The classical D1 dopamine receptor antagonist SCH23390 is a functional sigma-1 receptor allosteric modulator.

SCH23390 is a widely used D1 dopamine receptor (D1R) antagonist that also elicits some D1R-independent effects. We previously found that the benzazepine, SKF83959, an analog of SCH23390, produces positive allosteric modulation of the Sigma-1 receptor (Sig1R). SCH23390 does not bind to the orthodoxic site of Sig1R but enhances the binding of 3H (+)-pentazocine to Sig1R. In this study, we investigated whether SCH23390 functions as an allosteric modulator of Sig1R. We detected increased Sig1R dissociation from binding immunoglobulin protein (BiP) and translocation of Sig1R to the plasma membrane in response to SCH23390 in transfected HEK293T and SH-SY5Y cells, respectively. Activation of Sig1R by SCH23390 was further confirmed by inhibition of GSK3ß activity in a time- and dose-dependent manner; this effect was blocked by pretreatment with the Sig1R antagonist, BD1047, and by knockdown of Sig1R. SCH23390 also inhibited GSK3ß in wild-type mice but not in Sig1R knockout mice. Finally, we showed that SCH23390 allosterically modulated the effect of the Sig1R agonist SKF10047 on inhibition of GSK3ß. This positive allosteric effect of SCH23390 was further confirmed via promotion of neuronal protection afforded by SKF10047 in primary cortical neurons challenged with MPP+. These results provide the first evidence that SCH23390 elicits functional allosteric modulation of Sig1R. Our findings not only reveal novel pharmacological effects of SCH23390 but also indicate a potential mechanism for SCH23390-mediated D1R-independent effects. Therefore, attention should be paid to these Sig1R-mediated effects when explaining pharmacological responses to SCH23390.

4'-Hydroxywogonin suppresses lipopolysaccharide-induced inflammatory responses in RAW 264.7 macrophages and acute lung injury mice.

4'-Hydroxywogonin (4'-HW), a flavonoid, has been isolated from various plants and shown to inhibit NO production in macrophages. However, the molecular mechanisms and its in vivo activity have not been determined. Our study aimed to investigate the mechanisms underlying the anti-inflammatory effects of 4'-HW in vitro and in vivo. We showed that 4'-HW potently reduced the expression levels of COX-2 and iNOS as well as their products, prostaglandin E2 (PGE2) and nitric oxide (NO) respectively, in LPS-stimulated RAW 264.7 macrophages. 4'-HW also suppressed LPS-induced pro-inflammatory cytokines at mRNA and protein levels. Moreover, 4'-HW blocked the interaction of TAK1 and TAB1 in LPS-stimulated RAW 264.7 macrophages, resulting in an inhibition of the TAK1/IKK/NF-κB signaling pathway. Furthermore, 4'-HW also reduced the phosphorylation of MAPKs and PI3/Akt signaling pathways in LPS-stimulated RAW 264.7 macrophages. 4'-HW was also significantly decreased the intracellular reactive oxygen species (ROS) level. The effect of 4'-HW was confirmed in vivo. 4'-HW exhibited potent protective effect against LPS-induced ALI in mice. These findings indicate that 4'-HW suppresses the LPS-induced response in vitro and in vivo. It is likely that the inhibition of the TAK1/IKK/NF-κB, MAPKs and PI3/AKT signaling pathways contribute to the anti-inflammatory effects of 4'-HW. Our study suggests that 4'-HW may be an important functional constituent in the plants and has the potential value to be developed as a novel anti-inflammatory agent.

The protective action of ketanserin against lipopolysaccharide-induced shock in mice is mediated by inhibiting inducible NO synthase expression via the MEK/ERK pathway.

Nitric oxide (NO) plays an important role in the pathogenesis of endotoxic shock. This work tested the hypothesis that ketanserin could attenuate endotoxic shock by inhibiting the expression of inducible NO synthase (iNOS). The results demonstrated that ketanserin could inhibit iNOS expression in the heart, lungs, liver, and kidneys and nitrate production in the serum upon endotoxic shock in mice. In RAW264.7 cells, ketanserin significantly inhibited the expression of iNOS and decreased the production of NO, TNFα, IL-6, and reactive oxygen species upon lipopolysaccharide (LPS) challenge. Ketanserin also increased the level of ATP and mitochondrial membrane potential in RAW264.7 cells upon LPS exposure. LPS-induced iNOS expression was inhibited by the 5-HT2A receptor antagonist ritanserin and not the α1 receptor antagonist prazosin. Knockdown of 5-HT2A receptor by siRNA abolished the inhibitory effect of ketanserin on the expression of iNOS. These results indicated that the inhibitory effect of ketanserin on the expression of iNOS is mediated by blocking the 5-HT2A receptor. Furthermore, ketanserin significantly inhibited the activation of ERK1/2 and NF-κB signal. Pretreatment with PD184352, a specific inhibitor of ERK1/2, blocked the inhibitory effect of ketanserin on the expression of iNOS and NO production, indicating a critical role for the MEK/ERK1/2 signaling pathway. Collectively, our findings indicate that inhibition of the expression of iNOS via the MEK/ERK pathway mediates the protective effects of ketanserin against LPS-induced shock in mice.

Combined administration of anisodamine and neostigmine produces anti-shock effects: involvement of α7 nicotinic acetylcholine receptors.

AIM: To evaluate the anti-effects of anisodamine and neostigmine in animal models of endotoxic and hemorrhagic shock. METHODS: Kunming mice were injected with lipopolysaccharide (LPS 30 mg/kg, ip) to induce endotoxic shock. Anisodamine (12.5, 25, and 50 mg/kg, ip) and neostigmine (12.5, 25, and 50 µg/kg, ip) were administered immediately after LPS injection. Survival rate was monitored, and the serum levels of TNF-α and IL-1ß were analyzed using ELISA assays. The effects of anisodamine and neostigmine were also examined in α7 nicotinic acetylcholine receptor (α7 nAChR) knockout mice with endotoxic shock and in Beagle dogs with hemorrhagic shock. RESULTS: In mice with experimental endotoxemia, combined administration of anisodamine and neostigmine significantly increased the survival rate and decreased the serum levels of inflammatory cytokines, as compared to those produced by either drug alone. The anti-shock effect of combined anisodamine and neostigmine was abolished in α7 nAChR knockout mice. On the other hand, intravenous injection of the combined anisodamine and neostigmine, or the selective α7 nAChR agonist PNU282987 exerted similar anti-shock effects in dogs with hemorrhagic shock. CONCLUSION: The results demonstrate that combined administration of anisodamine and neostigmine produces significant anti-shock effects, which involves activation of α7 nAChRs.

Effects of ketanserin on endotoxic shock and baroreflex function in rodents.

BACKGROUND: Ketanserin, a 5-hydroxytryptamine receptor antagonist, is clinically used as an antihypertensive agent and could enhance baroreflex function. The present work tested the hypothesis that restoration of baroreflex function is an effective treatment for lipopolysaccharide (LPS)-induced shock. METHODS: Kunming mice were injected with LPS (30 mg/kg; intraperitoneal) to induce endotoxic shock. Ketanserin (0.3, 1, 3, or 10 mg/kg; intraperitoneal) was administered immediately after LPS injection. Survival time was monitored, and serum cytokines were analyzed after the onset of LPS. Effects of ketanserin were also examined in IL-10-deficient mice and mice with sinoaortic denervation. Finally, effects of ketanserin on blood pressure, heart rate, and baroreflex sensitivity were examined in Wistar-Kyoto (WKY) rats with endotoxic shock. RESULTS: Ketanserin significantly increased survival time and decreased serum levels of tumor necrosis factor α and interleukin (IL) 1ß in mice with endotoxic shock. At a dose of 10 mg/kg, ketanserin also significantly increased serum IL-10 concentration. The antishock effect of ketanserin was also apparent in IL-10-knockout mice. In mice with sinoaortic denervation, however, ketanserin had little antishock effects. In WKY rats, ketanserin significantly prevented the baroreflex impairment induced by LPS and prolonged the survival time. CONCLUSIONS: Ketanserin could ameliorate endotoxic shock by restoring baroreflex function.