GZ coupling to the rat κ-opioid receptor

AbstractWe have expressed the cloned rat κ-opioid receptor in human embryonic kidney 293 cells and studied the ability of κ-selective ligands to inhibit adenylyl cyclase. In transfected 293 cells, activation of the κ-opioid receptor by U50,488 and the dynorphins resulted in the inhibition of cAMP accumulation. The inhibitory response was sensitive to pertussis toxin and highly selective for κ-agonists; neither μ- nor θ-opioids were able to activate the κ-opioid receptor. Upon co-transfection with the α subunit of Gz, inhibition of cAMP accumulation by κ-agonist became refractory to pertussis toxin, indicating that the κ-opioid receptor can couple to both Gi and Gz proteins

Evidence for receptor-mediated inhibition of intrinsic activity of GTP-binding protein, Gi1 and Gi2, but not G0 in reconstitution experiments

AbstractThe receptor-mediated inhibition of intrinsic activities of GTP-binding proteins (G-proteins) was studied. Pertussis toxin (IAP)-substrate G-protein, Gi1, Gi2 or G0, was prelabeled with [α-32P]GDP and reconstituted with synaptic membranes of the guinea pig cerebellum in the presence of 0.02% of Chaps. Intrinsic activities of G-proteins were evaluated by the release of [α-32P]GDP in exchange for added GppNHp or GDP in reconstituted preparations. U-50,488H (1 nM-10 μM), a specific ϰ-subtype of opioid receptor agonist, inhibited the [α-32P]GDP release in exchange for added 1 μM GppNHp in Gi1-reconstituted preparations in a concentration-dependent manner. On the other hand, the ϰ-opioid agonist at 10 μM increases the Km values of GppNHp, but not GDP in exchange for [α-32P]GDP release in preparations reconstituted with Gi1 or Gi2, but not with G0. These findings indicate that ϰ-opioid receptor is coupled to inhibition of intrinsic activities of Gi1 and Gi2, but not G0, in guinea pig cerebellar membranes. In addition, it was revealed that the mode of action is mediated by a decrease in affinity of GTP (or its analog) for G-proteins, but not by a change in affinity of GDP

(-)-Pentazocine induces visceral chemical antinociception, but not thermal, mechanical, or somatic chemical antinociception, in μ-opioid receptor knockout mice

<p>Abstract</p> <p>Background</p> <p>(-)-Pentazocine has been hypothesized to induce analgesia via the κ-opioid (KOP) receptor, although the involvement of other opioid receptor subtypes in the effects of pentazocine remains unknown. In this study, we investigated the role of the μ-opioid (MOP) receptor in thermal, mechanical, and chemical antinociception induced by (-)-pentazocine using MOP receptor knockout (MOP-KO) mice.</p> <p>Results</p> <p>(-)-Pentazocine-induced thermal antinociception, assessed by the hot-plate and tail-flick tests, was significantly reduced in heterozygous and abolished in homozygous MOP-KO mice compared with wildtype mice. The results obtained from the (-)-pentazocine-induced mechanical and somatic chemical antinociception experiments, which used the hind-paw pressure and formalin tests, were similar to the results obtained from the thermal antinociception experiments in these mice. However, (-)-pentazocine retained its ability to induce significant visceral chemical antinociception, assessed by the writhing test, in homozygous MOP-KO mice, an effect that was completely blocked by pretreatment with nor-binaltorphimine, a KOP receptor antagonist. <it>In vitro </it>binding and cyclic adenosine monophosphate assays showed that (-)-pentazocine possessed higher affinity for KOP and MOP receptors than for δ-opioid receptors.</p> <p>Conclusions</p> <p>The present study demonstrated the abolition of the thermal, mechanical, and somatic chemical antinociceptive effects of (-)-pentazocine and retention of the visceral chemical antinociceptive effects of (-)-pentazocine in MOP-KO mice. These results suggest that the MOP receptor plays a pivotal role in thermal, mechanical, and somatic chemical antinociception induced by (-)-pentazocine, whereas the KOP receptor is involved in visceral chemical antinociception induced by (-)-pentazocine.</p

Support vector machines as multivariate calibration model for prediction of blood glucose concentration using a new non-invasive optical method named pulse glucometry

金沢大学理工研究域機械工学系A novel optical non-invasive in vivo blood glucose concentration (BGL) measurement technique, named "Pulse Glucometry", was combined with a kernel method; support vector machines. The total transmitted radiation intensity (Iλ) and the cardiac-related pulsatile changes superimposed on Iλ in human adult fingertips were measured over the wavelength range from 900 to 1700 nm using a very fast spectrophotometer, obtaining a differential optical density (ΔOD λ) related to the blood component in the finger tissues. Subsequently, a calibration model using paired data of a family of ΔODλs and the corresponding known BGLs was constructed with support vector machines regression instead of using calibration by a conventional partial least squares regression (PLS). Our results show that the calibration model based on the support vector machines can provide a good regression for the 183 paired data, in which the BGLs ranged from 89.0-219 mg/dl (4.94-12.2 mmol/l). The resultant regression was evaluated by the Clarke error grid analysis and all data points fell within the clinically acceptable regions (region A: 93%, region B: 7%). © 2007 IEEE.

Shared and Distinct Functions of the Transcription Factors IRF4 and IRF8 in Myeloid Cell Development

Interferon regulatory factor (IRF) 8 and IRF4 are structurally-related, hematopoietic cell-specific transcription factors that cooperatively regulate the differentiation of dendritic cells and B cells. Whilst in myeloid cells IRF8 is known to modulate growth and differentiation, the role of IRF4 is poorly understood. In this study, we show that IRF4 has activities similar to IRF8 in regulating myeloid cell development. The ectopic expression of IRF4 in myeloid progenitor cells in vitro inhibits cell growth, promotes macrophages, but hinders granulocytic cell differentiation. We also show that IRF4 binds to and activates transcription through the IRF-Ets composite sequence (IECS). Furthermore, we demonstrate that Irf8-/-Irf4-/- mice exhibit a more severe chronic myeloid leukemia (CML)-like disease than Irf8-/- mice, involving a disproportionate expansion of granulocytes at the expense of monocytes/macrophages. Irf4-/- mice, however, display no obvious abnormality in myeloid cell development, presumably because IRF4 is expressed at a much lower level than IRF8 in granulocyte-macrophage progenitors. Our results also suggest that IRF8 and IRF4 have not only common but also specific activities in myeloid cells. Since the expression of both the IRF8 and IRF4 genes is downregulated in CML patients, these results may add to our understanding of CML pathogenesis

Inhibitory role of supraspinal P2X3/P2X2/3 subtypes on nociception in rats.

Extracellular ATP is known to mediate synaptic transmission as a neurotransmitter or a neuromodulator via ionotropic P2X and metabotropic P2Y receptors. Several lines of evidence have suggested that ATP facilitates pain transmission at peripheral and spinal sites via the P2X receptors, in which the P2X3 subtype is considered as an important candidate for the effect. Conversely, we previously found that the activation of supraspinal P2X receptors evoked antinociception. However, the subtypes responsible for the antinociception via supraspinal P2X receptors remain unclear. In the present study, we showed that intracerebroventricular (i.c.v.) pretreatment with A-317491 (1 nmol), the novel non-nucleotide antagonist selective for P2X3 and P2X2/3 receptors, attenuated the antinociceptive effect produced by i.c.v. administered alpha,beta-methylene-ATP (10 nmol), the P2X receptor agonist, in rats. Similarly, the abolishment of the P2X3 receptor mRNA in the brainstem by repeated i.c.v. pretreatments with antisense oligodeoxynucleotide for P2X3 gene once a day for 5 consecutive days diminished the antinociceptive effect of alpha,beta-methylene-ATP. Furthermore, i.c.v. administration of A-317491 (1 and 10 nmol) significantly enhanced the inflammatory nociceptive behaviors induced by the intraplantar injection of formalin and intraperitoneal injection of acetic acid. Taken together, these results suggest that supraspinal P2X3/P2X2/3 receptors play an inhibitory role in pain transmission