TRP) cation channels, like the TRP Vanilloid 1 
(TRPV1) and TRP Ankyrin 1 (TRPA1), are expressed on primary sensory neurons. 
These thermosensor channels play a role in pain processing. We have provided 
evidence previously that lipid raft disruption influenced the TRP channel 
activation, and a carboxamido-steroid compound (C1) inhibited TRPV1 activation. 
Therefore, our aim was to investigate whether this compound exerts its effect 
through lipid raft disruption and the steroid backbone (C3) or whether altered 
position of the carboxamido group (C2) influences the inhibitory action by 
measuring Ca2+ transients on isolated neurons and calcium-uptake on 
receptor-expressing CHO cells. Membrane cholesterol content was measured by 
filipin staining and membrane polarization by fluorescence spectroscopy. Both 
the percentage of responsive cells and the magnitude of the intracellular Ca2+ 
enhancement evoked by the TRPV1 agonist capsaicin were significantly inhibited 
after C1 and C2 incubation, but not after C3 administration. C1 was able to 
reduce other TRP channel activation as well. The compounds induced cholesterol 
depletion in CHO cells, but only C1 induced changes in membrane polarization. 
The inhibitory action of the compounds on TRP channel activation develops by 
lipid raft disruption, and the presence and the position of the carboxamido 
group is essential.