Basis of the hyperexcitability of nociceptive neurons induced by activation of protease- activated receptor-2

Abstract

Proteases released during inflammation cleave a portion of the external segment of the Protease-Activated Receptors-2 (PAR2) and the distal portion of the receptor can then activate itself. The neuronal fibers of dorsal root ganglia (DRG) that innervate the intestine express PAR2 and its activation has been related with visceral hyperalgesia. In the first set of experiments, we analized some mechanisms underlying PAR2-evoked hyperexcitability of mouse colonic DRG neurons. To identify DRG neurons that inervate the colon we injected Fast Blue and DiI retrograde tracers into the mouse colon. We later used epi-fluorescence, to identify DiI-labelled neurons. Whole-cell current-clamp recordings of acutely dissociated neurons demonstrated that PAR2 activation with a brief application (2 min) of PAR2 agonists (SLIGRL-NH2 and trypsin) evoked sustained membrane depolarizations, which occurred concomitantly to an increased in input membrane resistance and a marked reduction in rheobase. In voltage clamp, SLIGRL-NH2 markedly suppressed delayed rectifier IK currents, but had no effect on the transient IA current or TTX-resistant sodium currents. The hyperexcitability induced by PAR2 activation was blocked with Calphostin C (a PKC inhibitor), and PD98059 (a ERK1/2 inhibitor). Our conclusion is that activation of PAR2 receptors on colonic nociceptive neurons causes hyperexcitability that is mediated by suppression of delayed rectifier IK currents. Both PKC and ERK1/2 mediate this hyperexcitability. These studies describe a novel mechanism of sensitization of colonic nociceptive neurons that may be implicated in conditions of visceral hyperalgesia such as irritable bowel syndrome. In a second set of experiments, carried out in HEK cells, activation of PAR2 sensitizes TRPV1 currents by a PKC���� and PKA-dependent mechanism. Activation of these receptors also potentiates TRPV4-mediated currents, effect that appears to require PKC and PKA activity.