Transient receptor potential ion channel, vanilloid subfamily, type 1 (TRPV1) cation channel, and cannabinoid receptor 1 (CB1) are essential in the modulation of nociceptive signaling in the spinal cord dorsal horn that underlies different pathological pain states. TRPV1 and CB1 receptors share the endogenous agonist anandamide (AEA), produced from N-arachidonoylphosphatidylethanolamine (20:4-NAPE). We investigated the effect of the anandamide precursor 20:4-NAPE on synaptic activity in naive and inflammatory conditions. Patch-clamp recordings of miniature excitatory postsynaptic currents (mEPSCs) from superficial dorsal horn neurons in rat acute spinal cord slices were used. Peripheral inflammation was induced by subcutaneous injection of carrageenan. Under naive conditions, mEPSCs frequency (0.96 ± 0.11 Hz) was significantly decreased after 20 μM 20:4-NAPE application (55.3 ± 7.4%). This 20:4-NAPE-induced inhibition was blocked by anandamide-synthesizing enzyme N-acyl phosphatidylethanolamine phospholipase D (NAPE-PLD) inhibitor LEI-401. In addition, the inhibition was prevented by the CB1 receptor antagonist PF 514273 (0.2 μM) but not by the TRPV1 receptor antagonist SB 366791 (10 μM). Under inflammatory conditions, 20:4-NAPE (20 μM) also exhibited a significant inhibitory effect (74.5 ± 8.9%) on the mEPSCs frequency that was prevented by the TRPV1 receptor antagonist SB 366791 but not by PF 514273 application. Our results show that 20:4-NAPE application has a significant modulatory effect on spinal cord nociceptive signaling that is mediated by both TRPV1 and CB1 presynaptic receptors, whereas peripheral inflammation changes the underlying mechanism. The switch between TRPV1 and CB1 receptor activation by the AEA precursor 20:4-NAPE during inflammation may play an important role in nociceptive processing, hence the development of pathological pain.

• Keywords
• 20:4-NAPE, CB1, NAPE-PLD, TRPV1, anandamide, inflammation, spinal cord,
• Publication type
• Journal Article MeSH

Protease-activated receptors 2 (PAR2) and transient receptor potential vanilloid 1 (TRPV1) receptors in the peripheral nerve endings are implicated in the development of increased sensitivity to mechanical and thermal stimuli, especially during inflammatory states. Both PAR2 and TRPV1 receptors are co-expressed in nociceptive dorsal root ganglion (DRG) neurons on their peripheral endings and also on presynaptic endings in the spinal cord dorsal horn. However, the modulation of nociceptive synaptic transmission in the superficial dorsal horn after activation of PAR2 and their functional coupling with TRPV1 is not clear. To investigate the role of spinal PAR2 activation on nociceptive modulation, intrathecal drug application was used in behavioural experiments and patch-clamp recordings of spontaneous, miniature and dorsal root stimulation-evoked excitatory postsynaptic currents (sEPSCs, mEPSCs, eEPSCs) were performed on superficial dorsal horn neurons in acute rat spinal cord slices. Intrathecal application of PAR2 activating peptide SLIGKV-NH2 induced thermal hyperalgesia, which was prevented by pretreatment with TRPV1 antagonist SB 366791 and was reduced by protein kinases inhibitor staurosporine. Patch-clamp experiments revealed robust decrease of mEPSC frequency (62.8 ± 4.9%), increase of sEPSC frequency (127.0 ± 5.9%) and eEPSC amplitude (126.9 ± 12.0%) in dorsal horn neurons after acute SLIGKV-NH2 application. All these EPSC changes, induced by PAR2 activation, were prevented by SB 366791 and staurosporine pretreatment. Our results demonstrate an important role of spinal PAR2 receptors in modulation of nociceptive transmission in the spinal cord dorsal horn at least partially mediated by activation of presynaptic TRPV1 receptors. The functional coupling between the PAR2 and TRPV1 receptors on the central branches of DRG neurons may be important especially during different pathological states when it may enhance pain perception.

• MeSH
• Hypersensitivity metabolism   pathology   MeSH
• Anilides pharmacology   MeSH
• Posterior Horn Cells drug effects   physiology   MeSH
• Behavior, Animal drug effects   MeSH
• Cinnamates pharmacology   MeSH
• Excitatory Postsynaptic Potentials drug effects   MeSH
• Hyperalgesia etiology   prevention & control   MeSH
• Protein Kinase Inhibitors pharmacology   MeSH
• TRPV Cation Channels antagonists & inhibitors   metabolism   MeSH
• Rats MeSH
• Patch-Clamp Techniques MeSH
• Spinal Cord metabolism   MeSH
• Synaptic Transmission physiology   MeSH
• Oligopeptides pharmacology   MeSH
• Rats, Wistar MeSH
• Receptor, PAR-2 agonists   metabolism   MeSH
• Staurosporine pharmacology   MeSH
• In Vitro Techniques MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Anilides MeSH
• Cinnamates MeSH
• Protein Kinase Inhibitors MeSH
• TRPV Cation Channels MeSH
• N-(3-methoxyphenyl)-4-chlorocinnamanilide MeSH Browser
• Oligopeptides MeSH
• Receptor, PAR-2 MeSH
• seryl-leucyl-isoleucyl-glycyl-lysyl-valinamide MeSH Browser
• Staurosporine MeSH
• TRPV1 receptor MeSH Browser

BACKGROUND: Acute postoperative pain is one of the frequent reasons for pain treatment. However, the exact mechanisms of its development are still not completely clear. Transient receptor potential vanilloid 1 (TRPV1) receptors are involved in nociceptive signaling in various hypersensitive states. Here we have investigated the contribution of TRPV1 receptors expressed on cutaneous peripheral nociceptive fibers and in the spinal cord on the development and maintenance of hypersensitivity to thermal and mechanical stimuli following surgical incision. A rat plantar incision model was used to test paw withdrawal responses to thermal and mechanical stimuli. The effect of the TRPV1 receptor antagonist SB366791 was investigated 1) by intrathecal injection 15 min before incision and 2) intradermal injection before (30 min) and immediately after the surgery. Vehicle-injected rats and naïve animals treated identically were used as controls. RESULTS: Plantar incision induced mechanical allodynia and hyperalgesia and thermal hyperalgesia. A single intrathecal administration of SB366791 significantly reduced postincisional thermal hyperalgesia and also attenuated mechanical allodynia, while mechanical hyperalgesia remained unaffected. Local intradermal SB366791 treatment reduced thermal hyperalgesia and mechanical allodynia without affecting mechanical hyperalgesia. CONCLUSIONS: Our experiments suggest that both peripheral and spinal cord TRPV1 receptors are involved in increased cutaneous sensitivity following surgical incision. The analgesic effect of the TRPV1 receptor antagonist was especially evident in the reduction of thermal hyperalgesia. The activation of TRPV1 receptors represents an important mechanism in the development of postoperative hypersensitivity.

• MeSH
• Anilides pharmacology   therapeutic use   MeSH
• Time Factors MeSH
• Cinnamates pharmacology   therapeutic use   MeSH
• Physical Stimulation adverse effects   MeSH
• Touch MeSH
• Hyperalgesia drug therapy   etiology   MeSH
• TRPV Cation Channels antagonists & inhibitors   metabolism   MeSH
• Rats MeSH
• Pain Measurement drug effects   MeSH
• Spinal Cord drug effects   metabolism   MeSH
• Disease Models, Animal MeSH
• Pain, Postoperative complications   drug therapy   MeSH
• Rats, Wistar MeSH
• Pain Threshold drug effects   MeSH
• Reaction Time drug effects   MeSH
• Hot Temperature MeSH
• Drug Administration Routes MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Anilides MeSH
• Cinnamates MeSH
• TRPV Cation Channels MeSH
• N-(3-methoxyphenyl)-4-chlorocinnamanilide MeSH Browser
• Trpv1 protein, rat MeSH Browser

BACKGROUND: The cytokine tumor necrosis factor α (TNFα) is an established pain modulator in both the peripheral and central nervous systems. Modulation of nociceptive synaptic transmission in the spinal cord dorsal horn (DH) is thought to be involved in the development and maintenance of several pathological pain states. Increased levels of TNFα and its receptors (TNFR) in dorsal root ganglion (DRG) cells and in the spinal cord DH have been shown to play an essential role in neuropathic pain processing. In the present experiments the effect of TNFα incubation on modulation of primary afferent synaptic activity was investigated in a model of peripheral neuropathy. METHODS: Spontaneous and miniature excitatory postsynaptic currents (sEPSC and mEPSCs) were recorded in superficial DH neurons in acute spinal cord slices prepared from animals 5 days after sciatic nerve transection and in controls. RESULTS: In slices after axotomy the sEPSC frequency was 2.8 ± 0.8 Hz, while neurons recorded from slices after TNFα incubation had significantly higher sEPSC frequency (7.9 ± 2.2 Hz). The effect of TNFα treatment was smaller in the slices from the control animals, where sEPSC frequency was 1.2 ± 0.2 Hz in slices without and 2.0 ± 0.5 Hz with TNFα incubation. Tetrodotoxin (TTX) application in slices from axotomized animals and after TNFα incubation decreased the mEPSC frequency to only 37.4 ± 6.9% of the sEPSC frequency. This decrease was significantly higher than in the slices without the TNFα treatment (64.4 ± 6.4%). TTX application in the control slices reduced the sEPSC frequency to about 80% in both TNFα untreated and treated slices. Application of low concentration TRPV1 receptors endogenous agonist N-oleoyldopamine (OLDA, 0.2 μM) in slices after axotomy induced a significant increase in mEPSC frequency (175.9 ± 17.3%), similar to the group with TNFα pretreatment (158.1 ± 19.5%). CONCLUSIONS: Our results indicate that TNFα may enhance spontaneous transmitter release from primary afferent fibres in the spinal cord DH by modulation of TTX-sensitive sodium channels following sciatic nerve transection. This nerve injury also leads to enhanced sensitivity of presynaptic TRPV1 receptors to endogenous agonist. Modulation of presynaptic receptor activity on primary sensory terminals by TNFα may play an important role in neuropathic pain development.

• MeSH
• Excitatory Postsynaptic Potentials MeSH
• Rats MeSH
• Patch-Clamp Techniques MeSH
• Spinal Cord pathology   physiology   ultrastructure   MeSH
• Peripheral Nervous System Diseases chemically induced   pathology   physiopathology   MeSH
• Synaptic Transmission physiology   MeSH
• Neurons, Afferent cytology   drug effects   physiology   MeSH
• Nociception drug effects   physiology   MeSH
• Rats, Wistar MeSH
• Sodium Channels metabolism   MeSH
• Ganglia, Spinal cytology   physiology   MeSH
• Synapses physiology   MeSH
• Tumor Necrosis Factor-alpha pharmacology   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Sodium Channels MeSH
• Tumor Necrosis Factor-alpha MeSH

Modulation of synaptic transmission in the spinal cord dorsal horn is thought to be involved in the development and maintenance of different pathological pain states. The proinflamatory cytokine, tumor necrosis factor alpha (TNFalpha), is an established pain modulator in both the peripheral and the central nervous system. Up-regulation of TNFalpha and its receptors (TNFR) in dorsal root ganglion (DRG) cells and in the spinal cord has been shown to play an important role in neuropathic and inflammatory pain conditions. Transient receptor potential vanilloid 1 (TRPV1) receptors are known as molecular integrators of nociceptive stimuli in the periphery, but their role on the spinal endings of nociceptive DRG neurons is unclear. The endogenous TRPV1 receptor agonist N-oleoyldopamine (OLDA) was shown previously to activate spinal TRPV1 receptors. In our experiments the possible influence of TNFalpha on presynaptic spinal cord TRPV1 receptor function was investigated. Using the patch-clamp technique, miniature excitatory postsynaptic currents (mEPSCs) were recorded in superficial dorsal horn neurons in acute slices after incubation with 60 nM TNFalpha. A population of dorsal horn neurons with capsaicin sensitive primary afferent input recorded after the TNFalpha pretreatment had a basal mEPSC frequency of 1.35 +/- 0.20 Hz (n = 13), which was significantly higher when compared to a similar population of neurons in control slices (0.76 +/- 0.08 Hz; n = 53; P < 0.01). In control slices application of a low concentration of OLDA (0.2 uM) did not evoke any change in mEPSC frequency. After incubation with TNFalpha, OLDA (0.2 uM) application to slices induced a significant increase in mEPSC frequency (155.5 +/- 17.5%; P < 0.001; n = 10). Our results indicate that TNFalpha may have a significant impact on nociceptive signaling at the spinal cord level that could be mediated by increased responsiveness of presynaptic TRPV1 receptors to endogenous agonists. This could be of major importance, especially during pathological conditions, when increased levels of TNFalpha and TNFR are present in the spinal cord.

• MeSH
• Analysis of Variance MeSH
• Posterior Horn Cells drug effects   metabolism   MeSH
• Dopamine analogs & derivatives   pharmacology   MeSH
• Excitatory Postsynaptic Potentials drug effects   physiology   MeSH
• TRPV Cation Channels metabolism   MeSH
• Rats MeSH
• Patch-Clamp Techniques MeSH
• Spinal Cord drug effects   metabolism   MeSH
• Miniature Postsynaptic Potentials drug effects   physiology   MeSH
• Synaptic Transmission drug effects   physiology   MeSH
• Rats, Wistar MeSH
• Tumor Necrosis Factor-alpha pharmacology   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Dopamine MeSH
• TRPV Cation Channels MeSH
• N-oleoyldopamine MeSH Browser
• Tumor Necrosis Factor-alpha MeSH
• TRPV1 receptor MeSH Browser