ABSTRACT
Eugenol is widely used in dentistry to relieve pain. We have recently demonstrated voltage-gated Na+ and Ca2+ channels as molecular targets for its analgesic effects, and hypothesized that eugenol acts on P2X3, another pain receptor expressed in trigeminal ganglion (TG), and tested the effects of eugenol by whole-cell patch clamp and Ca2+ imaging techniques. In the present study, we investigated whether eugenol would modulate 5'-triphosphate (ATP)-induced currents in rat TG neurons and P2X3-expressing human embryonic kidney (HEK) 293 cells. ATP-induced currents in TG neurons exhibited electrophysiological properties similar to those in HEK293 cells, and both ATP- and alpha,beta-meATP-induced currents in TG neurons were effectively blocked by TNP-ATP, suggesting that P2X3 mediates the majority of ATP-induced currents in TG neurons. Eugenol inhibited ATP-induced currents in both capsaicin-sensitive and capsaicin-insensitive TG neurons with similar extent, and most ATP-responsive neurons were IB4-positive. Eugenol inhibited not only Ca2+ transients evoked by alpha,beta-meATP, the selective P2X3 agonist, in capsaicin-insensitive TG neurons, but also ATP-induced currents in P2X3-expressing HEK293 cells without co-expression of transient receptor potential vanilloid 1 (TRPV1). We suggest, therefore, that eugenol inhibits P2X3 currents in a TRPV1-independent manner, which contributes to its analgesic effect.
Subject(s)
Animals , Humans , Rats , Adenosine Triphosphate , Dentistry , Eugenol , HEK293 Cells , Kidney , Neurons , Nociceptors , Trigeminal GanglionABSTRACT
The effects of phorbol-12,13-dibuterate (PDBu) on total sodium current (INa-total), tetrodotoxin-resistant sodium current (INa-TTXr), 4-AP-sensitive potassium current (IA) and TEA-sensitive potassium current (IK) in trigeminal ganglion (TG) neurons were investigated.Whole-cell patch clamp techniques were used to record ion currents in cultured TG neurons of rats. Results revealed that 0.5 μmol/L PDBu reduced the amplitude of INa-total by (38.3±4.5)% (n=6, P＜0.05), but neither the G-V curve (control: V0.5 =-17.1±4.3 mV, k=7.4±1.3; PDBu: V0.5=-15.9±5.9 mV, k=5.9±1.4; n=6, P＞0.05) nor the inactivation rate constant (control: 3.6±0.9 ms; PDBu: 3.6±0.8 ms; n=6, P＞0.05) was altered. 0.5 μmol/L PDBu could significantly increase the amplitude of INa-TTXr by (37.2± 3.2)% (n=9, P＜0.05) without affecting the G-V curve (control: V0.5=-14.7±6.0 mV, k=6.9±1.4; PDBu: V0.5=-11.1±±5.3 mV, k=8.1±1.5; n=5, P＞0.05) or the inactivation rate constant (control: 4.6±±0.6 ms; PDBu: 4.2±0.5 ms; n=5, P＞0.05). 0.5 μmol/L PDBu inhibited IK by (15.6±5.0) % (n=16, P＜0.05), and V0.5 was significantly altered from - 4.7±1.4 mV to -7.9 ±1.8 mV (n=16, P＜0.05). IA was not significantly affected by PDBu, 0.5 μmol/L PDBu decreased IA by only (0.3±3.2)% (n=5, P＞0.05). It was concluded that PDBu inhibited INa-total but enhanced INa-TTXr, and inhibited IK without affecting IA. These data suggested that the activation of PKC pathway could exert the actions.
ABSTRACT
To investigate the effect of interleukin-1β (IL-1β) on IA and IK currents in cultured murine trigeminal ganglion (TG) neurons, whole-cell patch clamp technique was used to record the IA and IKcurrents before and after 20 ng/mL IL-1β perfusion. Our results showed that 20 ng/mL IL-1β inhibited IA currents (18.3±10.7)% (n=6, P＜0.05). IL-1β at 20 ng/mL had no effect on G-V curve of IA but moved the H-infinity curve V0.5 from -36.6 ± 6.1 mV to-42.4 ±5.2 m V (n=5, P＜0.01). However, 20 ng/mL IL-1β had effect on neither the amplitude nor the G-V curve of IK. IL-1β was found to selectively inhibit IA current in TG neurons and the effect may contribute to hyperalgesia under various inflammatory conditions.
ABSTRACT
We investigated the effect of alpha-adrenergic and cholinergic receptor agonists on Ca2+ current in adult rat trigeminal ganglion neurons using whole-cell patch clamp methods. The application of acetylcholine, carbachol, and oxotremorine (50 muM each) produced a rapid and reversible reduction of the Ca2+ current by 17+/-6%, 19+/-3% and 18+/-4%, respectively. Atropine, a muscarinic antagonist, blocked carbachol-induced Ca2+ current inhibition to 3 +/- 1%. Norepinephrine (50 muM) reduced Ca2+ current by 18 +/- 2%, while clonidine (50 muM), an alpha2-adrenergic receptor agonist, inhibited Ca2+ current by only 4 +/- 1%. Yohimbine, an alpha2-adrenergic receptor antagonist, did not block the inhibitory effect of norepinephrine on Ca2+ current, whereas prazosin, an alpha1-adrenergic receptor antagonist, attenuated the inhibitory effect of norepinephrine on Ca2+ current to 6 +/- 1%. This pharmacology contrasts with alpha2-adrenergic receptor modulation of Ca2+ channels in rat sympathetic neurons, which is sensitive to clonidine and blocked by yohimbine. Our data suggest that the modulation of voltage dependent Ca2+ channel by norepinephrine is mediated via an alpha1-adrenergic receptor. Pretreatment with pertussis toxin (250 ng/ml) for 16 h greatly reduced norepinephrine- and carbachol-induced Ca2+ current inhibition from 17 +/- 3% and 18 +/- 3% to 2 +/- 1% and 2 +/- 1%, respectively. These results demonstrate that norepinephrine, through an alpha1-adrenergic receptor, and carbachol, through a muscarinic receptor, inhibit Ca2+ currents in adult rat trigeminal ganglion neurons via pertussis toxin sensitive GTP-binding proteins.