Calcium Increase and Substance P Release Induced by the Neurotoxin Brevetoxin-1 in Sensory Neurons: Involvement of PAR2 Activation through Both Cathepsin S and Canonical Signaling

Abstract

Red tides involving Karenia brevis expose humans to brevetoxins (PbTxs). Oral exposition triggers neurotoxic shellfish poisoning, whereas inhalation induces a respiratory syndrome and sensory disturbances. No curative treatment is available and the pathophysiology is not fully elucidated. Protease-activated receptor 2 (PAR2), cathepsin S (Cat-S) and substance P (SP) release are crucial mediators of the sensory effects of ciguatoxins (CTXs) which are PbTx analogs. This work explored the role of PAR2 and Cat-S in PbTx-1-induced sensory effects and deciphered the signaling pathway involved. We performed calcium imaging, PAR2 immunolocalization and SP release experiments in monocultured sensory neurons or co-cultured with keratinocytes treated with PbTx-1 or P-CTX-2. We demonstrated that PbTx-1-induced calcium increase and SP release involved Cat-S, PAR2 and transient receptor potential vanilloid 4 (TRPV4). The PbTx-1-induced signaling pathway included protein kinase A (PKA) and TRPV4, which are compatible with the PAR2 biased signaling induced by Cat-S. Internalization of PAR2 and protein kinase C (PKC), inositol triphosphate receptor and TRPV4 activation evoked by PbTx-1 are compatible with the PAR2 canonical signaling. Our results suggest that PbTx-1-induced sensory disturbances involve the PAR2-TRPV4 pathway. We identified PAR2, Cat-S, PKA, and PKC that are involved in TRPV4 sensitization induced by PbTx-1 in sensory neurons.

Pacific-Ciguatoxin-2 and Brevetoxin-1 Induce the Sensitization of Sensory Receptors Mediating Pain and Pruritus in Sensory Neurons

“…PAR2 is mainly localized in small and medium DRG sensory neurons [ 79 ], particularly in peptidergic neurons [ 57 ] which are the sensory neurons mainly activated by CTX [ 46 ]. In previous work, we showed that P-CTX-2 and PbTx-1 activate PAR2 and increase the activity of the PAR2-activating protease cathepsin S in sensory neurons [ 26 , 28 ]. P-CTX-2 and PbTx-1 induced PAR2 internalization 20 min after exposure [ 26 , 28 ].…”

“…In previous work, we showed that P-CTX-2 and PbTx-1 activate PAR2 and increase the activity of the PAR2-activating protease cathepsin S in sensory neurons [ 26 , 28 ]. P-CTX-2 and PbTx-1 induced PAR2 internalization 20 min after exposure [ 26 , 28 ]. The calcium response to trypsin resensitized after 60−90 min suggests that PAR2 is re-expressed at the plasmic membrane from preformed pools of receptors after its rapid internalization [ 80 ].…”

“…PAR2 activation leads to TRPV4 sensitization [ 55 , 84 ] by both biased and canonical pathways. We showed that P-CTX-2 activates PAR2 [ 26 ], PbTx-1 activates PAR2 by both biased and canonical pathways, and PbTx-1 activates both PAR2 and TRPV4 in a common pathway [ 28 ]. The latter finding suggests a PAR2-dependent TRPV4 sensitization by PbTx-1.…”

“…Toxin binding to NaV channels enhances a sodium influx which leads to membrane hyperexcitability [ 22 , 23 , 24 , 25 ] that can lead to sensory disturbance. Previously, we have shown that Pacific-ciguatoxin-2 (P-CTX-2) and brevetoxin-1 (PbTx-1) induce neuropeptide (substance P (SP) and calcitonin gene-related peptide (CGRP)) release from a co-culture of sensory neurons and differentiated keratinocytes [ 26 , 27 , 28 ]. Other studies demonstrated a neuropeptide release from murine skin treated with Pacific-ciguatoxin-1 (P-CTX-1) [ 12 , 29 ].…”

“…Protease-activated receptor 2 (PAR2), mainly expressed by small diameter fibers [ 57 ], is involved in pruritus [ 58 ] and peripheral neuropathy induced by chemotherapy [ 59 , 60 , 61 ]. Previously, we have shown that PAR2 is activated by P-CTX-2 and PbTx-1 in sensory neurons [ 26 , 28 ]. PAR2 is known to sensitize transient receptor potential (TRP) channels [ 53 , 54 , 55 , 62 ].…”