Opposing Effects on Descending Control of Nociception by µ and κ Opioid Receptors in the Anterior Cingulate Cortex.

BACKGROUND: The efficiency of descending pain modulation, commonly assessed with the conditioned pain modulation procedure, is diminished in patients with chronic pain. The authors hypothesized that the efficiency of pain modulation is controlled by cortical opioid circuits. METHODS: This study evaluated the effects of µ opioid receptor activation in the anterior cingulate cortex on descending control of nociception, a preclinical correlate of conditioned pain modulation, in male Sprague-Dawley rats with spinal nerve ligation-induced chronic pain or in sham-operated controls. Additionally, the study explored the consequences of respective activation or inhibition of κ opioid receptor in the anterior cingulate cortex of naive rats or animals with neuropathic pain. Descending control of nociception was measured as the hind paw withdrawal response to noxious pressure (test stimulus) in the absence or presence of capsaicin injection in the forepaw (conditioning stimulus). RESULTS: Descending control of nociception was diminished in the ipsilateral, but not contralateral, hind paw of rats with spinal nerve ligation. Bilateral administration of morphine in the anterior cingulate cortex had no effect in shams but restored diminished descending control of nociception without altering hypersensitivity in rats with neuropathic pain. Bilateral anterior cingulate cortex microinjection of κ opioid receptor antagonists, including nor-binaltorphimine and navacaprant, also re-established descending control of nociception in rats with neuropathic pain without altering hypersensitivity and with no effect in shams. Conversely, bilateral injection of a κ opioid receptor agonist, U69,593, in the anterior cingulate cortex of naive rats inhibited descending control of nociception without altering withdrawal thresholds. CONCLUSIONS: Anterior cingulate cortex κ opioid receptor activation therefore diminishes descending control of nociception both in naive animals and as an adaptive response to chronic pain, likely by enhancing net descending facilitation. Descending control of nociception can be restored by activation of µ opioid receptors in the anterior cingulate cortex, but also by κ opioid receptor antagonists, providing a nonaddictive alternative to opioid analgesics. Navacaprant is now in advanced clinical trials.

Endogenous opioid activity in the anterior cingulate cortex is required for relief of pain.

Pain is aversive, and its relief elicits reward mediated by dopaminergic signaling in the nucleus accumbens (NAc), a part of the mesolimbic reward motivation pathway. How the reward pathway is engaged by pain-relieving treatments is not known. Endogenous opioid signaling in the anterior cingulate cortex (ACC), an area encoding pain aversiveness, contributes to pain modulation. We examined whether endogenous ACC opioid neurotransmission is required for relief of pain and subsequent downstream activation of NAc dopamine signaling. Conditioned place preference (CPP) and in vivo microdialysis were used to assess negative reinforcement and NAc dopaminergic transmission. In rats with postsurgical or neuropathic pain, blockade of opioid signaling in the rostral ACC (rACC) inhibited CPP and NAc dopamine release resulting from non-opioid pain-relieving treatments, including peripheral nerve block or spinal clonidine, an α2-adrenergic agonist. Conversely, pharmacological activation of rACC opioid receptors of injured, but not pain-free, animals was sufficient to stimulate dopamine release in the NAc and produce CPP. In neuropathic, but not sham-operated, rats, systemic doses of morphine that did not affect withdrawal thresholds elicited CPP and NAc dopamine release, effects that were prevented by blockade of ACC opioid receptors. The data provide a neural explanation for the preferential effects of opioids on pain affect and demonstrate that engagement of NAc dopaminergic transmission by non-opioid pain-relieving treatments depends on upstream ACC opioid circuits. Endogenous opioid signaling in the ACC appears to be both necessary and sufficient for relief of pain aversiveness.

Pain relief produces negative reinforcement through activation of mesolimbic reward-valuation circuitry.

Relief of pain is rewarding. Using a model of experimental postsurgical pain we show that blockade of afferent input from the injury with local anesthetic elicits conditioned place preference, activates ventral tegmental dopaminergic cells, and increases dopamine release in the nucleus accumbens. Importantly, place preference is associated with increased activity in midbrain dopaminergic neurons and blocked by dopamine antagonists injected into the nucleus accumbens. The data directly support the hypothesis that relief of pain produces negative reinforcement through activation of the mesolimbic reward-valuation circuitry.

Decreased dopaminergic inhibition of pyramidal neurons in anterior cingulate cortex maintains chronic neuropathic pain.

Pyramidal neurons in the anterior cingulate cortex (ACC), a prefrontal region involved in processing the affective components of pain, display hyperexcitability in chronic neuropathic pain conditions, and their silencing abolishes hyperalgesia. We show that dopamine, through D1 receptor (D1R) signaling, inhibits pyramidal neurons of mouse ACC by modulation of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels. Activation of Gs-coupled D1R by dopamine induces the opening of HCN channels at physiological membrane potentials, driving a significant decrease in input resistance and excitability. Systemic L-DOPA in chronic neuropathic mice rescues HCN channel activity, normalizes pyramidal excitability in ACC, and blocks mechanical and thermal allodynia. Moreover, microinjection of a selective D1R agonist in the ACC relieves the aversiveness of ongoing neuropathic pain, while an ACC D1R antagonist blocks gabapentin- and lidocaine-evoked antinociception. We conclude that dopaminergic inhibition via D1R in ACC plays an analgesic role in physiological conditions and is decreased in chronic pain.

Synaptic connections between GABAergic elements and serotonergic terminals or projecting neurons in the ventrolateral orbital cortex.

The ventrolateral orbital cortex (VLO) is part of an endogenous analgesic system, consisting of the spinal cord-thalamic nucleus submedius-VLO periaqueductal gray (PAG)-spinal cord loop. The present study examined morphological connections of GABAergic (gamma-aminobutyric acidergic) neurons and serotonergic projection terminals from the dorsal raphe nucleus (DR), as well as the relationship between GABAergic terminals and VLO neurons projecting to the PAG, by using anterograde and retrograde tracing combined with immunofluorescence, immunohistochemistry, and electron microscopy methods. Results indicate that the majority (93%) of GABAergic neurons in the VLO also express the 5-HT(1A) (5-hydroxytryptamine 1A) receptor, and serotonergic terminals originating from the DR nucleus made symmetrical synapses with GABAergic neuronal cell bodies and dendrites within the VLO. GABAergic terminals also made symmetrical synapses with neurons expressing GABA(A) receptors and projecting to the PAG. These results suggest that a local neuronal circuit, consisting of 5-HTergic terminals, GABAergic interneurons, and projection neurons, exists in the VLO, and provides morphological evidence for the hypothesis that GABAergic modulation is involved in 5-HT(1A) receptor activation-evoked antinociception.

Antihyperalgesic effects of Meteorin in the rat chronic constriction injury model: a replication study.

Data from preclinical research have been suggested to suffer from a lack of inherent reproducibility across laboratories. The goal of our study was to replicate findings from a previous report that demonstrated positive effects of Meteorin, a novel neurotrophic factor, in a rat model of neuropathic pain induced by chronic constriction injury (CCI). Notably, 5 to 6 intermittent subcutaneous (s.c.) injections of Meteorin had been reported to produce reversal of mechanical allodynia/thermal hyperalgesia after injury, wherein maximum efficacy of Meteorin was reached slowly and outlasted the elimination of the compound from the blood by several weeks. Here, we evaluated the efficacy of Meteorin in reversing hindpaw mechanical hyperalgesia and cold allodynia in male, Sprague-Dawley rats with CCI. Nociceptive behavior was monitored before and after CCI, and after drug treatment until day 42 after injury. Systemic administration of recombinant mouse Meteorin (0.5 and 1.8 mg/kg, s.c.) at days 10, 12, 14, 17, and 19 after CCI produced a prolonged reversal of neuropathic hypersensitivity with efficacy comparable with that obtained with gabapentin (100 mg/kg, orally). Despite some protocol deviations (eg, nociceptive endpoint, animal vendor, testing laboratory, investigator, etc.) being incurred, these did not affect study outcome. By paying careful attention to key facets of study design, using bioactive material, and confirming drug exposure, the current data have replicated the salient findings of the previous study, promoting confidence in further advancement of this novel molecule as a potential therapy for neuropathic pain.

Positive allosteric modulators of nonbenzodiazepine γ-aminobutyric acidA receptor subtypes for the treatment of chronic pain.

Chronic neuropathic pain may be caused, in part, by loss of inhibition in spinal pain processing pathways due to attenuation of local GABAergic tone. Nociception and nocifensive behaviors are reduced after enhancement of tonically activated extrasynaptic GABAAR-mediated currents by agonist ligands for δ subunit-containing GABAARs. However, typical ligands that target δ subunit-containing GABAARs are limited due to sedative effects at higher doses. We used the spinal nerve ligation (SNL) and gp120 models of experimental neuropathic pain to evaluate compound 2-261, a nonbenzodiazepine site positive allosteric modulator of α4ß3δ GABAARs optimized to be nonsedative by selective activation of ß2/3-subunit-containing GABAARs over receptor subtypes incorporating ß1 subunits. Similar levels of 2-261 were detected in the brain and plasma after intraperitoneal administration. Although systemic 2-261 did not alter sensory thresholds in sham-operated animals, it significantly reversed SNL-induced thermal and tactile hypersensitivity in a GABAAR-dependent fashion. Intrathecal 2-261 produced conditioned place preference and elevated dopamine levels in the nucleus accumbens of nerve-injured, but not sham-operated, rats. In addition, systemic pretreatment with 2-261 blocked conditioned place preference from spinal clonidine in SNL rats. Moreover, 2-261 reversed thermal hyperalgesia and partially reversed tactile allodynia in the gp120 model of HIV-related neuropathic pain. The effects of 2-261 likely required interaction with the α4ß3δ GABAAR because 2-301, a close structural analog of 2-261 with limited extrasynaptic receptor efficacy, was not active. Thus, 2-261 may produce pain relief with diminished side effects through selective modulation of ß2/3-subunit-containing extrasynaptic GABAARs.

Kappa opioid signaling in the central nucleus of the amygdala promotes disinhibition and aversiveness of chronic neuropathic pain.

Chronic pain is associated with neuroplastic changes in the amygdala that may promote hyper-responsiveness to mechanical and thermal stimuli (allodynia and hyperalgesia) and/or enhance emotional and affective consequences of pain. Stress promotes dynorphin-mediated signaling at the kappa opioid receptor (KOR) in the amygdala and mechanical hypersensitivity in rodent models of functional pain. Here, we tested the hypothesis that KOR circuits in the central nucleus of the amygdala (CeA) undergo neuroplasticity in chronic neuropathic pain resulting in increased sensory and affective pain responses. After spinal nerve ligation (SNL) injury in rats, pretreatment with a long-acting KOR antagonist, nor-binaltorphimine (nor-BNI), subcutaneously or through microinjection into the right CeA, prevented conditioned place preference (CPP) to intravenous gabapentin, suggesting that nor-BNI eliminated the aversiveness of ongoing pain. By contrast, systemic or intra-CeA administration of nor-BNI had no effect on tactile allodynia in SNL animals. Using whole-cell patch-clamp electrophysiology, we found that nor-BNI decreased synaptically evoked spiking of CeA neurons in brain slices from SNL but not sham rats. This effect was mediated through increased inhibitory postsynaptic currents, suggesting tonic disinhibition of CeA output neurons due to increased KOR activity as a possible mechanism promoting ongoing aversive aspects of neuropathic pain. Interestingly, this mechanism is not involved in SNL-induced mechanical allodynia. Kappa opioid receptor antagonists may therefore represent novel therapies for neuropathic pain by targeting aversive aspects of ongoing pain while preserving protective functions of acute pain.

[Roles of ventrolateral orbital cortex in pain modulation and acupuncture analgesia].

The ventrolateral orbital cortex (VLO) is a major component of orbital cortex, which has extensive connections with periaqueductal gray (PAG), thalamus and other cortical regions. Researches suggest that the VLO is involved not only in nociception, but also in pain modulation, through activation of PAG brainstem descending antinociceptive pathway to inhibit the nociceptive inputs at the spinal/trigeminal level. Furthermore, many results demonstrate that opioid, 5-HT, GABA and their receptors are involved in the VLO antinociception. VLO plays an important role in acupuncture analgesia. In this review we summarized the roles of ventrolateral orbital cortex in pain modulation and acupuncture antinociception.

Selective deficiencies in descending inhibitory modulation in neuropathic rats: implications for enhancing noradrenergic tone.

Pontine noradrenergic neurones form part of a descending inhibitory system that influences spinal nociceptive processing. Weak or absent descending inhibition is a common feature of chronic pain patients. We examined the extent to which the descending noradrenergic system is tonically active, how control of spinal neuronal excitability is integrated into thalamic relays within sensory-discriminative projection pathways, and how this inhibitory control is altered after nerve injury. In vivo electrophysiology was performed in anaesthetised spinal nerve-ligated (SNL) and sham-operated rats to record from wide dynamic range neurones in the ventral posterolateral thalamus (VPL). In sham rats, spinal block of α2-adrenoceptors with atipamezole resulted in enhanced stimulus-evoked and spontaneous firing in the VPL, and produced conditioned place avoidance. However, in SNL rats, these conditioned avoidance behaviours were absent. Furthermore, inhibitory control of evoked neuronal responses was lost, but spinal atipamezole markedly increased spontaneous firing. Augmenting spinal noradrenergic tone in neuropathic rats with reboxetine, a selective noradrenergic reuptake inhibitor, modestly reinstated inhibitory control of evoked responses in the VPL but had no effect on spontaneous firing. By contrast, clonidine, an α2 agonist, inhibited both evoked and spontaneous firing, and exhibited increased potency in SNL rats compared with sham controls. These data suggest descending noradrenergic inhibitory pathways are tonically active in sham rats. Moreover, in neuropathic states, descending inhibitory control is diminished, but not completely absent, and distinguishes between spontaneous and evoked neuronal activity. These observations may have implications for how analgesics targeting the noradrenergic system provide relief.

Multiple sites and actions of gabapentin-induced relief of ongoing experimental neuropathic pain.

Gabapentin (GBP) is a first-line therapy for neuropathic pain, but its mechanisms and sites of action remain uncertain. We investigated GBP-induced modulation of neuropathic pain following spinal nerve ligation (SNL) in rats. Intravenous or intrathecal GBP reversed evoked mechanical hypersensitivity and produced conditioned place preference (CPP) and dopamine (DA) release in the nucleus accumbens (NAc) selectively in SNL rats. Spinal GBP also significantly inhibited dorsal horn wide-dynamic-range neuronal responses to a range of evoked stimuli in SNL rats. By contrast, GBP microinjected bilaterally into the rostral anterior cingulate cortex (rACC), produced CPP, and elicited NAc DA release selectively in SNL rats but did not reverse tactile allodynia and had marginal effects on wide-dynamic-range neuronal activity. Moreover, blockade of endogenous opioid signaling in the rACC prevented intravenous GBP-induced CPP and NAc DA release but failed to block its inhibition of tactile allodynia. Gabapentin, therefore, can potentially act to produce its pain relieving effects by (a) inhibition of injury-induced spinal neuronal excitability, evoked hypersensitivity, and ongoing pain and (b) selective supraspinal modulation of affective qualities of pain, without alteration of reflexive behaviors. Consistent with previous findings of pain relief from nonopioid analgesics, GBP requires engagement of rACC endogenous opioid circuits and downstream activation of mesolimbic reward circuits reflected in learned pain-motivated behaviors. These findings support the partial separation of sensory and affective dimensions of pain in this experimental model and suggest that modulation of affective-motivational qualities of pain may be the preferential mechanism of GBP's analgesic effects in patients.

CRISPR/Cas9 editing of Nf1 gene identifies CRMP2 as a therapeutic target in neurofibromatosis type 1-related pain that is reversed by (S)-Lacosamide.

Neurofibromatosis type 1 (NF1) is a rare autosomal dominant disease linked to mutations of the Nf1 gene. Patients with NF1 commonly experience severe pain. Studies on mice with Nf1 haploinsufficiency have been instructive in identifying sensitization of ion channels as a possible cause underlying the heightened pain suffered by patients with NF1. However, behavioral assessments of Nf1 mice have led to uncertain conclusions about the potential causal role of Nf1 in pain. We used the clustered regularly interspaced short palindromic repeats (CRISPR)-associated 9 (CRISPR/Cas9) genome editing system to create and mechanistically characterize a novel rat model of NF1-related pain. Targeted intrathecal delivery of guide RNA/Cas9 nuclease plasmid in combination with a cationic polymer was used to generate allele-specific C-terminal truncation of neurofibromin, the protein encoded by the Nf1 gene. Rats with truncation of neurofibromin, showed increases in voltage-gated calcium (specifically N-type or CaV2.2) and voltage-gated sodium (particularly tetrodotoxin-sensitive) currents in dorsal root ganglion neurons. These gains-of-function resulted in increased nociceptor excitability and behavioral hyperalgesia. The cytosolic regulatory protein collapsin response mediator protein 2 (CRMP2) regulates activity of these channels, and also binds to the targeted C-terminus of neurofibromin in a tripartite complex, suggesting a possible mechanism underlying NF1 pain. Prevention of CRMP2 phosphorylation with (S)-lacosamide resulted in normalization of channel current densities, excitability, as well as of hyperalgesia following CRISPR/Cas9 truncation of neurofibromin. These studies reveal the protein partners that drive NF1 pain and suggest that CRMP2 is a key target for therapeutic intervention.

Involvement of GABAergic modulation of antinociception induced by morphine microinjected into the ventrolateral orbital cortex.

Previous studies have shown that microinjection of morphine into the prefrontal ventrolateral orbital cortex (VLO) produces antinociception. The current study examined whether gamma-aminobutyric acid (GABA) containing neurons in the VLO were involved in this antinociception. Under light anesthesia, the GABA(A) receptor antagonist bicuculline and picrotoxin or agonist muscimol and THIP was microinjected into the VLO in non-morphine-treated (control) and morphine-treated (microinjection into the VLO) rats. Noxious heat-evoked tail flick (TF) latencies (TFLs) were measured in all of these groups of rats every 5 min. Bicuculline or picrotoxin (100, 200, 500 ng in 0.5 microl) depressed the TF reflex in a dose-related fashion. A smaller dose (100 ng) of bicuculline or picrotoxin microinjected into VLO significantly enhanced the VLO morphine-evoked inhibition of the TF reflex. In contrast, administration of muscimol (250 ng) or THIP (1.0 microg) significantly attenuated the morphine-induced antinociception in the VLO morphine-treated rats. These results suggest that the GABA(A) receptor is involved in the modulation of VLO morphine-induced antinociception, and provide a behavioral support for the hypothesis that morphine may directly inhibit the GABAergic inhibitory interneurons leading to indirect activation of the descending antinociceptive pathway through a disinhibitory effect on the VLO output neurons and depression of the nociceptive inputs at the spinal cord level.

Discovery of Stable Non-opioid Dynorphin A Analogues Interacting at the Bradykinin Receptors for the Treatment of Neuropathic Pain.

Dynorphin A (Dyn A) is a unique endogenous ligand that possesses well-known neuroinhibitory effects via opioid receptors with a preference for the kappa receptor but also neuroexcitatory effects, which cause hyperalgesia. We have shown that the neuroexcitatory effects are mediated through bradykinin (BK) receptors and that intrathecal (i.th.) administration of our lead ligand 1, [des-Arg7]-Dyn A-(4-11), which shows good binding affinity (IC50 = 150 nM) at the BK receptors, blocks Dyn A-induced hyperalgesia in naïve animals and reverses thermal and tactile hypersensitivities in a dose-dependent manner in nerve-injured animals. However, 1 has a serious drawback as a potential drug candidate for the treatment of neuropathic pain because of its susceptibility to enzymatic degradation. In an effort to increase its stability, we modified ligand 1 using non-natural amino acids and found that analogues substituted at or near the N-terminus with a d-isomer retain binding at the receptor and provide a large increase in stability. In particular when Leu5 was modified, with either the d-isomer or N-methylation, there was a large increase in stability (t1/2 = 0.7-160 h in rat plasma) observed. From these studies, we have developed a very stable Dyn A analogue 16, [d-Leu5,des-Arg7]-Dyn A-(4-11), that binds to BK receptors (IC50 = 130 nM) in the same range as ligand 1 and shows good antihyperalgesic effects in both naïve rats and L5/L6 spinal nerve ligation rats.

Activation of α1 adrenoceptors in ventrolateral orbital cortex attenuates allodynia induced by spared nerve injury in rats.

Recent studies have demonstrated that noradrenaline acting in the ventrolateral orbital cortex (VLO) can potentially reduce allodynia induced by spared nerve injury (SNI), and this effect is mediated by α2 adrenoceptor. The present study examined the effect of the α1 adrenoceptors in the VLO on allodynia induced by SNI in the rats. The mechanical paw withdrawal threshold (PWT) was measured using von-Frey filaments. Microinjection of selective α1 adrenoceptor agonist methoxamine (20, 50, 100 µg in 0.5 µl) into the VLO, contralateral to the site of nerve injury, increased PWT in a dose-dependent manner. This effect was antagonized by pre-microinjection of the selective α1 adrenoceptor antagonist benoxathian into the same VLO site, and blocked by electrolytic lesion of the ventrolateral periaqueductal gray (PAG). Furthermore, pre-administration of non-selective glutamate receptor antagonist kynurenic acid, phospholipase C (PLC) inhibitor U73122, and protein kinase C (PKC) inhibitor chelerythrine to the VLO also blocked methoxamine-induced inhibition of allodynia. These results suggest that activation of α1 adrenoceptors in the VLO can potentially reduce allodynia induced by SNI. This effect may be direct excitation of the VLO neurons, via PLC-PKC signaling pathway, projecting to the PAG or facilitating glutamate release and then indirectly exciting the VLO output neurons projecting to the PAG, leading to activation of the PAG-brainstem descending inhibitory system which depresses the nociceptive transmission at the spinal cord level.

(S)-lacosamide inhibition of CRMP2 phosphorylation reduces postoperative and neuropathic pain behaviors through distinct classes of sensory neurons identified by constellation pharmacology.

Chronic pain affects the life of millions of people. Current treatments have deleterious side effects. We have advanced a strategy for targeting protein interactions which regulate the N-type voltage-gated calcium (CaV2.2) channel as an alternative to direct channel block. Peptides uncoupling CaV2.2 interactions with the axonal collapsin response mediator protein 2 (CRMP2) were antinociceptive without effects on memory, depression, and reward/addiction. A search for small molecules that could recapitulate uncoupling of the CaV2.2-CRMP2 interaction identified (S)-lacosamide [(S)-LCM], the inactive enantiomer of the Food and Drug Administration-approved antiepileptic drug (R)-lacosamide [(R)-LCM, Vimpat]. We show that (S)-LCM, but not (R)-LCM, inhibits CRMP2 phosphorylation by cyclin dependent kinase 5, a step necessary for driving CaV2.2 activity, in sensory neurons. (S)-lacosamide inhibited depolarization-induced Ca influx with a low micromolar IC50. Voltage-clamp electrophysiology experiments demonstrated a commensurate reduction in Ca currents in sensory neurons after an acute application of (S)-LCM. Using constellation pharmacology, a recently described high content phenotypic screening platform for functional fingerprinting of neurons that uses subtype-selective pharmacological agents to elucidate cell-specific combinations (constellations) of key signaling proteins that define specific cell types, we investigated if (S)-LCM preferentially acts on certain types of neurons. (S)-lacosamide decreased the dorsal root ganglion neurons responding to mustard oil, and increased the number of cells responding to menthol. Finally, (S)-LCM reversed thermal hypersensitivity and mechanical allodynia in a model of postoperative pain, and 2 models of neuropathic pain. Thus, using (S)-LCM to inhibit CRMP2 phosphorylation is a novel and efficient strategy to treat pain, which works by targeting specific sensory neuron populations.

Sustained relief of ongoing experimental neuropathic pain by a CRMP2 peptide aptamer with low abuse potential.

Uncoupling the protein-protein interaction between collapsin response mediator protein 2 (CRMP2) and N-type voltage-gated calcium channel (CaV2.2) with an allosteric CRMP2-derived peptide (CBD3) is antinociceptive in rodent models of inflammatory and neuropathic pain. We investigated the efficacy, duration of action, abuse potential, and neurobehavioral toxicity of an improved mutant CRMP2 peptide. A homopolyarginine (R9)-conjugated CBD3-A6K (R9-CBD3-A6K) peptide inhibited the CaV2.2-CRMP2 interaction in a concentration-dependent fashion and diminished surface expression of CaV2.2 and depolarization-evoked Ca influx in rat dorsal root ganglia neurons. In vitro studies demonstrated suppression of excitability of small-to-medium diameter dorsal root ganglion and inhibition of subtypes of voltage-gated Ca channels. Sprague-Dawley rats with tibial nerve injury had profound and long-lasting tactile allodynia and ongoing pain. Immediate administration of R9-CBD3-A6K produced enhanced dopamine release from the nucleus accumbens shell selectively in injured animals, consistent with relief of ongoing pain. R9-CBD3-A6K, when administered repeatedly into the central nervous system ventricles of naive rats, did not result in a positive conditioned place preference demonstrating a lack of abusive liability. Continuous subcutaneous infusion of R9-CBD3-A6K over a 24- to 72-hour period reversed tactile allodynia and ongoing pain, demonstrating a lack of tolerance over this time course. Importantly, continuous infusion of R9-CBD3-A6K did not affect motor activity, anxiety, depression, or memory and learning. Collectively, these results validate the potential therapeutic significance of targeting the CaV-CRMP2 axis for treatment of neuropathic pain.

Administration of somatostatin analog octreotide in the ventrolateral orbital cortex produces sex-related antinociceptive effects on acute and formalin-induced nociceptive behavior in rats.

The present study was designed to examine whether somatostatin analog octreotide (OCT) was involved in antinociception in the ventrolateral orbital cortex (VLO) and determine whether this effect had a sex difference between male and female rats. The radiant heat-evoked tail flick (TF) reflex was used as an index of acute nociceptive response in lightly anesthetized rats. The number of flinches evoked by formalin injection into the hindpaw was used to evaluate inflammatory persistent pain in conscious rats. Administration of OCT (2.0, 5.0 10.0 ng in 0.5 µl) into the VLO depressed the TF reflex in a dose-dependent manner only in female rats, but not male rats. Pretreatment with a nonselective somatostatin receptor antagonist cyclo-somatostatin (c-SOM) (25.0 µg in 0.5 µl) into the VLO antagonized 10.0 ng OCT-induced inhibition of the TF reflex in female rats. Similarly, application of high dose of OCT (10.0 ng in 0.5 µl) into the VLO depressed formalin-induced flinching response in the early and late phases only in female rats, and had no any effects in male rats. Pretreatment with c-SOM (25.0 µg in 0.5 µl) into the VLO totally antagonized the 10 ng OCT-induced inhibition of the flinches in both phases in female rats. Additionally, single administration of c-SOM into the VLO failed to alter tail reflex latencies and formalin-induced nociceptive behaviors in female rats. The results provide the first valuable evidence that somatostatin and its receptors are involved in antinociception in acute heat-evoked nociception and inflammatory persistent pain only in female rats, not male rats, in the VLO.

Activation of mu-opioid receptors in the ventrolateral orbital cortex inhibits the GABAergic miniature inhibitory postsynaptic currents in rats.

Previous studies have indicated that mu-opioid receptors in the ventrolateral orbital cortex (VLO) are involved in antinociception in tail flick tests and GABAergic neurons or terminals express mu-opioid receptors in the VLO. The current study examined the effect of selective mu-opioid receptor agonist DAMGO on the GABAergic miniature inhibitory postsynaptic currents (mIPSCs) in the VLO in rats using the whole-cell patch clamp. The results demonstrated that 5 µM DAMGO application into the rat VLO slices significantly reduced the GABAergic mIPSCs frequency, without any effect on its amplitude, and this effect of DAMGO was reversed by pretreatment with selective mu-opioid receptor antagonist 1 µM CTOP. Importantly, application of CTOP alone into the VLO slices did not produce any effect on the frequency and amplitude of GABAergic mIPSCs. These results indicate a presynaptic effect of mu-opioid receptor activation on the GABAergic neurons in the VLO. The current data suggests that a presynaptic inhibition of the GABA release may contribute to the mu-opioid receptor mediated effects in the VLO and provides novel electrophysiological evidence for the underlying mechanisms of mu-opioid receptors in the VLO.

Activation of mesocorticolimbic reward circuits for assessment of relief of ongoing pain: a potential biomarker of efficacy.

Preclinical assessment of pain has increasingly explored operant methods that may allow behavioral assessment of ongoing pain. In animals with incisional injury, peripheral nerve block produces conditioned place preference (CPP) and activates the mesolimbic dopaminergic reward pathway. We hypothesized that activation of this circuit could serve as a neurochemical output measure of relief of ongoing pain. Medications commonly used clinically, including gabapentin and nonsteroidal anti-inflammatory drugs (NSAIDs), were evaluated in models of post-surgical (1 day after incision) or neuropathic (14 days after spinal nerve ligation [SNL]) pain to determine whether the clinical efficacy profile of these drugs in these pain conditions was reflected by extracellular dopamine (DA) release in the nucleus accumbens (NAc) shell. Microdialysis was performed in awake rats. Basal DA levels were not significantly different between experimental groups, and no significant treatment effects were seen in sham-operated animals. Consistent with clinical observation, spinal clonidine produced CPP and produced a dose-related increase in net NAc DA release in SNL rats. Gabapentin, commonly used to treat neuropathic pain, produced increased NAc DA in rats with SNL but not in animals with incisional, injury. In contrast, ketorolac or naproxen produced increased NAc DA in animals with incisional but not neuropathic pain. Increased extracellular NAc DA release was consistent with CPP and was observed selectively with treatments commonly used clinically for post-surgical or neuropathic pain. Evaluation of NAc DA efflux in animal pain models may represent an objective neurochemical assay that may serve as a biomarker of efficacy for novel pain-relieving mechanisms.