ADX71943 and ADX71441, novel positive allosteric modulators of the GABAB receptor with distinct central/peripheral profiles, show efficacy in the monosodium iodoacetate model of chronic osteoarthritis pain in the rat.

We tested novel positive allosteric modulators (PAMs) of the γ-aminobutyric acid receptor B (GABAB), ADX71943 and ADX71441in the monosodium iodoacetate model of chronic osteoarthritis pain in rats with the objective to delineate the role of peripheral versus central GABAB receptor populations in modulation of chronic pain. Anesthetized Sprague-Dawley rats received an injection of monosodium iodoacetate into the knee and were tested for hyperalgesia starting post-MIA day 14. Effects of compounds on ipsilateral joint compression threshold were evaluated on post-MIA day 14 (after acute treatment), as well as after repeated, daily treatment on days 21 and 28 (ADX71943 only) and were compared to those of celecoxib (30mg/kg, p.o.). The PAMs were also tested in the rat rotarod test for potential muscle-relaxant effects. Acutely, ADX71943 (1-30mg/kg, p.o.), the peripherally restricted PAM, resulted in similar increases in pain threshold across the doses on day 14, while showing reduced efficacy on day 21 and no efficacy on day 28. A clear reduction in the efficacy of celecoxib across testing was also noted in this experiment. Acutely ADX71441 (0.3-15mg/kg, p.o.), the central-peripheral PAM, resulted in over 2-fold increases in pain threshold at 15mg/kg (but not at lower doses) on day 14, while causing more modest effects on day 21. Celecoxib increased pain threshold after both acute and daily treatment, showing overall similar efficacy. Thus, early, presumably more inflammatory phase of osteoarthritis pain in more sensitive to GABAB PAMs with peripherally restricted profile, while later, presumably more neuropathic phase is more sensitive to PAMs with central-peripheral profile.

Variable threshold of trigeminal cold-thermosensitive neurons is determined by a balance between TRPM8 and Kv1 potassium channels.

Molecular determinants of threshold differences among cold thermoreceptors are unknown. Here we show that such differences correlate with the relative expression of I(KD), a current dependent on Shaker-like Kv1 channels that acts as an excitability brake, and I(TRPM8), a cold-activated excitatory current. Neurons responding to small temperature changes have high functional expression of TRPM8 (transient receptor potential cation channel, subfamily M, member 8) and low expression of I(KD). In contrast, neurons activated by lower temperatures have a lower expression of TRPM8 and a prominent I(KD). Otherwise, both subpopulations have nearly identical membrane and firing properties, suggesting that they belong to the same neuronal pool. Blockade of I(KD) shifts the threshold of cold-sensitive neurons to higher temperatures and augments cold-evoked nocifensive responses in mice. Similar behavioral effects of I(KD) blockade were observed in TRPA1(-/-) mice. Moreover, only a small percentage of trigeminal cold-sensitive neurons were activated by TRPA1 agonists, suggesting that TRPA1 does not play a major role in the detection of low temperatures by uninjured somatic cold-specific thermosensory neurons under physiological conditions. Collectively, these findings suggest that innocuous cooling sensations and cold discomfort are signaled by specific low- and high-threshold cold thermoreceptor neurons, differing primarily in their relative expression of two ion channels having antagonistic effects on neuronal excitability. Thus, although TRPM8 appears to function as a critical cold sensor in the majority of peripheral sensory neurons, the expression of Kv1 channels in the same terminals seem to play an important role in the peripheral gating of cold-evoked discomfort and pain.

Transient receptor potential channels in sensory neurons are targets of the antimycotic agent clotrimazole.

Clotrimazole (CLT) is a widely used drug for the topical treatment of yeast infections of skin, vagina, and mouth. Common side effects of topical CLT application include irritation and burning pain of the skin and mucous membranes. Here, we provide evidence that transient receptor potential (TRP) channels in primary sensory neurons underlie these unwanted effects of CLT. We found that clinically relevant CLT concentrations activate heterologously expressed TRPV1 and TRPA1, two TRP channels that act as receptors of irritant chemical and/or thermal stimuli in nociceptive neurons. In line herewith, CLT stimulated a subset of capsaicin-sensitive and mustard oil-sensitive trigeminal neurons, and evoked nocifensive behavior and thermal hypersensitivity with intraplantar injection in mice. Notably, CLT-induced pain behavior was suppressed by the TRPV1-antagonist BCTC [(N-(-4-tertiarybutylphenyl)-4-(3-cholorpyridin-2-yl)tetrahydropyrazine-1(2H)-carboxamide)] and absent in TRPV1-deficient mice. In addition, CLT inhibited the cold and menthol receptor TRPM8, and blocked menthol-induced responses in capsaicin- and mustard oil-insensitive trigeminal neurons. The concentration for 50% inhibition (IC50) of inward TRPM8 current was approximately 200 nM, making CLT the most potent known TRPM8 antagonist and a useful tool to discriminate between TRPM8- and TRPA1-mediated responses. Together, our results identify TRP channels in sensory neurons as molecular targets of CLT, and offer means to develop novel CLT preparations with fewer unwanted sensory side effects.

Contribution of TRPM8 channels to cold transduction in primary sensory neurons and peripheral nerve terminals.

Transient receptor potential melastatin 8 (TRPM8) is the best molecular candidate for innocuous cold detection by peripheral thermoreceptor terminals. To dissect out the contribution of this cold- and menthol-gated, nonselective cation channel to cold transduction, we identified BCTC [N-(4-tert-butylphenyl)-4-(3-chloropyridin-2-yl)piperazine-1-carboxamide] as a potent and full blocker of recombinant TRPM8 channels. In cold-sensitive trigeminal ganglion neurons of mice and guinea pig, responses to menthol were abolished by BCTC. In contrast, the effect of BCTC on cold-evoked responses was variable but showed a good correlation with the presence or lack of menthol sensitivity in the same neuron, suggesting a specific blocking action of BCTC on TRPM8 channels. The biophysical properties of native cold-gated currents (I(cold)), and the currents blocked by BCTC were nearly identical, consistent with a role of this channel in cold sensing at the soma. The temperature activation threshold of native TRPM8 channels was significantly warmer than those reported in previous expression studies. The effect of BCTC on native I(cold) was characterized by a dose-dependent shift in the temperature threshold of activation. The role of TRPM8 in transduction was further investigated in the guinea pig cornea, a peripheral territory densely innervated with cold thermoreceptors. All cold-sensitive terminals were activated by menthol, suggesting the functional expression of TRPM8 channels in their membrane. However, the spontaneous activity and firing pattern characteristic of cold thermoreceptors was totally immune to TRPM8 channel blockade with BCTC or SKF96365 (1-[2-(4-methoxyphenyl)-2-[3-(4-methoxyphenyl)propoxy]ethyl-1H-imidazole hydrochloride). Cold-evoked responses in corneal terminals were also essentially unaffected by these drugs, whereas responses to menthol were completely abolished. The minor impairment in the ability to transduce cold stimuli by peripheral corneal thermoreceptors during TRPM8 blockade unveils an overlapping functional role for various thermosensitive mechanisms in these nerve terminals.

Evidence of a role for descending serotonergic facilitation in a rat model of cancer-induced bone pain.

Descending modulation of spinal processing plays an important role in chronic pain states. Monoamine pathways comprise a major component of descending controls from the brainstem to the spinal cord. Recent emphasis has been on facilitatory actions mediated by the 5-HT3 receptor. We investigated the effects of spinally administered ondansetron, a selective 5-HT3 receptor antagonist, on electrical- and natural-evoked dorsal horn (DH) neuronal responses in a rat model of cancer-induced bone pain (CIBP). Injection of MRMT-1 cells into the tibiae of Sprague-Dawley rats was used to model CIBP, whilst sham-operated rats were injected with the cell medium alone. Behavioural testing at regular intervals monitored the development of mechanical allodynia, cold allodynia, and ambulatory-evoked pain. In vivo electrophysiology experiments were carried out 15-17 days after surgery, when there were significant behavioural and neuronal alterations in the cancer animals. Spinally administered ondansetron (10, 50, and 100 microg) had no effect on electrical-evoked neuronal responses, but significantly reduced mechanical- and thermal-evoked responses in both the groups of animals. Furthermore, the effects of ondansetron were significantly greater in cancer animals compared to shams. These results therefore suggest a role for descending serotonergic facilitation in CIBP.

Efficacy of chronic morphine in a rat model of cancer-induced bone pain: behavior and in dorsal horn pathophysiology.

UNLABELLED: Morphine is one of the main analgesics in cancer-induced bone pain (CIBP). To investigate the efficacy of morphine in CIBP and alteration in dorsal horn pathophysiology, systemic morphine was administered (3 mg/kg) bi-daily between days 11 and 15 after MRMT-1 carcinoma cell injections (compared with a single injection (3 mg/kg) of morphine on day 15, and acute spinal morphine (0.1, 1, 10 microg/50 microL). The chronic systemic morphine schedule significantly attenuated pain behavior (von Frey 15 g; P < .01) to a greater extent than acute systemic morphine (von Frey 15 g; P < .05). In vivo electrophysiology (day 15 chronic systemic morphine) showed an attenuation of hyperexcitable wide dynamic range (WDR) neurons, but the abnormal raised WDR to nociceptive specific neuronal ratio remained. Acute spinal morphine attenuated electrical and natural WDR neuronal response in shams at a lower dose (1 microg) compared with cancer (10 microg). Chronic morphine is more effective at attenuating pain-related behaviors than single doses, although the dorsal horn retains a pathophysiologic characterization. PERSPECTIVE: This study confirms the resemblance of the rat model to human CIBP with respect to the efficacy of morphine and further suggests that adjuvant therapy is required to reverse the dorsal horn pathophysiology.

Gabapentin normalizes spinal neuronal responses that correlate with behavior in a rat model of cancer-induced bone pain.

BACKGROUND: Cancer-induced bone pain is a major clinical problem for which current treatments lack full efficacy. Gabapentin is licensed for use in neuropathic pain yet is also effective against inflammatory stimuli in animals. METHODS: A rat model of cancer-induced bone pain using the MRMT-1 cell line injected into the tibia was established to investigate the efficacy of acute (10, 30, 100 mg/kg) and chronic (30 mg/kg) systemic gabapentin on electrophysiological superficial dorsal horn neuronal responses to natural and noxious electrical stimuli, as well as on pain-related behavior. RESULTS: In electrophysiological studies gabapentin worked both acutely (100 mg/kg) and chronically (30 mg/kg) to normalize the hyperexcitable superficial dorsal horn neuronal response, significantly reducing electrical-evoked and mechanical-evoked but not thermal-evoked responses. The behavioral study showed that chronic gabapentin (30 mg/kg) significantly attenuated pain behavior in MRMT-1 rats, restoring responses to preoperative baseline degrees, and that this attenuation was accompanied by a reversion to normal (non-MRMT-1) wide-dynamic-range: nociceptive specific superficial dorsal horn neuronal profiles. CONCLUSIONS: Pain-related behavior in this rat model of cancer-induced bone pain is strongly linked to hyperexcitability of a population of superficial dorsal horn neurones. Gabapentin normalizes the cancer-induced bone pain induced dorsal horn neuronal changes and attenuates pain behavior. It may therefore provide a novel clinical treatment for cancer-induced bone pain.