The Ca²⁺ signal is the primary determinant of the contraction of the vascular smooth muscle. However, the alteration of the Ca²⁺ sensitivity of the contractile apparatus also plays an essential role. The regulation of the myosin light chain phosphatase (MLCP) activity is considered to be the most important mechanism underlying the regulation of Ca²⁺ sensitivity. The investigations during the last 15 years have identified many proteins that participate in the regulation of the MLCP activity. Recently, the Ca²⁺ signal has also been shown to cross-talk with the mechanisms regulating the Ca²⁺ sensitivity. Consequently, Rho kinase, protein kinase C, CPI-17, and MYPT1 have all been suggested to play a physiologically important role in the regulation of the MLCP activity. We are now close to elucidating the major rules regulating the MLCP activity and the Ca²⁺ sensitivity during vascular contractions. This article will give an overview of the current understanding of the biochemical basis for the regulation of the MLCP activity, while also discussing their functional roles from a physiological point of view. I hope this article will help to develop new pharmacological strategies for the prevention and treatment of the pathological vasoconstriction often seen in vascular diseases.

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There are few efficacious medications for drug dependence at present. Recent evidence has suggested that various cytokines are involved in the effects of abused drugs, suggesting that these factors play a role in drug dependence. In this article, the roles of glial cell line-derived neurotrophic factor (GDNF) and tumor necrosis factor-α (TNF-α) in drug dependence are discussed. GDNF inhibits the cocaine-induced upregulation of tyrosine hydroxylase activity in the ventral tegmental area and blocks behavioral responses to cocaine. TNF-α attenuates rewarding effects and locomotor sensitization induced by methamphetamine (METH) and morphine (MOR). Moreover, we mentioned the potential of Leu-Ile, which induces the expression of GDNF and TNF-α, as a novel therapeutic agent for drug dependence. Leu-Ile inhibits not only the development but also the maintenance of METH- or MOR-induced place preference and locomotor sensitization in mice. The inhibitory effect of Leu-Ile on METH- or MOR-induced place preference is not observed in GDNF heterozygous and TNF-α knockout mice. Leu-Ile inhibits METH- or MOR-induced place preference and sensitization by attenuating the METH- or MOR-induced increase in extracellular dopamine levels in the nucleus accumbens via the induction of GDNF and TNF-α expression. These findings suggest that Leu-Ile could be a novel therapeutic agent for drug dependence.

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Taking into consideration that the search for drugs capable of modifying blood flow through human radial artery (RA) is warranted, the present study was designed to examine the vasodilatatory effects of the potassium channel opener, pinacidil on the RA and to define the contribution of different K⁺-channel subtypes in the endothelium-independent pinacidil action on this blood vessel. Pinacidil relaxed the RA rings with endothelium and without endothelium with comparable potency. N-nitro-L-arginine methyl ester (L-NAME) and methylene blue did not affect the pinacidil-induced vasorelaxation in rings with endothelium. In the rings without endothelium, the K⁺-channel blockers glibenclamide and tetraethylammonium (TEA) moderately antagonized the pinacidil-induced relaxation, while charybdotoxin and 4-aminopiridine did not. In endothelium-denuded rings, precontracted with 100 mM K⁺, the relaxant responses to pinacidil were highly significantly shifted to the right compared to those obtained in RA precontracted with phenylephrine, but pinacidil-induced maximal relaxation was not affected. Addition of nifedipine did not but addition of nifedipine and nickel (Na⁺-Ca²⁺ exchanger inhibitor) did cause a statistically significant rightward shift of the pinacidil concentration-relaxation curve, although the effect 0.1 mM pinacidil was preserved. Thus, pinacidil induces relaxation of the human RA in endothelium-independent manner, and glibenclamide- and TEA-sensitive vascular smooth muscle K⁺ channels are probably involved. Its ability to completely relax the RA precontracted with K⁺-rich solution suggests that pinacidil has additional K⁺ channel-independent mechanism(s) of action. It seems that stimulation of the forward mode of the Na⁺-Ca²⁺ exchanger plays a part in this K⁺ channel-independent effect of pinacidil.

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The purpose of the current study was to explore the effects of N-methyl-D-aspartate (NMDA)-receptor antagonists (MK-801 and LY235959) administered intracerebroventricularly on the changes of both core temperature and hypothalamic levels of 2,3-dihydroxybenzoic acid (2,3-DHBA) induced by intracerebroventricular injection of glutamate (100 – 400 μg at 10 μl/rabbit) or intravenous administration of lipopolysaccharide (LPS) (2 μg/kg) in rabbits. The measurements of 2,3-DHBA were used as an index of the intrahypothalamic levels of hydroxyl radicals. The rise in both the core temperature and hypothalamic 2,3-DHBA could be induced by intracerebroventricular injection of glutamate or intravenous administration of LPS. The glutamate- or LPS-induced fever and increased hypothalamic levels of 2,3-DHBA were significantly antagonized by pretreatment with injection of MK-801 or LY235959 1 h before glutamate or LPS injection. The increased levels of prostaglandin E₂ in the hypothalamus induced by glutamate or LPS could be suppressed by MK-801 or LY235959. The data demonstrate that prior antagonism of NMDA receptors in the brain, in addition to reducing prostaglandin E₂ production in the hypothalamus, suppresses both the glutamate- and LPS-induced fever and increased hypothalamic hydroxyl radicals.

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The effects of a peroxisome proliferator activated receptor γ (PPARγ) agonist on hepatic stearoyl-CoA desaturase (SCD) in insulin-resistant and obese Zucker fa/fa rats were studied. The administration of pioglitazone, a PPARγ agonist, to Zucker obese rats greatly improved their insulin sensitivity. The treatment of Zucker obese rats with pioglitazone did not affect the index of fatty acid desaturation of either serum or liver. Hepatic SCD activity and the mRNA level of SCD1 were not changed by treatment of the rats with pioglitazone. The activity of palmitoly-CoA chain elongase, which is involved in the biosynthesis of oleic acid in concert with SCD, was not significantly altered when Zucker obese rats received pioglitazone. Although neither the activity nor mRNA expression of acyl-CoA oxidase was changed by treatment of Zucker obese rats with pioglitazone, the mRNA expressions of both sterol regulatory element-binding protein-1c and acetyl-CoA carboxylase sensitively responded to the challenge by pioglitazone. These results suggest that the insulin sensitivity of insulin-resistant and obese Zucker fa/fa rats is improved by pioglitazone independently of SCD activity.

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Clenbuterol [CLE: 4-amino-α(t-butyl-amino)methyl-3,5-dichlorobenzyl alcohol] is well known as a potent β₂-adrenergic agonist and non-steroidal anabolic drug, and thus it is generally used for sports doping and asthma therapy. Although the functions of immune cells such as white blood cells (WBCs) have shown to be modulated through β₂-adrenoceptors, the effects of CLE on immune-responsive systems have not been elucidated systematically. Therefore, the effects of CLE on the number of WBCs were studied in rats. Male adult rats were divided into CLE-administered group and the control group to compare the number of total WBCs, neutrophils, monocytes, lymphocytes, eosinophils, and basophils. The administration (dose = 1.0 mg · kg⁻¹ body weight · day⁻¹, s.c.) of CLE was maintained for 30 days. CLE did not change the number of total WBCs during the experimental period. However, CLE increased significantly the number of neutrophils and monocytes, while CLE decreased drastically the number of lymphocytes and eosinophils. There was no significant change in the number of basophils between both groups. These results suggest that the administration of CLE induces drastic redistribution of WBCs in circulation without changing the number of total WBCs, and these responses of WBCs during the administration of CLE are sustained for at least 30 days.

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The behavior of isolation-reared mice has not yet been studied in the staircase test. The present study examined the effects of anxiolytic or anxiogenic agents and isolation rearing on the behavior of ddY (outbred) strain mice in the staircase test. Diazepam and phenobarbital increased the number of steps climbed, but did not affect rearing behavior in group-reared mice. FG-7142, a benzodiazepine inverse agonist, significantly increased the number of rearing with no changes in the number of steps climbed in group-reared mice. Methamphetamine increased the number of steps climbed and decreased the number of rearing in group-reared mice. Although isolation-reared mice showed hyperactivity, there was no difference in locomotor activity for the test period of 3 min between isolation- and group-reared mice. Under these conditions, isolation rearing increased the numbers of steps climbed and rearing compared to group-reared controls. Microanalysis of locomotor patterns of group-reared mice in the staircase test showed that anxiolytic drugs increased the number of climbing to the top step of the staircase and methamphetamine increased the number of climbing to the first to third step. These results suggest that isolation rearing causes an anxiety-like state with increased exploratory behavior in mice.

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We previously demonstrated that electroacupuncture (EA) stimulation both produced antinociception and attenuated intrathecal (i.t.) morphine analgesia, suggesting that EA is capable of inducing two opposing systems, that is, opioid and anti-opioid mechanisms. This study examined the involvement of cholecystokinin (CCK) in the anti-opioid effects following EA in the spinal cord. EA was applied to commonly used acupoints for antinociception, ST-36 located 5-mm lateral to the anterior tubercle of the tibia, and analgesia was assessed by the hind-paw pressure test in male Sprague-Dawley rats. I.t. administration of CCK (0.01 – 10 μg) attenuated i.t. morphine analgesia (10 μg) dose-dependently. The attenuation of morphine analgesia following EA was reversed by i.t. proglumide, a CCK-receptor antagonist (0.01 μg). CCK-like immunoreactivity was increased in lamina I and II in the dorsal horn, and expression of spinal CCK mRNA increased after EA. Moreover, i.t. pretreatment with the neurokinin-1 (NK1)-receptor antagonist L-703,606 (18 μg) reversed both EA- and CCK-induced attenuation of morphine analgesia. These results suggest that CCK-mediated neural systems in the spinal cord may be involved in the attenuation of morphine analgesia following EA and that substance P-induced activation of NK1 receptors may be responsible for the downstream neuronal transmission of the CCK-mediated neuronal system.

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The aim of the present study was performed to determine whether a novel histone deacetylase (HDAC) inhibitor, N-(2-aminophenyl)-4-{[benzyl(2-hydroxyethyl)amino]methyl} benzamide (K-183), prevents a reversible cardiac hypertrophy induced by isoproterenol and improves left ventricular (LV) dysfunction in rats. Either isoproterenol or vehicle was infused for 3 days by osmotic minipump. One hour prior to the implantation of isoproterenol, K-183 or trichostatin A (TSA) was injected twice a day for 3 days. We recorded continuous LV pressure-volume (P-V) loops of in situ hearts one hour after removal of the osmotic minipump. LV work capability (systolic P-V area at midrange LV volume: PVA_mLVV) and hemodynamics were evaluated. K-183 per se induced neither cardiac hypertrophy nor collagen production. Although K-183 did not prevent the hypertrophy, where PVA_mLVV remained decreased, K-183, differently from TSA, significantly attenuated the decrease of cardiac output and the increase of effective arterial elastance in the hypertrophied heart. These results indicate that the novel HDAC inhibitor K-183 has some beneficial effects on hemodynamics, although K-183 has no effects of anti-hypertrophic modalities.

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Oleoyl-estrone (OE) decreases appetite, maintains energy expediture, induces lipolysis (sparing protein), and decreases cholesterolemia and insulin resistance. Rimonabant (SR141716) is a cannabinoid-receptor inhibitor that decreases appetite and mobilizes fat. We studied whether their combination improves their slimming effects. Male overweight rats received daily gavages of 5.3 mg/kg OE, 10 mg/kg rimonabant, or both drugs during 10 days. Body weight and composition, energy balance, adipose tissue weight, and serum hormones and metabolites were measured. OE halved food intake and maintained energy expenditure at the expense of body fat. Rimonabant effects on appetite and energy balance were less marked, resulting in lower lipid mobilization. OE and rimonabant followed the OE pattern, with no additive or synergic effects. Glycemia was maintained, but OE decreased insulin, GLP-1, and cholesterol, whilst rimonabant increased cholecystokinin and cholesterol, and decreased NEFA. Both drugs decreased leptin and triacylglycerols; ghrelin was unchanged. The results hint at different mechanisms of action of both drugs: we can assume that OE effects do not involve the cannabinoid pathway. OE does not seem to act, either, after 10 days, through the secretion of ghrelin or the intestinal appetite-controlling peptides tested.

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Adenosine inhibited interleukin (IL)-18 production in lipopolysaccharide (LPS)-stimulated monocytes. The action of adenosine was antagonized by an adenosine A₂A-receptor (A2AR) antagonist and was mimicked by an A2AR agonist, suggesting that the stimulation of A2AR may be involved in the actions of adenosine. On the other hand, the stimulation of A1R and A3R inhibited the actions of A2AR stimulation, whereas the stimulation of A2BR had no effect on them. Activation of A2AR is known to increase cyclic adenosine monophosphate (cAMP) levels and to activate protein kinase A (PKA). A PKA inhibitor prevented the actions of A2AR stimulation, indicating that the action mechanism of A2AR stimulation may be via the activation of the cAMP/PKA pathway.

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A possible involvement of histamine in acute radiation dermatitis in mice was investigated. The dose of 40 Gy of gamma irradiation induced erythema and edema in C57BL/6 mice treated with vehicle. However, in C57BL/6 mice treated with chlorpheniramine and WBB6F1-W/W^v mice, erythema and edema were not observed. In all of these mice, epilation and dry desquamation were induced, but bepotastine significantly reduced the extent of these areas. These results suggest that gamma irradiation-induced erythema and edema were caused by histamine released from mast cells via histamine H₁ receptor, and epilation was induced by other inflammatory mediators.

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Low-cost, simple procedures for organotypic tissue cultures are desirable for high-throughput biological experiments such as large-scale medical/drug screening. We present a practical and economical method to cultivate brain slices using hydrophilic filtration membranes. With a cost reduction of more than 90%, this technique allows us to prepare hippocampal slice cultures that are morphologically and functionally indistinguishable from those obtained by the widely used Millicell-CM^® method.

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