Distinctive Changes in Plasma Membrane Phosphoinositides Underlie Differential Regulation of TRPV1 in Nociceptive Neurons

Abstract

Transient Receptor Potential Vanilloid 1 (TRPV1) is a polymodal, Ca 2ϩ-permeable cation channel crucial to regulation of nociceptor responsiveness. Sensitization of TRPV1 by G-protein coupled receptor (GPCR) agonists to its endogenous activators, such as low pH and noxious heat, is a key factor in hyperalgesia during tissue injury as well as pathological pain syndromes. Conversely, chronic pharmacological activation of TRPV1 by capsaicin leads to calcium influx-induced adaptation of the channel. Paradoxically, both conditions entail activation of phospholipase C (PLC) enzymes, which hydrolyze phosphoinositides. We found that in sensory neurons PLC␤ activation by bradykinin led to a moderate decrease in phosphatidylinositol-4,5-bisphosphate (PI(4,5)P 2 ), but no sustained change in the levels of its precursor PI(4)P. Preventing this selective decrease in PI(4,5)P 2 inhibited TRPV1 sensitization, while selectively decreasing PI(4,5)P 2 independently of PLC potentiated the sensitizing effect of protein kinase C (PKC) on the channel, thereby inducing increased TRPV1 responsiveness. Maximal pharmacological TRPV1 stimulation led to a robust decrease of both PI(4,5)P 2 and its precursor PI(4)P in sensory neurons. Attenuating the decrease of either lipid significantly reduced desensitization, and simultaneous reduction of PI(4,5)P 2 and PI(4)P independently of PLC inhibited TRPV1. We found that, on the mRNA level, the dominant highly Ca 2ϩ -sensitive PLC isoform in dorsal root ganglia is PLC␦4. Capsaicin-induced desensitization of TRPV1 currents was significantly reduced, whereas capsaicininduced nerve impulses in the skin-nerve preparation increased in mice lacking this isoform. We propose a comprehensive model in which differential changes in phosphoinositide levels mediated by distinct PLC isoforms result in opposing changes in TRPV1 activity.

Physiological and behavioral evidence of a capsaicin sensitive TRPV-like channel in the medicinal leech

“…Third, SB366791 also blocked the polymodal cell response to noxious thermal stimulation. Fourth, the response of the lN cell to capsaicin exhibited desensitization, a property observed in other capsaicinsensitive TRPV channels (Joseph et al, 2013;Lukacs et al, 2013). Traditionally, it has been thought that invertebrates were not sensitive to capsaicin and thus that they do not have a TRPV channel that is functionally similar to mammalian TRPV1 channels.…”

“…However, lower concentrations of SB366791 (10 μmol l −1 ) have been shown previously to be effective at blocking the effects of capsaicin (10 μmol l −1 ) on central synapses (Yuan and Burrell, 2012). These results are consistent with the hypothesis that peripheral and central responses to capsaicin are mediated by a TRPV-like receptor.Mammalian TRPV channels undergo desensitization after exposure to capsaicin (Joseph et al, 2013;Lukacs et al, 2013). To determine whether desensitization of capsaicin-elicited responses occurs in the leech, experiments were conducted in which the initial response of lN cells to 100 μmol l −1 capsaicin was compared with that of a subsequent treatment with capsaicin (5 min inter-treatment interval).…”

Mechanism for phosphoinositide selectivity and activation of TRPV1 ion channels

“…PI(4,5)P 2 has been proposed to inhibit TRPV1, mediating the sensitizing actions of bradykinin and nerve growth factor (Chuang et al, 2001;Cao et al, 2013a). In contrast, others (including us) have proposed that PI(4,5)P 2 activates TRPV1 (Stein et al, 2006;Lukacs et al, 2007Lukacs et al, , 2013aKlein et al, 2008), and the depletion of PI(4,5)P 2 by phospholipase C may play a role in Ca 2+ -dependent desensitization (Mercado et al, 2010;Lukacs et al, 2013b). However, a recent study measured the apparent affinity of TRPV1 for diC8-PI(4,5)P 2 and found that the activating effects occurred with a K 1/2 of only 0.03 mol % in the plasma membrane, raising the question of whether PI(4,5)P 2 levels in the plasma membrane ever get sufficiently low selectivity must involve multiple, specific inter actions between TRPV1 and the phosphoinositide ligand.…”

“…In addition, specific dephosphorylation of PI(4,5)P 2 by a VSP in whole-cell patch-clamp experiments with HEK293 cells transfected with TRPV1 showed that the VSP did not inhibit TRPV1 activity induced by extracellular protons although it inhibited Kir2.1 channels in the same cells (Lukacs et al, 2013b). Together these data suggest that, even in TRPV1-WT, PI(4)P may play a role in translocation of PH-YFP, and presumably depletion of PI(4,5)P 2 , was not different for TRPV1-WT and TRPV1-R721A, a significant attenuation of channel inhibition was observed in TRPV1-R721A compared with TRPV1-WT.…”

“…4). Using excised patches was critical, as it has been previously shown in HEK293 cells examined with wholecell patch-clamp that VSP activation is insufficient to deplete PI(4,5)P 2 below the threshold for activation of TRPV1, albeit using another species of VSP (Lukacs et al, 2013b). In addition to removing the channels from many cellular enzymes, patch excision allowed us to perform our experiments in the absence of ATP, so that new synthesis of PI(4)P and PI(4,5)P 2 by lipid kinases did not occur.…”