• The Korean Journal of Physiology and Pharmacology
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• v.14 no.1
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• pp.21-28
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• 2010

Phenolic compounds affect intracellular free $Ca^{2+}$ concentration ($[Ca^{2+}]_i$) signaling. The study examined whether the simple phenolic compound octyl gallate affects ATP-induced $Ca^{2+}$ signaling in PC12 cells using fura-2-based digital $Ca^{2+}$ imaging and whole-cell patch clamping. Treatment with ATP ($100\;{\mu}M$) for 90 s induced increases in $[Ca^{2+}]_i$ in PC12 cells. Pretreatment with octyl gallate (100 nM to $20\;{\mu}M$) for 10 min inhibited the ATP-induced $[Ca^{2+}]_i$ response in a concentration-dependent manner ($IC_{50}=2.84\;{\mu}M$). Treatment with octyl gallate ($3\;{\mu}M$) for 10 min significantly inhibited the ATP-induced response following the removal of extracellular $Ca^{2+}$ with nominally $Ca^{2+}$-free HEPES HBSS or depletion of intracellular $Ca^{2+}$ stores with thapsigargin ($1\;{\mu}M$). Treatment for 10 min with the L-type $Ca^{2+}$ channel antagonist nimodipine ($1\;{\mu}M$) significantly inhibited the ATP-induced $[Ca^{2+}]_i$ increase, and treatment with octyl gallate further inhibited the ATP-induced response. Treatment with octyl gallate significantly inhibited the $[Ca^{2+}]_i$ increase induced by 50 mM KCI. Pretreatment with protein kinase C inhibitors staurosporin (100 nM) and GF109203X (300 nM), or the tyrosine kinase inhibitor genistein ($50\;{\mu}M$) did not significantly affect the inhibitory effects of octyl gallate on the ATP-induced response. Treatment with octyl gallate markedly inhibited the ATP-induced currents. Therefore, we conclude that octyl gallate inhibits ATP-induced $[Ca^{2+}]_i$ increase in PC12 cells by inhibiting both non-selective P2X receptor-mediated influx of $Ca^{2+}$ from extracellular space and P2Y receptor-induced release of $Ca^{2+}$ from intracellular stores in protein kinase-independent manner. In addition, octyl gallate inhibits the ATP-induced $Ca^{2+}$ responses by inhibiting the secondary activation of voltage-gated $Ca^{2+}$ channels.

• The Korean Journal of Physiology and Pharmacology
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• v.17 no.1
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• pp.57-64
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• 2013

Cells can resist and even recover from stress induced by acute hypoxia, whereas chronic hypoxia often leads to irreversible damage and eventually death. Although little is known about the response(s) to acute hypoxia in neuronal cells, alterations in ion channel activity could be preferential. This study aimed to elucidate which channel type is involved in the response to acute hypoxia in rat pheochromocytomal (PC12) cells as a neuronal cell model. Using perfusing solution saturated with 95% $N_2$ and 5% $CO_2$, induction of cell hypoxia was confirmed based on increased intracellular $Ca^{2+}$ with diminished oxygen content in the perfusate. During acute hypoxia, one channel type with a conductance of about 30 pS (2.5 pA at -80 mV) was activated within the first 2~3 min following onset of hypoxia and was long-lived for more than 300 ms with high open probability ($P_o$, up to 0.8). This channel was permeable to $Na^+$ ions, but not to $K^+$, $Ca^+$, and $Cl^-$ ions, and was sensitively blocked by amiloride (200 nM). These characteristics and behaviors were quite similar to those of epithelial sodium channel (ENaC). RT-PCR and Western blot analyses confirmed that ENaC channel was endogenously expressed in PC12 cells. Taken together, a 30-pS ENaC-like channel was activated in response to acute hypoxia in PC12 cells. This is the first evidence of an acute hypoxia-activated $Na^+$ channel that can contribute to depolarization of the cell.

• Journal of Oriental Neuropsychiatry
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• v.17 no.1
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• pp.37-57
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• 2006

Objective : This research investigates the effect of the CMT and MCMT on Alzheimer‘s disease. Methods : The effects of the CMT and MCMT extract on (1) amyloid precursor proteins(APP), acetylcholinesterase(AChE) mRNA of PC-12 cells treated with CT-105; (2) the AChE activity and the APP production of PC-12 cell treated with CT-105; (3) the behavior; (4) expression of $IL-1{\beta}$, $TNF-{\alpha}$, MDA, GFAP, CD68 abd CD11b; (5) the infarction area of the hippocampus in Alzheimer's diseased mice induced with ${\beta}A$ were investigated. Results : 1. The CMT and MCMT extract suppressed the expression of APP, AChE, and mRNA in PC-12 cells treated with CT-105. 2. The CMT and MCMT extract suppressed the AChE activity, and the production of APP significantly in PC-12 cells treated with CT-105. 3. For the CMT and MCMT extract group a significant inhibitory effect on the memory deficit was shown for the mice with Alzheimer's disease induced by ${\beta}A$ in the Morris water maze experiment, which measured stop-through latency, and distance movement-through latency. 4. The CMT and MCMT extract suppressed the over-expression of $IL-1{\beta}$, $TNF-{\alpha}$, MDA, GFAP, CD68 abd CD11bCD68/GFAP, in the mice with Alzheimer's disease induced by ${\beta}A$. 5. The CMT and MCMT extract reduced the infarction area of hippocampus with Alzheimer's disease induced by ${\beta}A$ 6. The MCMT showed more excellent effects than CMT in the every experiments except PC-12 cells. Conclusions : These results suggest that the CMT and MCMT extract may be effective for the prevention and treatment of Alzheimer's disease. Investigation into the clinical use of the CMT and MCMT extract for Alzheimer's disease is suggested for future research.

• Journal of Oriental Neuropsychiatry
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• v.15 no.2
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• pp.119-139
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• 2004

This research investigates the effect of the Hibiscus syriacus(HSS) on Alzheimer's disease. Specifically, the effects of the HSS extract on (1) $IL-1{\beta}$, IL-6, and $TNF-{\alpha}$ mRNA of PC-12 cells treated with LPS; (2) amyloid precursor proteins(APP), acetylcholinesterase(AChE), and glial fibrillary acidic protein(GFAP) mRNA of PC-12 cells treated with CT-105; (3) the AChE activity and the APP production of PC-12 cell treated with CT-105; (4) the behavior; (4) expression of $IL-1{\beta}$, $TNF-{\alpha}$, $IL-1{\beta}$ mRNA, $TNF-{\alpha}$ mRNA, and reactive oxygen species(ROS); (5) the infarction area of the hippocampus, and brain tissue injury in Alzheimer's diseased mice induced with ${\beta}A$ were investigated. The results were summarized below ; 1. The HSS extract suppressed the expression of $IL-1{\beta}$, IL-6 and $TNF-{\alpha}$ mRNA in THP-l cells treated with LPS. 2. The HSS extract suppressed the expression of APP, AChE, and GFAP mRNA in PC-12 cells treated with CT-105. 3. The HSS extract suppressed the AChE activity, and the production of APP significantly in PC-12 cells treated with CT-105. 4. For the HSS extract group a significant inhibitory effect on the memory deficit was shown for the mice with Alzheimer's disease induced by ${\beta}A$ in the Morris water maze experiment, which measured stop-through latency, and distance movement-through latency. 5. The HSS extract suppressed the over-expression of $IL-1{\beta}$, $TNF-{\alpha}$, $IL-1{\beta}$ and $TNF-{\alpha}$ mRNA, CD68/GFAP, ROS in the mice with Alzheimer's disease induced by ${\beta}A$. 6. The HSS extract reduced the infarction area of hippocampus, and controlled the injury of brain tissue in the mice with Alzheimer's disease induced by ${\beta}A$. These results suggest that the HSS extract may be effective for the prevention and treatment of Alzheimer's disease. Investigation into the clinical use of the HSS extract for Alzheimer's disease is suggested for future research.

• Biomedical Science Letters
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• v.13 no.3
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• pp.197-206
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• 2007

P2X receptors are membrane-bound ion channels that conduct $Na^+,\;K^+$, and $Ca^{2+}$ in response to ATP and its analogs. There are seven subunits identified so far ($P2X_1-P2X_7$). $P2X_2$ receptors are known to be expressed in a wide range of organs including brains and adrenal grands. PC12 cells are originated from adrenal grand and differentiated by nerve growth factor or pituitary adenylate cyclase activating poly peptide (PACAP). Previous studies indicate that $P2X_2$ receptor activation in PC12 cells couples to $Ca^{2+}-dependent$ release of catecholamine and ATP. It is known that acidic pH potentiates ATP currents at $P2X_2$ receptors. This leads to a hypothesis that $P2X_2$ receptors may play an important role in PC12 cell differentiation, one of the characteristics of which is neurite outgrowth, induced by the hormones under lower pH. In the present study, we isolated several clones which potentiate neurite outgrowth by PACAP in acidic pH (6.8), but not in alkaline pH (7.6). RT-PCR and electrophysiology data indicate that these clones express only functional $P2X_2$ receptors in the absence or presence of PACAP for 3 days. Potentiation of neurite outgrowth resulted from PACAP (100 nM) in acidic pH is inhibited by the two P2X receptor antagonists, suramin and PPADS ($100\;{\mu}M)$ each), and exogenous exprerssion of ATP-binding mutant $P2X_2$ receptor subunit ($P2X_2[K69A]$). However, acid sensing ion channels (ASICs) are not involved in PACAP-induced neurite outgrowth potentiation in lower pH since treatments of an inhibitor of ASICs, amyloride ($10\;{\mu}M$), did not give any effects to neurite extension. The vesicular proton pump ($H^+-ATPase$) inhibitor, bafilomycin (100 nM), reduced neurite extension indicating that ATP release resulted from $P2X_2$ receptor activation in PC12 cells is needed for neurite outgrowth. These were confirmed by activation of mitogen activated protein kinases, such as ERKs and p38. These results suggest roles of ATP and $P2X_2$ receptors in hormone-induced cell differentiation or neuronal synaptogenesis in local acidic environments.

• Korean Journal of Food Science and Technology
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• v.36 no.6
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• pp.1026-1031
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• 2004

Acetylcholinesterase inhibitory activity and protective effect against cytotoxicity of PC 12 cell induced by beta-amyloid protein and glutamate were examined in perilla seed methanol extract and its solvent fractions. Methanol extract of perilla seed showed dose-dependent acetylcholinesterase inhibitory activity, with n-butanol fraction showing strongest activity. Perilla seed methanol extract also decreased glutamate- and ${\beta}-amyloid$ protein $(A{\beta})-induced$ cytotoxicities of PC 12 cells dose-dependently. Formation of TBARS induced by $FeSO_{4^-}H_2O_2$ in rat brain was significantly reduced by perilla seed methanol extract, with strongest protective activity formation of TBARS shown in n-butanol fraction. Results suggest perilla seed methanol extract may attenuate actylcholinesterase activity and cytotoxicity induced by glutamate and ${\beta}-amyloid$ protein through suppression of oxidative stress.

• Korean Journal of Food Science and Technology
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• v.49 no.5
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• pp.538-543
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• 2017

Astringent persimmon (Diospyros kaki Thunb. cv. Cheongdo-Bansi) peel with the astringency removed, which is a by-product of dried persimmon (gotgam), was investigated for its antioxidant and neuroprotective properties. A mixture of peel and 40% (v/v) aqueous ethanol was subjected to ultrasonication and then thermal and nonthermal treatments, to produce thermally-treated and nonthermally-treated persimmon peel extracts (TPE and NTPE, respectively). The total phenolic and flavonoid contents and the antioxidant capacity of TPE was approximately 1.3-1.8 times higher than those of NTPE. TPE resulted in the increased viability of neuronal PC-12 cells compared with NTPE. Furthermore, intracellular oxidative stress in PC-12 cells was more decreased by treatment with TPE than NTPE. Cholinesterases, such as acetylcholinesterase and butyrylcholinesterase, were more inhibited by treatment with TPE than NTPE. These results suggest that TPE is useful as a functional material to decrease oxidative stress in neuronal cells and to inhibit cholinesterases.

• Korean Journal of Plant Resources
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• v.26 no.4
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• pp.417-425
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• 2013

The aim of this study was to examine improving effect of Platycodon extracts (PE) and/or Platycodon extracts jelly (PEJ) on cognitive impairment in vitro and in vivo. PC12 (Pheochromocytoma) cells were pretreated with PE for 1hr and than incubated with $50{\mu}M$ amyloid ${\beta}(A{\beta})_{25-35}$ for additional 48hr. Cell viability was assessed by WST-1. Animals for Morris water test and passive avoidance test were divided into normal, control and two Platycodon extracts treated groups that were named Normal (n=7), Control (0 mg/kg, n=7), PE (300 mg/kg, n=7), PEJ (10 g/kg, n=7). Cognitive impairment was induced by scopolamine (1 mg/kg/body weight, i.p.) in the three experimental groups but not the normal group. Pretretment of PE (0.01-1 mg/mL) were not induced cytotoxicity but observed in high dose-treated group (5 and 10 mg/mL) in PC12 cells. Protective effects of PE against $A{\beta}$-induced cytotoxicity were increased in dose dependent manner in PC12 cells. Administration of PE and PEJ were significantly reduced escape latency time on Morris water maze test and passive avoidance test in copolamine-induced cognitive impairment animal model. These results suggest that Platycodon extracts and its related product available to ameliorative purpose for cognitive ability impairments.

• Animal cells and systems
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• v.14 no.4
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• pp.261-266
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• 2010

PC12 cells were differentiated into the cells of chromaffin phenotype by butyrate treatment. Cells were aggregated and formed tight cell adhesion. To investigate the molecular change in this differentiation, we examined expression levels of cell adhesion proteins and extracellular proteins during butyrate induced-differentiation of PC12 cells. Integrin ${\beta}1$, integrin ${\alpha}7$, E cadherin, VCAM, collagen-I, fibronectin, desmoglein and connexin were increased during differentiation. The levels of clusterin and secreted clusterin were also increased. These increased levels of cell adhesion proteins and extracellular proteins appear to induce cell aggregation and tight cell adhesion. The levels of p21, p27 and p16 were increased probably because of differentiation-related growth arrest during differentiation. Prolonged incubation of butyrate up to 1 day was required for differentiation. Signal transduction pathways for this differentiatiom could not be identified since various inhibitors had no effect. The results showed that butyrateinduced differentiation of PC12 cells to chromaffin cells involves tight cell adhesion and induction of extracellular proteins and cell adhesion proteins.

• Journal of Oriental Neuropsychiatry
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• v.16 no.1
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• pp.97-117
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• 2005

This research investigates the effect of the Chaenomelis fructus(CMF) on Alzheimer's disease. Specifically, the effects of the CMF extract on (1) >$IL-1{\beta}$, IL-6, and $TNF-{\alpha}$ mRNA of PC-12 cells treated with LPS; (2) amyloid precursor proteins(APP), acetylcholinesterase(AChE), and glial fibrillary acidic protein(GFAP) mRNA of PC-12 cells treated with CT-105; (3) the AChE activity and the APP production of PC-12 cell treated with CT-105; (4) the behavior of AD mice with ${\beta}A$; (5) expression of $IL-1{\beta}$, $TNF-{\alpha}$, MDA, $IL-1{\beta}$ mRNA, $TNF-{\alpha}$ mRNA, and ROS; (6) the infarction area of the hippocampus, and brain tissue injury in Alzheimer's diseased mice induced with ${\beta}A$ were investigated. The results were summarized as follows; 1. The CMF extract suppressed the expression of $IL-1{\beta}$, IL-6 and $TNF-{\alpha}$ mRNA in THP-1 cells treated with LPS. 2. The CMF extract suppressed the expression of APP, AChE, and GFAP mRNA in PC-12 cells treated with CT-105. 3. The CMF extract suppressed the AChE activity, and the production of APP significantly in PC-12 cells treated with CT-105. 4. A significant inhibitory effect on the memory deficit was shown on the CMF extract group of the mice with Alzheimer's disease induced by ${\beta}A$ in the Morris water maze experiment, which measured stop-through latency, and distance movement-through latency. 5. The CMF extract suppressed the over-expression of $IL-1{\beta}$ protein, $TNF-{\alpha}$ protein, MDA, $IL-1{\beta}$ protein, mRNA, $TNF-{\alpha}$ mRNA, CD68/GFAP, and ROS in the mice with Alzheimer's disease induced by ${\beta}A$. 6. The CMF extract reduced the infarction area of hippocampus, and controlled the injury of brain tissue in the mice with Alzheimer’s disease induced by ${\beta}A$. These results suggest that the CMF extract may be effective for the treatment of Alzheimer’s disease. Investigation into the clinical use of the CMF extract for Alzheimer's disease is suggested for future research.