• Biomedical Science Letters
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• v.23 no.3
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• pp.194-200
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• 2017

Bone-resorbing osteoclasts play a major role in maintaining bone homeostasis with bone-forming osteoblasts. Although it has been reported that A2B adenosine receptor (A2BAR) regulates osteoclast differentiation, its effects on apoptosis or proliferation of osteoclasts have been less-defined. Here, we demonstrate that A2BAR stimulation regulates macrophage-colony stimulating factor (M-CSF)-mediated osteoclast proliferation. Stimulation with a specific agonist of A2BAR, BAY 60-6583, significantly reduced M-CSF-mediated osteoclast proliferation in a time- and dose-dependent manner. In addition, A2BAR stimulation induced both apoptosis of the cells and cell arrest in the G1 phase with a decrease of cell number in the G2/M phase. Stimulation with BAY 60-6583 inhibited the activation of Akt by M-CSF, whereas M-CSF-induced ERK1/2 activation was not affected. These results suggest that the inhibition of M-CSF-mediated Akt activation by A2BAR stimulation increases apoptotic response of osteoclasts and induces cell cycle arrest in the G1 phase, thus contributing to the down-regulation of osteoclast proliferation.

• The Korean Journal of Pharmacology
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• v.28 no.2
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• pp.129-136
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• 1992

As it has been reported that the depolarization-induced acetylcholine (ACh) release is modulated by activation of presynaptic $A_1-adenosine$ heteroreceptor in hippocampus and various lines of evidence indicate the involvement of adenylate cyclase system in $A_1-adenosine$ post-receptor mechanism in hippocampus, it was attempted to delineate the role of adenylate cyclase system in the $A_1-receptor-mediated$ control of ACh release in this study. Slices from rat hippocampus were incubated with $[^3H]-choline$ and the release of the labelled products was evoked by electrical stimulation $(3\;Hz,\;5\;Vcm^{-1},\;2\;ms,\;rectangular\;pulses)$, and the influence of various agents on the evoked tritium-outflow was investigated. $N^6-cyclopentyladenosine$ (CPA), a specific $A_1-adenosine$ receptor agonist, in concentrations ranging from 0.1 to $10\;{\mu}M$, decreased the $[^3H]-ACh$ release in a dose-dependent manner without the changes of basal rate of release. 8-cyclopentyl-1,3-dipropylxanthine $(DPCPX,\;1{\sim}10\;{\mu}M)$, a selective $A_1-receptor$ antagonist, increased the $[^3H]-ACh$ release in a dose-related fashion with slight increase of basal tritium-release. And the CPA effects were significantly inhibited by DPCPX $(2\;{\mu}M)$ pretreatment and the dose-response curve produced by CPA was shifted to the right. The responses to N-ethylmaleimide $(NEM,\;10\;&\;30\;{\mu}M)$, a SH-alkylating agent of G-protein, were characterized by increments of the evoked ACh-release and the basal release, and the CPA effect were completely abolished by NEM pretreatment. Forskolin, a specific adenylate cyclase activator, in concentrations ranging from 0.3 to $10\;{\mu}M$, increased the evoked ACh-release in a dose-dependent manner and the CPA effects were inhibited by forskolin. These results indicate that the $A_1-adenosine$ heteroreceptor plays an important role in ACh-release via nucleotide-binding protein Gi in the rat hippocampus and that the adenylate cyclase system might be participated in this process.

• The Korean Journal of Physiology and Pharmacology
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• v.2 no.2
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• pp.147-154
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• 1998

It was attempted to clarify the participation of $K^+-channels$ in the post-receptor mechanisms of the muscarinic and $A_1-adenosine$ receptor- mediated control of acetylcholine (ACh) release in the present study. Slices from the rat hippocampus were equilibrated with $[^3H]$choline and the release of the labelled products was evoked by electrical stimulation (3 Hz, 5 V/cm, 2 ms, rectangular pulses), and the influence of various agents on the evoked tritium-outflow was investigated. Oxotremorine (Oxo, $0.1{\sim}10\;{\mu}M$), a muscarinic agonist, and $N^6-cyclopentyladenosine$ (CPA, $1{\sim}30\;{\mu}M$), a specific $A_1-adenosine$ agonist, decreased the ACh release in a dose-dependent manner, without affecting the basal rate of release. 4-aminopyridine (4AP), a specific A-type $K^+-channel$ blocker ($1{\sim}100\;{\mu}M$), increased the evoked ACh release in a dose-related fashion, and the basal rate of release is increased by 3 and $100\;{\mu}M$. Tetraethylammonium (TEA), a non-specific $K^+-channel$ blocker ($0.1{\sim}10\;{\mu}M$), increased the evoked ACh release in a dose-dependent manner without affecting the basal release. The effects of Oxo and CPA were not affected by $3\;{\mu}M$ 4AP co-treatment, but 10 mM TEA significantly inhibited the effects of Oxo and CPA. 4AP ($10\;{\mu}M$)- and TEA (10 mM)-induced increments of evoked ACh release were completely abolished in Ca^{2+}-free$ medium, but these were recoverd in low Ca^{2+}$ medium. And the effects of $K^+-channel$ blockers in low Ca^{2+}$ medium were inhibited by $Mg^{2+}$ (4 mM) and abolished by $0.3\;{\mu}M$ tetrodotoxin (TTX). These results suggest that the changes in TEA-sensitive potassium channel permeability and the consequent limitation of Ca^{2+}$ influx are partly involved in the presynaptic modulation of the evoked ACh-release by muscarinic and $A_1-adenosine$ receptors of the rat hippocampus.

• The Korean Journal of Physiology
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• v.30 no.1
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• pp.1-9
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• 1996

The objective of the present study was to characterize the role of adenosine in regulation of ATP-sensitive $K^+\;channel\;(K_{ATP}\;channel)$ activity in isolated rabbit ventricular myocytes using the patch clamp technique. Internal adenosine had little effects on KaTr channel activity. In an outside-out patch with intrapipette GTP and ATP, external adenosine stimulated $K_{ATP}\;channel$ activity. In an inside-out Patch with intrapipette adenosine, ATP reduced $K_{ATP}\;channel$ activity, and GTP stimulated $K_{ATP}\;channel$ activity. Adenosine receptor activation shifted the half-maximal inhibition Of $K_{ATP}\;channel\;from\;70\;to\;241\;{\mu}m$. These results Suggest that activation of adenosine receptors stimulates $K_{ATP}\;channels$ in rabbit ventricular myocytes by reducing the apparent affinity of the channel for ATP. The effect may be important for activating $K_{ATP}\;channels$ during early phase of myocardial ischemia.

• Proceedings of the PSK Conference
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• 2003.10b
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• pp.189.1-189.1
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• 2003

4-(4-Methoxyphenyl)-2-aminothiazole and 3-(4-methoxyphenyl)-5-aminothiadiazole derivatives have been synthesized and evaluated as selective antagonists for human adenosine A$_3$ receptors. A methoxy group in the 4-position of the phenyl ring and N-acetyl or propionyl substitutions of the aminothiazole and aminothiadiazole templates displayed great increases of binding affinity and selectivity for human adenosine A$_3$ receptors. The most potent A$_3$ antagonist of the present series, N-［3-(4-methoxy-phenyl)-［1,2,4］thiadiazol-5-yl］-acetamide exhibiting a K$\_$i/ value of 0.79 nM at human adenosine A$_3$ receptors, showed antagonistic property in functional assay of cAMP biosynthesis involved in one of the signal transduction pathways of adenosine A$_3$ receptors. (omitted)

• The Korean Journal of Pharmacology
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• v.31 no.2
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• pp.145-152
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• 1995

The effects and interactions of $4{\beta}-phorbol$ 12,13-dibutyrate(PDB) and polymyxin B(PMB) with adenosine on the electrically-evoked norepinephrine (NE) release were studied in the rat hippocampus. Slices from the rat hippocampus were equilibrated with $^3H-noradrenaline$ and the release of the labelled product, $^3H-NE$, which evoked by electrical stimulation$(3\;Hz,\;2\;ms,\;5\;VCm^{-1},\;rectangular\;pulses)$ was measured. PDB$(0.3{\sim}10\;{\mu}M)$, a selective protein kinase C(PKC) activator, increased the evoked NE release in a dose related fashion while increasing the basal rate of release. And the effects of $1\;{\mu}M$ PDB were significantly inhibited by $0.3\;{\mu}M$ tetrodotoxin(TTX) pretreatment or $Ca^{++}-free$ medium. $PMB(0.03{\sim}1\;mg)$, a specific PKC inhibitor, decreased the NE release in a dose dependent manner while increasing the basal rate of release. Adenosine $(1{\sim}10\;{\mu}M)$ decreased the NE release without changing the basal rate of release, and this effect was significantly inhibited by 8-cyclopentyl-1,3-dipropylxanthine$(2\;{\mu}M)$, a selective $A_1-receptor$ antagonist, treatment. Also, adenosine effects were significantly inhibited by PDB-and PMB-pretreatment. These results suggest that the PKC plays a role in the NE release in the rat hippocampus and might be participated in a post-receptor mechanism of the $A_1-adenosine$ receptor.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1997.11a
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• pp.27-33
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• 1997

• Biomedical Science Letters
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• v.23 no.4
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• pp.327-332
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• 2017

A2B adenosine receptor (A2BAR) is known to be a regulator of bone homeostasis, but the regulatory mechanism of A2BAR on the osteoclast proliferation are poorly explored. Recently, we have shown that stimulation with BAY 60-6583, a specific agonist of A2BAR, significantly reduced macrophage-colony stimulating factor (M-CSF)-induced osteoclast proliferation by inducing cell cycle arrest at G1 phase and increasing the apoptosis of osteoclasts. The objective of this study was to investigate the regulatory mechanisms of cell cycle and apoptosis by A2BAR stimulation. The expression of A2BAR and M-CSF receptor, c-Fms, was not changed by A2BAR stimulation whereas M-CSF effectively induced c-Fms expression during osteoclast proliferation. Interestingly, A2BAR stimulation remarkably increased the expression of $p27^{Kip-1}$, a cell cycle inhibitor, but the expression of Cyclin D1 and cdk4 was not affected. In addition, while BAY 60-6583 treatment reduced the expression of Bcl2, an anti-apoptotic oncogene, it failed to regulate the expression of Bax, a pro-apoptotic marker. Taken together, these results imply that the increase of $p27^{Kip-1}$ inducing cell cycle arrest at G1 phase and the decrease of Bcl2 inducing anti-apoptotic response by A2BAR stimulation contribute to the down-regulation of osteoclast proliferation.

• YAKHAK HOEJI
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• v.44 no.6
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• pp.499-504
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• 2000

We previously demonstrated that 6-paradol, a compound structurally related to capsaicin, showed to produce prolonged analgesia in experimental animals. The effects of 6-paradol on the release of adenosine were investigated in the rat spinal cord synaptosomes by high performance liquid chromatography. In the presence of $Ca^{++}$, adenosine was released from synaptosomes of rat spinal cord by 6-paradol and capsaicin in a dose dependent manner. Nifedifine, L-type voltage sensitive calcium channel blocker, was found to be ineffective in releasing adenosine by $10\;{\mu}M$ 6-paradol. After exposure to $10\;{\mu}M$ capsazepine, a novel capsaicin selective antagonist, the level of adenosine evoked by $10\;{\mu}M$ 6-paradol was decreased by 75%, and that evoked by $10\;{\mu}M$ capsaicin was blocked completely. These results suggest that the analgesic effect of 6-paradol might be mediated by the vanilloid (capsaicin) sensitive pathway, or the direct binding to the vanilloid receptor.

• The Korean Journal of Pharmacology
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• v.29 no.2
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• pp.175-182
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• 1993

As it has been reported that the depolarization-induced ACh release is modulated by activation of presynaptic $A_1-adenosine$ heteroreceptor in hippocampus and various lines of evidence indicate the adenosine effect is magnesium dependent, the present study was undertaken to delineate the role of endogenus adenosine as a modulator of hippocampal acetylcholine release in this study. Slices from the rat hippocampus were equilibrated with $[^3H]-choline$ and the release of the labelled product, $[^3H]-ACh$, was evoked by electrical stimulation(3Hz, $5\;V\;cm^{-1},$ 2ms, rectangular pulses), and the influence of various agents on the evoked tritium outflow was investigated. Adenosine, in concentrations ranging from $0.3\;to\;100\;{\mu}M$, decreased the $[^3H]-ACh$ release in a dose-dependent manner without changing the basal rate of release. $DPCPX(1{\sim}10{\mu}M)$, a selective $A_1-receptor$ antagonist, increased the $[^3H]-ACh$ release in a dose-related fashion with slight increase of basal tritium release. And the effects of adenosine were significantly inhibited by $DPCPX(2{\mu}M)$ treatment. CPCA, a specific $A_2-agonist$, in concentration ranging from $0.3\;to\;30\;{\mu}M$ decreased evoked tritium outflow with increase of basal rate of tritium release, and these effects were also abolished by $DPCPX(2{\mu}M)$ pretreatment. But, $CGS(0.1{\sim}10{\mu}M)$, a recently introduced potent $A_2-agonist$, did not alter the evoked tritium outflow. When the magnesium concentration of the medium was reduced to 0 mM, there was no change in evoked ACh release by adenosine. In contrast, increasing the magnesium concentration to 4 mM, the inhibitory effects of adenosine were significantly potentiated. These results indicate that $A_1-adenosine$ heteroreceptor is involved in ACh-release in the rat hippocampus and the inhibitory effects of adenosine mediated by $A_1-receptor$ is magnesium-dependent.