The medical mechanism against type I allergies is to block the release or production of chemical mediators from mast cells or to block the H₁-receptor signaling. We previously reported that the anti-allergic action of the dry powder from Bidens pilosa L. var. radiata SCHERFF treated with the enzyme cellulosine (eMMBP) was dependent on the inhibition of histamine release from mast cells. Here, we investigate that the effect of fractions in eMMBP on the histamine-induced contraction in guinea pig ileum and on the release of histamine in rat peritoneal mast cells. The histamine-induced contraction in guinea pig ileum is dose-dependently inhibited by ketotifen, an antagonist of H₁-receptor. Fractions contained caffeic acid, caffeoylquinic acid and fractions contained flavonoids such as hyperin and isoquercitrin in eMMBP inhibit histamine release from mast cells, but only flavonoids such as hyperin, isoquercitrin and rutin suppress the histamine-induced contraction in guinea pig ileum. Moreover, the histamine-induced contraction was not affected by caffeic acid, however, such contraction was significantly inhibited by rutin. These results suggest that the primary antagonists of H₁- receptor are different from the components in eMMBP that inhibit histamine release, and that these components participate in the anti-allergic activity of eMMBP.

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Cooling-induced reduction of skin blood flow results from a reflex increase in sympathetic output and an enhanced vasoconstrictor activity of skin vessels. The latter has been proposed to be mediated by increased reactivity of α_2C-adrenoceptors during cooling in studies with isolated cutaneous vessels in vitro. We have previously shown in studies with tetrodotoxin-treated mice in vivo that reduction of plantar skin blood flow (PSBF) induced by local cooling results primarily from increased reactivity of α_2C-adrenoceptors. In addition, we showed that part of the cooling-induced response was also mediated by α₁-adrenoceptors. However, the mechanisms involved in the cooling-induced responses mediated by α₁-adrenoceptors have not been elucidated. The present study is an investigation seeking to clarify the mechanisms involving α₁-adrenoceptors. Medial plantar arteries were isolated from male ddY mice and changes in vessel diameter were measured in vitro using pressurized arteriography. In vivo measurements of PSBF were performed on artificially ventilated tetrodotoxin treated mice, anaesthetized with pentobarbital sodium, using laser Doppler flowmetry, with the probe positioned above the medial plantar artery. In the in vitro studies with isolated plantar arteries, cooling from 37 to 28°C did not affect the constrictor potency of phenylephrine, an α₁-adrenoceptor agonist, and the threshold concentration to evoke constriction was rather higher at 28°C than it was at 37°C. The cooling also suppressed the constrictor efficacy of UK14,304, an α₂-adrenoceptor agonist. In contrast, cooling the air temperature around the foot from 25 to 10°C in vivo decreased PSBF, which was significantly inhibited by phentolamine, an α-adrenoceptor antagonist, although MK-912, an α_2C-adrenoceptor antagonist, had no effect on it. These results suggest that although α₁-adrenoceptors are involved in cooling-induced reduction of PSBF in mice, the response is unlikely to result from an enhancement of α₁-adrenoceptor-mediated vasoconstriction of plantar arteries during cooling.

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The contractile capacity of smooth muscle cells depends on the cytoskeletal framework of the cell. The aim of this study was to determine the functional importance of the microtubule, actin filament and intermediate filament components of the cytoskeleton in acetylcholine (ACh)-induced contractile responses of the rat isolated bronchial smooth muscle. The expressions of α-actin, β-actin, α-tubulin, desmin and vimentin were observed by immunoblotting in rat bronchial tissues. α-Actin and desmin were immunohistochemicaly observed in smooth muscle layer. Cytochalasin D, latrunculin A (inhibitors of the actin cytoskeleton) and acrylamide (an inhibitor of the intermediate filament) significantly decreased the contractions induced by ACh in concentration-dependent manners. On the other hand, colchicine or nocodazole (inhibitors of the microtubule cytoskeleton) had no effect on the ACh-induced contraction. These findings suggest that the contraction induced by ACh is highly dependent on polymerization of actin and intermediate filament, such as desmin, but not on the polymerization of microtubule in rat bronchial smooth muscle.

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Modulation of slow waves in response to transmural nerve stimulation (TNS) was investigated in smooth muscle preparations isolated from the corpus of the guinea-pig stomach. Single TNS evoked an inhibitory junction potential (i.j.p.) and enhanced the amplitude of the following slow wave. Effects of atropine, N^ω-nitro-L-arginine (L-NA) and apamin revealed that corpus smooth muscle was innervated by cholinergic excitatory, nitrergic inhibitory and apamin-sensitive inhibitory nerves. In preparations isolated from the upper corpus which generated slow waves of 5-15 mV amplitude, a 1 min train of TNS (0.5 or 1 Hz frequency) increased the amplitude, with further enhancement by L-NA, but inhibition by atropine. In the lower corpus, larger amplitude (20-30 mV) slow waves were generated but these were not altered by a TNS train. However, application of L-NA and neostigmine, or often L-NA alone, resulted in increased frequency and decreased amplitude of slow waves during TNS, with an associated depolarization of the membrane. These changes were inhibited by atropine. In the presence of atropine, TNS reduced slow wave amplitude in an L-NA-sensitive manner. Acetylcholine (ACh) at 1 nM increased the amplitude of slow waves in the upper corpus. In the lower corpus, while low concentrations of ACh (<10 nM) did not increase the frequency and decrease the amplitude of slow waves with an associated depolarization of the membrane, this occurred at high concentrations of ACh (>10 nM). Application of the NO donor, sodium nitroprusside (SNP, 10 nM-1 μM), reduced the amplitude of slow waves. The changes in amplitude of slow waves elicited by ACh or SNP were not associated with a significant change in frequency. These results indicate that in the corpus circular smooth muscle, neural modulation of slow waves appeared to be exerted mainly on the amplitude, but not on the frequency.

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Advanced glycation end products (AGEs) derived from glucose are implicated in the pathogenesis of diabetic vascular disease. However, their direct modulatory effects on coronary vascular tone remain unclear. We previously reported that coronary vasoconstriction was induced by acetylcholine (ACh) infusion of the isolated perfused rat heart and that sensitivity was greater in perfused hearts from streptozotocin (STZ)-induced diabetic rats than in those from age-matched controls (Kamata et al., 2008). Here, we investigated the effect of N^ε-(carboxymethyl)lysine (CML), which has one of the main AGE structures, on ACh-induced vasoconstriction in perfused hearts isolated from control and diabetic rats. ACh-induced vasoconstriction was significantly greater in the STZ-induced diabetic group than in the age-matched controls. CML enhanced the ACh-induced vasoconstriction in coronary arteries from control rats, but not in those from STZ-induced diabetic rats. In the controls, the vasoconstriction induced by the calcium-channel activator Bay K 8644 was also enhanced by CML. These CML-mediated enhancements of the vasoconstrictions induced by ACh and Bay K 8644 were significantly suppressed by tempol, a superoxide dismutase mimetic. The plasma CML and glucose levels were each significantly elevated in STZ-induced diabetic rats. These findings suggest (a) that CML augments ACh-induced coronary vasoconstriction, an effect that may be attributable to increased superoxide and to activation of voltage-gated Ca²⁺ channels and (b) that this modulating effect may be desensitized in the STZ-induced diabetic heart.

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