Endocannabinoid Signaling

Moriello, Aniello Schiano, De Petrocellis, Luciano, Aniello Schiano Moriello, Luciano De Petrocellis

Mauro Maccarrone

The transient receptor potential vanilloid-1 ion channel (TRPV1) is a non-selective ligand-gated cation channel. It is an integrator of a wide variety of exogenous and endogenous physical and chemical stimuli, including capsaicin, noxious heat (>42 °C), and protons (pH < 5.2). TRPV1 is expressed predominantly in primary sensory neurons involved in pain sensation, but also in other neuronal cell types, in the plasma membrane of different non-neuronal cells such as immune cells, keratinocytes, smooth muscle cells, and in the urothelium. Some of these cell types are involved in inflammation. When activated, TRPV1 leads to the gating of cations, including Ca(2+), thus generating changes in intracellular Ca(2+) concentration. Calcium ions play fundamental roles in many cellular processes, virtually in all cells. The use of Ca(2+) fluorescent indicators is a tool for monitoring intracellular Ca(2+) concentration.In this chapter, we describe a method for recording and monitoring Ca(2+) signals through the single wavelength fluorescent indicator Fluo-4 acetoxymethyl (AM), and the ratiometric fluorescent indicator Fura-2 AM in HEK-293 cells transfected with TRPV1 and other TRP channels. TRPV1 pharmacological modulation may potentially represent a strategy for the control of pain and inflammatory conditions in a variety of diseases and injury states.