• 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.

• Biomolecules & Therapeutics
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• v.32 no.5
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• pp.531-539
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• 2024

Sleep is one of the most essential physiological phenomena for maintaining health. Sleep disturbances, such as insomnia, are often accompanied by psychiatric or physical conditions such as impaired attention, anxiety, and stress. Medication used to treat insomnia have concerns about potential side effects with long-term use, so interest in the use of alternative medicine is increasing. In this study, we investigated the hypnotic effects of β-lapachone (β-Lap), a natural naphthoquinone compound, using pentobarbital-induced sleep test, immunohistochemistry, real-time PCR, and western blot in mice. Our results indicated that β-Lap exerts a significant hypnotic effect by showing a decrease in sleep onset latency and an increase in total sleep time in pentobarbital-induced sleep model. The results of c-Fos immunostaining showed that β-Lap decreased neuronal activity in the basal forebrain and lateral hypothalamus, which are wakefulness-promoting brain regions, while increasing in the ventrolateral preoptic nucleus, a sleep-promoting region; all these effects were significantly abolished by 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), an adenosine A₁ receptor (A₁R) antagonist. Western blot analysis showed that β-Lap increased extracellular signal-regulated kinase phosphorylation and nuclear factor-kappa B translocation from the cytoplasm to the nucleus; these effects were inhibited by DPCPX. Additionally, β-Lap increased the mRNA levels of A₁R. Taken together, these results suggest that β-Lap exerts hypnotic effects, potentially through A₁R.

• The Korean Journal of Physiology and Pharmacology
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• v.1 no.6
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• pp.657-664
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• 1997

The effects of adenosine, adenosine A1 receptor antagonist (DPCPX), or NMDA receptor antagonist (APV) on the spontaneous release of $[^3H]-5-hydroxytryptamine$ ($[^3H]-5-HT$) during normoxic/normoglycemic or hypoxic/hypoglycemic period were studied in the rat hippocampal slices. The hippocampus was obtained from the rat brain and sliced $400\;{\mu}m$ thickness with the tissue slicer. After 30 min's preincubation in the normal buffer, the slices were incubated for 30 min in a buffer containing $[^3H]-5-HT$ ($0.1\;{\mu}M,\;74{\mu}Ci/8\;ml$) for uptake, and washed. To measure the release of $[^3H]-5-HT$ into the buffer, the incubation medium was drained off and refilled every ten minutes through sequence of 14 tubes. Induction of glucose/oxygen deprivation (GOD; medium depleting glucose and gassed with 95% $N_2/5%\;CO_2$) was done in 6th and 7th tube. The radioactivities in each buffer and the tissue were counted using liquid scintillation counter and the results were expressed as a percentage of the total radioactivities. When slices were exposed to GOD for 20 mins, the spontaneous release of $[^3H]-5-HT$ was markedly increased and this increase of $[^3H]-5-HT$ release was blocked by adenosine ($10\;{\mu}M$) or DL-2-amino-5-phosphonovaleric acid (APV; $30\;{\mu}M$). Adenosine $A_1$ receptor specific antagonist, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) exacerbate GOD-induced increase of spontaneous release of $[^3H]-5-HT$. These results suggest that Adenosine may play a role in the GOD-induced spontaneous release of $[^3H]-5-HT$ through adenosine $A_1$ receptor activity.

• 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.

• The Korean Journal of Pharmacology
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• v.32 no.1
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• pp.1-11
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• 1996

Since it has been reported that the depolarization-induced norepinephrine (NE) release is inhibited by activation of presynaptic $A_1-adenosine$ heteroreceptor in hippocampus, a large body of experimental data on the post-receptor mechanism of this process has been accumulated. But, the post-receptor mechanism of presynaptic $A_1-adenosine$ receptor on the NE release has not been clearly elucidated yet. Therefore, it was attempted to clarify the post-receptor mechanisms of the $A_1-adenosine$ receptor-mediated control of NE release in this study. Slices from rat hippocampus were equilibrated with $^3H-norepinephrine$ 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. Adenosine, in concentrations ranging from $1{\sim}30{\mu}M$, decreased the NE release in a dose-dependent manner, without affecting the basal rate of release. The adenosine effects were significantly inhibited by 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, $2{\mu}M$), a selective $A_1-receptor$ antagonist. The responses to N-ethylmaleimide (NEM, 10 & $30{\mu}M$), a SH-alkylating agent of G-protein, were characterized by increments of the evoked NE-release and the basal release, and the adenosine effects were completely abolished by NEM pretreatment. $4{\beta}-Phorbol$ 12,13-dibutyrate (PDB, $1{\mu}M$), a specific protein kinase C (PKC) activator, increased the evoked NE release, whereas polymyxin B sulfate (PMB,0.1 mg), a PKC inhibitor, decreased the release, and the adenosine effects were inhibited by these agents. Nifedipine $(1{\mu}M)$, a $Ca^{2+}-channel$ blocker of dihydropyridine analogue, did not affect the adenosine effect. Tetraethylammonium (TEA, 3 mM) increased the evoked NE release, and inhibited the adenosine effects, but glibenclamide, a ATP dependent $K^+-channel$ blocker, did not. Finally, 8-bromo cyclic AMP (100 & $300{\mu}M$), a membrane-permeable analogue of cAMP, did not alter the NE release, but adenosine effects were inhibited by pretreatment with 8br-cAMP. These results suggest that the decrement of the evoked NE-release by $A_1-adenosine$ receptor is mediated by the C-protein, which is coupled to protein kinase C, adenylate cyclase system and TEA sensitive $K^+-channel$, and that nifedipine-sensitive $Ca^{2+}-channel$ and glibenclamide-sensitive $K^+-channel$ are not involved in this process.

• Biomolecules & Therapeutics
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• v.27 no.6
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• pp.584-590
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• 2019

Luteolin, a widespread flavonoid, has been known to have neuroprotective activity against various neurologic diseases such as epilepsy, and Alzheimer's disease. However, little information is available regarding the hypnotic effect of luteolin. In this study, we evaluated the hypnotic effect of luteolin and its underlying mechanism. In pentobarbital-induced sleeping mice model, luteolin (1, and 3 mg/kg, p.o.) decreased sleep latency and increased the total sleep time. Through electroencephalogram (EEG) and electromyogram (EMG) recording, we demonstrated that luteolin increased non-rapid eye movement (NREM) sleep time and decreased wake time. To evaluate the underlying mechanism, we examined the effects of various pharmacological antagonists on the hypnotic effect of luteolin. The hypnotic effect of 3 mg/kg of luteolin was not affected by flumazenil, a GABAA receptorbenzodiazepine (GABAAR-BDZ) binding site antagonist, and bicuculine, a GABAAR-GABA binding site antagonist. On the other hand, the hypnotic effect of 3 mg/kg of luteolin was almost completely blocked by caffeine, an antagonist for both adenosine A1 and A2A receptor (A1R and A2AR), 8-Cyclopentyl-1,3-dipropylxanthine (DPCPX), an A1R antagonist, and SCH-58261, an A2AR antagonist. From the binding affinity assay, we have found that luteolin significantly binds to not only A1R but also A2AR with $IC_{50}$ of 1.19, $0.84{\mu}g/kg$, respectively. However, luteolin did not bind to either BDZ-receptor or GABAAR. From these results, it has been suggested that luteolin has hypnotic efficacy through A1R and A2AR binding.

• The Korean Journal of Physiology and Pharmacology
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• v.23 no.6
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• pp.483-491
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• 2019

Cordycepin exerts neuroprotective effects against excitotoxic neuronal death. However, its direct electrophysiological evidence in Alzheimer's disease (AD) remains unclear. This study aimed to explore the electrophysiological mechanisms underlying the protective effect of cordycepin against the excitotoxic neuronal insult in AD using whole-cell patch clamp techniques. ${\beta}$-Amyloid ($A{\beta}$) and ibotenic acid (IBO)-induced injury model in cultured hippocampal neurons was used for the purpose. The results revealed that cordycepin significantly delayed $A{\beta}$ + IBO-induced excessive neuronal membrane depolarization. It increased the onset time/latency, extended the duration, and reduced the slope in both slow and rapid depolarization. Additionally, cordycepin reversed the neuronal hyperactivity in $A{\beta}$ + IBO-induced evoked action potential (AP) firing, including increase in repetitive firing frequency, shortening of evoked AP latency, decrease in the amplitude of fast afterhyperpolarization, and increase in membrane depolarization. Further, the suppressive effect of cordycepin against $A{\beta}$ + IBO-induced excessive neuronal membrane depolarization and neuronal hyperactivity was blocked by DPCPX (8-cyclopentyl-1,3-dipropylxanthine, an adenosine $A_1$ receptor-specific blocker). Collectively, these results revealed the suppressive effect of cordycepin against the $A{\beta}$ + IBO-induced excitotoxic neuronal insult by attenuating excessive neuronal activity and membrane depolarization, and the mechanism through the activation of $A_1R$ is strongly recommended, thus highlighting the therapeutic potential of cordycepin in AD.

• The Korean Journal of Physiology and Pharmacology
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• v.2 no.1
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• pp.31-39
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• 1998

As it has been reported that the depolarization induced acetylcholine (ACh) release is modulated by activation of presynaptic $A_1$ adenosine heteroreceptor and various lines of evidence suggest the $A_2$ adenosine receptor is present in the hippocampus. The present study was undertaken to delineate the role of adenosine receptors on the hippocampal ACh release. Slices from the rat hippocampus were equilibrated with $[^3H]choline$ and then the release amount of the labelled product, $[^3H]ACh$, which was evoked by electrical stimulation (rectangular pulses, 3 Hz, 2 ms, 24 mA, $5\;V/cm^{-1}$, 2 min), was measured, and the influence of various adenosine receptor-related agents on the evoked tritium outflow was investigated. And also, the drug-receptor binding assay was performed in order to confirm the presence of $A_1$ and $A_2$ adenosine receptors in the rat hippocampus. N-ethylcarboxamidoadenosine (NECA), a potent adenosine receptor agonist with nearly equal affinity at $A_1$ and $A_2$ adenosine receptors, in concentrations ranging from $1{\sim}30\;{\mu}M$, decreased the electrically-evoked $[^3H]ACh$ release in a concentration-dependent manner without affecting the basal rate of release. And the effect of NECA was significantly inhibited by 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, 2 ${\mu}M$), a selective $A_1$ adenosine receptor antagonist, but was not influenced by 3,7-dimethyl-1-propargylxanthine (DMPX, 5 ${\mu}M$), a specific $A_2$ adenosine receptor antagonist. $N^6-cyclopentyladenosine$ (CPA), a selective $A_1$ adenosine receptor agonist, in doses ranging from 0.1 to 10 ${\mu}M$, reduced evoked $[^3H]ACh$ release in a dose-dependent manner without the change of the basal release. And the effect of CPA was significantly inhibited by 2 ${\mu}M$ DPCPX treatment. 2-P-(2-carboxyethyl)-phenethylamino-5'-N-ethylcarboxamidoadenosine hydrochloride (CGS-21680C), a potent $A_2$ adenosine receptor agonist, in concentrations ranging from 0.1 to 10 ${\mu}M$, did not alter the evoked ACh release. In the drug-receptor binding assay, the binding of $[^3H]2-chloro-N^6-cyclopentyladenosine$ ($[^3H]$CCPA) to the $A_1$ adenosine receptor of rat hippocampal membranes was inhibited by CPA ($K_i$ = 1.22 nM), NECA ($K_i=10.17 nM$) and DPCPX ($K_i=161.86 nM$), but not by CGS-21680C ($K_i=2,380 nM$) and DMPX ($K_i=22,367 nM$). However, the specific binding of $[^3H]CGS-21680C$ to the $A_2$ adenosine receptor was not observed. These results suggest that the $A_1$ adenosine heteroreceptor play an important role in evoked ACh release, but the presence of $A_2$ adenosine receptor is not confirmed in this study.

• The Korean Journal of Physiology and Pharmacology
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• v.1 no.3
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• pp.225-231
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• 1997

As it has been reported that the depolarization-induced norepinephrine (NE) release is modulated by activation of presynaptic $A_1-adenosine$ heteroreceptor 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 NE release in this study. Slices from rat hippocampus were equilibrated with $[^3H]-NE$ and the release of the labelled products was evoked by electrical stimulation.(3 Hz, $5Vcm^{-1}$, 2 ms, rectangular pulses). 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 Tanging from 0.1 to $10{\mu}M$ decreased the $[^3H]-NE$ release in a dose-dependent mauler without any change of basal rate of release. 8-Cyclopentyl-1,3-dipropylxanthine (DPCPX, $2{\mu}M$), a selective $A_1-receptor$ antagonist, inhibited the CPA effect. The responses to N-ethylmaleimide $(3&10{\mu}M)$, a SH-alkylating agent of G-protein, were characterized by increments of the evoked NE-release and the CPA effects were completely abolished by NEM pretreatment. Forskolin, a specific adenylate cyclase activator, in concentrations ranging from 0.1 to $30{\mu}M$ increased the evoked and basal rate of NE release in a dose-dependent manner and the CPA effects were inhibited by forskolin pretreatment. Rolipram $(1&10{\mu}M)$, a phosphodiesterase inhibitor, did not affect the evoked NE release but reduced the CPA effect. And 8-bromo-cAMP $(100&300{\mu}M)$, a membrane permeable cAMP analogue inhibited the CPA effect significantly. These results suggest that the $A_1-adenosine$ heteroreceptor plays an important role in NE-release via nucleotide-binding protein $G_i$ 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.1 no.2
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• pp.135-142
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• 1997

As it has been reported that the depolarization-induced norepinephrine (NE) release is modulated by activation of presynaptic $A_1$-adenosine heteroreceptor and various lines of evidence indicate that $A_2$-adenosine receptor also presents in hippocampus, and that the adenosine effect is magnesium dependent, the present study was undertaken to delineate the role of adenosine receptors in the modulation of hippocampal NE release. Slices from the rat hippocampus were equilibrated with $[^3H]-NE$ and the release of the labelled product, $[^3H]-NE$, was evoked by electrical stimulation (3 Hz, 5 V $cm^{-1}$, 2 ms, rectangular pulses), and the influence of various agents on the evoked tritium outflow was investigated. $N^6-cyclo-pentyladenosine$ (CPA), in concentrations ranging from 0.1 to 10 ${\mu}M$, decreased the $[^3H]-NE$ release in a dose-dependent manner without changing the basal rate of release, and these effects were significantly inhibited by 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, 2 ${\mu}M$) treatment. When the magnesium concentration was reduced to 0.4 mM or completely removed, the evoked NE release increased along with decreased basal rate of release. In contrast, increasing the magnesium concentrations to 2.4 and 4 mM, decreased the evoked NE release. The CPA effects on evoked NE release were reducedby magnesium removal, but potentiated by 2.4 mM magnesium in the medium. 5-(N-cyclopropyl)-carboxamodiadenosine (CPCA, 1 & 10 ${\mu}M$), an $A_2$-agonist, decreased the evoked tritium outflow, and this effect was also abolished by DPCPX pretreatment. CGS, a powerful $A_2$-agonist, did not affect the evoked NE release. However, the effects of CPCA and CGS on evoked NE release were significantly increased by pretreatment of DPCPX in the magnesium-free medium. These results indicate that inhibitory effect of $A_1$-adenosine receptor on NE release is magnesium-dependent, and $A_2$-receptor may be present in the rat hippocampus.