Ultra-low Dose Naloxone Research Articles

μ-Opioid Receptor Activation at the Dorsal Reticular Nucleus Shifts Diffuse Noxious Inhibitory Controls to Hyperalgesia in Chronic Joint Pain in Male Rats.

The dorsal reticular nucleus is a pain facilitatory area involved in diffuse noxious inhibitory control (DNIC) through opioidergic mechanisms that are poorly understood. The hypothesis was that signaling of μ-opioid receptors is altered in this area with prolonged chronic inflammatory pain and that this accounts for the loss of DNICs occurring in this condition. Monoarthritis was induced in male Wistar rats (n = 5 to 9/group) by tibiotarsal injection of complete Freund's adjuvant. The immunolabeling of µ-opioid receptors and the phosphorylated forms of µ-opioid receptors and cAMP response element binding protein was quantified. Pharmacologic manipulation of μ-opioid receptors at the dorsal reticular nucleus was assessed in DNIC using the Randall-Selitto test. At 42 days of monoarthritis, μ-opioid receptor labeling decreased at the dorsal reticular nucleus, while its phosphorylated form and the phosphorylated cAMP response element binding protein increased. [d-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin acetate (DAMGO) enhanced DNIC analgesia in normal animals (means ± SD: pre-DNIC: 126.9 ± 7.0 g; DNIC - DAMGO: 147.5 ± 8.0 g vs. DNIC + DAMGO: 198.1 ± 19.3 g; P < 0.001), whereas it produced hyperalgesia in monoarthritis (pre-DNIC: 67.8 ± 7.5 g; DNIC - DAMGO: 70.6 ± 7.7 g vs. DNIC + DAMGO: 32.2 ± 2.6 g; P < 0.001). An ultra-low dose of naloxone, which prevents the excitatory signaling of the μ-opioid receptor, restored DNIC analgesia in monoarthritis (DNIC - naloxone: 60.0 ± 6.1 g vs. DNIC + naloxone: 98.0 ± 13.5 g; P < 0.001), compared to saline (DNIC - saline: 62.5 ± 5.2 g vs. DNIC + saline: 64.2 ± 3.8 g). When injected before DAMGO, it restored DNIC analgesia and decreased the phosphorylated cAMP response element binding protein in monoarthritis. The dorsal reticular nucleus is likely involved in a facilitatory pathway responsible for DNIC hyperalgesia. The shift of μ-opioid receptor signaling to excitatory in this pathway likely accounts for the loss of DNIC analgesia in monoarthritis.

Evaluation of the Effects of Ultra-Low Dose Naloxone on Brachial Plexus Block with Bupivacaine

Introduction: Adjuvants are used in conjunction with local anaesthetics for brachial plexus block to quicken onset, prolong duration and augment the quality of anaesthesia.&#x0D; Objectives: The study aims to evaluate the effect of adding ultra-low dose naloxone to bupivacaine on the duration of postoperative analgesia.&#x0D; Methodology: A prospective, randomized, double-blind study was conducted including forty American Society of Anesthesiologists Physical Status (ASA PS) I and II patients of either sex between 15 and 65 years of age scheduled for upper limb surgery. They were randomized into two groups. Group BN received a mixture of 0.33% plain bupivacaine with ultra-low dose naloxone (100 ng) and group BS received 0.33% plain bupivacaine with saline. The procedure was standardized in both groups. The onset of sensory and motor block were assessed with pinprick sensation and muscle power respectively. Visual Analogue Score (VAS) was used to assess the pain at 1, 2, 4, 8 and 12 hours postoperatively.&#x0D; Result: The patients were comparable with respect to age, sex, ASA PS, baseline hemodynamic and respiratory parameters and duration of surgery. The mean duration of analgesia in group BN was significantly longer (610.7±125.4 mins) as compared to group BS (354.3±59.6 mins) (p&lt;0.001). No complications were noted in both the groups.&#x0D; Conclusion: This study demonstrated that 100 ng naloxone, when used with 0.33% plain bupivacaine for supraclavicular brachial plexus block, prolongs postoperative analgesia significantly after upper limb surgeries, without any significant side effects.

Effect of ultra-low-dose naloxone during supraclavicular brachial plexus block on the antinociceptive criteria of postoperative opioid in orthopedic upper limb surgeries

Background Brachial plexus block is a popular and widely used regional nerve block technique for perioperative anesthesia and analgesia for upper extremity surgeries. Many drugs such as morphine and midazolam have been used as an adjuvant with local anesthetics to achieve quick, dense, and prolonged block. Drugs with minimal side effects are always looked for. Naloxone as opioid antagonists could selectively block excitatory effects of opioids, stimulate release and displace endorphins from receptor sites, and reduce the side effects of opioids. An ultra-low-dose naloxone, added as an adjuvant to local anesthetic, may prolong sensory and motor blockades with enhanced opioid effect or direct antagonism of its excitatory receptors. Patients and methods After approval by Institutional Research Board, clinical trial registration, and obtaining informed consents, 64 patients scheduled for orthopedic upper limb surgeries under supraclavicular brachial plexus block were enrolled into two equal groups: group B (bupivacaine 0.5%) and group BN (bupivacaine 0.5% plus 100 ng naloxone). Results There was significant decrease in fentanyl consumption in GBN (216.6±76.10) versus group B (507.5±117.50). Also, the number of fentanyl requests was lesser in group BN (3.0±1.11) than in group B (7.3±1.73). The onset time for sensory and motor blocks showed no significant difference among groups; the duration of sensory and motor blocks was prolonged in GBN (17.2±2.96–7.7±1.93 h) versus GB (7.3±1.22–4.7±.79 h). The time to first rescue fentanyl was prolonged in GBN (18.3±3.21 h) than in GB (8.0±1.41 h). There was significant reduction in visual analog scale score at 8, 12, and 18 h postoperatively in GBN versus GB. There were no significant changes in Ramsay sedation scale or complications within 48 h postoperatively. Conclusion Ultra-low dose of naloxone (100 ng) added to 20 ml bupivacaine 0.5% for supraclavicular brachial plexus block enhanced the postoperative opioid analgesia by reducing the total postoperative opioid consumption, prolonging the interval between opioid doses and the duration of sensory and motor block.

Ultra-low-dose Naloxone as an Adjuvant to Patient Controlled Analgesia (PCA) With Morphine for Postoperative Pain Relief Following Lumber Discectomy: A Double-blind, Randomized, Placebo-controlled Trial.

Lumbar discectomy is one of the most commonly performed neurosurgical procedures. Many patients experience postoperative pain after lumbar discectomy. This study evaluated the effect of ultra-low-dose naloxone infusion on pain intensity after lumbar discectomy in individuals receiving patient-controlled analgesia (PCA) with morphine. In a double-blind, randomized, controlled trial, a total of 80 patients scheduled for open discectomy was randomly assigned to receive naloxone (group N) or placebo (group P). After surgery, all patients were connected to a morphine PCA pump. Both groups received 500 mL of normal saline using a continuous infusion pump through a separate intravenous line for 24 hours. However, group N received a total dose of 0.25 μg/kg/h naloxone, which was added to the normal saline infusion. All patients were asked to grade the intensity of their pain, severity of nausea, vomiting, and pruritus on a 0 to 10 visual analog scale before being discharged from the postanesthesia care unit and at 1, 6, 12, and 24 hours postoperatively. It was observed that both groups had a statistically significant (P<0.01) time trend difference for pain, nausea, and pruritus scores. A significant difference was found between the 2 groups in terms of intensity of pain, nausea, and pruritus, with the naloxone group experiencing a lower level in comparison with the placebo group. Moreover, the median (interquartile range) of morphine consumption after surgery for patients who received naloxone was 26 (24.25 to 28) mg, which is significantly (P<0.001) lower than for the placebo group, which had a median (interquartile range) of 34 (32 to 36) mg. It is concluded that infusion of ultra-low-dose naloxone (0.25 μg/kg/h) along with morphine PCA can significantly reduce pain intensity, morphine consumption, and opioid-induced nausea and pruritus after lumbar discectomy.

Ultra-low Dose Naloxone Added to 0.5% Bupivacaine Significantly Prolongs the Duration of Analgesia Following Supraclavicular Brachial Plexus Block

In this prospective, randomized, double-blind study, we evaluated the effect of ultra-low dose of naloxone on duration of supraclavicular brachial plexus block. It was hypothesized that naloxone can prolong the duration of sensory block. Following approval by Hospital Ethical Issues Committee, eighty patients scheduled for upper limb surgery under supraclavicular brachial plexus block were randomly allocated into control group who received 20ml bupivacaine with 3 ml normal saline (Group B) or study group that received 20ml bupivacaine with 100 mcg. of naloxone in 3 ml saline, (Group BN). Onsets of sensory and motor blockade were assessed at an interval of 3 min following the block. Duration of sensory and motor block was considered to be the time interval between the complete block and the first post operative pain reported by patient and complete recovery of motor functions respectively. The difference in onset time for sensory and motor block was statistically significant between two groups but clinically it may be considered insignificant. The recovery of sensory block was slower in group BN (15.6 ± 3.2 hr) compared to group B (13.3 ± 2.4 hr) [p=0.0001]. The recovery of motor block was slower in group B (13.3 ± 2.5 hr) compared to group BN (11.6 ± 4.3) [P=0.03]. In conclusion, addition of ultra-low dose of naloxone to bupivacaine in supraclavicular block prolongs the duration of sensory block and reduces duration of motor block significantly as compared to bupivacaine alone.

The Effects of the Toll-Like Receptor 4 Antagonist, Ibudilast, on Sevoflurane's Minimum Alveolar Concentration and the Delayed Remifentanil-Induced Increase in the Minimum Alveolar Concentration in Rats.

Ultralow doses of naloxone, an opioid and toll-like receptor 4 antagonist, blocked remifentanil-induced hyperalgesia and the associated increase in the minimum alveolar concentration (MAC), but not tolerance. The aim was to determine the effects of the toll-like receptor 4 antagonist, ibudilast, on the MAC in the rat and how it might prevent the effects of remifentanil. Male Wistar rats were randomly allocated to 5 treatment groups (n = 7 per group): 10 mg/kg ibudilast intraperitoneally, 240 µg/kg/h remifentanil IV, ibudilast plus remifentanil, remifentanil plus naloxone IV, or saline. The sevoflurane MAC was determined 3 times in every rat and every day (days 0, 2, and 4): baseline (MAC-A) and 2 further determinations were made after treatments, 1.5 hours apart (MAC-B and MAC-C). A reduction in baseline MAC was produced on day 0 by ibudilast, remifentanil, remifentanil plus ibudilast, remifentanil plus naloxone (P < 0.01), but not saline. Similar effects were found on days 2 and 4. A tolerance to remifentanil was found on days 0, 2, and 4, which neither ibudilast nor naloxone prevented. The MAC increase produced by remifentanil on day 4 (P = 0.001) was prevented by either ibudilast or naloxone. Ibudilast, besides reducing the MAC, prevented the delayed increase in baseline MAC produced by remifentanil but not the increase in MAC caused by tolerance to remifentanil.

A randomized controlled study of the effects of adding ultra-low dose naloxone to lidocaine for intravenous regional anesthesia

ObjectiveThis study was designed to evaluate the effect of adding ultra-low dose of naloxone as an adjuvant to lidocaine for intravenous regional anesthesia (IVRA).MethodForty patients undergoing elective short procedures in the upper limb were randomly and blindly divided into two groups of twenty patients each. Group L (n = 20) received 3 mg/kg of 2% lidocaine diluted with normal saline to 30 ml. Group LN (n = 20) received 3 mg/kg of 2% lidocaine and naloxone 100 ng (1 ml) diluted with normal saline to 30 ml. Onset and recovery time of sensory and motor block, intraoperative and post-operative pain were measured by visual analog score (VAS), and also intraoperative analgesic requirement, time of first requirement of diclofenac postoperatively, total amount of diclofenac needed in 24 h, patient’s satisfaction and surgeon’s satisfaction scores were measured.ResultsRecovery of sensory block was significantly longer in group LN (26.7 ± 2.8 min) compared to group L (16.3 ± 0.6 min); p value (0.000). Also the recovery of motor block was significantly longer in group LN (19.1 ± 1.0 min) compared to group L (17.9 ± 1.2 min), p value (0.002). Intraoperative fentanyl requirement was significantly less in group LN (15.8 ± 5.0 mcg) compared to group L (40.0 ± 10.5 mcg), p value (0.000). 1st fentanyl requirement time was significantly longer in group LN (22.4 ± 3.1 min) than in group L (14.5 ± 6.1 min), p value (0.000). Time of first analgesic requirement post-operative was longer in group LN (78.5 ± 6.8 min) compared to group L (40.5 ± 2.0 min), p value (0.000). Total amount of diclofenac needed in 24 h was significantly less in group LN (57 ± 50 mg) compared to group L (120 ± 45 mg), p value (0.000).ConclusionThe addition of ultra-low-dose naloxone 100 ng to lidocaine for IVRA in upper limb surgery, prolonged the duration of sensory and motor block, and reduced tourniquet pain, as well as intraoperative and postoperative analgesic consumption.

Ultralow Dose of Naloxone as an Adjuvant to Intrathecal Morphine Infusion Improves Perceived Quality of Sleep but Fails to Alter Persistent Pain: A Randomized, Double-blind, Controlled Study.

This randomized, cross-over, double-blind, controlled study of continuous intrathecal morphine administration in patients with severe, long-term pain addresses whether the supplementation of low doses of naloxone in this setting is associated with beneficial clinical effects. All of the study subjects (n=11) provided informed consent and were recruited from a subset of patients who were already undergoing long-term treatment with continuous intrathecal morphine because of difficult-to-treat pain. The patients were (in a randomized order) also given intrathecal naloxone (40 ng/24 h or 400 ng/24 h). As control, the patients' ordinary dose of morphine without any additions was used. The pain (Numeric Rating Scale, NRS) during activity, perceived quality of sleep, level of activity, and quality of life as well as the levels of several proinflammatory and anti-inflammatory cytokines in the blood were assessed. The prestudy pain (NRS during activity) in the study group ranged from 3 to 10. A total of 64% of the subjects reported improved quality of sleep during treatment with naloxone at a dose of 40 ng per 24 hours as compared with 9% with sham treatment (P=0.024). Although not statistically significant, pain was reduced by 2 NRS steps or more during supplemental treatment with naloxone in 36% of subjects when using the 40 ng per 24 hours dose and in 18% of the subjects when using naloxone 400 ng per 24 hours dose. The corresponding percentage among patients receiving unaltered treatment was 27%. To conclude, the addition of an ultralow dose of intrathecal naloxone (40 ng/24 h) to intrathecal morphine infusion in patients with severe, persistent pain improved perceived quality of sleep. We were not able to show any statistically significant effects of naloxone on pain relief, level of activity, or quality of life.

Opioid Antagonists

Opioid Antagonists

Intrathecal Ultra-Low Dose Naloxone Enhances the Antihyperalgesic Effects of Morphine and Attenuates Tumor Necrosis Factor-α and Tumor Necrosis Factor-α Receptor 1 Expression in the Dorsal Horn of Rats with Partial Sciatic Nerve Transection

Glutamate homeostasis and microglia activation play an important role in the development and maintenance of neuropathic pain. We designed this investigation to examine whether ultra-low dose naloxone administered alone or in combination with morphine could alter the concentration of the excitatory amino acids (EAAs) glutamate and aspartate, as well as the expression of tumor necrosis factor-α (TNF-α) and its receptors (TNFR1 and TNFR2) in the spinal cord dorsal horn of rats with partial sciatic nerve transection (PST). Male Wistar rats underwent intrathecal catheter implantation for drug delivery and were divided in 7 groups: sham-operated + saline (sham), PST + saline (S), PST + 15 ng naloxone (n), PST + 15 µg naloxone (N), PST + 10 µg morphine (M), PST + 15 ng naloxone + 10 µg morphine (Mn), PST + 15 µg naloxone + 10 µg morphine (MN). Thermal withdrawal latency and mechanical withdrawal threshold, TNF-α and TNFR expression in the spinal cord and dorsal root ganglia, and EAAs glutamate and aspartate concentration in cerebrospinal fluid dialysates were measured. Ten days after PST, rats developed hyperalgesia (P < 0.0001) and allodynia (P < 0.0001), and increased TNF-α (P < 0.0001) and TNFR1 expression (P = 0.0009) were measured in the ipsilateral spinal cord dorsal horn. The antihyperalgesic and antiallodynic effects of morphine (10 μg) were abolished by high-dose naloxone (15 μg; P = 0.0031) but enhanced by ultra-low dose naloxone (15 ng; P = 0.0015), and this was associated with a reduction of TNF-α (P < 0.0001) and TNFR1 (P = 0.0009) expression in the spinal cord dorsal horn and EAAs concentration (glutamate: P = 0.0001; aspartate: P = 0.004) in cerebrospinal fluid dialysate. Analysis of variance (ANOVA) or Student t test with Bonferroni correction were used for statistical analysis. Ultra-low dose naloxone enhances the antihyperalgesia and antiallodynia effects of morphine in PST rats, possibly by reducing TNF-α and TNFR1 expression, and EAAs concentrations in the spinal dorsal horn. Ultra-low dose naloxone may be a useful adjuvant for increasing the analgesic effect of morphine in neuropathic pain conditions.

Effect of ultra-low dose naloxone on postoperative hyperalgesia induced by large dose remifentanil

Objective To evaluate the effect of ultra-low dose naloxone on postoperative hyperalgesia caused by large-dose remifentanil.Methods Forty ASA Ⅰ-Ⅲ adult patients,scheduled for gastrointestinal surgery,were randomly assigned into 2 groups (n =20 each):large dose remifentail group (group R) and ultra-low dose naloxone group (group N).Anesthesia was induced with iv injection of remifentanil,propofol and cisatracurium and maintained with inhalation of sevoflurane and infusion of remifentanil.The patients were tracheal intubated and mechanically ventilated.In group R,remifentanil was infused at a rate of 0.25 μg· kg-1 · min-1 starting from the beginning of skin incision.The infusion rate was adjusted according to hemodynamics during operation and subsequently increased/decreased by 0.05 μg· kg-1· min-1 each time.In group N,naloxone was infused at 0.1 μg·kg-1· h-1 while infusing remifentanil,naloxone infusion was stopped at the beginning of peritoneum closure and the other treatments were similar to those previously described in group R.All patients were sent to post-anesthesia care unit after surgery and stayed there for 90 min.Morphine was given when need.The patient-controlled intravenous analgesia was used for postoperative analgesia after leaving post-anesthesia care unit.The first pain time was calculated.The morphine consumption and complications such as nausea,vomiting and pruritus were recorded at 15,30,60 and 90 min and 2,6,24,48 and 72 h after surgery.Results Compared with group R,the morphine consumption was significantly reduced at each time point after surgery,the first pain time was prolonged,and incidence of nausea was decreased (P ＜ 0.05),while no significant change was found in the incidence of vomiting and prutirus in group N (P ＞ 0.05).Conclusion Infusing ultra-low dose naloxone (0.1μg· kg-1 ·h-1) during operation can attenuate postoperative hyperalgesia caused by large-dose remifentanil in patients. Key words: Naloxone; Piperidines; Hyperalgesia; Pain, postoperative

Naloxone infusion and post‐hysterectomy morphine consumption: a double‐blind, placebo‐controlled study

The evidence that an infusion of a low dose of naloxone reduces post-operative pain and opioid analgesic consumption is somewhat conflicting. Thus, the aim of the present study was to investigate the effect of an ultra-low dose of naloxone on patient-controlled morphine analgesia. Ninety patients, 35-55 years old, scheduled for total abdominal hysterectomy, were enrolled in this prospective, randomized, double-blind and placebo-controlled study. Post-operatively, they received either saline (n = 45) or naloxone (n = 45) for 24 h. A standard general anesthesia was administered in both groups. In the recovery room, patients received morphine by a patient-controlled analgesia device. An ultra-low dose of naloxone was infused intravenously at 0.25 μg/kg/h for 24 h in the intervention group. Saline was infused in the control group. Following the surgery, morphine consumption, numeric rating score for pain intensity, nausea and vomiting, pruritus, and requests for antiemetic were recorded at baseline, 30 min, 1, 4, 8,16, 20, and 24 h following their discharge from recovery. Naloxone reduced morphine consumption over the first 24 post-operative hours significantly compared with the controls (saline) {19.5 [standard deviation (SD) 3.4] mg vs. 27.5 [SD 5.9] mg; P < 0.001}. The incidence and severity of nausea and vomiting was significantly reduced in the naloxone group. The incidence of pruritus and the pain scores at rest and activity were not significantly different. Following hysterectomy, an ultra-low dose of naloxone infusion proved to reduce morphine consumption as well as the incidence and severity of opioid-induced nausea and vomiting.

Ultra-low dose naloxone restores the antinocicepitve effect of morphine in PTX-treated rats: Association of IL-10 upregulation in the spinal cord

AimsUltra-low dose naloxone has been shown to restore the antinociceptive effect of morphine in pertussis toxin (PTX)-treated rats by suppressing spinal microglia activation and inhibiting inflammatory cytokine expression. This study was further investigated the mechanism by which ultra-low dose naloxone promotes analgesia in pertussis toxin-treated rats. Main methodsMale Wistar rats were implanted with an intrathecal (i.t.) catheter and injected either saline or PTX (1μg). Four days later, rats randomly received either saline, or ultra-low dose naloxone, or recombinant rat interleukin-10 (rrIL-10) (1μg) injection followed by saline or morphine (10μg) 30min later. In some experiments, mouse anti-rat IL-10 antibody (10μg) was injected intrathecally into PTX injected rats daily on days 4, 5, 6, and 7. On day 7, ultra-low dose naloxone was given 1h after antibody injection with or without subsequent morphine injection. Key findingsPTX injection induced notable thermal hyperalgesia and mechanical allodynia. Injection of ultra-low dose naloxone preserved the antinociceptive effect of morphine in PTX-treated rats and associated an increasing of IL-10 protein expression. Intrathecal injection rrIL-10 alone or in combination with morphine, not only reversed mechanical allodynia but also partially restored the antinociceptive effect of morphine; injection of anti-rat IL-10 antibody attenuated the effect of morphine plus ultra-low dose naloxone on mechanical allodynia and completely inhibited the antinociceptive effect of morphine. SignificanceThese results indicate that intrathecal ultra-low dose naloxone induces IL-10 expression in spinal neuron and microglia, which suppresses PTX-induced neuroinflammation and restores the antinociceptive effect of morphine.

Intrathecal Ultra-Low Dose Naloxone Enhances the Antinociceptive Effect of Morphine by Enhancing the Reuptake of Excitatory Amino Acids from the Synaptic Cleft in the Spinal Cord of Partial Sciatic Nerve–Transected Rats

In this study, we examined the effects of ultra-low dose naloxone on the antinociceptive effect of morphine and on spinal cord dorsal horn glutamate transporter expression in rats with neuropathic pain. Neuropathic pain was induced in male Wistar rats by partial transection of the left sciatic nerve and an intrathecal catheter was implanted for drug administration; in some rats, an intrathecal microdialysis probe for cerebrospinal fluid (CSF) dialysate collection was also implanted. Nociception was assessed using the plantar test, a Hargreaves radiant heat apparatus, and by the von Frey test, using a dynamic plantar anesthesiometer. Glutamate transporter protein expression in the left spinal cord dorsal horn was examined by Western blotting and immunohistochemistry. Levels of the excitatory amino acids (EAAs) glutamate and aspartate in the CSF dialysate were measured using high-performance liquid chromatography. Reduced astrocyte expression of glutamate transporters (GLT-1 and GLAST levels were 55% and 53%, respectively, of that in sham-operated rats) in laminae I and II of the spinal cord dorsal horn ipsilateral to the partial sciatic nerve transection (PST), and hyperalgesia and allodynia in the PST hindlimb were observed. High-dose naloxone (15 μg) attenuated the antihyperalgesia and antiallodynia effects of the morphine (10 μg). In contrast, ultra-low dose (15 ng) naloxone enhanced the antinociceptive effect of morphine (10 μg), with an increase in the paw withdrawal threshold to thermal stimulus (from 19% to 35%) and to tactile stimulus (from 33% to 55%) compared with morphine treatment alone, and this was associated with restoration of GLAST and GLT-1 expression to control levels (102% and 114%, respectively) in the astrocytes of laminae I and II in the spinal cord dorsal horn ipsilateral to the PST hindlimb and a decrease in EAA levels in the CSF dialysate (glutamate: 10.0 μM; aspartate: 1.1 μM). Ultra-low dose naloxone enhanced the antinociceptive effect of morphine in PST rats, possibly by restoration of GLAST and GLT-1 expression in astrocytes, which inhibited the accumulation of EAAs in the synapses, resulting in a neuroprotective effect.

Co-administration of ultra-low dose naloxone attenuates morphine tolerance in rats via attenuation of NMDA receptor neurotransmission and suppression of neuroinflammation in the spinal cords

Although mechanisms underlying ultra-low dose naloxone-induced analgesia have been proposed, possible interactions with glutamatergic transmission and glial cell activation have not been addressed. In the present study, we examined the effect of ultra-low dose naloxone on spinal glutamatergic transmission and glial cell activity in rats chronically infused with morphine. In male Wistar rats, intrathecal morphine infusion (15 μg/h) for 5 days induced (1) antinociceptive tolerance, (2) downregulation of glutamate transporters (GTs) GLT-1, GLAST, and EAAC1, (3) increasing of NMDA receptor (NMDAR) NR1 subunit expression and phosphorylation, (4) upregulation of protein kinase C gamma (PKCγ) expression, and (5) glial cell activation. On day 5, morphine challenge (15 μg/10 μl) caused a significant increase in the concentration of the excitatory amino acids (EAAs) aspartate and glutamate in the spinal CSF dialysates of morphine-tolerant rats. Intrathecal co-infusion of ultra-low dose naloxone (15 pg/h) with morphine attenuated tolerance development, reversed GTs expression, inhibited the NMDAR NR1 subunit expression and phosphorylation, and PKCγ expression, inhibited glial cell activation, and suppressed the morphine-evoked EAAs release. These effects may result in preservation of the antinociceptive effect of acute morphine challenge in chronic morphine-infused rats. Ultra-low dose naloxone infusion alone did not produce an antinociceptive effect. These findings demonstrated that attenuation of glutamatergic transmission and neuroinflammation by ultra-low dose naloxone co-infusion preserves the lasting antinociceptive effect of morphine in rats chronically infused with morphine.

Ultra-low dose naloxone upregulates interleukin-10 expression and suppresses neuroinflammation in morphine-tolerant rat spinal cords

Co-infusion of ultra-low dose naloxone and morphine attenuates morphine tolerance through the prevention of mu opioid receptor-Gs protein coupling. We previously demonstrated that chronic intrathecal infusion of morphine leads to tolerance and spinal neuroinflammation. The aim of present study was to examine the possible mechanisms by which ultra-low dose naloxone modulates spinal neuroinflammation, particularly the role of anti-inflammatory cytokine interleukin 10 (IL-10). Morphine tolerance was induced in male Wistar rats by intrathecal infusion of morphine (15 μg/h) for 5 days, and co-infusion of naloxone (15 pg/h) was used to evaluate the impact on spinal cytokine expression. Recombinant rat IL-10 (rrIL-10) or anti-rat IL-10 antibody was injected to elucidate the effect of IL-10 on morphine tolerance. Our results showed that co-infusion of naloxone (15 pg/h) with morphine not only attenuated tolerance, shifting the AD 50 from 89.2 to 11.7 μg but also inhibited the increased expression of pro-inflammatory cytokine (TNF-α, IL-1β, and IL-6) caused by chronic intrathecal morphine infusion. The increase of IL-10 protein and mRNA were 1.5- and 3-fold, respectively, compared to that in morphine-infused rat spinal cords. A combination of daily rrIL-10 (1 μg) injection with morphine infusion produced, in a less potent, preservative antinociception and inhibited pro-inflammatory cytokine production compared to ultra-low dose naloxone co-infusion, and the effect of ultra-low dose naloxone co-infusion was inhibited by daily intrathecal anti-rat IL-10 antibody injection. These results demonstrate that IL-10 contributes to the attenuation of pro-inflammatory cytokine expression caused by ultra-low dose naloxone/morphine co-infusion and thus the attenuation of morphine tolerance.

Ultra-low dose naloxone restores the antinociceptive effect of morphine in pertussis toxin-treated rats by reversing the coupling of μ-opioid receptors from Gs-protein to coupling to Gi-protein

Pertussis toxin (PTX) treatment results in ADP-ribosylation of Gi-protein and thus in disruption of μ-opioid receptor signal transduction and loss of the antinociceptive effect of morphine. We have previously demonstrated that pretreatment with ultra-low dose naloxone preserves the antinociceptive effect of morphine in PTX-treated rats. The present study further examined the effect of ultra-low dose naloxone on μ-opioid receptor signaling in PTX-treated rats and the underlying mechanism. Male Wistar rats implanted with an intrathecal catheter received an intrathecal injection of saline or PTX (1 μg in 5 μl of saline), then, 4 days later, were pretreated by intrathecal injection with either saline or ultra-low dose naloxone (15 ng in 5 μl of saline), followed, 30 min later, by saline or morphine (10 μg in 5 μl of saline). Four days after PTX injection, thermal hyperalgesia was observed, together with increased coupling of excitatory Gs-protein to μ-opioid receptors in the spinal cord. Ultra-low dose naloxone pretreatment preserved the antinociceptive effect of morphine, and this effect was completely blocked by the μ-opioid receptor antagonist CTOP, but not by the κ-opioid receptor antagonist nor-BNI or the δ-opioid receptor antagonist naltrindole. Moreover, a co-immunoprecipitation study showed that ultra-low dose naloxone restored μ-opioid receptor/Gi-protein coupling and inhibited the PTX-induced μ-opioid receptor/Gs-protein coupling. In addition to the anti-neuroinflammatory effect and glutamate transporter modulation previously observed in PTX-treated rats, the re-establishment of μ-opioid receptor Gi/Go-protein coupling is involved in the restoration of the antinociceptive effect of morphine by ultra-low dose naloxone pretreatment by normalizing the balance between the excitatory and inhibitory signaling pathways. These results show that ultra-low dose naloxone preserves the antinociceptive effect of morphine, suppresses spinal neuroinflammation, and reduces PTX-elevated excitatory Gs-coupled opioid receptors in PTX-treated rats. We suggest that ultra-low dose naloxone might be clinically valuable in pain management.

Ultra-low dose naloxone restores the antinociceptive effect of morphine in pertussis toxin–treated rats and prevents glutamate transporter downregulation by suppressing the p38 mitogen-activated protein kinase signaling pathway

We previously demonstrated that ultra-low dose naloxone restores the antinociceptive effect of morphine in rats with pertussis toxin (PTX)–induced thermal hyperalgesia by reversing the downregulation of glutamate transporter (GT) expression and suppressing spinal neuroinflammation. In the present study, we examined the underlying mechanisms of this anti-inflammatory effect in PTX-treated rats, particularly on the expression of GTs. Male Wistar rats were implanted with an intrathecal catheter and, in some cases, with a microdialysis probe. All rats were injected intrathecally with saline (5 μl) or PTX (1 μg), then, 4 days later, were randomly assigned to receive a single injection of saline, ultra-low dose naloxone (15 ng), or the p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 (5 μg), followed by morphine injection (10 μg) 30 min later. Our results showed that PTX injection induced activation of microglia and a significant increase in P-p38 MAPK expression in the spinal cord. Ultra-low dose naloxone plus morphine significantly inhibited the effect of PTX on P-p38 MAPK expression in the spinal cord, while the p38 MAPK inhibitor SB203580 attenuated the PTX-induced mechanical allodynia, thermal hyperalgesia, increase in spinal cerebrospinal fluid excitatory amino acids, and downregulation of GTs. These results show that the restoration of the antinociceptive effect of morphine and GT expression in PTX-treated rats by ultra-low dose naloxone involves suppression of the p38 MAPK signal transduction cascade.

Buprenorphine analgesia "may be significantly enhanced" with the addition of ultra-low doses of naloxone,

Buprenorphine analgesia "may be significantly enhanced" with the addition of ultra-low doses of naloxone,

Naloxone rapidly evokes endogenous kappa opioid receptor-mediated hyperalgesia in naïve mice pretreated briefly with GM1 ganglioside or in chronic morphine-dependent mice

Low-dose naloxone-precipitated withdrawal hyperalgesia is a reliable indicator of physical dependence after chronic morphine treatment. A remarkably similar long-lasting (> 3–4 h) hyperalgesia is evoked by injection of a low dose of naloxone (10 μg/kg, s.c.) in naïve mice after acute pretreatment with the glycolipid, GM1 ganglioside (1 mg/kg) (measured by warm-water-immersion tail-flick assays). GM1 treatment markedly increases the efficacy of excitatory Gs-coupled opioid receptor signaling in nociceptive neurons. Co-treatment with an ultra-low-dose (0.1 ng/kg, s.c.) of the broad-spectrum opioid receptor antagonist, naltrexone or the selective kappa opioid receptor antagonist, nor-binaltorphimine, blocks naloxone-evoked hyperalgesia in GM1-pretreated naïve mice and unmasks prominent, long-lasting (> 4 h) inhibitory opioid receptor-mediated analgesia. This unmasked analgesia can be rapidly blocked by injection after 1–2 h of a high dose of naltrexone (10 mg/kg) or nor-binaltorphimine (0.1 mg/kg). Because no exogenous opioid is administered to GM1-treated mice, we suggest that naloxone may evoke hyperalgesia by inducing release of endogenous bimodally acting opioid agonists from neurons in nociceptive networks by antagonizing putative presynaptic inhibitory opioid autoreceptors that “gate” the release of endogenous opioids. In the absence of exogenous opioids, the specific pharmacological manipulations utilized in our tail-flick assays on GM1-treated mice provide a novel bioassay to detect the release of endogenous bimodally acting (excitatory/inhibitory) opioid agonists. Because mu excitatory opioid receptor signaling is blocked by ultra-low doses of naloxone, the higher doses of naloxone that evoke hyperalgesia in GM1-treated mice cannot be mediated by activation of mu opioid receptors. Co-treatment with ultra-low-dose naltrexone or nor-binaltorphimine may selectively block signaling by endogenous GM1-sensitized excitatory kappa opioid receptors, unmasking inhibitory kappa opioid receptor signaling, and converting endogenous opioid receptor-mediated hyperalgesia to analgesia. Co-treatment with kelatorphan stabilizes putative endogenous opioid peptide agonists released by naloxone in GM1-treated mice, so that analgesia is evoked rather than hyperalgesia. Acute treatment of chronic morphine-dependent mice with ultra-low-dose naltrexone (0.1 ng/kg) results in remarkably similar rapid blocking of naloxone (10 μg/kg)-precipitated withdrawal hyperalgesia and unmasking of prominent opioid analgesia. These studies may clarify complex mechanisms underlying opioid physical dependence and opioid addiction.