Adenosine, a metabolite of ATP, serves a number of important physiological roles in the body. These actions contribute to sedation, bradycardia, vasorelaxation, inhibition of lipolysis and regulation of the immune system and are mediated, in part, through activation of three distinct adenosine receptor (AR) subtypes. To date, four receptor types have been cloned: A₁, A_2A, A_2B and A₃. It is becoming increasing clear that adenosine contributes significantly to cytoprotection, a function mediated principally by the A₁AR and A₃AR. In this review, we survey the literature on the role of adenosine and the mechanisms underlying cytoprotection and ischemic preconditioning, a process characterized by cytoprotection derived from repeated brief ischemic challenges. An important recent observation is that the expression of several AR subtypes could be regulated by oxidative stress to provide a greater cytoprotective role. Thus, like other proteins known to be regulated during ischemia, the A₁AR and A₃AR can be considered as being inducible receptors.

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Many kinds of drugs are used for the treatment of allergic diseases. Glucocorticoids are the most efficacious drugs and widely used for the treatment of allergic diseases. Recently, effectiveness of inhaled glucocorticoids for the treatment of bronchial asthma has been established. Beclomethasone dipropionate and fluticasone propionate, which are degraded easily after absorption, are applied by inhalation. Histamine is one of the most important mediators in allergic reactions and antihistamines have widely been applied for the treatment of allergic skin diseases. In Japan, over 20 antiallergic drugs, such as mediator release inhibitors, mediator antagonists and mediator synthesis inhibitors, have been developed. Recently developed compounds such as pranlukast and suplatast are very effective. To relieve the asthmatic attack, bronchodilators such as β₂-adrenoceptor agonists, theophylline and anti-cholinergic drugs are used. Clinical application of tacrolimus ointment has just started for the treatment of atopic dermatitis. Recently the number of allergic patients has increased. The onset and development of allergic diseases are considered to be dependent on both the genetic factors and the environmental factors. For the successful treatment of patients with allergic diseases, it is also important to consider the control of environmental factors.

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We investigated the effects of YM905 [(+)-(1S,3'R)-quinuclidin-3'-yl 1-phenyl-1,2,3,4-tetrahydroisoquinoline-2-carboxylate monosuccinate], a new orally active muscarinic M₃-receptor antagonist, on bowel dysfunction in vivo using experimental models that reproduce the symptoms present in irritable bowel syndrome (IBS). YM905 potently inhibited restraint stress-induced fecal pellet output in fed rats (ED₅₀: 4.0 mg/kg) and diarrhea in fasted rats (ED₅₀: 1.7 mg/kg), with similar potencies to the inhibition of bethanechol-, neostigmine- and nicotine-induced fecal pellet output in rats (ED₅₀: 3.3, 7.9 and 4.5 mg/kg, respectively). YM905 also inhibited 5-hydroxytryptamine (5-HT)-, prostaglandin E₂- and castor oil-induced secretory diarrhea in mice (ED₅₀: 5.5, 14 and 6.3 mg/kg, respectively), but showed no significant effect on cholera toxin-induced intestinal secretion in mice. In addition, YM905 (3, 10 mg/kg) reversed morphine-decreased postprandial defecation in ferrets, a model of spastic constipation, whereas remosetron, a 5-HT₃-receptor antagonist, was not effective. The mode of YM905 action was similar to that of darifenacin, a selective M₃-receptor antagonist, with equivalent potencies. By contrast, propantheline, an antimuscarinic drug that has been used for IBS, was much less potent. These results show that YM905 ameliorates a wide spectrum of bowel dysfunctions through the blockade of M₃ receptors, suggesting its therapeutic potential for treating IBS.

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We investigated effects of fluvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, and its major metabolites, M2 and M4, on CuSO₄-induced low-density lipoprotein (LDL) oxidation and cholesteryl ester accumulation in mouse peritoneal macrophages. All the test compounds inhibited LDL oxidation, and M2 had the most potent effect comparable to vitamin E. When LDL was previously incubated with the test compounds in the presence of CuSO₄, the pre-treatment resulted in a marked reduction of facilitated cholesteryl ester accumulation in macrophages. Supplementation of mevalonate did not overcome the inhibitory effects of fluvastatin and its metabolites on both LDL oxidation and facilitated cholesterol esterification. Pravastatin, another HMG-CoA reductase inhibitor, did not show any inhibitory effect. Consequently, these effects of fluvastatin and its metabolites are considered to be derived from their own unique chemical structures. Moreover, fluvastatin and M2 directly inhibited cholesterol esterification induced by oxidized LDL in macrophages, but pravastatin was also found to have a weak effect. As their inhibitory effects were overcome by addition of mevalonate, the direct inhibitory effect on cholesterol esterification would be a common property of HMG-CoA reductase inhibitors. The inhibitory effects of fluvastatin and its metabolites on both LDL oxidation and cholesterol esterification in macrophages may contribute to the antiatherogenic action in vivo.

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It has been suggested that nitric oxide (NO) is involved in the pathophysiology of epilepsy. Data are, however controversial because it is not clear whether NO has pro- or anticonvulsant effects. The aim of this study was to investigate the effects of NO on penicillin G-induced epileptiform activity. The left cerebral cortex was exposed by craniotomy in urethane-anesthetized Wistar rats. The epileptic activity was produced by intraperitoneal injection of penicillin G (3 million U/kg, i.p.). The ECoG (electrocorticogram) activity was displayed on a four-channel recorder. At 39.7 ± 5.4 min after penicillin administration, large amplitude sharp waves appeared in the ECoG. Mean spike frequency and mean spike amplitude were calculated as 29.5 ± 3.2/min and 865 ± 91 μV, respectively, at the 55th min. 7-Nitroindazole (60 mg/kg, i.p.) injection 30 min before penicillin G administration significantly reduced the latency of epileptiform activity. Intracerebroventricular administration of L-arginine (300 μg/2 μl, i.c.v.) and sodium nitroprusside (100 μg /2 μl, i.c.v.) suppressed epileptiform activity. Saline (2 μl) and D-arginine (300 μg/2 μl, i.c.v.) administration into the cerebral ventricle were completely ineffective on epileptiform activity (P<0.01). These findings suggest that NO may be an endogenous antiepileptic substance.

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The polycyclic aromatic hydrocarbon, 7,12-dimethylbenz[a]anthracene (DMBA), is an immunosuppressor as well as a potent organ-specific carcinogen. To understand the organ-specific mechanism of DMBA-induced lymphoid toxicity, aryl hydrocarbon-nonresponsive mice and microsomal epoxide hydrolase (mEH)-null mice were analyzed. DMBA caused a dose-dependent decrease in spleen weights, but not the thymus weights in aryl hydrocarbon-nonresponsive mice. On the other hand, both spleen and thymus weights were decreased to less than a half in wild-type mice exposed to 30 mg/kg of DMBA. In contrast, no decrease was detected in spleen weights of mEH-null mice exposed to up to 100 mg/kg of DMBA, while thymus weights were markedly lower. Responses to the B-cell mitogen lipopolysaccharide and to T-cell mitogen phytohemagglutinin were nearly completely abolished in splenocytes isolated from wild-type mice treated with 100 mg/kg of DMBA. These responses were decreased, but maintained in splenocytes isolated from mEH-null mice treated with DMBA. Two DMBA metabolites dependent on mEH including DMBA-3,4-diol were detected in an HPLC chromatogram of spleen microsomes isolated from wild-type mice, but not those from mEH-null mice. These results suggest the involvement of mEH in splenic activation of DMBA for immunotoxicity and the difference for the DMBA-induced lymphoid toxicity between spleen and thymus.

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In monocrotaline-treated rat pulmonary artery from which endothelium was removed, greater spontaneous muscular tone was observed under resting conditions than in vehicle-treated artery. The aim of the present study was to show the possible contribution of Cl⁻ channels in the mechanism of the elevated tone. Verapamil almost completely inhibited the elevated spontaneous muscular tone by decreasing [Ca²⁺]_i. The elevated muscular tone was also inhibited by 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid (DIDS), a Cl⁻ channel inhibitor. After the inhibition of muscular tone by DIDS, verapamil did not induce further relaxation. Quantitative RT-PCR analysis indicated that the mRNA levels of ClC3 and Ca²⁺-activated Cl⁻ channels did not change in the pulmonary hypertensive pulmonary artery from those of vehicle-treated rats. These results suggest that the elevated muscular tone observed in the monocrotaline-induced hypertensive pulmonary artery is due to membrane depolarization of smooth muscle cells and that this phenomenon might be mediated by the activation of DIDS-sensitive Cl⁻ channels.

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We have conducted this study to elucidate the influence of GABAergic systems on manifestation of pharmacological activity of desipramine using both pharmacological and electrophysiological methods. Desipramine (20 mg/kg, i.p.) significantly blocked the adjuvant-induced thermal hyperalgesia, which was facilitated by treatment with the GABA_A antagonist picrotoxin (2 mg/kg, i.p.) or the GABA_B antagonist saclofen (2 mg/kg, i.p.). This analgesic effect of desipramine was antagonized by post-treatment with picrotoxin or saclofen. However, none of these compounds showed any effect in normal animals without adjuvant-induced inflammation. In a slice preparation of the hippocampus, treatment with GABA (10⁻⁵ - 5 × 10⁻⁴ M), baclofen (10⁻⁵ - 10⁻⁴ M) or muscimol (10⁻⁵ - 10⁻⁴ M) inhibited the field potential evoked in pyramidal neurons by Schaffer collateral stimulation. The inhibitory effect of GABA was facilitated by concurrent application of desipramine, carbamazepine or diazepam at a concentration of 5 × 10⁻⁵ - 2 × 10 ⁻⁴ M. The rank of order of facilitation is: desipramine > carbamazepine > diazepam. Desipramine also enhanced the inhibitory effect of baclofen and muscimol. These results suggest that desipramine causes GABAergic systems to activate still more, and this phenomenon appears to be involved in manifestation of the pharmacological activity of desipramine such as antinociception.

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We examined the hormonal effects of Z-350, (S)-4-[3-(4-{1-(4-methylphenyl)-3-[4-(2-methoxyphenyl)piperazine-1-yl]propoxy}benzoyl)indole-1-yl]butyric acid hydrochloride, which has both α₁-adrenoceptor blocking activity and steroid 5α-reductase inhibitory activity, in male and female rats. Z-350 administered orally for 14 days at a dose of 30 mg/kg to normal male rats significantly reduced the weight of the prostate and seminal vesicles without affecting the weight of the testis, epididymis, adrenals, kidney or liver. Prostatic levels of dihydrotestosterone decreased dose-dependently, with a slight increase in the level of testosterone at a Z-350 dose of 100 mg/kg. We observed no effects on the weight of the prostate in castrated rats or on the weight of the uterus in normal or 17β-estradiol-treated female rats. These results suggest that Z-350 inhibits prostatic growth via inhibition of steroid 5-reductase without other hormonal effects.

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Neurotropin is a non-protein extract from the dermis of rabbits inoculated with vaccinia virus and has been clinically used as an analgesic in Japan. We present in the current report evidence for its potential therapeutic value against endotoxin shock. Administration of this compound prior to lipopolysaccharide (LPS) challenge resulted in a reversal of a decrease of the mean arterial pressure in rats and also amelioration of lethality in mice. Anti-inducible nitric oxide synthase (iNOS) Western blotting of tissue extracts from LPS-treated mice revealed almost complete suppression of iNOS induction by Neurotropin. The findings in vivo were reproduced in in vitro experiments in which cultured human umbilical vascular endothelial cells were challenged with LPS. Simultaneous treatment of the cells with Neurotropin resulted in complete suppression of iNOS induction and significant reduction of cell death. These results suggested a therapeutic value of Neurotropin in the treatment of endotoxin shock that was linked, at least in part, to suppression of iNOS induction and reduced cell damage in vascular endothelial cells.

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The intrathecal injection of fenvalerate, a sodium channel activator, at doses of 0.01 to 3 μg, dose-dependently induced the duration of a characteristic behavioral syndrome mainly consisting of reciprocal hind limb scratching directed towards caudal parts of the body and biting or licking of the hind legs in mice. Fenvalerate-induced behavior was inhibited by morphine (1 - 10 mg/kg, i.p.). The characteristic behavior was also inhibited by mexiletine, a sodium channel blocker; MK-801, a N-methyl-D-aspartate ion-channel blocker; and GR82334, a neurokinin-1-receptor antagonist. Calphostin C (3 pmol, i.t.), a protein kinase C inhibitor, inhibited fenvalerate-induced behavior. On the other hand, phorbol-12, 13-dibutyrate (50 pmol, i.t.), a protein kinase C activator, markedly enhanced the fenvalerate-induced behavior. The present results also showed that fenvalerate produced thermal allodynia and hyperalgesia in the tail-flick test. Furthermore, fenvalerate-induced thermal allodynia and hyperalgesia were inhibited by the pretreatment with calphostin C. These results suggest that the intrathecal administration of fenvalerate induces a marked nociceptive response and thermal allodynia/hyperalgesia, and they suggest that tetrodotoxin-resistant sodium channels may play an important role in this effect.

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The usefulness of bis-(1,3-dibutylbarbituric acid)-trimethine oxonol (DiBAC₄(3)), a voltage-sensitive fluorescent dye, for the measurement of membrane potentials (MPs) was evaluated in HEK293 cells, where α or α plus β1 subunits of large conductance Ca²⁺-activated K⁺ (BK) channels were expressed (HEKBKα and HEKBKαβ). The fluorescent intensity of DiBAC₄(3) was measured at various potentials under voltage-clamp for calibration to estimate the absolute MP semi-quantitatively. The resting MPs measured with DiBAC₄(3) were roughly comparable to those recorded with a microelectrode; the MP in HEKBKαβ was 10 - 20 mV more negative than that in native HEK. In HEKBKα, the membrane hyperpolarization induced by 10 μM Evans blue, a BK channel opener, was detected with DiBAC₄(3). NS-1619, another BK channel opener, induced gradual but substantial change in F/F_K even in native HEK, while the BK channel opening effect was detected. Oscillatory membrane hyperpolarization was induced in HEKBKαβ by application of 10 μM acetylcholine via increase in intracellular Ca²⁺ concentration. The oscillatory hyperpolarization was, however, detected only as a slow hyperpolarization with DiBAC₄(3). It can be concluded that relatively slow effects of BK channel modulators can be semi-quantitatively measured by use of DiBAC₄(3) in HEKBK, while the limited temporal resolution and possible artifacts should be taken into account.

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Pre-exposure of the rat gastric mucosa to capsaicin reduced the mucosal lesion by 50% ethanol to 1/4. Treatment with an antagonist of calcitonin gene-related peptide (CGRP), CGRP (8 - 37), nullified the effect of capsaicin. During constant perfusion of the gastric lumen with physiological saline + pepstatin, the CGRP level was not increased by 50% ethanol, but it showed a peak (802.5 ± 145.7 pg/2 min) after 1.6 mM capsaicin. Four minutes after capsaicin, the CGRP level was kept at a high level and the gastric lesion was markedly reduced by re-exposure of the mucosa to 50% ethanol. At 20 - 30 min after capsaicin, the CGRP levels returned to the resting level and the reddened area by 50% ethanol was not reduced. It was concluded that capsaicin transiently prevented the mucosal lesion through CGRP release.

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We investigated the apoptotic development and apoptotic-related gene expression after myocardial infarction (MI) at different time points in the current study. Bax gene expression was increased at 12 h after MI and peaked at 24 h. Fas gene started to over-express at 12 h after MI as well but it reached maximum at 72 h. In the MI groups, strongest staining of apoptosis was detected in rats 3 days post operation. Our results demonstrate that apoptotic development after MI is time dependent in the ischemic area and there could be some linkage with the over expression of angiotensin II receptors post MI.

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The effects of a novel tachykinin NK₁-receptor antagonist HSP-117 {(2S,3S)-3-[(5-isopropyl-2,3-dihydrobenzofuran-7-yl)methyl]amino-2-phenylpiperidine dihydrochloride} on cisplatin-induced pica, i.e., the eating of nonnutritive substances such as kaolin were examined in rats. HSP-117 inhibited kaolin intake in a dose-dependent manner for 2 days. The 5-HT₃-receptor antagonist ondansetron inhibited only on the first day, but not on the second day. These results indicate that the cisplatin-induced kaolin intake on the first day is related to both 5-HT₃- and NK₁ receptors, while only the NK₁ receptor is involved on the second day. Thus, cisplatin-induced continuous pica in rats represents a useful model of not only acute but also delayed emesis.

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We studied the effects of β-casomorphins (β-CMs, μ-acting opioid peptides from milk protein β-casein) on survival of primary-cultured chick dorsal root ganglion neurons in the presence of nerve growth factor. β-CM-5 and β-CM-7 had potent neuronal survival-promoting activities. β-CM-4 amide (morphiceptin) and des-Tyr¹-β-CM-7 also exhibited the similar promoting effects, although their effects were very weak. The promoting effect of β-CM-5 was prevented by co-administration of naloxone, or pretreatment with pertussis toxin. These results suggest that the neuronal survival-promoting effects of β-CMs might be mediated through opioid receptors coupled to G proteins.

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The effects of intracerebroventricular administration of calcium or magnesium on the blood pressure regulation in the brain were investigated. The systolic blood pressure in spontaneously hypertensive rats (male, 13-week-old) was decreased by calcium chloride (100 μg/rat) and increased by magnesium chloride (20, 100 or 500 μg/rat). The depressor response induced by calcium was inhibited by magnesium chloride in a dose-dependent manner. Combining these results with those previously reported, it is suggested that magnesium inhibits the ability of calcium to reduce blood pressure through calmodulin- and dopamine-dependent functions in the brain.

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