NMP-7 inhibits chronic inflammatory and neuropathic pain via block of Cav3.2 T-type calcium channels and activation of CB2 receptors.
TL;DR: This work shows that NMP-7 mediates a significant analgesic effect in a model of persistent inflammatory and chronic neuropathic pain by way of T-type channel modulation and CB2 receptor activation, and provides a novel therapeutic avenue for managing chronic pain conditions via mixed CB ligands/T-type channels.
Abstract: Background: T-type calcium channels and cannabinoid receptors are known to play important roles in chronic pain, making them attractive therapeutic targets. We recently reported on the design, synthesis and analgesic properties of a novel T-type channel inhibitor (NMP-7), which also shows mixed agonist activity on CB1 and CB2 receptors in vitro. Here, we analyzed the analgesic effect of systemically delivered NMP-7 (intraperitoneal (i.p.) or intragstric (i.g.) routes) on mechanical hypersensitivity in inflammatory pain induced by Complete Freund’s Adjuvant (CFA) and neuropathic pain induced by sciatic nerve injury. Results: NMP-7 delivered by either i.p. or i.g. routes produced dose-dependent inhibition of mechanical hyperalgesia in mouse models of inflammatory and neuropathic pain, without altering spontaneous locomotor activity in the open-field test at the highest active dose. Neither i.p. nor i.g. treatment reduced peripheral inflammation per se ,a s evaluated by examining paw edema and myeloperoxidase activity. The antinociception produced by NMP-7 in the CFA test was completely abolished in CaV3.2-null mice, confirming CaV 3.2 as ak ey target. The analgesic action of intraperitoneally delivered NMP-7 was not affected by pretreatment of mice with the CB1 antagonist AM281, but was significantly attenuated by pretreatment with the CB2 antagonist AM630, suggesting that CB2 receptors, but not CB1 receptors are involved in the action of NMP-7 in vivo. Conclusions: Overall, our work shows that NMP-7 mediates a significant analgesic effect in a model of persistent inflammatory and chronic neuropathic pain by way of T-type channel modulation and CB2 receptor activation. Thus, this study provides a novel therapeutic avenue for managing chronic pain conditions via mixed CB ligands/ T-type channel blockers.
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TL;DR: An overview of calcium channels as drug targets for nervous system disorders is provided, and potential challenges and opportunities for the development of new clinically effective calcium channel inhibitors are discussed.
Abstract: Voltage-gated calcium channels are important regulators of brain, heart and muscle functions, and their dysfunction can give rise to pathophysiological conditions ranging from cardiovascular disorders to neurological and psychiatric conditions such as epilepsy, pain and autism. In the nervous system, calcium channel blockers have been used successfully to treat absence seizures, and are emerging as potential therapeutic avenues for pathologies such as pain, Parkinson disease, addiction and anxiety. This Review provides an overview of calcium channels as drug targets for nervous system disorders, and discusses potential challenges and opportunities for the development of new clinically effective calcium channel inhibitors.
301 citations
TL;DR: It is suggested that drugs that affect multiple processes, rather than a single specific target, show the greatest promise for future therapeutic development.
Abstract: Injury to or disease of the nervous system can invoke chronic and sometimes intractable neuropathic pain. Many parallel, interdependent, and time-dependent processes, including neuroimmune interactions at the peripheral, supraspinal, and spinal levels, contribute to the etiology of this "disease of pain." Recent work emphasizes the roles of colony-stimulating factor 1, ATP, and brain-derived neurotrophic factor. Excitatory processes are enhanced, and inhibitory processes are attenuated in the spinal dorsal horn and throughout the somatosensory system. This leads to central sensitization and aberrant processing such that tactile and innocuous thermal information is perceived as pain (allodynia). Processes involved in the onset of neuropathic pain differ from those involved in its long-term maintenance. Opioids display limited effectiveness, and less than 35% of patients derive meaningful benefit from other therapeutic approaches. We thus review promising therapeutic targets that have emerged over the last 20 years, including Na+, K+, Ca2+, hyperpolarization-activated cyclic nucleotide-gated channels, transient receptor potential channel type V1 channels, and adenosine A3 receptors. Despite this progress, the gabapentinoids retain their status as first-line treatments, yet their mechanism of action is poorly understood. We outline recent progress in understanding the etiology of neuropathic pain and show how this has provided insights into the cellular actions of pregabalin and gabapentin. Interactions of gabapentinoids with the α2δ-1 subunit of voltage-gated Ca2+ channels produce multiple and neuron type-specific actions in spinal cord and higher centers. We suggest that drugs that affect multiple processes, rather than a single specific target, show the greatest promise for future therapeutic development.
240 citations
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...2 channels (Berger et al., 2014)....
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TL;DR: Recent developments in the discovery of novel classes of T‐type calcium channel blockers, and their analgesic effects in animal models of pain and in clinical trials are reviewed.
Abstract: Cav3.2 T-type calcium channels are important regulators of pain signals in afferent pain pathway, and their activities are dysregulated during various chronic pain states. Therefore it stands to reason that inhibiting T-type calcium channels in dorsal root ganglion neurons and in the spinal dorsal horn can be targeted for pain relief. This is supported by early pharmacological studies with T-type channel blockers such as ethosuximide, and by analgesic effects of siRNA depletion of Cav3.2 channels. In the past five years, considerable effort has been applied towards identifying novel classes of T-type calcium channel blockers. Here we review recent developments in the discovery of novel classes of T-type calcium channel blockers, and their analgesics effects in animal models of pain and in clinical trials.
86 citations
Cites background from "NMP-7 inhibits chronic inflammatory..."
...More recently, a series of novel DHP derivatives with a condensed hexahydroquinoline 1,4-DHP ring system were identified (Bladen et al., 2014)....
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TL;DR: It is concluded that activation of CB2R ameliorates DSS-induced colitis through enhancing autophagy that may inhibit NLRP3 inflammasome activation in macrophages and it is demonstrated that AMPK-mTOR-P70S6K signaling pathway was involved in thisCB2R-mediated process.
Abstract: Activation of cannabinoid receptor 2 (CB2R) ameliorates inflammation, but the underlying mechanism remains unclear. In the present study, we examined whether activation of CB2R could suppress the nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome. In peritoneal macrophages isolated from C57BL/6 mice, LPS/DSS challenge for 24 h increased the expression of the components of NLRP3 inflammasome NLRP3, Casp-1 p20/Casp-1 p45 ratio, proIL-1β and IL-1β and also enhanced autophagy (LC3-II/LC3-I ratio, Beclin-1 and SQSTM1). Pretreatment of peritoneal macrophages with HU 308, a selective CB2R agonist, attenuated LPS/DSS-induced NLRP3 inflammasome activation, but further enhanced autophagy. In comparison with wild-type (WT) control, peritoneal macrophages from CB2R knockout (KO) mice had more robust NLRP3 inflammasome activation and attenuated autophagy upon LPS/DSS challenge. Knockdown autophagy-related gene 5 (Atg5) with a siRNA in peritoneal macrophages attenuated the inhibitory effects of HU 308 on LPS/DSS-induced NLRP3 inflammasome activation in vitro. In vivo, HU308 treatment attenuated DSS-induced colitis mice associated with reduced colon inflammation and inhibited NLRP3 inflammasome activation in wild-type mice. In CB2R KO mice, DSS-induced inflammation and NLRP3 inflammasome activation were more pronounced than those in WT control. Finally, we demonstrated that AMPK-mTOR-P70S6K signaling pathway was involved in this CB2R-mediated process. We conclude that activation of CB2R ameliorates DSS-induced colitis through enhancing autophagy that may inhibit NLRP3 inflammasome activation in macrophages.
67 citations
TL;DR: The anatomical physiology, underlying mechanisms of neuropathic pain is overviewed to provide a better understanding in the initiation, development, maintenance, and modulation of this pervasive disease, and inspire research in the unclear mechanisms as well as potential targets.
46 citations
Cites background from "NMP-7 inhibits chronic inflammatory..."
...2 blocker SNI model analgesic effect (Berger et al., 2014) Suramin and the flavonoid gossypetin USP5-Cav3....
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...2 T-type channel blocker, was shown to mediate a significant analgesic effect in the model of neuropathic pain (Berger et al. , 2014)....
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References
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TL;DR: Treatment of diabetic rats with daily insulin injections reversed T‐current alterations in DRG neurons in parallel with reversal of thermal and mechanical hypersensitivities in vivo, confirming that CaV3.2 T‐channels, important signal amplifiers in peripheral sensory neurons, may contribute to the cellular hyperexcitability that ultimately leads to the development of painful PDN.
Abstract: Earlier, we showed that streptozocin (STZ)-induced type 1 diabetes in rats leads to the development of painful peripheral diabetic neuropathy (PDN) manifested as thermal hyperalgesia and mechanical allodynia accompanied by significant enhancement of T-type calcium currents (T-currents) and cellular excitability in medium-sized dorsal root ganglion (DRG) neurons. Here, we studied the in vivo and in vitro effects of gene-silencing therapy specific for the Ca(V)3.2 isoform of T-channels, on thermal and mechanical hypersensitivities, and T-current expression in small- and medium-sized DRG neurons of STZ-treated rats. We found that silencing of the T-channel Ca(V)3.2 isoform using antisense oligonucleotides, had a profound and selective anti-hyperalgesic effect in diabetic rats and is accompanied by significant down-regulation of T-currents in DRG neurons. Anti-hyperalgesic effects of Ca(V)3.2 antisense oligonucleotides in diabetic rats were similar in models of rapid and slow onset of hyperglycemia following intravenous and intraperitoneal injections of STZ, respectively. Furthermore, treatments of diabetic rats with daily insulin injections reversed T-current alterations in DRG neurons in parallel with reversal of thermal and mechanical hypersensitivities in vivo. This confirms that Ca(V)3.2 T-channels, important signal amplifiers in peripheral sensory neurons, may contribute to the cellular hyperexcitability that ultimately leads to the development of painful PDN.
169 citations
TL;DR: Assessment of agonist potency and efficacy of cannabinoids at rat brain CB1 receptors using agonist‐stimulated [35S]‐GTPγS binding to cerebellar membranes and whole brain sections establishes that 2‐AG is a full efficacy agonist, clearly more potent than AEA, in mediating CB1 receptor‐dependent G‐protein activity in native membranes.
Abstract: Two endocannabinoids, arachidonoyl ethanolamide (AEA) and 2-arachidonoylglycerol (2-AG) bind and activate G-protein-coupled cannabinoid receptors, but limited data exist on their relative ability to activate G-proteins.
Here we assess agonist potency and efficacy of various cannabinoids, including 2-AG, HU-310 (2-arachidonoyl glyceryl ether, a third putative endocannabinoid), HU-313 (another ether analogue of 2-AG), AEA, R-methanandamide (an enzymatically stable analogue of AEA), and CP-55,940 at rat brain CB1 receptors using agonist-stimulated [35S]-GTPγS binding to cerebellar membranes and whole brain sections. Degradation of endocannabinoids under experimental conditions was monitored by HPLC.
To enhance efficacy differences, agonist dose-response curves were generated using increasing GDP concentrations. At 10−6 M GDP, all compounds, except HU-313, produced full agonists responses ∼2.5 fold over basal. The superior efficacy of 2-AG over all other compounds became evident by increasing GDP (10−5 and 10−4 M).
In membrane incubations, 2-AG was degraded by 85% whereas AEA and HU-310 were stable. Pretreatment of membranes with phenylmethylsulphonyl fluoride inhibited 2-AG degradation, resulting in 2 fold increase in agonist potency. Such pretreatment had no effect on AEA potency.
Responses in brain sections were otherwise consistent with membrane binding data, but 2-AG evoked only a weak signal in brain sections, apparently due to more extensive degradation.
These data establish that even under conditions of substantial degradation, 2-AG is a full efficacy agonist, clearly more potent than AEA, in mediating CB1 receptor-dependent G-protein activity in native membranes.
British Journal of Pharmacology (2001) 134, 664–672; doi:10.1038/sj.bjp.0704297
161 citations
"NMP-7 inhibits chronic inflammatory..." refers methods in this paper
...Yet, AM281 reversed the antinociceptive action of JZL184 (16 mg/kg, i.p., an irreversible inhibitor of monoacyglycerol lipase, the primary degradatory enzyme for endocannabinoid 2-arachidonoylglycerol) (Two-way ANOVA, p 0.05) (Figure 6B), thus confirming that AM281 was indeed active in vivo....
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...JZL184 was used as a positive control, and was administered 15 minutes post-AM281 treatment in the same way as NMP-7....
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...%) and PBS. Selective CB1 antagonist AM281, irreversible inhibitor of monoacylglycerol lipase JZL184 [32,33], selective CB2 antagonist AM630, and the irreversible inhibitor of fatty acid amide hydrolase URB597 [34] were provided by Cayman Chemical, and dissolved in phosphate buffered saline (PBS) and dimethyl sulfoxide (DMSO) to 5%....
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...Selective CB1 antagonist AM281, irreversible inhibitor of monoacylglycerol lipase JZL184 [32,33], selective CB2 antagonist AM630, and the irreversible inhibitor of fatty acid amide hydrolase URB597 [34] were provided by Cayman Chemical, and dissolved in phosphate buffered saline (PBS) and dimethyl sulfoxide (DMSO) to 5%....
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TL;DR: Recent advances in understanding of T-type channel regulation are highlighted, and their implications for brain function are highlighted.
153 citations
"NMP-7 inhibits chronic inflammatory..." refers background in this paper
...Nociceptive transmission relies in part on low-voltageactivated T-type calcium channels that open in response to small membrane depolarizations [3]....
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TL;DR: It is reported that T-type calcium channels encoded by the CaV3.2 isoform are expressed in colonic nociceptive primary afferent neurons and that they contribute to the exaggerated pain perception in a butyrate-mediated rodent model of IBS.
Abstract: The symptoms of irritable bowel syndrome (IBS) include significant abdominal pain and bloating. Current treatments are empirical and often poorly efficacious, and there is a need for the development of new and efficient analgesics aimed at IBS patients. T-type calcium channels have previously been validated as a potential target to treat certain neuropathic pain pathologies. Here we report that T-type calcium channels encoded by the CaV3.2 isoform are expressed in colonic nociceptive primary afferent neurons and that they contribute to the exaggerated pain perception in a butyrate-mediated rodent model of IBS. Both the selective genetic inhibition of CaV3.2 channels and pharmacological blockade with calcium channel antagonists attenuates IBS-like painful symptoms. Mechanistically, butyrate acts to promote the increased insertion of CaV3.2 channels into primary sensory neuron membranes, likely via a posttranslational effect. The butyrate-mediated regulation can be recapitulated with recombinant CaV3.2 channels expressed in HEK cells and may provide a convenient in vitro screening system for the identification of T-type channel blockers relevant to visceral pain. These results implicate T-type calcium channels in the pathophysiology of chronic visceral pain and suggest CaV3.2 as a promising target for the development of efficient analgesics for the visceral discomfort and pain associated with IBS.
141 citations
"NMP-7 inhibits chronic inflammatory..." refers background in this paper
...knockdown [7,8] or inhibition of the T-type calcium channel by T-type channel modulators produce significant antinociceptive effects in vivo [9-13]....
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...Conversely, increased T-type channel activity in the primary pain pathway occurs in models of chronic pain such as spinal nerve injury [25], peripheral nerve injury [26], colonic hypersensitivity [8] and diabetic neuropathy [27]....
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...Inhibition of T-type channels or in vivo antisense-mediated knockdown produces antinociception in these and other models of chronic pain [7-16]....
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TL;DR: A presynaptic location for CB(1) receptors whose activation results in inhibition of glutamate release in the spinal dorsal horn is suggested, thus producing analgesia at the spinal level.
Abstract: The effect of cannabinoids on excitatory transmission in the substantia gelatinosa was investigated using intracellular recording from visually identified neurons in a transverse slice preparation ...
139 citations
"NMP-7 inhibits chronic inflammatory..." refers background in this paper
...CB1 receptors are expressed throughout the brain and the spinal cord, where they modulate neurotransmitter release such as inhibition of glutamate release by spinal cord interneurons [29,30]....
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