Models of cardiac cells are sufficiently well developed to answer questions concerning the actions of drugs on repolarization and the initiation of arrhythmias. These models can be used to characterize drug-receptor action profiles that would be expected to avoid arrhythmia and so help to identify drugs that may be safer. Several examples of such action profiles are presented here, including a recently-developed blocker of persistent sodium current, ranolazine. The models have also been incorporated into tissue and organ models that enable arrhythmia to be modelled also at these levels. Work at these levels can reproduce both re-entrant arrhythmia and fibrillation.

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Resveratrol is a stilbene compound found in grapes and other sources. In this study we examined the effects of trans-resveratrol (4.38 – 438 μM/implant) in the vasculogenesis of yolk-sac membranes and its capacity to improve chick embryo growth. High concentrations of the stilbene (43.8 – 438 μM) significantly inhibited early vessel formation, decreasing the percentage vitelline vessels of 3.5-day embryos by 50% compared to the control. In addition, basic fibroblast growth factor–stimulated vasculogenesis (140% of vessels as compared to control) was partially reversed by t-resveratrol (35% of inhibition) and treatments with cyclooxygenase inhibitors (acetylsalicylic acid and indomethacin) as well a protein-kinase C (PKC) activator (phorbol-12,13-dibutyrate) decreased the vessel number to 60%, 50%, and 44%, respectively. Treatments with t-resveratrol (4.38 – 43.8 μM/implant) significantly increased the body length of embryos incubated in vitro uncoupled from any impairment in the body shape or detectable embryotoxic effect. We suggest that the antivasculogenic activity and the enhancement in embryonic growth promoted by non acute treatments with t-resveratrol were, at least in part, due to PKC inhibition. We suggest that t-resveratrol can be usable not only as a reliable functional nutriment, but also is useful for the development of prophylactic and/or therapeutic agents for treatment of angiogenic-degenerative diseases.

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Metabolic syndrome (MetS) is a group of cardiovascular risk factors, including visceral obesity, glucose intolerance, hypertension, and dyslipidemia. Increased oxidative and nitrative stress and inflammation and decreased endothelial function occur in an animal model of metabolic syndrome, SHR/NDmcr-cp (SHR/cp) rats. The present study investigated the effects of coenzyme Q10 (CoQ10), one of the important antioxidants, on the abnormal oxidative condition and characteristic components of metabolic syndrome in SHR/cp rats by maintaining them on a diet supplemented with 0.07% – 0.7% CoQ10 for 26 weeks. We determined serum levels of oxidatively modified low-density lipoprotein (Ox-LDL) and 8-hydroxy-2'-deoxyguanosine (8-OHdG) as oxidative stress markers, 3-nitrotyrosine as a nitrative stress marker, 3-chlorotyrosine as a marker of myeloperoxidase (MPO)-catalyzed oxidation and high-sensitivity C-reactive protein (hsCRP) as an inflammatory marker. The administration of CoQ10 significantly attenuated the increase of oxidative and nitrative stress markers and inflammatory markers in a dose-dependent manner. CoQ10 prevented the elevated serum insulin levels, although it did not affect the elevated glucose level and dyslipidemia. CoQ10 also reduced elevated blood pressure, but did not affect body weight gain. In addition, CoQ10 improved endothelial dysfunction in the mesenteric arteries. These findings suggest that the antioxidant properties of CoQ10 can be effective for ameliorating cardiovascular risk in MetS.

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The inositol 1,4,5-trisphosphate receptors (IP₃Rs) form clusters following agonist stimulation, but its mechanism remains controversial. In this study, we visualized the clustering of green fluorescent protein (GFP)-tagged type 3 IP₃R (GFP-IP₃R3) in cultured living cells using confocal microscopy. Stimulation with ATP evoked GFP-IP₃R3 clustering not only in cells with replete Ca²⁺-stores but also in cells with depleted Ca²⁺ stores. Thapsigargin (ThG) and ionomycin failed to mimic the ATP-induced cluster formation despite the continuous elevation of intracellular Ca²⁺ concentration ([Ca²⁺]_i). Application of IP₃ caused GFP-IP₃R3 clustering in permeabilized cells, and the response was completely inhibited by heparin, a competitive inhibitor of IP₃R. Experiments using LIBRAv, an IP₃ biosensor, showed that ATP significantly stimulated IP₃ generation even in store-depleted cells. We also found that pretreatment with ThG accelerated or enhanced the ATP-induced clustering in both the presence and absence of extracellular Ca²⁺. When permeabilized cells were stimulated with the threshold of IP₃, the GFP-IP₃R3 clustering clearly occurred in Ca²⁺-free medium but not in Ca²⁺-containing medium. These results strongly support the hypothesis that the agonist-induced clustering of IP₃R is triggered by IP₃ binding, rather than [Ca²⁺]_i elevation. Although depletion of the Ca²⁺ store by itself does not cause the clustering, it may increase the sensitivity of IP₃R to cluster formation, leading to facilitation of IP₃-triggered clustering.

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The effect of T-2328 {2-fluoro-4'-methoxy-3'-[[[(2S,3S)-2-phenyl-3-piperidinyl]amino]methyl]-[1,1'-biphenyl]-4-carbonitrile dihydrochloride}, a novel tachykinin NK₁–receptor antagonist, was examined on cisplatin-induced emesis in ferrets. Cisplatin induced acute emesis in 24 h and delayed emesis during 24 and 72 h, respectively. Ondansetron, a 5-HT₃ antagonist, almost completely blocked the acute emesis and transiently reduced the delayed emesis. In contrast, T-2328 elicited long-lasting anti-emetic effects on both acute and delayed phases by a single intravenous administration. Suppression of delayed emesis was not due to elimination of the acute phase because the delayed emesis was also suppressed by administration after the onset of delayed emesis. Persistent blockade of NK₁ receptors in the brain was demonstrated by inhibition of the NK₁ agonist–induced foot tapping response for over 24 h. An appreciable amount of T-2328 was present in the brain 32 and 72 h after the injection. The NK₁ agonist–induced contractions of isolated ileum in guinea pigs was antagonized with IC₅₀ values of 1.4 nM in an insurmountable manner. It is likely that T-2328 exerts the long-lasting anti-emetic effect by not only long-term presence in the brain but also its insurmountable inhibition of NK₁ receptors.

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It has been shown that probiotic bacteria are effective for the treatment of allergic diseases. As histamine plays a central role in allergic diseases, it is possible that probiotic bacteria affect the allergy-related histamine signaling. Here, we investigated the effect of Lac-B, a mixture of freeze-dried Bifidobacterium infantis and Bifidobacterium longum, on the allergy-related histamine signaling. In the nasal allergy model rats made by sensitization and provocation with toluene 2,4-diisocyanate (TDI) for 3 weeks, TDI provocation caused acute allergy–like behaviors along with significant up-regulation of histamine H₁ receptor (H1R) and histidine decarboxylase (HDC) mRNA expression, increased HDC activity, histamine content, and [³H]mepyramine binding activity in nasal mucosa. Prolonged treatment with Lac-B (40 mg/rat, p.o.) significantly suppressed both the allergy-like behaviors and all of the above mentioned factors involved in histamine signaling. Our findings indicate that oral administration of Lac-B showed significant anti-allergic effect through suppression of both H1R and HDC gene expression followed by decrease in H1R, HDC protein level, and histamine content. Suppression of histamine signaling may be a novel target of probiotics in preventing allergic diseases.

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The aim of this study was to investigate the effects of histamine H₁ and H₃ antagonists on learning and mnemonic dysfunction in mice. Two H₁ antagonists, pyrilamine and clozapine, and the prototypic H₃ antagonist thioperamide were used to study the role of histamine in mice with social isolation and repeated methamphetamine administration. Mice with social isolation and repeated methamphetamine administration showed significant disruption of prepulse inhibition as compared to both the socially-housed mice and isolation-housing mice. Furthermore, social isolation and repeated methamphetamine administration caused significant learning and mnemonic dysfunctions. Treatment with clozapine improved learning and mnemonic ability in all of the tasks. Pyrilamine treatment ameliorated performance in all the tests examined except for the passive avoidance test. Thioperamide, however, did not change the learning and mnemonic ability. Donepezil, an acetylcholinesterase inhibitor, reversed the learning and mnemonic dysfunction in all four tasks. The present study has shown that blockade of histamine H₁ receptor improved the learning and mnemonic ability in mice, raising the possibility that treatment with clozapine or pyrilamine may improve learning and mnemonic performance in certain patients with psychiatric diseases such as schizophrenic patients with cognitive dysfunction.

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To compare the mechanisms underlying mechanical responses to ephedrine and Ephedra herb, a main component of Kakkon-to, in isolated male and female rabbit urinary bladder and urethral strips, responses of isolated strips to the agents were recorded in organ bath systems. Ephedrine and Ephedra herb relaxed the female urinary bladder to the similar extent. These relaxations are reversed to contractions by timolol. In the presence of timolol, ephedrine produced less contraction of urethral strips in the female than those in the male; this contraction was abolished by prazosin. Ephedra herb contracted the female urethra less than that of the male, and the contraction was stronger than that by ephedrine. The contraction caused by Ephedra herb in strips treated with timolol was significantly inhibited by prazosin. The prazosin-resistant contraction of the female urethra was greater than that of the male. Quinacrine, a phospholipase A₂ inhibitor, indomethacin, and AA861, a 5-lipoxygenase inhibitor, inhibited the contraction. The contraction was inhibited by ZK 158252, a leukotriene (LT) B₄-receptor antagonist. These findings suggest that Ephedra herb contracts the urethra via arachidonic acid metabolites together with α₁-adrenoceptor stimulation. The metabolites produced by 5-lipoxygenase may stimulate LTB₄, but not CysLt₁, receptors. These contractile components induced by Ephedra herb and Kakkon-to might be effective for the treatment of stress urinary incontinence.

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The present study investigates the mechanisms related to the endogenous nitric oxide synthase (eNOS) activation in the relaxant effects of a proanthocyanidin-rich fraction (PRF), obtained from Croton celtidifolius Baill barks, in rat thoracic aorta rings with endothelium. In vessels pre-contracted with phenylephrine (Phe), PRF (0.1 – 100 μg/mL) induced a concentration-dependent relaxation. This effect was significantly reduced by endothelium denudation, by N^ω-nitro-L-arginine, and by 1H[1,2,3]oxadiazolo[4,3-alpha]quinoxalin. However, the vasorelaxant effect was not altered by indomethacin, atropine, tetraethylammonium, and charybdotoxin plus apamin. In thoracic aorta rings pre-contracted with phorbol-12,13-dibuyrate, PRF also induced a concentration-dependent relaxation. The PRF-induced relaxation disappeared in the absence of extracellular calcium in the medium and decreased significantly in the presence of lanthanum. A sulfhydryl alkylating agent, N-ethylmaleimide, and a phospholipase C (PLC) blocker, neomycin, significantly decreased PRF-induced vasorelaxation. In vessels pre-contracted with Phe, the PRF-induced vasorelaxant effect was not altered by quinacrine and ONO-RS-082, genistein and thyrphostin A-23, GF109203, and pertussis toxin and cholera toxin. The results suggest that the PRF-induced vasorelaxant effect is endothelium-dependent and involves the NO/cGMP pathway. We hypothesize that the activation of eNOS is due to an increase of intracellular calcium derived from PLC activation and an N-ethylmaleimide sensitive pathway.

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The aim of this study was to determine whether the pathway of inducible NO synthase (iNOS) in blood vessels is changed by type 2 diabetes. Lipopolysaccharide (LPS)-induced nitric oxide (NO) production and expression of iNOS and effects of LPS on phenylephrine-induced contractile force were compared in aortae isolated from Goto-Kakizaki (G-K) diabetes rats and aortae isolated from control Wistar rats. Both LPS-stimulated nitrite generation and iNOS expression levels were significantly higher in aortae from G-K rats than in those from control rats. Phenylephrine-induced contractile force in the presence of LPS was significantly lower in aortae from G-K rats than in those from control rats, while contractile force in the absence of LPS was comparable in the diabetic and control groups. On the other hand, incubation of aortae in high glucose-containing medium did not affect the LPS-stimulated nitrite accumulation and iNOS expression and the phenylephrine-induced contractile force, regardless of the presence of LPS. These results suggest that LPS-induced NO production through the iNOS pathway is increased and subsequent attenuation of contractile force by excess NO is enhanced in arteries of rats with type 2 diabetes.

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We performed this study to demonstrate the applicability of the microelectrode array (MEA) to study electrophysiological changes of rat peritoneal mast cells in the presence of compound 48/80 under normal, Ca²⁺-free, Ca²⁺-free with EDTA, and Cl⁻-free conditions. The use of high extracellular K⁺ (KCl, 150 mM), charybdotoxin (ChTX, 100 nM), and Cl⁻-free containing ChTX buffers verified that the hyperpolarizing signal was due to the activation of mainly K⁺ and, to a lesser extent, Cl⁻ channels. Compound 48/80 concentration-dependently shortened the latent periods (the onset of response) and increased both the spatial (the K⁺ and Cl⁻ hyperpolarizing field potentials, HFP) and temporal measurements (the duration of response). Ca²⁺-free buffer had no effect on the latent period of compound 48/80 but increased the HFP at high concentrations. The latent period increased while the HFP diminished when cells were equilibrated in Ca²⁺-free buffer containing EDTA. Durations of the HFP were generally longer when cells were in either Ca²⁺-free or Ca²⁺-free containing EDTA buffers than when cells were in normal buffer. The EC₅₀ values confirmed that effects were only affected in Ca²⁺-free buffer containing EDTA but not in Ca²⁺-free or Cl⁻-free buffers, further reinforcing the hypothesis that the presence of Ca²⁺ is not essential to the action of compound 48/80. The present study is the first application of MEA to study rat peritoneal mast cells, and our results indicate that it could be of value in future pharmacological research on other non-excitable cells.

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We evaluated the effects of zonisamide on inflammatory and neuropathic pain using the mouse formalin test and streptozotocin (STZ)-induced diabetic mice with a reduced withdrawal threshold to mechanical stimuli, respectively. When administered systemically (subcutaneously, s.c.), intracerebroventricularly (i.c.v.) or intrathecally (i.t.) before formalin injection, zonisamide (3 and 10 mg/kg, s.c., 10 and 30 μg, i.c.v., or i.t.) significantly reduced licking/biting behavior during the second phase of the formalin test in a dose-dependent manner. However, zonisamide (30 μg, i.t.) did not affect the second phase of the formalin test when given after the first phase, suggesting that it can prevent the development of injury-induced hyperexcitability of the spinal dorsal horn triggered by the repetitive nociceptive input during the first phase. Moreover, zonisamide administered into the dorsal hindpaw ipsilateral but not contralateral to the formalin injection partly reduced the second phase. Thus it is likely that zonisamide generates analgesic effects in the formalin test via both central and peripheral mechanisms. In mice with STZ-induced diabetes, zonisamide (10 and 30 mg/kg, s.c. or 10 and 30 μg, i.t.) reversed the mechanical hyperexcitability. Our results suggest that zonisamide can be a useful therapeutic agent, presumably for both prevention and reversal of pathophysiologic pain.

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To evaluate the feasibility of two kinds of pterin derivatives, 2-(N,N-dimethylaminomethyleneamino)-6-formyl-3-pivaloylpteridin-4-one (DFP) and 2-(N,N-dimethylaminomethyleneamino)-3-pivaloylpteridin-4-one (DP), as anticancer drugs, their photodynamic and non-photodynamic effects on pancreatic cancer cell line Panc-1 cells were examined. For photodynamic effects, cell death 48 h after UV-A irradiation was more prominent in cells preloaded with DP than DFP. When cells were simply incubated for 96 h without irradiation, DFP induced cell death, while DP suppressed cell proliferation. Furthermore, DP was much more soluble in water than DFP. These findings collectively indicated that DP is more feasible as an anticancer drug than DFP.

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Tamoxifen (Tam) decreased the clearance of L-glutamate (L-Glu) by cultured astrocytes at 1 pM, 1 nM, and 1 μM, but became toxic at 10 μM. When L-Glu transporters were mostly inhibited by threo-β-benzyloxyaspartate (TBOA) (1 mM) or D,L-threo-β-hydroxyaspartate (THA) (1 mM), Tam (1 nM) did not change extracellular L-Glu concentration, confirming that Tam attenuates L-Glu transport through L-Glu transporters. ICI182,780, LY294002, and U0126 inhibited the effect of Tam dose-dependently, suggesting the involvement of estrogen receptors (ERs), the phosphatidylinositol 3-kinase (PI3K) cascade, and the mitogen-activated protein kinase (MAPK) cascade in the effect of Tam.

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