The role of carbon monoxide, heme oxygenase 1, and the Nrf2 transcription factor in the modulation of chronic pain and their interactions with opioids and cannabinoids.

Chronic pain and its associated comorbidities are difficult to treat, even when the most potent analgesic compounds are used. Thus, research on new strategies to effectively relieve nociceptive and/or emotional disorders accompanying chronic pain is essential. Several studies have demonstrated the anti-inflammatory and antinociceptive effects of different carbon monoxide-releasing molecules (CO-RMs), inducible heme oxygenase 1 (HO-1), and nuclear factor-2 erythroid factor-2 (Nrf2) transcription factor activators in several models of acute and chronic pain caused by inflammation, nerve injury or diabetes. More recently, the antidepressant and/or anxiolytic effects of several Nrf2 transcription factor inducers were demonstrated in a model of chronic neuropathic pain. These effects are mainly produced by inhibition of oxidative stress, inflammation, glial activation, mitogen-activated protein kinases and/or phosphoinositide 3-kinase/phospho-protein kinase B phosphorylation in the peripheral and/or central nervous system. Other studies also demonstrated that the analgesic effects of opioids and cannabinoids are improved when these drugs are coadministered with CO-RMs, HO-1 or Nrf2 activators in different preclinical pain models and that these improvements are generally mediated by upregulation or prevention of the downregulation of µ-opioid receptors, δ-opioid receptors and/or cannabinoid 2 receptors in the setting of chronic pain. We reviewed all these studies as well as studies on the mechanisms of action underlying the effects of CO-RMs, HO-1, and Nrf2 activators in chronic pain. In summary, activation of the Nrf2/HO-1/carbon monoxide signaling pathway alone and/or in combination with the administration of specific analgesics is a valid strategy for the treatment of chronic pain and some associated emotional disorders.

Treatment with 5-fluoro-2-oxindole Increases the Antinociceptive Effects of Morphine and Inhibits Neuropathic Pain.

The efficacy of µ-opioid receptors (MOR) in neuropathic pain is low and with numerous side effects that limited their use. Chronic neuropathic pain is also linked with emotional disorders that aggravate the sensation of pain and which treatment has not been resolved. This study investigates whether the administration of an oxindole, 5-fluoro-2-oxindole, could inhibit the nociceptive and emotional behaviors and increase the effectiveness of morphine via modulating the microglia and activating the nuclear factor erythroid-2 related factor 2 (Nrf2) signaling pathway and MOR expression. In C57BL/6 mice with neuropathic pain provoked by the total constriction of sciatic nerve we studied the effects of 10 mg/kg 5-fluoro-2-oxindole in: (i) the allodynia and hyperalgesia caused by the injury; (ii) the anxiety- and depressive-like behaviors; (iii) the local antinociceptive actions of morphine; (iv) the expression of CD11b/c (a microglial marker), the antioxidant and detoxificant enzymes Nrf2, heme oxygenase 1 (HO-1) and NAD(P)H:quinone oxidoreductase-1 (NQO1), and of MOR in the spinal cord and hippocampus. Results showed that the inhibition of the main nociceptive symptoms and the anxiety- and depressive-like behaviors induced by 5-fluoro-2-oxindole were accompanied with the suppression of microglial activation and the activation of Nrf2/HO-1/NQO1 signaling pathway in the spinal cord and/or hippocampus. This treatment also potentiated the pain-relieving activities of morphine by normalizing the reduced MOR expression. This work demonstrates the antinociceptive, anxiolytic and antidepressant effects of 5-fluoro-2-oxindole, suggests a new strategy to enhance the antinociceptive actions of morphine and proposes a new mechanism of action of oxindoles during chronic neuropathic pain.

The Effects of Cobalt Protoporphyrin IX and Tricarbonyldichlororuthenium (II) Dimer Treatments and Its Interaction with Nitric Oxide in the Locus Coeruleus of Mice with Peripheral Inflammation.

Heme oxygenase 1 (HO-1) and carbon monoxide were shown to normalize oxidative stress and inflammatory reactions induced by neuropathic pain in the central nervous system, but their effects in the locus coeruleus (LC) of animals with peripheral inflammation and their interaction with nitric oxide are unknown. In wild-type (WT) and knockout mice for neuronal (NOS1-KO) or inducible (NOS2-KO) nitric oxide synthases with inflammatory pain induced by complete Freund's adjuvant (CFA), we assessed: 1) antinociceptive actions of cobalt protoporphyrin IX (CoPP), an HO-1 inducer; 2) effects of CoPP and tricarbonyldichlororuthenium(II)dimer (CORM-2), a carbon monoxide-liberating compound, on the expression of HO-1, NOS1, NOS2, CD11b/c, GFAP,and mitogen-activated protein kinases (MAPK)in the LC. CoPP reduced inflammatory pain in different time-dependent manners in WT and KO mice. Peripheral inflammation activated astroglia in the LC of all genotypes and increased the levels of NOS1 and phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK 1/2) in WT mice. CoPP and CORM-2 enhanced HO-1 and inhibited astroglial activationin all genotypes. Both treatments blocked NOS1 overexpression,and CoPP normalized ERK 1/2 activation. This study reveals an interaction between HO-1 and NOS1/NOS2 during peripheral inflammation andshows that CoPP and CORM-2 improved HO-1 expression and modulated the inflammatory and/or plasticity changes caused by peripheral inflammation in the LC.

Treatment with Sulforaphane Produces Antinociception and Improves Morphine Effects during Inflammatory Pain in Mice.

The activation of nuclear factor erythroid 2-related factor 2 (Nrf2) exerts potent antioxidative and anti-inflammatory effects; however, its participation in the modulation of chronic inflammatory pain and on the antinociceptive effects of µ-opioid receptor (MOR) agonists has not been evaluated. We investigated whether the induction of Nrf2 could alleviate chronic inflammatory pain and augment the analgesic effects of morphine and mechanisms implicated. In male C57BL/6 mice with inflammatory pain induced by complete Freund's adjuvant (CFA) subplantarly administered, we assessed: 1) antinociceptive actions of the administration of 5 and 10 mg/kg of a Nrf2 activator, sulforaphane (SFN); and 2) effects of SFN on the antinociceptive actions of morphine and on protein levels of Nrf2, heme oxygenase 1 (HO-1), and NAD(P)H: quinone oxidoreductase 1 (NQO1) enzymes, microglial activation and inducible nitric oxide synthase (NOS2) overexpression, as well as on mitogen-activated protein kinase (MAPK) and MOR expression in the spinal cord and paw of animals with inflammatory pain. Results showed that treatment with SFN inhibited allodynia and hyperalgesia induced by CFA and increased the local antinociceptive actions of morphine. This treatment also augmented the expression of Nrf2, HO-1, NQO1, and MOR, and inhibited NOS2 and CD11b/c overexpression and MAPK phosphorylation induced by inflammation. Thus, this study shows that the induction of Nrf2 might inhibit inflammatory pain and enhance the analgesic effects of morphine by inhibiting oxidative stress and inflammatory responses induced by peripheral inflammation. This study suggests the administration of SFN alone and in combination with morphine are potential new ways of treating chronic inflammatory pain.

The Inhibitory Effects of Cobalt Protoporphyrin IX and Cannabinoid 2 Receptor Agonists in Type 2 Diabetic Mice.

The activation of the transcription factor Nrf2 inhibits neuropathy and modulates the activity of delta-opioid receptors (DOR) in type 2 diabetic mice but the impact of Nrf2/HO-1 pathway on the antinociceptive actions of cannabinoid 2 receptors (CB2R) has not been assessed. Using male mice BKS.Cg-m+/+Leprdb/J (db/db) we investigated if treatment with cobalt protoporphyrin IX (CoPP), an HO-1 inductor, inhibited mechanical allodynia, hyperglycemia and obesity associated to type 2 diabetes. The antinociceptive effects of JWH-015 and JWH-133 (CB2R agonists) administered with and without CoPP or sulforaphane (SFN), a Nrf2 transcription factor activator, have been also evaluated. The expression of Nrf2, HO-1, NAD(P)H: quinone oxidoreductase 1 (NQO1) and c-Jun N-terminal kinase (JNK) in sciatic nerve and that of the CB2R on the dorsal root ganglia from animals treated with CoPP and/or SFN were assessed. CoPP treatment inhibited allodynia, hyperglycemia and body weight gain in db/db mice by enhancing HO-1/NQO1 levels and reducing JNK phosphorylation. Both CoPP and SFN improved the antiallodynic effects of JWH-015 and JWH-133 and expression of CB2R in db/db mice. Therefore, we concluded that the activation of antioxidant Nrf2/HO-1 pathway potentiate the effects of CB2R agonists and might be suitable for the treatment of painful neuropathy linked to type 2 diabetes.

Treatment with a heme oxygenase 1 inducer enhances the antinociceptive effects of µ-opioid, δ-opioid, and cannabinoid 2 receptors during inflammatory pain.

The administration of µ-opioid receptor (MOR), δ-opioid receptor (DOR), and cannabinoid 2 receptor (CB2R) agonists attenuates inflammatory pain. We investigated whether treatment with the heme oxygenase 1 (HO-1) inducer, cobalt protoporphyrin IX (CoPP), could modulate the local effects and expression of MOR, DOR, or CB2R during chronic inflammatory pain. In mice with inflammatory pain induced by the subplantar administration of complete Freund's adjuvant, we evaluated the effects of the intraperitoneal administration of 10 mg/kg CoPP on the antiallodynic and antihyperalgesic actions of locally administered MOR (morphine), DOR (DPDPE {[d-Pen(2),d-Pen(5)]-enkephalin}), or CB2R [JWH-015 {(2-methyl-1-propyl-1H-indol-3-yl)-1-naphthalenylmethanone}] agonists and its reversion with the HO-1 inhibitor, tin protoporphyrin IX (SnPP). The effect of CoPP treatment on the dorsal root ganglia expression of HO-1, MOR, DOR, and CB2R was also assessed. The results show that treatment with CoPP increased the local antinociceptive effects produced by morphine, DPDPE, or JWH-015 during chronic inflammatory pain, and these effects were blocked by the subplantar administration of SnPP, indicating the participation of HO-1 in the antinociceptive actions. CoPP treatment, apart from inducing the expression of HO-1, also enhanced the expression of MOR, did not alter CB2R, and avoided the decreased expression of DOR induced by inflammatory pain. This study shows that the HO-1 inducer (CoPP) increased the local antinociceptive effects of MOR, DOR, and CB2R agonists during inflammatory pain by altering the peripheral expression of MOR and DOR. Therefore, the coadministration of CoPP with local morphine, DPDPE, or JWH-015 may be a good strategy for the management of chronic inflammatory pain.

The Combination of Molecular Hydrogen and Heme Oxygenase 1 Effectively Inhibits Neuropathy Caused by Paclitaxel in Mice.

Chemotherapy-provoked peripheral neuropathy and its associated affective disorders are important adverse effects in cancer patients, and its treatment is not completely resolved. A recent study reveals a positive interaction between molecular hydrogen (H2) and a heme oxygenase (HO-1) enzyme inducer, cobalt protoporphyrin IX (CoPP), in the inhibition of neuropathic pain provoked by nerve injury. Nevertheless, the efficacy of CoPP co-administered with hydrogen-rich water (HRW) on the allodynia and emotional disorders related to paclitaxel (PTX) administration has not yet been assessed. Using male C57BL/6 mice injected with PTX, we examined the effects of the co-administration of low doses of CoPP and HRW on mechanical and thermal allodynia and anxiodepressive-like behaviors triggered by PTX. Moreover, the impact of this combined treatment on the oxidative stress and inflammation caused by PTX in the amygdala (AMG) and dorsal root ganglia (DRG) were studied. Our results indicated that the antiallodynic actions of the co-administration of CoPP plus HRW are more rapid and higher than those given by each of them when independently administered. This combination inhibited anxiodepressive-like behaviors, the up-regulation of the inflammasome NLRP3 and 4-hydroxynonenal, as well as the high mRNA levels of some inflammatory mediators. This combination also increased the expression of NRF2, HO-1, superoxide dismutase 1, glutathione S-transferase mu 1, and/or the glutamate-cysteine ligase modifier subunit and decreased the protein levels of BACH1 in the DRG and/or AMG. Thus, it shows a positive interaction among HO-1 and H2 systems in controlling PTX-induced neuropathy by modulating inflammation and activating the antioxidant system. This study recommends the co-administration of CoPP plus HRW as an effective treatment for PTX-provoked neuropathy and its linked emotive deficits.

Treatment with carbon monoxide-releasing molecules and an HO-1 inducer enhances the effects and expression of µ-opioid receptors during neuropathic pain.

BACKGROUND: The administration of µ-opioid receptors (MOR) and δ-opioid receptors (DOR) as well as cannabinoid-2 receptor (CB2R) agonists attenuates neuropathic pain. We investigated if treatment with two carbon monoxide-releasing molecules (CORM-2 and CORM-3) or an inducible heme oxygenase inducer (cobalt protoporphyrin IX, CoPP) could modulate the local and systemic effects and expression of MOR, DOR, and CB2R during neuropathic pain. METHODS: In C57BL/6 mice, at 10 days after the chronic constriction of sciatic nerve, we evaluated the effects of the intraperitoneal administration of 10 mg/kg of CORM-2, CORM-3, or CoPP on the antiallodynic and antihyperalgesic actions of a locally or systemically administered MOR (morphine), DOR ([d-Pen(2),d-Pen(5)]-enkephalin) or CB2R ((2-methyl-1-propyl-1H-indol-3-yl)-1-naphthalenylmethanone ) agonist. The effects of CORM-2 and CoPP treatments on the expression of MOR, DOR, CB2R, inducible and constitutive heme oxygenases, microglia activation marker (CD11b/c), and neuronal and inducible nitric oxide synthases were also assessed. RESULTS: Treatments with CO-RMs and CoPP reduced the mechanical and thermal hypersensitivity induced by sciatic nerve injury, increased the local, but not systemic, antinociceptive effects of morphine, and decreased those produced by DPDPE and JWH-015. Both CORM-2 and CoPP treatments enhanced MOR and inducible heme oxygenase expression, unaltered DOR and constitutive heme oxygenase expression, and decreased the overexpression of CB2R, CD11b/c, and neuronal and inducible nitric oxide synthases induced by sciatic nerve injury. CONCLUSIONS: This study shows that CO-RMs and CoPP treatments increase the local antinociceptive effects of morphine through enhancing MOR peripheral expression and inhibiting spinal microglial activation and overexpression of neuronal/inducible nitric oxide synthases.

A Novel Therapy for Cisplatin-Induced Allodynia and Dysfunctional and Emotional Impairments in Male and Female Mice.

Patients undergoing chemotherapy with cisplatin (CIS) develop neuropathy in addition to other symptoms such as, anxiety, depression, muscle wasting and body weight loss. This symptomatology greatly weakens patients and may even lead to adjournment of chemotherapy. The protecting actions of molecular hydrogen in many neurological illnesses have been described, but its effect on the functional and emotional deficiencies caused by CIS has not been assessed. In C57BL/6J male and female mice injected with CIS, we examined the impact of the prophylactic treatment with hydrogen-rich water (HRW) on: (i) the tactile and cold allodynia, (ii) the deficits of grip strength and weight loss, (iii) the anxiodepressive-like behaviors and (iv) the inflammatory and oxidative reactions incited by CIS in the dorsal root ganglia (DRG) and prefrontal cortex (PFC). The results demonstrate that the mechanical allodynia and the anxiodepressive-like comportment provoked by CIS were similarly manifested in both sexes, whereas the cold allodynia, grip strength deficits and body weight loss produced by this chemotherapeutic agent were greater in female mice. Nonetheless, the prophylactic treatment with HRW prevented the allodynia and the functional and emotional impairments resulting from CIS in both sexes. This treatment also inhibited the inflammatory and oxidative responses activated by CIS in the DRG and PFC in both sexes, which might explain the therapeutic actions of HRW in male and female mice. In conclusion, this study revealed the plausible use of HRW as a new therapy for the allodynia and physical and mental impairments linked with CIS and its possible mechanism of action.

Hydrogen Sulfide Interacting with Cannabinoid 2 Receptors during Sciatic Nerve Injury-Induced Neuropathic Pain.

Hydrogen sulfide (H2S) donors make opioids more effective in inhibiting nociception during inflammatory and neuropathic pain. We examined whether the analgesic, anxiolytic and/or antidepressant actions of the cannabinoid 2 receptor (CB2R) agonist, JWH-133, might be improved by pretreatment with H2S donors, DADS and GYY4137 in mice with sciatic nerve injury-provoked neuropathy (CCI). The reversion of the antinociceptive effects of these treatments with the CB2R antagonist, AM630, and the regulatory actions of H2S in the phosphorylation of NF-κB inhibitor alpha (IKBα) and in the brain-derived neurotrophic factor (BDNF), CB2R, Nrf2 and heme oxygenase 1 (HO-1) levels in prefrontal cortex (PFC), ventral hippocampus (vHIP) and periaqueductal gray matter (PAG), were examined. Data showed that the analgesic effects of JWH-133, systemically and locally administered, were improved by the DADS or GYY4137 pretreatment. The co-treatment of GYY4137 with JWH-133 also stopped anxiodepressive-like activities that concur with neuropathy. Our data likewise showed that both H2S donors normalized the inflammatory (p-IKBα), neurotrophic (BDNF) variations caused by CCI, increased the expression of CB2R and activated the Nrf2/HO-1 antioxidant pathway in PFC, v-HIP and/or PAG of animals with neuropathic pain. In addition, the blockade of the analgesia produced by high doses of DADS and GYY4137 with AM630 indicated the contribution of the endocannabinoid system in the effects of H2S during neuropathic pain, thus supporting the positive interaction between H2S and CB2R. Therefore, this study demonstrates the potential use of CB2R agonists combined with H2S donors as a possible treatment for peripheral nerve injury-caused neuropathic pain and the associated emotional disturbances.

The Impact of UFP-512 in Mice with Osteoarthritis Pain: The Role of Hydrogen Sulfide.

The pain-relieving properties of opioids in inflammatory and neuropathic pain are heightened by hydrogen sulfide (H2S). However, whether allodynia and functional and/or emotional impairments related to osteoarthritis (OA) could be reduced by activating δ-opioid receptors (DOR) and the plausible influence of H2S on these actions has not been completely established. In female C57BL/6J mice with OA pain generated via monosodium acetate (MIA), we analyze: (i) the effects of UFP-512 (a DOR agonist), given alone and co-administered with two H2S donors, on the symptoms of allodynia, loss of grip strength (GS), and anxiodepressive-like comportment; (ii) the reversion of UFP-512 actions with naltrindole (a DOR antagonist), and (iii) the impact of UFP-512 on the expression of phosphorylated NF-kB inhibitor alpha (p-IKBα) and the antioxidant enzymes superoxide dismutase 1 (SOD-1) and glutathione sulfur transferase M1 (GSTM1); and the effects of H2S on DOR levels in the dorsal root ganglia (DRG), amygdala (AMG), and hippocampus (HIP) of MIA-injected animals. Results showed that systemic and local administration of UFP-512 dose-dependently diminished the allodynia and loss of GS caused by MIA, whose effects were potentiated by H2S and reversed by naltrindole. UFP-512 also inhibited anxiodepressive-like behaviors, normalized the overexpression of p-IKBα in DRG and HIP, and enhanced the expression of SOD-1 and GSTM1 in DRG, HIP, and/or AMG. Moreover, the increased expression of DOR triggered by H2S might support the improved analgesic actions of UFP-512 co-administered with H2S donors. This study proposes the use of DOR agonists, alone or combined with H2S donors, as a new treatment for OA pain.

Novel Heme Oxygenase-1 Inducers Palliate Inflammatory Pain and Emotional Disorders by Regulating NLRP3 Inflammasome and Activating the Antioxidant Pathway.

Chronic pain caused by persistent inflammation is current in multiple diseases and has a strong negative impact on society. It is commonly associated with several mental illnesses, which can exert a negative influence on pain perception, and needs to be eradicated. Nevertheless, actual therapies are not sufficiently safe and effective. Recent reports demonstrate that the induction of heme oxygenase-1 (HO-1) enzyme produces analgesic effects in animals with osteoarthritis pain and reverses the grip strength loss caused by sciatic nerve crush. In this research, we evaluated the potential use of three new HO-1 inducers, 1m, 1a, and 1b, as well as dimethyl fumarate (DMF), for treating persistent inflammatory pain induced by the subplantar injection of complete Freud's adjuvant and the functional deficits and emotional sickness associated. The modulator role of these treatments on the inflammatory and antioxidant pathways were also assessed. Our findings revealed that repeated treatment, for four days, with 1m, 1a, 1b, or DMF inhibited inflammatory pain, reversed grip strength deficits, and reversed the linked anxious- and depressive-like behaviors, with 1m being the most effective. These treatments also suppressed the up-regulation of the inflammasome NLRP3 and activated the expression of the Nrf2 transcription factor and the HO-1 and superoxide dismutase 1 enzymes in the paw and/or amygdala, thus revealing the anti-inflammatory and antioxidant capacity of these compounds during inflammatory pain. Results suggest the use of 1m, 1a, 1b, and DMF, particularly 1m, as promising therapies for inflammatory pain and the accompanying functional disabilities and emotional diseases.

The Inhibition of Neuropathic Pain Incited by Nerve Injury and Accompanying Mood Disorders by New Heme Oxygenase-1 Inducers: Mechanisms Implicated.

Neuropathic pain is a type of pain that persists for a long time and becomes pathological. Additionally, the anxiodepressive disorders derived from neuropathic pain are difficult to palliate with the current treatments and need to be resolved. Then, using male mice with neuropathic pain provoked by chronic constriction of the sciatic nerve (CCI), we analyzed and compared the analgesic actions produced by three new heme oxygenase 1 (HO-1) inducers, 1m, 1b, and 1a, with those performed by dimethyl fumarate (DMF). Their impact on the anxiety- and depressive-like comportments and the expression of the inflammasome NLRP3, Nrf2, and some antioxidant enzymes in the dorsal root ganglia (DRG) and amygdala (AMG) were also investigated. Results revealed that the administration of 1m, 1b, and DMF given orally for four days inhibited the allodynia and hyperalgesia caused by CCI, while 1a merely reduced the mechanical allodynia. However, in the first two days of treatment, the antiallodynic effects produced by 1m were higher than those of 1a and DMF, and its antihyperalgesic actions were greater than those produced by 1b, 1a, and DMF, revealing that 1m was the most effective compound. At four days of treatment, all drugs exerted anxiolytic and antidepressant effects, decreased the NLRP3 levels, and increased/normalized the Nrf2, HO-1, and superoxide dismutase 1 levels in DRG and AMG. Data indicated that the dual modulation of the antioxidant and inflammatory pathways produced by these compounds, especially 1m, is a new promising therapeutic approach for neuropathic pain and related emotional illnesses.

New Treatment for the Cognitive and Emotional Deficits Linked with Paclitaxel-Induced Peripheral Neuropathy in Mice.

Chemotherapy-provoked peripheral neuropathy and its linked comorbidities severely reduce the quality of a patient's life. Its therapy is not completely resolved and has become an important clinical challenge. The protective actions of molecular hydrogen (H2) in many neurological disorders have been described, but its effects on memory and the emotional deficits accompanying neuropathic pain induced by chemotherapy remain unknown. In this study, using male mice injected with paclitaxel (PTX), we examined the effects of systemic treatment with hydrogen-rich water (HRW) in: (i) the mechanical and thermal allodynia provoked by PTX and the pathways involved; (ii) the memory deficits, anxiety- and depressive-like behaviors associated with PTX-induced peripheral neuropathy (PIPN); and (iii) the plasticity (p-extracellular signal-regulated protein kinase; p-ERK ½), nociceptive (p-protein kinase B, p-Akt), inflammatory (p-nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha; p-IKBα), and oxidative (4-hydroxynonenal: 4-HNE) alterations provoked by PIPN in the prefrontal cortex (PFC). The results revealed: (1) the antiallodynic actions of HRW administered at one or two times per day during 7 and 3 consecutive days; (2) the participation of Kv7 potassium channels and the Nrf2-heme oxygenase 1-NAD(P)H: quinone oxidoreductase 1 pathway in the painkiller effects of HRW; (3) the inhibition of memory deficits and the anxiodepressive-like behaviors related with PIPN induced by HRW; and (4) the normalization of p-ERK ½, p-Akt and 4-HNE up-regulation and the activation of antioxidant enzymes produced by this treatment in PFC. This study proposes HRW as a possible effective and safe therapy for PIPN and its associated cognitive and emotional deficits.

The Interaction between Carbon Monoxide and Hydrogen Sulfide during Chronic Joint Pain in Young Female Mice.

A relationship between carbon monoxide (CO) and hydrogen sulfide (H2S) has been described in different pathological conditions, but their interaction in modulating joint pain has not yet been investigated. In young female mice with monosodium acetate-induced joint degeneration and pain, we assessed: (1) the effects of CORM-2 (tricarbonyldichlororuthenium(II)dimer), a CO-releasing molecule, and CoPP (cobalt protoporphyrin IX), an inducer of heme oxygenase 1 (HO-1), administered alone and combined with low doses of two slow-releasing H2S donors, DADS (diallyl disulfide) and GYY4137 (morpholin-4-ium 4-methoxyphenyl(morpholino) phosphinodithioate dichloromethane complex) on the mechanical allodynia and loss of grip strength provoked by joint degeneration; (2) the role of Nrf2, NAD(P)H: quinone oxidoreductase 1 (NQO1) and HO-1 in the antinociceptive actions of H2S donors; (3) the impact of DADS and GYY4137 treatment on the expression of Nrf2 and several antioxidant proteins in dorsal root ganglia (DRG) and periaqueductal gray matter (PAG). Our data showed that treatment with H2S donors inhibited allodynia and functional deficits, while CORM-2 and CoPP only prevented allodynia. The Nrf2 pathway is implicated in the analgesic actions of DADS and GYY4137 during joint degeneration. Moreover, the co-administration of low doses of CORM-2 or CoPP with DADS or GYY4137 produced higher antiallodynic effects and greater recovery of grip strength deficits than those produced by each of these compounds alone. The activation of the antioxidant system caused by H2S donors in DRG and/or PAG might explain the enhancement of antinociceptive effects. These data reveal a positive interaction between H2S and CO in modulating joint pain in female mice.

Hydrogen Sulfide Increases the Analgesic Effects of µ- and δ-Opioid Receptors during Neuropathic Pain: Pathways Implicated.

Recent studies have revealed that hydrogen sulfide (H2S) increases the analgesic actions of the δ-opioid receptor (DOR) in inflammatory pain. However, the possible improvement of the analgesia of µ-opioid receptor (MOR) and DOR agonists during neuropathic pain, through pretreatment with two slow-releasing H2S donors-DADS (diallyl disulfide) and GYY4137 (morpholin-4-ium 4-methoxyphenyl(morpholino) phosphinodithioate dichloromethane complex)-is still unknown. In male C57BL/6J mice with neuropathic pain incited by chronic constriction of the sciatic nerve (CCI), we evaluated: (1) the influence of DADS (3.5 mg/kg) and GYY4137 (0.7 mg/kg) on the inhibition of the allodynia and hyperalgesia produced by the systemic or local administration of morphine (3 mg/kg or 65 µg) and UFP-512 (1 mg/kg or 12.5 µg); (2) the reversion of the antinociceptive actions of high doses of DADS (30 mg/kg) and GYY4137 (24 mg/kg) with MOR and DOR antagonists; and (3) the effects of H2S donors on oxidative stress, apoptotic responses, and MOR and DOR expression in the medial septum (MS) and dorsal root ganglia (DRG). The results revealed that both DADS and GYY4137 improved the antiallodynic effects of morphine and UFP-512, possibly by up-regulating MOR and DOR expression in DRG. The administration of MOR and DOR antagonists blocked the analgesic properties of DADS and GYY4137, revealing the feasible participation of the endogenous opioid system in H2S analgesic effects. Moreover, both H2S donors inhibited oxidative stress and apoptosis generated by CCI in the MS and/or DRG. This study suggests the co-treatment of H2S donors with MOR or DOR agonists as a potential therapy for neuropathic pain.

Effects of heme oxygenase 1 in the molecular changes and neuropathy associated with type 2 diabetes in mice.

Painful diabetic neuropathy is one of the most common complications of diabetes in humans. The current treatments are not completely effective, and the main mechanisms implicated in the development of diabetic neuropathy are not completely elucidated. Thus, in male db/db mice, a murine model of type 2 diabetes, we investigated the effects of treatment with a heme oxygenase 1 (HO-1) inducer, cobalt protoporphyrin IX (CoPP), on the 1) hyperglycemia and mechanical allodynia associated with type 2 diabetes and 2) molecular changes induced by diabetic neuropathy in the central nervous system (CNS). Thus, we evaluated the effects of CoPP on the protein levels of 4-HNE (oxidative stress), Nrf2, superoxide dismutase 1 (SOD1), NAD(P)H quinone oxidoreductase 1 (NQO1), HO-1, glutathione S-transferase Mu 1 (GSTM1) (antioxidant enzymes), phosphatidylinositol 3-kinase/protein kinase B (nociceptive pathway), CD11b/c (microglial activation), and BAX (apoptosis) in the amygdala and spinal cord of db/db mice. Our results showed the antihyperglycemic and antiallodynic effects of CoPP treatment as well as the potent antioxidant, antinociceptive, anti-inflammatory, and antiapoptotic properties of this HO-1 inducer in the CNS of type 2 diabetic mice. Treatment with CoPP also prevented the downregulation of several antioxidant proteins (Nrf2, SOD-1, and NQO1) and/or enhanced the protein levels of HO-1 and GSTM1 in the spinal cord and/or amygdala of db/db mice. These effects might be implicated in the antiallodynic actions of CoPP. Our findings revealed the modulatory effects of CoPP in the CNS of db/db mice and provide new prospects for novel type 2 diabetes-associated neuropathy therapies.

The Beneficial Effects of Heme Oxygenase 1 and Hydrogen Sulfide Activation in the Management of Neuropathic Pain, Anxiety- and Depressive-like Effects of Paclitaxel in Mice.

Chemotherapy-induced peripheral neuropathy constitutes an unresolved clinical problem that severely decreases the quality of the patient's life. It is characterized by somatosensory alterations, including chronic pain, and a high risk of suffering mental disorders such as depression and anxiety. Unfortunately, an effective treatment for this neuropathology is yet to be found. We investigated the therapeutic potential of cobalt protoporphyrin IX (CoPP), a heme oxygenase 1 inducer, and morpholin-4-ium 4-methoxyphenyl(morpholino) phosphinodithioate dichloromethane complex (GYY4137), a slow hydrogen sulfide (H2S) donor, in a preclinical model of paclitaxel (PTX)-induced peripheral neuropathy (PIPN) in mice. At three weeks after PTX injection, we evaluated the effects of the repetitive administration of 5 mg/kg of CoPP and 35 mg/kg of GYY4137 on PTX-induced nociceptive symptoms (mechanical and cold allodynia) and on the associated emotional disturbances (anxiety- and depressive-like behaviors). We also studied the mechanisms that could mediate their therapeutic properties by evaluating the expression of key proteins implicated in the development of nociception, oxidative stress, microglial activation, and apoptosis in prefrontal cortex (PFC) and dorsal root ganglia (DRG) of mice with PIPN. Results demonstrate that CoPP and GYY4137 treatments inhibited both the nociceptive symptomatology and the derived emotional alterations. These actions were mainly mediated through potentiation of antioxidant responses and inhibiting oxidative stress in the DRG and/or PFC of mice with PIPN. Both treatments normalized some plasticity changes and apoptotic reactions, and GYY4137 blocked microglial activation induced by PTX in PFC. In conclusion, this study proposes CoPP and GYY4137 as good candidates for treating neuropathic pain, anxiety- and depressive-like effects of PTX.

Hydrogen-Rich Water as a Novel Therapeutic Strategy for the Affective Disorders Linked with Chronic Neuropathic Pain in Mice.

Neuropathic pain manifested with allodynia and hyperalgesia usually becomes a chronic condition accompanied with mood disorders. Clinical therapies for neuropathic pain are still unsatisfactory with notable side effects. Recent studies have reported the protective role of molecular hydrogen (H2) in different diseases including neurological disorders, such as Alzheimer's as well as its antidepressant activities in animals with chronic stress. This study explored the effects of treatment with hydrogen-rich water (HRW) in male mice with neuropathic pain induced by the chronic constriction of the sciatic nerve (CCI) and the accompanying affective deficits. The likely pathways implied in the HRW analgesic activity, as well as the interaction between heme oxygenase 1 (HO-1) enzyme and H2 during neuropathic pain were also studied. The results showed: (i) the inhibitory effects of the repetitive treatment with HRW on the allodynia and hyperalgesia provoked by CCI; (ii) the anxiolytic and antidepressant actions of HRW in animals with neuropathic pain; (iii) the contribution of the antioxidant enzymes (HO-1 and NAD(P)H: quinone oxidoreductase 1) and the ATP sensitive potassium channels in the painkiller activities of HRW during neuropathic pain; (iv) a positive interaction between the HO-1 and H2 systems in inhibiting the CCI-induced neuropathy; and (v) the antioxidant, antinociceptive, anti-inflammatory and/or antiapoptotic features of HRW treatment in the dorsal root ganglia and/or amygdala of sciatic nerve-injured mice. This study demonstrates new protective actions of H2 and suggests that treatment with HRW might be an interesting therapeutic strategy for chronic neuropathic pain and its associated mood disorders.

Effects of treatment with a carbon monoxide donor and an activator of heme oxygenase 1 on the nociceptive, apoptotic and/or oxidative alterations induced by persistent inflammatory pain in the central nervous system of mice.

The activation of heme oxygenase 1 (HO-1)/carbon monoxide (CO) inhibits chronic inflammatory pain, but its role in the central nervous system (CNS) is not entirely known. We evaluated whether the treatment with an HO-1 inducer, cobalt protoporphyrin IX (CoPP), or a CO-releasing molecule, tricarbonyldichlororuthenium(II)dimer (CORM-2), modulates the nociceptive, apoptotic and/or oxidative responses provoked by persistent inflammatory pain in the CNS. In C57BL/6 male mice with peripheral inflammation caused by complete Freund's adjuvant (CFA), we assessed the effects of CORM-2 and CoPP on the expression of protein kinase B (Akt), the apoptotic protein BAX, and the antioxidant enzymes HO-1 and NADPH quinone oxidoreductase 1 (NQO1) in the periaqueductal gray matter (PAG), amygdala (AMG), ventral hippocampus (VHPC) and medial septal area (MSA). Our results showed that the increased expression of p-Akt caused by peripheral inflammation in the four analyzed brain areas was reversed by CORM-2 and CoPP therapies. Both treatments also normalized the upregulation of BAX induced by CFA on the VHPC and MSA. Oxidative stress, demonstrated with the decreased expression of HO-1 on the PAG and AMG, was normalized in CORM-2 and CoPP treated animals. CoPP also increased the expression of HO-1 on VHPC, and both treatments up-regulated the NQO1 levels on the PAG of CFA-injected animals. In conclusion, both CORM-2 and CoPP treatments inhibited the nociceptive and apoptotic responses generated by peripheral inflammation and/or potentiated the antioxidant responses in several brain areas revealing the new modulatory effects of these treatments in the CNS of animals with chronic inflammatory pain.