• 한국한의학연구원논문집
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• 제9권1호
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• pp.157-178
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• 2003

Objective : This study was designed to assess the protective effects of Chengwhabosimtang on the animal model of depression, chronic mild stress(CMS). Method : Male Sprague-Dawley rats Were used for this experiment. The subjects Were divided into 3 groups (1. CMS-drug: Chengwhabosimtang administered during CMS treatment, 2. CMS-vehicle: water administered, 3. normal ). After 4 weeks of CMS treatment they were executed Forced swimming test(FST) and Elevated plus maze. Tyrosine hydroxylase(TH) in ventral tegmental area(VTA) and c-Fos in paraventricular nucleus(PVN) were measured. Result : 1. In FST, CMS-drug group showed significantly decreased immobility behavior. 2. CMS-drug group showed no significantly lower TH level in VTA than CMS-vehicle group. 3. CMS-drug group showed significantly less c-Fos expressed cell bodies in PVN than CMS-vehicle group. 4. In Elevated plus maze, CMS-drug group showed no significantly anxiety. Conclusion : These results suggest that Chengwhabosimtang may have protective antidepressant effects in CMS model rats. And these effects could be explained by the elevated stress-copying behaviors which are related with PVN of hypothalamus and dopaminergic neurons in VTA.

• 동의생리병리학회지
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• 제24권2호
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• pp.328-332
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• 2010

Acupuncture has been used for the treatment and prevention of stress-related disorders. In the present study, the effect of electroacupuncture on the behavioral and biochemical responses to restraint stress was evaluated in rats. Sprague-Dawley male rats underwent to immobilization stress for 21 days (6 hours/day). Electroacupuncture (2 Hz, 2 mA, and 10 minutes) was applied either to the acupuncture point HT7 (Shenmen) or the nonacupuncture point in the tail for the last 7 days. Rats were randomly divided into four groups: the normal group (n=10, without the restraint stress), the stress group (n=10, with restraint stress), the HT7 group (n=10, with restraint stress and electroacupuncture to HT7), and the NA group (n=10, with restraint stress and electroacupuncture to the nonacupuncture point). The anxiety-related behavior was tested using the elevated plus maze and the Vogel test on day 22. The expression of tyrosine hydroxylase in the locus coeruleus was measured by immunohistochemistry. R maze and the Vogecreased the response of the anxiety-related behavior. The number of tyrosine hydroxylase-immunoreactive cells were also increased. The HT7 group showed a significant decrease of anxiety-related behavioral response, compared to the stress group. The tyrosine hydroxylase-immunoreactive expression was also decreased in the HT7 group. These findings suggestthat electroacupuncture to HT7 might play a role in reducing the stress-related responses, which may be helpful for the treatment of stress-related disorders.

• Biomolecules & Therapeutics
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• 제16권2호
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• pp.106-112
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• 2008

The effects of harman and norharman on dopamine content and L-DOPA-induced cytotoxicity were investigated in PC12 cells. Harman and norharman decreased the intracellular dopamine content for 24 h. The $IC_{50}$ values of harman and norharman were 20.4 ${\mu}M$ and 95.8 ${\mu}M$, respectively. Tyrosine hydroxylase (TH) activity and TH mRNA levels were also decreased by 20 ${\mu}M$ harman and 100 ${\mu}M$ norharman. Under the same conditions, the intracellular cyclic AMP levels were decreased by harman and norharman. In addition, harman and norharman at concentrations higher than 80 ${\mu}M$ and 150 ${\mu}M$ caused cytotoxicity at 24 h in PC12 cells. Non-cytotoxic ranges of 10-30 ${\mu}M$ harman and 50-150 ${\mu}M$ norharman inhibited L-DOPA (20-50 ${\mu}M$)-induced increases of dopamine content at 24 h. Harman at 20-150 ${\mu}M$ and norharman at 100-300 ${\mu}M$ also enhanced LDOPA (20-100 ${\mu}M$)-induced cytotoxicity at 24 h. These results suggest that harman and norharman decrease dopamine content by reducing TH activity and aggravate L-DOPA-induced cytotoxicity in PC12 cells.

• 약학회지
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• 제50권2호
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• pp.105-110
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• 2006

The effects of tributyltin acetate (TBTA), one of the endocrine-disrupting organotin compounds, on dopamine biosynthesis in PC12 cells were investigated. Treatment of PC12 cells with TBTA at $0.05\sim0.25{\mu}M$ significantly decreased the intracellular dopamine content in a concentration-dependent manner ($IC_{50}$ value, $0.17{\mu}M$). Under these conditions, tyrosine hydroxylase (TH) activity and TH mRNA level were also decreased by $0.1{\mu}M$ TBTA at 24 h, and recovered there-after. In addition, treatment with L-DOPA at 20 and $50 {\mu}M$ increased the intracellular dopamine content in PC12 cells and the increase in dopamine content by L-DOPA was significantly abolished by TBTA at $0.1\sim0.2{\mu}M$. These results indicate that TBTA at $0.1\sim0.2{\mu}M$ causes the decrease in the basal dopamine content and abolishes the increase in dopamine content in L-DOPA-treated cells in part by the inhibition of TH gene expression and activity.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2003년도 제3회 국제심포지움 및 학술대회
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• pp.104-104
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• 2003

As an effort to direct differentiation of human embryonic stem (hES, MB03) cells to dopamine-producing neuronal cells, Nurr1 was transfected using conventional transfection protocol into MB03 and examined the expression of tyrosine hydroylase (TH) after differentiation induced by retinoic acid (RA) and ascorbic acid (AA). Experimentally, cells were transfected with linearized Nurr1 cDNA in pcDNA3.1 (+)-hygovernight followed by selection in medium containing hygromycin-B (150 $\mu$/ml). Expression of Nurr1 mRNA was confirmed by RT-PCR and protein by immunocytochemistry in the drug resistant clones. In order to study the effect of Nurr1 protein on the differentiation pattern of ES cells, one of the positive clones (MBNr24) was allowed to form embryoid body (EB) for 2 days and were induced to differentiate for another 4 days using RA (1 $\mu M$) and AA (50 mM) (2-/4+ protocol) followed by selection in N2 medium for 10 or 20 days. After 10 days in N2 medium, cells immunoreactive to anti-GFAP, anti-TH, or anti-NF200 antibodies were 38.8%, 11%, and 20.5%, respectively. After 20 days in N2 medium, cells expressing GFAP, TH, or NF200 were 28%, 15% and 44.8%, respectively but approximately 9% of MB03 expressed TH protein when the cells were induced to differentiate using a similar prorocol, These results suggest that ectopic expression of Nurr1 enhances generation of TH+ cells as well as neuronal cells when hES cells were differentiated by 2-/4+ protocol.

• 약학회지
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• 제49권2호
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• pp.156-161
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• 2005

Tetrahydropapaveroline (THP) at 5-15 ${\mu}$M has been found to induce L-DOPA-induced oxidative apoptosis in PC12 cells. In this study, the inhibitory effects of THP on dopamine bios ynthesis in PC12 cells and tyrosine hydroxylase (TH) activity in bovine adrenal were investigated. Treatment of PC12 cells with THP at 2.5-10 ${\mu}$M significantly decreased the intracellular dopamine content in a concentration-dependent manner (18.3% inhibition at 10 ${\mu}$M THP). In these conditions, TH activity was markedly inhibited by the treatment with THP at 2.5-10 ${\mu}$M in PC12 cells (23.4% inhibition at 10 $\mu$ M THP). In addition, THP had an inhibitory effect on bovine adrenal TH activity IC50 value, 153.9${\mu}$M). THP exhibited uncompetitive inhibition on bovine adrenal TH activity with a substrate L-tyrosine with the KI value of 0.30 mM. Treatment with L-DOPA at 20~50 ${\mu}$M increased the intracellular dopamine content in PC12 cells, and the increase in dopamine content by L-DOPA was inhibited in part when THP at non-cytotoxic (5-10 ${\mu}$M) or cytotoxic (15${\mu}$M) concentrations was associated with L-DOPA (20 and 50 ${\mu}$M) for 24 h incubation. These results suggest that THP at 5-10${\mu}$M decreases the basal dopamine content and reduces the increased dopamine content induced by L-DOPA in part by the inhibition of TH activity, and that THP at 15${\mu}$M also decreases dopamine content by oxidative stress in PC12 cells.

• Environmental Analysis Health and Toxicology
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• 제15권4호
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• pp.131-137
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• 2000

최근 독성 유해물질의 환경으로의 방출로 인해 인간 및 생태계에 대한 위해성 문제가 심각하게 제기되고 있다. 독성화학물질을 포함한 여러 환경 오염물질의 위해성평가는 화학물질의 유해성과 노출량 측정을 동시에 측정함으로써 가능한데 이 경우 생물지표(biomarker)가 최근 각광을 받고 있다. 본 연구에서는 동물의 행동에 관련된 신경전달물질의 생성에 결정적 역할을 하는 tyrosine hydroxylase(TH)및 그 유전자가 생물지표로서 이용 가능성이 있는지를 검토하였다. Ovary cDNA library의 PCR 스크리닝을 통한 송사리 TH유전자를 부분적으로 를론하였으며(327 bp), DNA염기서열 분석 결과 쥐 (rat)의 TH유전자와 동일한 염기서열을 보였다. 그리고 다이아지논 처리구 및 무처리구에서 송사리의 머리부분(head)및 몸통 부분(body)에서 추출된 총RNA에 TH mRNA가 존재함을 RT-PCR를 통하여 확인하였다. 그러나 다이아지논의 처리효과가 송사리의 행동에 미치는 영향을 보기 위해서는 TH의 발현을 보다 정량적으로 검토할 필요가 있을 것으로 판단된다. 생물지표로서 TH의 활성 및 mRNA의 기관별 또는 조직별 검출은 독성물질에 영향을 받는 어류 신경행동 변화를 모니터링 할 수 있는 유용한 수단이 될 것이다. 나아가 환경관리에 있어서 신경화학물질과 분자생물학적 상관관계를 통한 이상반응행동의 분석은 환경 위해성평가에 상당히 기여할 것이다.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2003년도 제10차 국제학술회의 및 추계정기 학술발표회
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• pp.17-17
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• 2003

Enzymes involved in catecholamine synthesis are present in the highest concentration in the adrenal medulla, however they were found also in other, mainly nervous tissues. Increased transcription of genes for catecholamine biosynthetic enzymes is an important mechanism to increase the capacity for epineprine/norepinephrine biosynthesis with stress. Gastrodia elata(Chinese name: Tienma), are very important Chinese herbal medicines used for the medical treatment of headaches, migraine, dizziness, epilepsy, rheumatism, neuralgia, paralysis and other neuralgic and nervous disorders. Immobilize stressed rat markedly increased tyrosine hydroxylase (TH) mRNA and dopamine-${\beta}$-hydroxylase (DBH) mRNA transcriptior level more than control group. But treated Gastrodia elata extracts in immobilized stressed rat slightly increased TH mRNA and DBH mRNA transcription level more than normal group. In addition, we are obtained identical results in PC12 cell line. Decrease of transcription level of TH mRNA and DBH mRNA is indicating that Gastrodia elata have a anti-stress effects which decrease the transcription level of TH and DBH mRNA on catecholamine biosynthesis pathway.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2003년도 심포지엄
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• pp.37-37
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• 2003

Enzymes involved in catecholamine synthesis are present in the highest concentration in the adrenal medulla, however they were found also in other, mainly nervous tissues. Increased transcription of genes for catecholamine biosynthetic enzymes is an important mechanism to increase the capacity for epineprine/norepinephrine biosynthesis with stress. Gastrodia elata(Chinese name: Tienma), are very important Chinese herbal medicines used for the medical treatment of headaches, migraine, dizziness, epilepsy, rheumatism, neuralgia, paralysis and other neuralgic and nervous disorders. Immobilize stressed rat markedly increased tyrosine hydroxylase (TH) mRNA and dopamine-$\beta$-hydroxylase (DBH) mRNA transcription level more than control group. But treated Gastrodia elata extracts in immobilized stressed rat slightly increased TH mRNA and DBH mRNA transcription level more than normal group. In addition, we are obtained identical results in PC12 cell line. Decrease of transcription level of TH mRNA and DBH mRNA is indicating that Gastrodia elata have a anti-stress effects which decrease the transcription level of TH and DBH mRNA on catecholamine biosynthesis pathway.

• Archives of Pharmacal Research
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• 제16권2호
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• pp.155-157
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• 1993

DNA transfection conditions were investigated by calcium phosphate-DNA co-precipitation in SK-N-BE(2)C human neuroblastoma cells. The DNA plasmid of TH2400CAT was used in which rat tyrosine hydroxylase gene was inserted into chloramphenicol acetyltransferase reporter gent. The transfection efficiency was 25-30% and the method was simple and reproducible. So, the method will be a good tool for transient transfection analysis.