CXCL1 is a chemokine with pleiotropic effects, including pain and itch. Itch, an unpleasant sensation that elicits the desire or reflex to scratch, it is evoked mainly from the skin and implicates activation of a specific subset of IB4+, C-type primary afferents. In previous studies we showed that acute application of CXCL1 induced a Ca2 + influx of low amplitude and slow kinetics in a subpopulation of transient receptor potential vanilloid type 1 (TRPV1)+/isolectin B4 (IB4) + dorsal root ganglia neurons which also responded to other itch-inducing agents. In this study we explored the mechanism behind the Ca2 + influx to better understand how CXCL1 acts on primary sensitive neurons to induce itch.Materials and methods:
Intracellular Ca2 + imaging and patch-clamp recordings on dorsal root ganglia neurons primary cultures and HEK293T cell transiently transfected with TRPV1 and CXCR2 plasmids were used to investigate the acute effect (12 min application) of 4 nM CXCL1. In primary cultures, the focus was on TRPV1+/IB4+ cells to which the itch-sensitive neurons belong.Key findings:
The results showed that the Ca2 + influx induced by the acute application of CXCL1 is mediated mainly by TRPV1 receptors and depends on extracellular Ca2 + not on intracellular stores. TRPV1 was activated, not sensitized by CXCL1, in a CXCR2 receptors- and actin filaments-dependent manner, since specific blockers and actin depolymerizing agents disrupted the CXCL1 effect.Significance:
This study brings additional data about the itch inducing mechanism of CXCL1 chemokine and about a new mechanism of TRPV1 activation via actin filaments.Graphical abstract:
The putative signaling pathway between CXCR2 receptors and TRPV1 upon acute application of CXCL1. After the ligand binding, the CXCR2 receptor couples to Gi/o protein and activates protein kinase pathways that can induce diverse intracellular effects, including phosphorylation of CXCR2. After activation, the receptor is internalized through a clathrin-mediated phosphorylation-dependent pathway and interacts with the actin filaments trough β-arrestin or another protein. Changes in the actin filaments are then transmitted to TRPV1 receptors which respond in a slow, low amplitude manner, allowing an influx of Na+ and Ca2 + ions. Questions marks indicate unknown molecules/mechanisms that mediate the interactions between actin filaments and CXCR2 or TRPV1.