Hydrogen sulfide functions as a micro-modulator bound at the copper active site of Cu/Zn-SOD to regulate the catalytic activity of the enzyme.

The present study examines whether there is a mechanism beyond the current concept of post-translational modifications to regulate the function of a protein. A small gas molecule, hydrogen sulfide (H2S), was found to bind at active-site copper of Cu/Zn-SOD using a series of methods including radiolabeled binding assay, X-ray absorption near-edge structure (XANES), and crystallography. Such an H2S binding enhanced the electrostatic forces to guide the negatively charged substrate superoxide radicals to the catalytic copper ion, changed the geometry and energy of the frontier molecular orbitals of the active site, and subsequently facilitated the transfer of an electron from the superoxide radical to the catalytic copper ion and the breakage of the copper-His61 bridge. The physiological relevance of such an H2S effect was also examined in both in vitro and in vivo models where the cardioprotective effects of H2S were dependent on Cu/Zn-SOD.

The role of Nav1.7 and methylglyoxal-mediated activation of TRPA1 in itch and hypoalgesia in a murine model of type 1 diabetes.

Methylglyoxal (MGO), an endogenous reactive carbonyl compound, plays a key role in the pathogenesis of diabetic neuropathy. The aim of this study is to investigate the role of MGO in diabetic itch and hypoalgesia, two common symptoms associated with diabetic neuropathy. Methods: Scratching behavior, mechanical itch (alloknesis), and thermal hypoalgesia were quantified after intradermal (i.d.) injection of MGO in naïve mice or in diabetic mice induced by intraperitoneal (i.p.) injection of streptozotocin (STZ). Behavioral testing, patch-clamp recording, transgenic mice, and gene expression analysis were used to investigate the mechanisms underlying diabetic itch and hypoalgesia in mice. Results: I.d. injection of MGO evoked dose-dependent scratching in normal mice. Addition of MGO directly activated transient receptor potential ankyrin 1 (TRPA1) to induce inward currents and calcium influx in dorsal root ganglia (DRG) neurons or in TRPA1-expressing HEK293 cells. Mechanical itch, but not spontaneous itch was developed in STZ-induced diabetic mice. Genetic ablation of Trpa1 (Trpa1-/- ), pharmacological blockade of TRPA1 and Nav1.7, antioxidants, and mitogen-activated protein kinase kinase enzyme (MEK) inhibitor U0126 abrogated itch induced by MGO or in STZ-induced diabetic mice. Thermal hypoalgesia was induced by intrathecal (i.t.) injection of MGO or in STZ-induced diabetic mice, which was abolished by MGO scavengers, intrathecal injection of TRPA1 blockers, and in Trpa1-/- mice. Conclusion: This study revealed that Nav1.7 and MGO-mediated activation of TRPA1 play key roles in itch and hypoalgesia in a murine model of type 1 diabetes. Thereby, we provide a novel potential therapeutic strategy for the treatment of itch and hypoalgesia induced by diabetic neuropathy.

Antioxidants Attenuate Acute and Chronic Itch: Peripheral and Central Mechanisms of Oxidative Stress in Pruritus.

Itch (pruritus) is one of the most disabling syndromes in patients suffering from skin, liver, or kidney diseases. Our previous study highlighted a key role of oxidative stress in acute itch. Here, we evaluated the effects of antioxidants in mouse models of acute and chronic itch and explored the potential mechanisms. The effects of systemic administration of the antioxidants N-acetyl-L-cysteine (NAC) and N-tert-butyl-α-phenylnitrone (PBN) were determined by behavioral tests in mouse models of acute itch induced by compound 48/80 or chloroquine, and chronic itch by treatment with a mixture of acetone-diethyl-ether-water. We found that systemic administration of NAC or PBN significantly alleviated compound 48/80- and chloroquine-induced acute itch in a dose-dependent manner, attenuated dry skin-induced chronic itch, and suppressed oxidative stress in the affected skin. Antioxidants significantly decreased the accumulation of intracellular reactive oxygen species directly induced by compound 48/80 and chloroquine in the cultured dorsal root ganglia-derived cell line ND7-23. Finally, the antioxidants remarkably inhibited the compound 48/80-induced phosphorylation of extracellular signal-regulated kinase in the spinal cord. These results indicated that oxidative stress plays a critical role in acute and chronic itch in the periphery and spinal cord and antioxidant treatment may be a promising strategy for anti-itch therapy.

Peripheral and spinal 5-HT receptors participate in cholestatic itch and antinociception induced by bile duct ligation in rats.

Although 5-HT has been implicated in cholestatic itch and antinociception, two common phenomena in patients with cholestatic disease, the roles of 5-HT receptor subtypes are unclear. Herein, we investigated the roles of 5-HT receptors in itch and antinociception associated with cholestasis, which was induced by common bile duct ligation (BDL) in rats. 5-HT-induced enhanced scratching and antinociception to mechanical and heat stimuli were demonstrated in BDL rats. 5-HT level in the skin and spinal cord was significantly increased in BDL rats. Quantitative RT-PCR analysis showed 5-HT1B, 5-HT1D, 5-HT2A, 5-HT3A, 5-HT5B, 5-HT6, and 5-HT7 were up-regulated in peripheral nervous system and 5-HT1A, 5-HT1F, 5-HT2B, and 5-HT3A were down-regulated in the spinal cord of BDL rats. Intradermal 5-HT2, 5-HT3, and 5-HT7 receptor agonists induced scratching in BDL rats, whereas 5-HT3 agonist did not induce scratching in sham rats. 5-HT1A, 5-HT2, 5-HT3, and 5-HT7 agonists or antagonists suppressed itch in BDL rats. 5-HT1A agonist attenuated, but 5-HT1A antagonist enhanced antinociception in BDL rats. 5-HT2 and 5-HT3 agonists or antagonists attenuated antinociception in BDL rats. Our data suggested peripheral and central 5-HT system dynamically participated in itch and antinociception under cholestasis condition and targeting 5-HT receptors may be an effective treatment for cholestatic itch.

Restoring Spinal Noradrenergic Inhibitory Tone Attenuates Pain Hypersensitivity in a Rat Model of Parkinson's Disease.

In the present study, we investigated whether restoring descending noradrenergic inhibitory tone can attenuate pain in a PD rat model, which was established by stereotaxic infusion of 6-hydroxydopamine (6-OHDA) into the bilateral striatum (CPu). PD rats developed thermal and mechanical hypersensitivity at the 4th week after surgery. HPLC analysis showed that NE content, but not dopamine or 5-HT, significantly decreased in lumbar spinal cord in PD rats. Additional noradrenergic depletion by injection of N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) aggravated pain hypersensitivity in PD rats. At the 5th week after injection of 6-OHDA, systemic treatment with pharmacological norepinephrine (NE) precursor droxidopa (L-DOPS) or α2 adrenoceptor agonist clonidine significantly attenuated thermal and mechanical pain hypersensitivity in PD rats. Furthermore, application of norepinephrine (NE) and 5-hydroxytryptamine (5-HT) reuptake inhibitors duloxetine, but not 5-HT selective reuptake inhibitors sertraline, significantly inhibited thermal and mechanical pain hypersensitivity in PD rats. Systemic administration of Madopar (L-DOPA) or the D2/D3 agonist pramipexole slightly inhibited the thermal, but not mechanical, hypersensitivity in PD rats. Thus, our study revealed that impairment of descending noradrenergic system may play a key role in PD-associated pain and restoring spinal noradrenergic inhibitory tone may serve as a novel strategy to manage PD-associated pain.