Prostaglandin E2 depolarises sensory axons in vitro in an ANO1 and Nav1.8 dependent manner

TY - JOUR

T1 - Prostaglandin E2 depolarises sensory axons in vitro in an ANO1 and Nav1.8 dependent manner

AU - Kimourtzis, Georgios

AU - Rangwani, Natasha

AU - Jenkins, Bethan J.

AU - Jani, Siddharth

AU - McNaughton, Peter A.

AU - Raouf, Ramin

N1 - Publisher Copyright: © The Author(s) 2024.

PY - 2024/7/16

Y1 - 2024/7/16

N2 - Prostaglandin E2 (PGE2) is a major contributor to inflammatory pain hyperalgesia, however, the extent to which it modulates the activity of nociceptive axons is incompletely understood. We developed and characterized a microfluidic cell culture model to investigate sensitisation of the axons of dorsal root ganglia neurons. We show that application of PGE2 to fluidically isolated axons leads to sensitisation of their responses to depolarising stimuli. Interestingly the application of PGE2 to the DRG axons elicited a direct and persistent spiking activity propagated to the soma. Both the persistent activity and the membrane depolarisation in the axons are abolished by the EP4 receptor inhibitor and a blocker of cAMP synthesis. Further investigated into the mechanisms of the spiking activity showed that the PGE2 evoked depolarisation was inhibited by Nav1.8 sodium channel blockers but was refractory to the application of TTX or zatebradine. Interestingly, the depolarisation of axons was blocked by blocking ANO1 channels with T16Ainh-A01. We further show that PGE2-elicited axonal responses are altered by the changes in chloride gradient within the axons following treatment with bumetanide a Na-K-2Cl cotransporter NKCC1 inhibitor, but not by VU01240551 an inhibitor of potassium-chloride transporter KCC2. Our data demonstrate a novel role for PGE2/EP4/cAMP pathway which culminates in a sustained depolarisation of sensory axons mediated by a chloride current through ANO1 channels. Therefore, using a microfluidic culture model, we provide evidence for a potential dual function of PGE2 in inflammatory pain: it sensitises depolarisation-evoked responses in nociceptive axons and directly triggers action potentials by activating ANO1 and Nav1.8 channels.

AB - Prostaglandin E2 (PGE2) is a major contributor to inflammatory pain hyperalgesia, however, the extent to which it modulates the activity of nociceptive axons is incompletely understood. We developed and characterized a microfluidic cell culture model to investigate sensitisation of the axons of dorsal root ganglia neurons. We show that application of PGE2 to fluidically isolated axons leads to sensitisation of their responses to depolarising stimuli. Interestingly the application of PGE2 to the DRG axons elicited a direct and persistent spiking activity propagated to the soma. Both the persistent activity and the membrane depolarisation in the axons are abolished by the EP4 receptor inhibitor and a blocker of cAMP synthesis. Further investigated into the mechanisms of the spiking activity showed that the PGE2 evoked depolarisation was inhibited by Nav1.8 sodium channel blockers but was refractory to the application of TTX or zatebradine. Interestingly, the depolarisation of axons was blocked by blocking ANO1 channels with T16Ainh-A01. We further show that PGE2-elicited axonal responses are altered by the changes in chloride gradient within the axons following treatment with bumetanide a Na-K-2Cl cotransporter NKCC1 inhibitor, but not by VU01240551 an inhibitor of potassium-chloride transporter KCC2. Our data demonstrate a novel role for PGE2/EP4/cAMP pathway which culminates in a sustained depolarisation of sensory axons mediated by a chloride current through ANO1 channels. Therefore, using a microfluidic culture model, we provide evidence for a potential dual function of PGE2 in inflammatory pain: it sensitises depolarisation-evoked responses in nociceptive axons and directly triggers action potentials by activating ANO1 and Nav1.8 channels.

KW - Chloride channels

KW - Dorsal root ganglia neurons

KW - Inflammation

KW - Microfluidic cultures

KW - Pain

KW - Prostaglandin E2

KW - Sodium channels

UR - http://www.scopus.com/inward/record.url?scp=85199994801&partnerID=8YFLogxK

U2 - 10.1038/s41598-024-67793-1

DO - 10.1038/s41598-024-67793-1

M3 - Article

C2 - 39075089

AN - SCOPUS:85199994801

SN - 2045-2322

VL - 14

JO - Scientific Reports

JF - Scientific Reports

IS - 1

M1 - 17360

ER -