Physiological, pharmacological, and behavioral evidence for a TRPA1 channel that can elicit defensive responses in the medicinal leech

Abstract

Transient receptor potential ankyrin subtype 1 (TRPA1) channels are chemosensitive to compounds such as allyl isothiocyanate (AITC, the active component of mustard oil) and other reactive electrophiles and may also be thermodetectors in many animal phyla. In this study, we provide the first pharmacological evidence of a putative TRPA1-like channel in the medicinal leech. The leech's polymodal nociceptive neuron was activated by both peripheral and central application of the TRPA1 agonist AITC in a concentration-dependent manner. Responses to AITC were inhibited by the selective TRPA1 antagonist HC030031, but also by the TRPV1 antagonist SB366791. Other TRPA1 activators -N-methylmaleimide (NMM) and cinnamaldehyde (CIN) -also activated this nociceptive neuron, although HC030031 only inhibited the effects of NMM. The polymodal nociceptive neurons responded to moderately cold thermal stimuli (<17°C) and these responses were blocked by HC030031. AITC sensitivity was also found in the pressure-sensitive sensory neurons and was blocked by HC030031, but not by SB366791. AITC elicited a nocifensive withdrawal of the posterior sucker in a concentration-dependent manner that could be attenuated with HC030031. Peripheral application of AITC in vivo also produced swimming-like behavior that was attenuated by HC030031. These results suggest the presence of a TRPA1-like channel in the medicinal leech nervous system that responds to cold temperatures and may interact with the leech TRPV-like channel.

Calmodulin is responsible for Ca2+-dependent regulation of TRPA1 Channels

“…Electrophilic agonists, such as AITC, open TRPA1 through covalent modification of cysteine residues on TRPA11011, while non-electrophilic agonists, such as carvacrol, are believed to activate TRPA1 through binding directly to the channel212. In addition to chemosensation, considerable evidence supports a role for TRPA1 in transducing noxious cold131415161718, though this remains controversial. Furthermore, TRPA1 transduces noxious mechanical stimuli in pathological conditions such as inflammatory pain21920.…”

Endocannabinoids Have Opposing Effects On Behavioral Responses To Nociceptive And Non-nociceptive Stimuli

“…AITC has been shown to elicit nocifensive responses in invertebrates possessing a TRPA1 channel homolog 39, 40 . Previous studies in our lab have found that the Hirudo polymodal N cell does respond to peripheral application of AITC, which elicits withdrawal of the posterior sucker from the site of application 25 . The magnitude of this withdrawal reflex was noticeably greater than the responses elicited by the von Frey fibers and likely corresponds to a whole-body shortening response 29 .…”

“…The techniques for testing Hirudo responses to nociceptive and non-nociceptive stimuli are based on previously published protocols 25, 36 . Individual Hirudo were placed in a testing arena consisting of a plastic petri dish (145 mm diameter) lined with filter paper that had been saturated with pond water and maintained at room temperature.…”

“…For a nociceptive stimuli, 800 μL of 250 μM allyl isothiocyanate (AITC, the active component of mustard oil) was applied to the external surface of the posterior sucker using a pipette similar to what we have previously reported 25 . AITC has been shown to elicit nocifensive responses in invertebrates possessing a TRPA1 channel homolog 39, 40 .…”

“…In terms of somatosensory signaling, the Hirudo CNS possesses three bilateral pairs of rapidly-adapting touch-sensitive neurons (T cells), two pairs of slow-adapting pressure-sensitive neurons (P cells) and two pairs of high-threshold nociceptive neurons (N cells) 21 . The N cells can be further divided into mechanical and polymodal nociceptors, with the latter being sensitive to noxious mechanical, thermal and chemical stimuli, e.g., H + , capsaicin and mustard oil 22–25 . P cell stimulation is capable of producing localized withdrawals from mechanical stimuli referred to as local bending and local shortening 26–28 .…”