Effects of probenecid and brilliant blue G on rat enteric glial cells following intestinal ischemia and reperfusion

The P2X7 receptor participates in several intracellular events and acts with the pannexin-1 channel. This study examined the effects of probenecid (PB) and brilliant blue G (BBG), which are antagonists of the pannexin-1 channel and P2X7 receptor, respectively, on rat ileum enteric glial cells after on ischemia and reperfusion. The ileal vessels were occluded for 45 min with nontraumatic vascular tweezers, and reperfusion was performed for periods of 24 h and 14 and 28 days. After ischemia (IR groups), the animals were treated with BBG (BG group) or PB (PB group). The double-labeling results demonstrated the following: the P2X7 receptor was present in enteric glial cells (S100β) and enteric neurons positive for HuC/D; enteric glial cells exhibited different phenotypes; some enteric glial cells were immunoreactive to only S100β or GFAP; and the pannexin-1 channel was present in enteric glial cells (GFAP). Density (in cells/cm²) analyses showed that the IR group exhibited a decrease in the number of cells immunoreactive for the P2X7 receptor, pannexin-1, and HuC/D and that treatment with BBG or PB resulted in the recovery of the numbers of these cells. The number of glial cells (S100β and GFAP) was higher in the IR group, and the treatments decreased the number of these cells to the normal value. However, the PB group did not exhibit recovery of S100β-positive glia. The cell profile area (μm²) of S100β-positive enteric glial cells decreased to the normal value after BBG treatment, whereas no recovery was observed in the PB group. The ileum contractile activity was decreased in the IR group and returned to baseline in the BG and PB groups. BBG and PB can effectively induce the recovery of neurons and glia cells and are thus potential therapeutic agents in the treatment of gastrointestinal tract diseases.     

Thermoresponsive Reversible Unimer Micelles of Amphiphilic Fluorinated Copolymers

Amphiphilic fluorinated copolymers PEGMAx-co-FAy and TEGMAx-co-FAy are prepared by activators regenerated by electron transfer atom transfer radical polymerization (ARGET-ATRP). All polymers present a reversible thermoresponsive lower critical solution temperature-type behavior, and a cloud point temperature (T_c) in the range of 30–60 °C strictly dependent on the length of the oxyethylene side chain, the content of the hydrophobic counits, and the concentration of the solution. Combined small angle X-ray scattering (SAXS) and dynamic light scattering measurements are used to study the self-assembly behavior in water, organic solvents (tetrahydrofuran [THF] and dimethylformamide [DMF]), and a fluorinated solvent (hexafluorobenzene [HFB]). SAXS confirms the formation of compact-globular single-chain self-folded unimer micelles in water below T_c, which generally presents small hydrodynamic diameters (D_h ≤ 8 nm) as a result of the folding of the hydrophobic perfluorohexylethyl acrylate counits. The copolymers are also able to form reverse unimer micelle in HFB. The copolymers are not able to self-assemble in unimer micelles in THF or DMF solutions, in which they adopt conventional random coil conformations.     

Evidence for the involvement of the nitric oxide–cGMP pathway in the antinociception of morphine in the formalin test

The effect of inhibition of nitric oxide synthesis and guanylate cyclase on the peripheral antinociceptive effect of morphine was assessed by using the formalin test in the rat. Saline, N^G-monomethyl-

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-arginine, a nitric oxide synthesis inhibitor (50 μg) and methylene blue, a guanylate cyclase inhibitor (500 μg), did not exhibit any antinociceptive activity. However, morphine (10 μg) produced a significant antinociceptive effect in phases 2a and 2b, which was reduced by pretreatment with either N^G-monomethyl-

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-arginine or methylene blue. These results suggest that the local administration of morphine induces antinociception by the activation of the

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-arginine–nitric oxide–cGMP pathway.     

Functional consequences of inhibition of nucleotide breakdown in rat vas deferens: a study with Evans blue

The effect of Evans blue on nucleotide breakdown, nucleotide-evoked contractions and electrically evoked contractions, overflow of ATP and overflow of tritium (after labelling with [³H]-noradrenaline) was studied in rat vas deferens. Pieces of vas deferens degraded 83 to 85% of added ATP, ADP and 2-methylthio ATP (all 100 M) over 30 min. Evans blue (100 M) reduced this degradation to 22 to 26%. Nucleotides elicited contraction with potency declining in the order , \-methylene ATP > 2-methylthio ATP > ATP > ADP. Evans blue (100 M) shifted the concentration-response curve of , \-methylene ATP to the right and increased the maximum. Concentration-response curves of ATP, ADP and 2-methylthio ATP, in contrast, were shifted to the left and responses were much potentiated. In the presence of Evans blue, the rank order of potency was ATP > 2-methylthio ATP > , \-methylene ATP > ADP. Electrical field stimulation (100 pulses at 10 Hz) elicited contraction and an overflow of tritium and ATP. Evans blue (100 M) did not alter the contraction and the evoked overflow of tritium but increased 24-fold the evoked overflow of ATP. The results indicate that Evans blue may serve as an — albeit impure — ecto-nucleotidase inhibitor in functional experiments. Such experiments demonstrate that the low potency of ATP (and also ADP and 2-methylthio ATP) in eliciting contraction, and the small size of the overflow of ATP upon sympathetic nerve stimulation, are due to rapid breakdown.     

Cultured chick sympathetic neurons: ATP-induced noradrenaline release and its blockade by nicotinic receptor antagonists

The ATP-induced increase in tritium outflow from cultured chick sympathetic neurons prelabelled with [³H]-noradrenaline was investigated.Seven days-old dissociated cell cultures of embryonic paravertebral ganglia, loaded with [³H]-noradrenaline (0.05 M), were superfused in the presence of (+)-oxaprotiline and exposed to ATP, ATP-analogues, or 1,1-dimethyl-4-piperazinium (DMPP) for 2 min. ATP (3 LM-3 mM), 2-methylthio-ATP (3–100 M), as well as DMPP (10 and 100 M) induced a significant overflow of tritium. The EC₅₀-value of ATP was 20 M. Both the ATP-induced and the DMPP-induced tritium overflow was Ca²⁺-dependent and sensitive to tetrodotoxin (0.3 M) and -conotoxin (0.1 M); in addition, it was inhibited by the ₂-adrenoceptor agonist 5-bromo-6-(2-imidazoline-2-ylamino)-quinoxaline (UK-14,304; 1 M). The effects of ATP and DMPP were not additive. The ATP-induced as well as the DMPP-induced overflow of tritium was diminished by the P₂-purinoceptor antagonists suramin (300 M) and reactive blue 2 (3 M); in all 4 cases, the inhibition amouted to approximately 40%. The tritium overflow induced by ATP or DMPP was almost abolished by the nicotinic receptor antagonist mecamylamine (10 M) and markedly inhibited by hexamethonium (100 M). Neither ATP nor electrical stimulation caused an overflow of tritium from cultures loaded with [³H]-choline.The results suggest that ATP at molar concentrations induces noradrenaline release from cultured chick sympathetic neurons via an action on a subclass of the nicotinic cholinoceptor.     

New aspects in the treatment of priapism

Recent advances in the understanding of erectile physiology have improved the prompt diagnosis and treatment of priapism. During initial assessment, the physician must distinguish between veno-occlusive low flow (ischemic) and arterial high flow (nonischemic) in order to choose the correct treatment option for each type of priapism. Patient history, physical examination, penile haemodynamics and corporeal metabolic blood quality assist the distinction between static and dynamic priapism. Normally, priapism is effectively treated with intracavernous vasoconstrictive agents or surgical shunting. However, when these two methods fail, subsequent treatment procedures are a matter for debate. Alternative options, such as intracavernous injection of methylene blue or selective penile arterial embolization, for the management of high and low flow priapism are described and a survey of current treatment modalities is presented.     

锥兰联合茜素红活细胞染色法观察角膜中央碱烧伤内皮细胞愈合过程

目的:研究角膜碱烧伤内皮细胞的愈合过程。方法:锥兰联合茜素红内皮细胞染色法对角膜中央碱烧伤内皮细胞形态学变化进行观察。结果:伤后20分钟内皮细胞已破坏。72小时,损伤区的周边内皮细胞变形向创面内迁移。第7天后缺损区大部分范围均可见梭形细胞覆盖。结论:兔碱烧伤损伤区内皮细胞的修复由邻近内皮细胞变形,增殖和移行长入创面内完成,修复的内皮细胞具有纤维细胞特征。眼科学报1999;15:218-220。     

Concomitant blockade of P2X-receptors and ecto-nucleotidases by P2-receptor antagonists: functional consequences in rat vas deferens

In order to assess the consequences of a concomitant blockade of P2X-receptors and ecto-nucleotidases, effects of 13 P2-receptor antagonists were investigated on contractions of the rat vas deferens elicited by α,β-methylene ATP (α,β-MeATP) and ATP and on the removal of ATP from the incubation medium by vas deferens tissue. Increasing concentrations of all antagonists reduced and finally abolished contractions elicited by α,β-MeATP (3 μM), with IC₅₀-values ranging from 1.1 to 100 μM. Pyridoxalphosphate-6-azophenyl-2’,4’-disulphonate (PPADS), 6-azophenyl-4-amino-5-hydroxy-naphthalene-1,3-disulphonate (NH02), 4,4’-diisothiocyanatostilbene-2,2’-disulphonate (DIDS) and uniblue A also progressively reduced and finally abolished contractions elicited by ATP (1 mM). 8,8’-[Carbonylbis(imino-3,1-phenylenecarbonyl-imino)]-bis-(1,3,5-naphthalenetrisulphonate) (NF023), sura- min, pyridoxalphosphate-6-azophenyl-2’,5’-disulphonate (iso-PPADS), trypan blue and reactive blue 19, in contrast, caused only partial blockade, by 34–43% maximally; reactive blue 2 and reactive red 2 had no effect; and 6,6’-(1,1’-biphenyl-4,4’-diylbisazo)-bis-4-amino-5-hydroxy-naphtha-lene-1,3-disulphonate (NH01) and Evans blue even enhan- ced the response to ATP. For antagonists causing full or partial inhibition, the IC₅₀-values against ATP were close to those against α,β-MeATP. All antagonists attenuated the removal of ATP, with IC_25%-values ranging from 0.8 μM to >320 μM. The results confirm the frequent combination, in one antagonist molecule, of P2-receptor blockade and blockade of ecto-nucleotidases. This dual action underlies the effect of such compounds on contractions of the vas deferens elicited by ATP which, for certain substances (e.g., suramin, reactive blue 2), can be explained by a simple model in which the antagonist simultaneously blocks the degradation of ATP and a single contraction-mediating receptor (P2X₁). Several observations, however, do not conform with this model, and the existence of multiple contraction-mediating receptors for ATP or multiple, pharmacologically distinct ecto-nucleotidases has to be considered. Received: 23 October 1998 / Accepted: 11 January 1999     

Liposomes and Nanoparticles in the Treatment of Intracellular Bacterial Infections

The treatment of infections caused by obligate or facultative intracellular microorganisms is difficult because most of the available antibiotics have either poor intracellular diffusion and retention or reduced activity at the acidic pH of the lysosomes. The need for antibiotics with greater intracellular efficacy led to the development of endocytosable drug carriers, such as liposomes and nanoparticles, which mimic the entry path of the bacteria by penetrating the cells into phagosomes or lysosomes. This Review assesses the potential of liposomes and nanoparticles in the targeted antibiotic therapy of intracellular bacterial infections and diseases and the pharmaceutical advantages and limitations of these submicron delivery systems.     

Photodynamic therapy in the bladder: Can the light dose be easily quantified?

The shape of the bladder and the optical characteristics of the tissue within the wall can be shown to play an important role in the amount of light actually received at the wall. The use of estimated doses even assuming spherical geometry, cannot therefore be relied upon. This paper describes some experimental work carried out on a glass model that was used to simulate the bladder. A specially constructed dual detector was used which consisted of an isotropic probe and semiconductor detector. This enabled measurements of irradiance and space irradiance (light energy fluence rate) to be made simultaneously. By changing the optical characteristics of the wall a four-fold increase in space irradiance was measured. Contamination of the water contained within the model by blood was also investigated and has shown that with concentrations as low as 0.5% the delivered light dose reaching the wall can be reduced by up to 50% at a wavelength of 510 nm. Some in vivo measurements are also presented together with some comments on the difficulties that have been encountered when transferring measurements from the model to the patient.