Use of a shuttle box model to assess the behavioral and analgesic effects of opioid injections in goldfish (Carassius auratus).

OBJECTIVE: To evaluate if opioid-induced behavioral effects, such as sedation, can be detected using a shuttle box experimental apparatus and whether thermal preference following noxious stimulation using mustard oil is reversed by morphine administration in fish. METHODS: 5 goldfish (Carassius auratus) underwent 2 randomized blinded experimental trials, with a crossover study design. First, opioid effects were tested in a shuttle box without painful stimulus. Fish were injected 5 days apart with butorphanol at 0.4 or 10 mg/kg, morphine at 5 or 10 mg/kg, or saline IM. After 30 minutes, each fish was placed in a shuttle box for 2 hours with a temperature gradient of 26 to 28 °C. Temperature preference, time spent immobile, and swimming velocity were assessed. The second trial consisted of cutaneous noxious stimulation using mustard oil immersion for 5 minutes followed by an assessment of thermal preference for 4 minutes in the shuttle box after either morphine at 10 mg/kg or saline IM injections. Linear mixed models were used to compare treatment groups. RESULTS: Before noxious stimulation, a low dose of morphine caused sedation compared with control group and high-dose morphine and butorphanol treatments. Immersion in mustard oil caused fish to spend more time in the cold area in the control group. Morphine administration reversed this pattern. CONCLUSIONS: The sedative and analgesic effects of opioids were detected through this model. CLINICAL RELEVANCE: The shuttle box model could be used to assess the analgesic effects of other opioids in goldfish while reducing biases associated with the sedative and stimulatory effects of these drugs.

Morphine promotes non-small cell lung cancer progression by downregulating E-cadherin via the PI3K/AKT/mTOR pathway.

Morphine has been suggested to affect cancer cell dynamics and decrease survival rates in lung cancer patients at specific doses, but the precise mechanisms poorly understood. In this study, we aimed to investigate the molecular mechanisms by which morphine modulates the malignant characteristics of non-small cell lung cancer. Cell proliferation was assessed via the Cell Counting Kit-8 assay, and cell migration and invasion were examined via wound healing and Transwell assays. We employed immunofluorescence staining to evaluate E-cadherin expression in A549 and Lewis lung cancer (LLC) cell lines and immunohistochemistry to evaluate E-cadherin expression in nude mice tumours. Additionally, the in vivo effects of morphine on lung cancer progression were explored in a xenograft tumour experiments, in which naloxone was used as a morphine antagonist. Western blot analysis was performed to detect E-cadherin, phosphorylated mTOR (p-mTOR), mTOR, phosphorylated AKT (p-AKT), AKT, phosphorylated PI3K (p-PI3K), and PI3K protein levels in A549 and LLC cells as well as in tumour samples. Morphine (10 µM) significantly increased the proliferation of A549 and LLC cells in vitro (p < 0.05). It also enhanced the migratory and invasive capacities of these cell lines (p < 0.01). Mechanistically, morphine treatment (10 µM) led to a reduction in the expression of E-cadherin, and an increase in the phosphorylation of PI3K, AKT, and mTOR in A549 and LLC cells (p < 0.01). Morphine treatment (1.5 mg/kg) also reduced E-cadherin expression in xenograft tumours and promoted tumour growth in vivo (p < 0.05). This effect was reversed by naloxone (0.1 mg/kg). The results demonstrated that morphine stimulates the malignant proliferation of A549 and LLC cell lines and promotes xenograft tumour growth. Perhaps by specifically targeting MOR, morphine triggers a signalling cascade that activates the PI3K/AKT/mTOR pathway while inhibiting the EMT marker E-cadherin, which may consequently promote the progression of lung cancer.

The cell-permeant antioxidant D-thiol ester D-cysteine ethyl ester overcomes physical dependence to morphine in male Sprague Dawley rats.

The ability of morphine to decrease cysteine transport into neurons by inhibition of excitatory amino acid transporter 3 (EAA3) may be a key molecular mechanism underlying the acquisition of physical and psychological dependence to morphine. This study examined whether co-administration of the cell-penetrant antioxidant D-thiol ester, D-cysteine ethyl ester (D-CYSee), with morphine, would diminish the development of physical dependence to morphine in male Sprague Dawley rats. Systemic administration of the opioid receptor antagonist, naloxone (NLX), elicited pronounced withdrawal signs (e.g., wet-dog shakes, jumps, rears, circling) in rats that received a subcutaneous depot of morphine (150 mg/kg, SC) for 36 h and continuous intravenous infusion of vehicle (20 µL/h, IV). The NLX-precipitated withdrawal signs were reduced in rats that received an infusion of D-CYSee, but not D-cysteine, (both at 20.8 µmol/kg/h, IV) for the full 36 h. NLX elicited pronounced withdrawal signs in rats treated for 48 h with morphine (150 mg/kg, SC), plus continuous infusion of vehicle (20 µL/h, IV) that began at the 36 h timepoint of morphine treatment. The NLX-precipitated withdrawal signs were reduced in rats that received a 12 h infusion of D-CYSee, but not D-cysteine, (both at 20.8 µmol/kg/h, IV) that began at the 36 h timepoint of morphine treatment. These findings suggest that D-CYSee may attenuate the development of physical dependence to morphine and reverse established dependence to the opioid in male Sprague Dawley rats. Alternatively, D-CYSee may simply suppress the processes responsible for NLX-precipitated withdrawal. Nonetheless, D-CYSee and analogues may be novel therapeutics for the treatment of opioid use disorders.

Neuraxial Analgesia for Scoliosis Correction: A Case Series in a Specialised Children's Centre.

BACKGROUND: Surgical correction of spinal deformities in children presents a challenge to the anaesthetist due to the extensive nature of the surgery, the co-morbidities of the patients and the constraints on aesthetic techniques of intraoperative neurophysiological monitoring of the spinal cord. Patients undergoing scoliosis surgery are considered to suffer severe pain, which may lead to a negative impact on patient psychology and physical well-being. By using effective postoperative pain regimens to enhance recovery after surgery, pain can be significantly reduced, leading to patient satisfaction, facilitating early mobilisation, promoting oral intake, lowering postoperative pain and shortening the length of hospital stay. Thus, the primary objectives of this study were to assess the postoperative pain management and first rescue analgesic medications, by using preservative-free morphine 50 mcg/kg and dexmedetomidine 4 mcg along with normal saline 0.5 ml kg caudally, as well as to look for the secondary objectives side effects, including respiratory depression, nausea, vomiting and pruritis, timing of postoperative ambulation and length of hospital stay.  Methods: In this study, we retrospectively included children under 14 years of age and above three years who underwent scoliosis surgery under a caudal epidural-general anaesthesia using caudal morphine and the dexmedetomidine technique in the period from January to May 2023 at the National Guard Health Affairs Hospital (NGHA), Riyadh. We collected the data of seven cases using the electronic chart system of the Best Care database to extract the specific cases that meet the inclusion criteria for the study, which are idiopathic scoliosis patients, aged 3-14 years, and primary correction procedures.  Results: The pain-free duration was between seven hours and 48 hours (about two days). There were four female cases (57.14%), and three cases were male (42.85%). The patients' American Society of Anesthesiologists (ASA) status were II to III. In all the cases, there was no documentation of any episodes of postoperative nausea and vomiting (PONV), respiratory distress or pruritus, except for one case where the patient had an episode of PONV.  Conclusion: In this study, our aim was to present our experience with dexmedetomidine as an efficient medication when coadministered with morphine to be used in the operating room. We found a high level of reliability in prolonging analgesia time and delaying the usage of rescue medication. We encourage more studies on caudal dexmedetomidine for patients undergoing scoliosis surgery.

Impact of Morphine withdrawal on genes related to the TLR2 pathway expressed in the Prefrontal Cortex.

Millions of people suffer from opioid use disorder, because of the ongoing opioid epidemic. The aversive symptoms of withdrawal are a leading factor for drug relapses, yet there are limited therapeutic options to minimize or prevent withdrawal symptoms. The mechanism behind opioid withdrawal is still not fully understood, thus preventing the development of new therapeutics. This study is an extension of our previously proposed mechanism of a toll-like receptor 2 (TLR2) mediated withdrawal response as a result of morphine induced microbial change that occurs during morphine withdrawal. Transcriptome analysis of the pre-frontal cortex indicated that there was increased expression of genes related to TLR2 signaling in morphine withdrawal treated animals compared to placebo controls. Antibiotic treatment further altered TLR2 related genes, recovering some of the morphine induced effect and leading to additional suppression of some genes related to the TLR2 pathway. Morphine withdrawal induced gene expression was attenuated in a whole body TLR2 knockout model. These results provide more support that TLR2 plays an integral role in morphine withdrawal mechanisms and could be a potential therapeutic target to minimize opioid withdrawal associated co-morbidities.

Morphine induces inflammatory responses via both TLR4 and cGAS-STING signaling pathways.

BACKGROUND: Opioid activation of the microglia or macrophage Toll-like receptor 4 (TLR4) and associated inflammatory cytokine release are implicated in opioid-induced hyperalgesia and tolerance. The cyclic GMP-AMP synthase/stimulator of interferon genes (cGAS-STING) signaling pathway, activated by double-stranded DNA including mitochondrial DNA (mtDNA), has emerged as another key mediator of inflammatory responses. This study tested the hypothesis that morphine induces immune inflammatory responses in microglia and macrophages involving TLR4 and cGAS-STING pathway. METHODS: BV2 microglia and Raw 264.7 (Raw) macrophage cells were exposed to morphine with and without a STING inhibitor (C176) for 6 h or TLR 4 inhibitor (TAK242) for 24 h. Western blotting and RT-qPCR analyses assessed TLR4, cGAS, STING, nuclear factor-kappa B (NF-κB), and pro-inflammatory cytokine expression. Morphine-induced mitochondria dysfunction was quantified by reactive oxygen species (ROS) release using MitoSOX, mtDNA release by immunofluorescence, and RT-qPCR. Polarization of BV2 and Raw cells was assessed by inducible nitric oxide (iNOS) and CD86 expression. The role of mtDNA on morphine-related inflammation was investigated by mtDNA depletion of the cells with ethidium bromide (EtBr) or cell transfection of mtDNA extracted from morphine-treated cells. RESULTS: Morphine significantly increased the expression of TLR4, cGAS, STING, p65 NF-κB, and cytokines (IL-6 and TNF-α) in BV2 and Raw cells. Morphine-induced mitochondrial dysfunction by increased ROS and mtDNA release; the increased iNOS and CD86 evidenced inflammatory M1-like phenotype polarization. TLR4 and STING inhibitors reduced morphine-induced cytokine release in both cell types. The transfection of mtDNA activated inflammatory signaling proteins, cytokine release, and polarization. Conversely, mtDNA depletion led to the reversal of these effects. CONCLUSION: Morphine activates the cGAS-STING pathway in macrophage cell types. Inhibition of the STING pathway can be an additional method to overcome immune cell inflammation-related morphine tolerance and opioid-induced hyperalgesia.

N-palmitoylethanolamide attenuates negative emotions induced by morphine withdrawal in mice.

Depression and anxiety are prominent symptoms of withdrawal syndrome, often caused by the abuse of addictive drugs like morphine. N-palmitoylethanolamide (PEA), a biologically active lipid, is utilized as an anti-inflammatory and analgesic medication. Recent studies have highlighted PEA's role in mitigating cognitive decline and easing depression resulting from chronic pain. However, it remains unknown whether PEA can influence negative emotions triggered by morphine withdrawal. This study seeks to explore the impact of PEA on such emotions and investigate the underlying mechanisms. Mice subjected to morphine treatment underwent a 10-day withdrawal period, followed by assessments of the effect of PEA on anxiety- and depression-like behaviors using various tests. Enzyme-linked immunosorbent assay was conducted to measure levels of monoamine neurotransmitters in specific brain regions. The findings indicate that PEA mitigated anxiety and depression symptoms and reduced 5-hydroxytryptamine, noradrenaline, and dopamine levels in the hippocampus and prefrontal cortex. In summary, PEA demonstrates a significant positive effect on negative emotions associated with morphine withdrawal, accompanied with the reduction in levels of monoamine neurotransmitters in key brain regions. These insights could be valuable for managing negative emotions arising from morphine withdrawal.

Disrupted neuregulin 1-ErbB4 signaling: Consequences of prenatal morphine exposure in rat pups and molecular gateway to neurological impairment.

Objective: Morphine exposure during pregnancy has detrimental effects on both the mother and her offspring, both during and after childbirth. This study aimed to investigate the impact of prenatal morphine exposure on rat pups and dams, specifically focusing on changes in Neuregulin-1 (Nrg-1)/ErbB4 gene expression, inflammation, and brain-derived neurotrophic factor (BDNF) levels. Materials and methods: Twenty female rats were randomized into two experimental groups:1-Morphine Group: Dams received morphine throughout pregnancy. 2-Control Group: Dams received no interventions.At the end of gestation, blood samples were collected from the dams. Subsequently, dams and their pups underwent tissue collection from the cortical area of the brain to evaluate the following parameters: Interleukin-6 (IL-6), Interleukin-10 (IL-10), total antioxidant capacity (TAC), Malondialdehyde (MDA), and Brain-derived neurotrophic factor (BDNF).Additionally, RNA was extracted from the pup's cortical brain tissue for the assessment of gene expression levels of Neuregulin-1 (NRG-1) and ErbB-4 using quantitative real-time polymerase chain reaction (qrt-PCR). Results: The molecular investigation revealed a decrease in NRG-1 and ErbB-4 expressions in the brain cortex of offspring exposed to morphine during prenatal development. Additionally, the levels of IL-6 and IL-10 in both the serum and brain of both the mothers and their offspring in the morphine group were significantly higher compared to the control group. The morphine-exposed group also exhibited significantly lower levels of TAC and higher levels of MDA, indicating increased oxidative stress. Furthermore, the levels of BDNF in the morphine group were significantly lower compared to the control group. Conclusion: Prenatal morphine exposure in rats has detrimental effects on both the dams and their offspring. This study demonstrates that prenatal morphine exposure disrupts critical molecular pathways involved in neurodevelopment, inflammation, oxidative stress, and neurotrophic signaling. These findings suggest that prenatal morphine exposure can have long-lasting consequences for the offspring, potentially contributing to neurodevelopmental disorders and other health issues later in life.

Comparison of Intrathecal Morphine Versus Erector Spinae Plane Block for Perioperative Analgesia in Patients Undergoing Lumbar Spine Surgery: A Randomized Control Trial.

BACKGROUND: Lumbar spine surgery is associated with a significant degree of moderate-to-severe perioperative pain which can be alleviated by using different pain-relieving modalities; of these, erector spinae plane block and intrathecal morphine have promise. PURPOSE: To compare the analgesic efficacy between intrathecal morphine (ITM) versus erector spinae plane block (ESPB) for perioperative analgesia in patients undergoing lumbar spine surgery. METHOD: A total of 74 patients aged between 20 and 65 years of either sex were posted for elective lumbar spine surgery. Patients were divided into two groups: Group A patients received 0.3 mg intrathecal morphine and Group B received bilateral erector spinae plane block at L3 level by using 30 mL of 0.5% ropivacaine before starting the surgery for perioperative analgesia. In the perioperative period, pain was assessed by hemodynamic parameters (heart rate and mean arterial pressure), numeric rating scale (NRS), and patient satisfaction score.  Result: The difference in heart rate and mean arterial pressure was found to be statistically significant between groups at three, six, 12, and 24 hours (p<0.05). The patients who required rescue analgesia in Group A and Group B were 23 (62.2%) and 37 (100%) patients in the first 24 hours. The rate of complication was higher in Group A than in Group B (45% vs 5.4%). The patient satisfaction score was found to be better in Group A than in Group B. CONCLUSION: Intrathecal morphine provides more substantial and extended analgesia up to 48 hours postoperatively as compared to erector spinae plane block.

Immobilisation and Epidural Anaesthesia in a Eurasian Lynx (Lynx lynx) Undergoing Pelvic Limb Orthopaedic Surgery.

Immobilisation and anaesthesia of wild felids may be complex and potentially dangerous events, making it difficult to implement more advanced anaesthetic techniques such as neuraxial anaesthesia. A Eurasian lynx was referred for femur fracture repair after it was seen with lameness of the left pelvic limb sustained in its natural environment. The animal was remotely darted using a combination of ketamine (5 mg/kg) and xylazine (5 mg/kg) intramuscularly. Once immobilised, the lynx was transported to the veterinary hospital in a restraining cage. After induction and endotracheal intubation, pelvic limb radiographs confirmed a closed, comminuted fracture of the left femur that required open reduction and internal stabilisation. A sacrococcygeal epidural was performed before surgery using lidocaine (2 mg/kg) and morphine (0.1 mg/kg) to complement the ketamine-xylazine-isoflurane anaesthesia, which allows a low-end-tidal isoflurane concentration. Clinical signs were continuously monitored and remained stable during the entire procedure, with the exception of a temperature that decreased to 35.8°C. No intraoperative analgesic rescues were necessary. Recovery was smooth and uneventful. The lynx showed no signs of motor weakness after surgery or other side effects related to the anaesthetic procedure. The successful management of this surgical case suggests that the described anaesthetic protocol could be recommended in orthopaedic procedures of the pelvic limbs in wild Felidae.

Non-pulmonary complications of intrathecal morphine administration: a systematic review and meta-analysis with meta-regression.

BACKGROUND: Intrathecal morphine provides effective analgesia for a range of operations. However, widespread implementation into clinical practice is hampered by concerns for potential side-effects. We undertook a systematic review, meta-analysis, and meta-regression with the primary objective of determining whether a threshold dose for non-pulmonary complications could be defined and whether an association could be established between dose and complication rates when intrathecal morphine is administered for perioperative or obstetric analgesia. METHODS: We systematically searched the literature for randomised controlled trials comparing intrathecal morphine vs control in patients undergoing any type of surgery under general or spinal anaesthesia, or women in labour. Primary outcomes were rates of postoperative nausea and vomiting, pruritus, and urinary retention within the first 24 postoperative hours, analysed according to doses (1-100 µg; 101-200 µg; 201-500 µg; >500 µg), type of surgery, and anaesthetic strategy. Trials were excluded if doses were not specified. RESULTS: Our analysis included 168 trials with 9917 patients. The rates of postoperative nausea and vomiting, pruritus, and urinary retention were significantly increased in the intrathecal morphine group, with an odds ratio (95% confidence interval) of 1.52 (1.29-1.79), P<0.0001; 6.11 (5.25-7.10), P<0.0001; and 1.73 (1.17-2.56), P=0.005, respectively. Meta-regression could not establish an association between dose and rates of non-pulmonary complications. There was no subgroup difference according to surgery for any outcome. The quality of evidence was low (Grading of Recommendations Assessment, Development, and Evaluation [GRADE] system). CONCLUSIONS: Intrathecal morphine significantly increased postoperative nausea and vomiting, pruritus, and urinary retention after surgery or labour in a dose-independent manner. SYSTEMATIC REVIEW PROTOCOL: PROSPERO (CRD42023387838).

Opioid-environment interaction: Contrasting effects of morphine administered in a novel versus familiar environment on acute and repeated morphine induced behavioral effects and on acute morphine ERK activation in reward associated brain areas.

We report that environmental context can have a major impact on morphine locomotor behavior and ERK effects. We manipulated environmental context in terms of an environmental novelty/ familiarity dimension and measured morphine behavioral effects in both acute and chronic morphine treatment protocols. Wistar rats (n=7 per group) were injected with morphine 10 mg/kg or vehicle (s.c.), and immediately placed into an arena for 5 min, and locomotor activity was measured after one or 5 days. The morphine treatments were initiated either when the environment was novel or began after the rats had been familiarized with the arena by being given 5 daily nondrug tests in the arena. The results showed that acute and chronic morphine effects were strongly modified by whether the environment was novel or familiar. Acute morphine administered in a novel environment increased ERK activity more substantially in several brain areas, particularly in reward-associated areas such as the VTA in comparison to when morphine was given in a familiar environment. Repeated morphine treatments initiated in a novel environment induced a strong locomotor sensitization, whereas repeated morphine treatments initiated in a familiar environment did not induce a locomotor stimulant effect but rather a drug discriminative stimulus dis-habituation effect. The marked differential effects of environmental novelty/familiarity and ongoing dopamine activity on acute and chronic morphine treatments may be of potential clinical relevance for opioid drug addiction.

Negative allosteric modulation of CB1 cannabinoid receptor signalling decreases intravenous morphine self-administration and relapse in mice.

The endocannabinoid system interacts with the reward system to modulate responsiveness to natural reinforcers, as well as drugs of abuse. Previous preclinical studies suggested that direct blockade of CB1 cannabinoid receptors (CB1R) could be leveraged as a potential pharmacological approach to treat substance use disorder, but this strategy failed during clinical trials due to severe psychiatric side effects. Alternative strategies have emerged to circumvent the side effects of direct CB1 binding through the development of allosteric modulators. We hypothesized that negative allosteric modulation of CB1R signalling would reduce the reinforcing properties of morphine and decrease behaviours associated with opioid misuse. By employing intravenous self-administration in mice, we studied the effects of GAT358, a functionally-biased CB1R negative allosteric modulator (NAM), on morphine intake, relapse-like behaviour and motivation to work for morphine infusions. GAT358 reduced morphine infusion intake during the maintenance phase of morphine self-administration under a fixed ratio 1 schedule of reinforcement. GAT358 also decreased morphine-seeking behaviour after forced abstinence. Moreover, GAT358 dose dependently decreased the motivation to obtain morphine infusions under a progressive ratio schedule of reinforcement. Strikingly, GAT358 did not affect the motivation to work for food rewards in an identical progressive ratio task, suggesting that the effect of GAT358 in decreasing opioid self-administration was reward specific. Furthermore, GAT58 did not produce motor ataxia in the rotarod test. Our results suggest that CB1R NAMs reduced the reinforcing properties of morphine and could represent a viable therapeutic route to safely decrease misuse of opioids.

Association Between Low-Dose Ketamine After Periacetabular and/or Femoral Osteotomy and Postoperative Opioid Requirements.

Background: Previous studies have sought to determine the effect of inpatient ketamine therapy on postoperative pain in a variety of surgical specialties. Purpose: To determine the effects of postoperative ketamine analgesia after periacetabular osteotomy (PAO) and/or derotational femoral osteotomy (DFO) on opioid requirements, pain, and discharge time. Study Design: Cohort study; Level of evidence, 3. Methods: Prospective data were collected on 145 patients who underwent PAO and/or DFO by the senior author between January 2021 and December 2022. Hip arthroscopy was performed 3 to 10 days before addressing any intra-articular pathology. In 2021, patients (n = 91 procedures; control group) received a traditional postoperative multimodal pain regimen. In 2022, postoperative low-dose ketamine (0.1-1 mg/kg/h) was added to the multimodal analgesic approach until 24 hours before discharge (n = 81 procedures; ketamine group). The ketamine and control groups were matched based on procedure type. Total opioid consumption was collected using milligram morphine equivalents (MMEs) for both groups. Postoperative pain was measured using the Defense and Veterans Pain Rating Scale (DVPRS), which was analyzed as the mean score per day. Data on the mean MME and DVPRS were analyzed for up to 7 days postoperatively. Linear mixed statistical analysis was performed to determine the significance of low-dose postoperative ketamine on postoperative pain and opioid utilization. Results: Patients who did not receive ketamine after PAO and/or DFO utilized a mean of 181 ± 335 MMEs and had a mean DVPRS score of 4.18 ± 1.63. Patients who received postoperative ketamine required a mean of 119 ± 291 MMEs and had a mean DVPRS score of 4.34 ± 1.61. The ketamine group was found to consume a significantly lower total MME dose per day (P < .001). No significant difference was found in the mean DVPRS score between the ketamine and control groups (P = .42). Also, no significant difference was found on the day of discharge (P = .79). Conclusion: Patients who received postoperative ketamine after PAO and/or DFO had a significant decrease in MME dose when compared with a control group of patients who did not receive ketamine. Surgeons should consider adding ketamine to their postoperative multimodal pain control protocol to decrease opioid consumption while adequately addressing postoperative pain.

Simultaneous use of venlafaxine and calcium channel blockers on tolerance to morphine: The role of mitochondrial damage and oxidative stress in the brain.

BACKGROUND: One of the reasons for tolerance to morphine is increased oxidative stress and dysfunction of cell mitochondria in the hippocampus. Venlafaxine and calcium channel blockers can protect mitochondrial function. The investigation of the role of mitochondrial damage and oxidative stress in the simultaneous use of venlafaxine and calcium channel blockers on the acute analgesic effects of morphine and the induction of tolerance to its effects in mice was assessed. METHOD: In this experimental study, to induce tolerance to morphine, NMRI mice were treated with 50 mg/kg morphine for three consecutive days and 5 mg/kg morphine on the fourth day. Venlafaxine (20 mg/kg) alone or in combination with calcium channel blockers, nimodipine (10 mg/kg), and diltiazem (40 mg/kg) was administered 30 min before morphine, and the hot plate test was used. Then, hippocampal mitochondria were isolated by differential centrifugation method, and the levels of mitochondrial dehydrogenase activity, mitochondrial membrane potential, mitochondrial ROS production rate, as well as the content of glutathione and malondialdehyde in hippocampal mitochondria, were measured. RESULTS: The administration of venlafaxine-nimodipine and venlafaxine-diltiazem increased morphine's acute analgesic effects (P < 0.05) and reduced the induction and expression of tolerance to the analgesic effects of morphine (P < 0.05). Morphine significantly decreased MTT and GSH and increased MDA, mitochondrial membrane damage, and ROS compared to the control group (P < 0.01). Injection of venlafaxine-nimodipine and also venlafaxine-diltiazem 30 min before morphine can improve these alterations (P < 0.05). DISCUSSION AND CONCLUSION: Our data showed that the simultaneous use of venlafaxine with calcium channel blockers could increase the acute analgesic effects of morphine and reduce the induction and expression of tolerance to it. Also, the preventive and protective roles of simultaneous administration of venlafaxine and calcium channel blockers on morphine-induced mitochondrial oxidative stress and damage during the tolerance test were achieved.

Exploring the Role of Bergamot Polyphenols in Alleviating Morphine-Induced Hyperalgesia and Tolerance through Modulation of Mitochondrial SIRT3.

Morphine is an important pain reliever employed in pain management, its extended utilize is hindered by the onset of analgesic tolerance and oxidative stress. Long-term morphine administration causes elevated production of reactive oxygen species (ROS), disrupting mitochondrial function and inducing oxidation. Sirtuin 3 (SIRT3), a mitochondrial protein, is essential in modulating ROS levels by regulating mitochondrial antioxidant enzymes as manganese superoxide dismutase (MnSOD). Our investigation focused on the impact of SIRT3 on hyperalgesia and morphine tolerance in mice, as evaluating the antioxidant effect of the polyphenolic fraction of bergamot (BPF). Mice were administered morphine twice daily for four consecutive days (20 mg/kg). On the fifth day, mice received an acute dose of morphine (3 mg/kg), either alone or in conjunction with BPF or Mn (III)tetrakis (4-benzoic acid) porphyrin (MnTBAP). We evaluated levels of malondialdehyde (MDA), nitration, and the activity of SIRT3, MnSOD, glutamine synthetase (GS), and glutamate 1 transporter (GLT1) in the spinal cord. Our findings demonstrate that administering repeated doses of morphine led to the development of antinociceptive tolerance in mice, accompanied by increased superoxide production, nitration, and inactivation of mitochondrial SIRT3, MnSOD, GS, and GLT1. The combined administration of morphine with either BPF or MnTBAP prevented these effects.

Ketamine and major ketamine metabolites function as allosteric modulators of opioid receptors.

Ketamine is a glutamate receptor antagonist that was developed over 50 years ago as an anesthetic agent. At subanesthetic doses, ketamine and some metabolites are analgesics and fast-acting antidepressants, presumably through targets other than glutamate receptors. We tested ketamine and its metabolites for activity as allosteric modulators of opioid receptors expressed in recombinant receptors in heterologous systems and native receptors in rodent brain; signaling was examined by measuring GTP binding, b-arrestin recruitment, MAPK activation and neurotransmitter release. While micromolar concentrations of ketamine alone had weak agonist activity at mu opioid receptors, the combination of submicromolar concentrations of ketamine with endogenous opioid peptides produced robust synergistic responses with statistically significant increases in efficacies. All three opioid receptors (mu, delta, and kappa) showed synergism with submicromolar concentrations of ketamine and either Met-enkephalin, Leu-enkephalin, and/or dynorphin A17, albeit the extent of synergy was variable between receptors and peptides. S-ketamine exhibited higher modulatory effect compared to R-ketamine or racemic ketamine with nearly ~100% increase in efficacy. Importantly, the ketamine metabolite 6-hydroxynorketamine showed robust allosteric modulatory activity at mu opioid receptors; this metabolite is known to have analgesic and antidepressant activity but does not bind to glutamate receptors. Ketamine enhanced potency and efficacy of Met-enkephalin signaling both in mouse midbrain membranes and in rat ventral tegmental area neurons, as determined by electrophysiology recordings in brain slices. Taken together, these findings support the hypothesis that some of the therapeutic effects of ketamine and its metabolites are mediated by directly engaging the endogenous opioid system. Significance Statement We found that ketamine and its major biologically-active metabolites function as potent allosteric modulators of mu, delta, and kappa opioid receptors, with submicromolar concentrations of these compounds synergizing with endogenous opioid peptides such as enkephalin and dynorphin. This allosteric activity may contribute to ketamine's therapeutic effectiveness for treating acute and chronic pain and as a fast-acting antidepressant drug.

Effective Treatment of Methotrexate Induced Oral Mucositis With a Morphine Mouthwash Solution: A Case Report.

Introduction: Methotrexate (MTX) is a common medication used to treat rheumatoid arthritis (RA). MTX inhibits rapid cell turnover throughout the body which can lead to significant side effects. Patients who present with oral lesions may have suffered severe acute toxicity from MTX. Supportive pain treatment includes magic mouthwash solution and/or oral viscous lidocaine to manage pain and allow for healing. We report a case of MTX induced oral mucositis that did not respond to magic mouthwash but did improve with a morphine mouthwash solution. Case: A 67-year-old female with RA presented with worsening oral lesions over 2 weeks. She reported non-compliance with folic acid for 2 weeks while on MTX. Physical exam revealed ulcerating oral lesions on the mucous membranes consistent with mucositis. Pain treatment was initiated with magic mouthwash, but her pain was not well controlled after 24 hours, and still unable to swallow. An oral morphine mouthwash solution was initiated, and patient reported improved pain control over the next 48 hours. She was on the morphine mouthwash for 6 days during which improvement in the lesions was noted. Discussion: Pain management is imperative for oral mucositis. When traditional therapies do not provide adequate control, morphine mouthwash can be considered. It is a safer alternative to systemic opioids and topical opioids may influence cell proliferation and migration, which can positively impact healing of oral lesions. Conclusion: A morphine mouthwash solution can provide effective pain management for oral mucositis lesions in patients who do not respond adequately to magic mouthwash.

A novel intervention of molecular hydrogen on the unbalance of the gut microbiome in opioid addiction: Experimental and human studies.

The gut-brain axis mediates the interaction pathway between microbiota and opioid addiction. In recent years, many studies have shown that molecular hydrogen has therapeutic and preventive effects on various diseases. This study aimed to investigate whether molecular hydrogen could serve as pharmacological intervention agent to reduce risks of reinstatement of opioid seeking and explore the mechanism of gut microbiota base on animal experiments and human studies. Morphine-induced conditioned place preference (CPP) was constructed to establish acquisition, extinction, and reinstatement stage, and the potential impact of H2 on the behaviors related to morphine-induced drug extinction was determined using both free accessible and confined CPP extinction paradigms. The effects of morphine on microbial diversity and composition of microbiota, as well as the subsequent changes after H2 intervention, were assessed using 16 S rRNA gene sequencing. Short-Chain Fatty Acids (SCFAs) in mice serum were detected by gas chromatography-mass spectrometry (GC-MS). Meanwhile, we also conducted molecular hydrogen intervention and gut microbiota testing in opioid-addicted individuals. Our results revealed that molecular hydrogen could enhance the extinction of morphine-related behavior, reducing morphine reinstatement. Gut microbes may be a potential mechanism behind the therapeutic effects of molecular hydrogen on morphine addiction. Additionally, molecular hydrogen improved symptoms of depression and anxiety, as well as gut microbial features, in individuals with opioid addiction. This study supports molecular hydrogen as a novel and effective intervention for morphine-induced addiction and reveals the mechanism of gut microbiota.