Up-Regulation of Regulator of G Protein Signaling 4 Expression in a Model of Neuropathic Pain and Insensitivity to Morphine (original) (raw)

Contribution of G inhibitory protein alpha subunits in paradoxical hyperalgesia elicited by exceedingly low doses of morphine in mice

Life sciences, 2011

Although morphine, at higher doses, induces analgesia, it may also enhance sensitivity to pain at extremely low doses as shown in studies for testing an animal's sensitivity to pain. We used an antisense approach capable of selectively down-regulating in vivo G(i)(G inhibitory protein),G(o) and G(s) members of the G(α) sub-family protein subunits in order to establish if these proteins might be implicated in the effects induced by extremely low morphine doses on acute thermonociception. Mice pretreated with a morphine hyperalgesic dose (1μg/kg) were submitted to hot plate test after pre-treatment with antisense oligodeoxynucleotides (aODNs) targeting G(iα), G(oα) and G(sα) regulatory proteins. The association of G-protein (guanine nucleotide-binding regulatory protein) coupled receptors with G protein was investigated using co-immunoprecipitation procedure. The downregulation of the G(iα1-3) and G(oα1) proteins reversed the licking latency responses induced by 1μg/kg morphine ad...

Inhibition of the regulator of G protein signalling RGS4 in the spinal cord decreases neuropathic hyperalgesia and restores cannabinoid CB1receptor signalling

British Journal of Pharmacology, 2015

BACKGROUND AND PURPOSE Regulators of G protein signalling (RGS) are major determinants of metabotropic receptor activity, reducing the lifespan of the GTP-bound state of G proteins. Because the reduced potency of analgesic agents in neuropathic pain may reflect alterations in RGS, we assessed the effects of CCG 63802, a specific RGS4 inhibitor, on pain hypersensitivity and signalling through cannabinoid receptors, in a model of neuropathic pain. EXPERIMENTAL APPROACH The partial sciatic nerve ligation (PSNL) model in male Sprague Dawley rats was used to measure paw withdrawal thresholds to mechanical (von Frey hairs) or thermal (Hargreaves method) stimuli, during and after intrathecal injection of CCG 63802. HEK293 cells expressing CB 1 receptors and conditional expression of RGS4 were used to correlate cAMP production and ERK phosphorylation with receptor activation and RGS4 action. KEY RESULTS Treatment of PSNL rats with CCG 63802, twice daily for 7 days after nerve injury, attenuated thermal hyperalgesia during treatment. Spinal levels of anandamide were higher in PSNL animals, irrespective of the treatment. Although expression of CB 1 receptors was unaffected, HU210-induced CB 1 receptor signalling was inhibited in PSNL rats and restored after intrathecal CCG 63802. In transfected HEK cells expressing CB 1 receptors and RGS4, inhibition of cAMP production, a downstream effect of CB 1 receptor signalling, was blunted after RGS4 overexpression. RGS4 expression also attenuated the CB 1 receptor-controlled activation of ERK1/2. CONCLUSIONS AND IMPLICATIONS Inhibition of spinal RGS4 restored endogenous analgesic signalling pathways and mitigated neuropathic pain. Signalling through CB 1 receptors may be involved in this beneficial effect Abbreviations GFAP, glial fibrillary acidic protein; Iba 1, ionized calcium-binding adapter molecule 1; PSNL, partial sciatic nerve ligation;

Effector antagonism by the regulators of G protein signalling (RGS) proteins causes desensitization of mu-opioid receptors in the CNS

Psychopharmacology, 2005

Rationale: In cell culture systems, agonists can promote the phosphorylation and internalization of receptors coupled to G proteins (GPCR), leading to their desensitization. However, in the CNS opioid agonists promote a profound desensitization of their analgesic effects without diminishing the presence of their receptors in the neuronal membrane. Recent studies have indicated that CNS proteins of the RGS family, specific regulators of G protein signalling, may be involved in mu-opioid receptor desensitization in vivo. Objective: In this work we review the role played by RGS proteins in the intensity and duration of the effects of mu-opioid receptor agonists, and how they influence the delayed tolerance that develops in response to specific doses of opioids. Results: RGS proteins are GTPase-activating proteins (GAP) that accelerate the hydrolysis of GαGTP to terminate signalling at effectors. The GAP activity of RGS-R4 and RGS-Rz proteins restricts the amplitude of opioid analgesia, and the efficient deactivation of GαzGTP subunits by RGS-Rz proteins prevents mu receptor desensitization. However, RGS-R7 proteins antagonize effectors by binding to and sequestering mu receptor-activated Gαi/o/z subunits. Thus, they reduce the pool of receptor-regulated G proteins and hence, the effects of agonists. The delayed tolerance observed following morphine administration correlates with the transfer of Gα subunits from mu receptors to RGS-R7 proteins and the subsequent stabilization of this association. Conclusion: In the CNS, the RGS proteins control the activity of mu opioid receptors through GAP-dependent (RGS-R4 and RGS-Rz) as well as by GAP-independent mechanisms (RGS-R7). As a result, they can both antagonize effectors and desensitize receptors under certain circumstances.

The GBeta5 subunit that associates with the R7 subfamily of RGS proteins regulates mu-opioid effects

Neuropharmacology, 2003

The Gb5 protein, which is similar in sequence to other G-protein beta subunits, mainly associates with the G-protein g-like (GGL) domains of the R7 subfamily of regulators of G-protein signalling (RGS) proteins. This paper reports the presence of the Gb5 protein and its mRNA in all areas of mouse CNS, and also its involvement in the cellular signals initiated at mu-and deltaopioid receptors. The expression of Gb5 and RGS9-2 proteins (member of the R7 subfamily of RGS) was reduced by blocking their mRNAs with antisense oligodeoxynucleotides (ODN). Knock-down of these proteins enhanced the potency and duration of antinociception promoted by morphine and [D-Ala 2 , N-MePhe 4 ,Gly-ol 5 ]-enkephalin (DAMGO), agonists at mu opioid receptors. However, the activity of the selective agonist at delta opioid receptors, [D-Pen 2,5 ]-encephalin (DPDPE), appeared to be reduced. A single intracerebroventricular (icv) ED 80 analgesic dose of morphine gave rise to acute tolerance in control mice, but did not promote tolerance in Gb5 or RGS9-2 knock-down animals. In a model of sustained morphine treatment, the impairment of Gb5 proteins facilitated the development of tolerance. This treatment did not alter the incidence of jumping behaviour precipitated by naloxone 3 days after commencing with chronic morphine. These results show differences in the signalling regulation of G-proteins when activated by mu or delta opioid agonists. For mu opioid receptors, acute tolerance, but probably not long-term tolerance, appears to depend on the function of Gb5 subunits and associated RGS proteins.

RGS9 proteins facilitate acute tolerance to mu-opioid effects

European Journal of Neuroscience, 2001

This paper reports that regulators of G-protein signalling (RGS) proteins modulate the timing and amplitude of opioid signals by a push±pull mechanism. This is achieved without noticeable changes in the binding properties of opioids, e.g. b-endorphin to muopioid receptors. The expression of RGS proteins was reduced by blocking their mRNA with antisense oligodeoxynucleotides (ODN). Knock down of RGS2 or RGS3 diminished morphine and b-endorphin analgesia, whereas that of RGS9 or RGS12 enhanced this activity. In mice with impaired RGS9, but not impaired RGS2, the potency and, in particular, the duration of opioid antinociception increased. Further, the animals did not exhibit acute tolerance generated by a single and ef®cacious dose of morphine, nor did they develop tolerance after a daily i.c.v. injection of the opioid for 4 days. In a model of sustained morphine treatment, the impairment of RGS9 proteins facilitated increases in the response to the delivered opioid. This was only effective for 2±3 h after the subcutaneous implantation of an oily morphine pellet; later, tolerance developed. To reduce the impact of the chronic morphine acting on opioid receptors, other RGS proteins presumably substitute the GTPase-activating function of RGS9 on morphine-activated Ga-GTP subunits. The desensitization of mu-opioid receptors appears to be a cell membrane-limited process facilitated by RGS9¢s sequestering of agonist-segregated Ga subunits.

Chronic morphine treatment enhances sciatic nerve stimulation-induced immediate early gene expression in the rat dorsal horn

PubMed, 2015

Synaptic plasticity is a property of neurons that can be induced by conditioning electrical stimulation (CS) of afferent fibers in the spinal cord. This is a widely studied property of spinal cord and hippocampal neurons. CS has been shown to trigger enhanced expression of immediate early gene proteins (IEGPs), with peak increases observed 2 hour post stimulation. Chronic morphine treatment has been shown to promoteinduce opioid-induced hyperalgesia, and also to increase CS-induced central sensitization in the dorsal horn. As IEGP expression may contribute to development of chronic pain states, we aimed to determine whether chronic morphine treatment affects the expression of IEGPs following sciatic nerve CS. Changes in expression of the IEGPs Arc, c-Fos or Zif268 were determined in cells of the lumbar dorsal horn of the spinal cord. Chronic Morphine pretreatment over 7 days led to a significant increase in the number of IEGP positive cells observed at both 2 h and 6 h after CS. The same pattern of immunoreactivity was obtained for all IEGPs, with peak increases occurring at 2 h post CS. In contrast, morphine treatment alone in sham operated animals had no effect on IEGP expression. We conclude that chronic morphine treatment enhances stimulus-induced expression of IEGPs in the lumbar dorsal horn. These data support the notion that morphine alters gene expression responses linked to nociceptive stimulation and plasticity.

RGS4 Maintains Chronic Pain Symptoms in Rodent Models

The Journal of Neuroscience, 2019

Regulator of G-protein signaling 4 (RGS4) is a potent modulator of G-protein-coupled receptor signal transduction that is expressed throughout the pain matrix. Here, we use genetic mouse models to demonstrate a role of RGS4 in the maintenance of chronic pain states in male and female mice. Using paradigms of peripheral inflammation and nerve injury, we show that the prevention of RGS4 action leads to recovery from mechanical and cold allodynia and increases the motivation for wheel running. Similarly, RGS4KO eliminates the duration of nocifensive behavior in the second phase of the formalin assay. Using the Complete Freud's Adjuvant (CFA) model of hindpaw inflammation we also demonstrate that downregulation of RGS4 in the adult ventral posterolateral thalamic nuclei promotes recovery from mechanical and cold allodynia. RNA sequencing analysis of thalamus (THL) from RGS4WT and RGS4KO mice points to many signal transduction modulators and transcription factors that are uniquely re...