한국발생생물학회지:발생과생식 (Development and Reproduction)

  • 제13권4호
  • /
  • Pages.257-264
  • /
  • 2009
  • /
  • 2465-9525(pISSN)
  • /
  • 2465-9541(eISSN)
한국발생생물학회 (The Korean Society of Developmental Biology)
권호 표지

수컷 흰쥐의 시상하부-뇌하수체 축 호르몬 유전자 발현에 미치는 6-Hydroxydopamine(6-OHDA)의 영향

Effect of 6-Hydroxydopamine (6-OHDA) on the Expression of Hypothalamus-Pituitary Axis Hormone Genes in Male Rats

  • 허현진 (상명대학교 생명과학) ;
  • 안련섭 (차의과대학 대체의학대학원) ;
  • 이성호 (상명대학교 생명과학)
  • Heo, Hyun-Jin (Dept. of Life Science, Sangmyung University) ;
  • Ahn, Ryun-Sup (Graduate School of Complementary and Alternative Medicine, CHA Medical University) ;
  • Lee, Sung-Ho (Dept. of Life Science, Sangmyung University)
  • 발행 : 2009.12.31
PDF 다운로드
⟨ 이전 논문 다음 논문 ⟩

초록

6-hydroxydopamine(6-OHDA)는 파킨슨 질환 동물 모델의 제조에 널리 사용되는 신경독소로 도파민성 뉴런에 대한 특이적인 독성을 나타낸다. 도파민 신호는 중추신경계의 광범위한 영역에서 생리 기능을 조절하는데, 이에 따라 파킨슨병 환자와 6-OHDA를 처리한 동물들의 신경내분비 활성에 극심한 변화가 있을 것으로 예상할 수 있다. 하지만 6-OHDA 주사 모델에서 시상하부-뇌하수체 신경내분비 회로에 관한 연구들은 전무한 실정이다. 본 연구는 6-OHDA에 의한 뇌 카테콜아민 합성의 차단이 성체 수컷 흰쥐의 시상하부-뇌하수체 호르몬 유전자들의 전사 활성에 일으키는 변화를 조사한 것이다. 생후 3개월의 수컷 흰쥐(SD strain)에 개체 당 $200{\mu}g$의 6-OHDA를 $10{\mu}\ell$의 생리식염수에 녹여 뇌실 내 주사(icv)하였고, 2주 후에 모든 실험동물들을 희생시켰다. 시상하부-뇌하수체 호르몬 유전자들의 mRNA 수준을 조사하기 위해 total RNA를 추출하여 반-정량적 RT-PCR을 시행하였다. 카테콜아민 생합성에서 속도조절효소로 작용하는 tyrosine hydroxylase(TH)의 경우 6-OHDA군에서 대조군에 비해 유의한 발현 감소가 나타났고(대조군:6-OHDA군=1:0.72${\pm}$0.02AU, p<0.001), 이를 통해 6-OHDA 주사의 효력을 확인 하였다. 시상하부에서 gonadotropin-releasing hormone(GnRH)과 corticotropin releasing hormone(CRH)의 mRNA 수준은 6-OHDA군이 대조군에 비해 유의하게 낮았다(GnRH, 대조군:6-OHDA군=1:0.39${\pm}$0.03AU, p<0.001; CRH, 대조군:6-OHDA군=1:0.76${\pm}$0.07AU, p<0.01). 뇌하수체에서 glycoprotein hormone들의 공통적인 alpha subunit(Cg$\alpha$)과 LH beta subunit(LH-$\beta$) 그리고 FSH beta subunit(FSH-$\beta$)의 mRNA 수준의 경우 모두 6-OHDA군에서 대조군에 비해 유의한 감소를 나타냈다(Cg$\alpha$, 대조군:6-OHDA군=1:0.81${\pm}$0.02AU, p<0.001; LH-$\beta$, 대조군:6-OHDA군=1:0.68${\pm}$0.04AU, p<0.001; FSH-$\beta$, 대조군:6-OHDA군=1:0.84${\pm}$0.05AU, p<0.01). 이와 유사하게, 6-OHDA군에서의 뇌하수체 adrenocorticotrophic hormone(ACTH) 전사 수준 역시 대조군에 비해 유의하게 낮았다(대조군:6-OHDA군=1:0.86${\pm}$0.04AU, p<0.01). 본 연구는 중추신경계로의 도파민 신경독소 주입에 의해 두 가지의 시상하부-뇌하수체 신경내분비 회로인 GnRH-성선자극호르몬 회로와 CRH-ACTH 회로의 전사 활성이 하향 조정됨을 증명하였다. 이러한 결과는 시상하부로의 CA 입력은 시상하부-뇌하수체 기능 조절을 통해 생식소와 부신의 활성에 영향을 미침을 시사하는 것으로, 파킨슨병 환자들에게서 빈번하게 발생하는 성 기능 장애와 열악한 스트레스 반응을 설명할 단서를 제공한다.

A neurotoxin, 6-hydroxydopamine (6-OHDA) has been widely used to create animal model for Parkinson's disease (PD) due to its specific toxicity against dopaminergic (DA) neurons. Since DA signals modulate a broad spectrum of CNS physiology, one can expect profound alterations in neuroendocrine activities of both PD patients and 6-OHDA treated animals. Limited applications of 6-OHDA injection model, however, have been made on the studies of hypothalamuspituitary neuroendocrine circuits. The present study was performed to examine whether blockade of brain catecholamine (CA) biosynthesis with 6-OHDA can make any alteration in the transcriptional activities of hypothalamus-pituitary hormone genes in adult male rats. Three-month-old male rats (SD strain) were received 6-OHDA ($200{\mu}g$ in $10{\mu}\ell$ of saline/animal) by intracerebroventricular (icv) injection, and sacrificed after two weeks. To determine the mRNA levels of hypothalamuspituitary hormone genes, total RNAs were extracted and applied to the semi-quantitative RT-PCRs. The mRNA levels of tyrosine hydroxylase (TH), the rate-limiting enzyme for the catecholamine biosynthesis, were significantly lower than those from the control group (control:6-OHDA=1:0.72${\pm}$0.02AU, p<0.001), confirming the efficacy of 6-OHDA injection. The mRNA levels of gonadotropin-releasing hormone (GnRH) and corticotropin releasing hormone (CRH) in the hypothalami from 6-OHDA group were significantly lower than those from the control group (GnRH, control:6-OHDA=1:0.39${\pm}$0.03AU, p<0.001; CRH, control:6-OHDA=1:0.76${\pm}$0.07AU, p<0.01). There were significant decreases in the mRNA levels of common alpha subunit of glycoprotein homones (Cg$\alpha$), LH beta subunit (LH-$\beta$), and FSH beta subunit (FSH-$\beta$) in pituitaries from 6-OHDA group compared to control values (Cg$\alpha$, control:6-OHDA=1:0.81${\pm}$0.02AU, p<0.001; LH-$\beta$, control:6-OHDA=1:0.68${\pm}$0.04AU, p<0.001; FSH-$\beta$, control:6-OHDA=1:0.84${\pm}$0.05AU, p<0.001). Similarly, the level of adrenocorticotrophic hormone (ACTH) transcripts from 6-OHDA group was significantly lower than that from the control group (control: 6-OHDA=1:0.86${\pm}$0.04AU, p<0.01). The present study demonstrated that centrally injected DA neurotoxin could downregulate the transcriptional activities of the two hypothalamus-pituitary neuroendocrine circuits, i.e., GnRH-gonadotropins and CRH-ACTH systems. These results suggested that hypothalamic CA input might affect on the activities of gonad and adrenal through modulation of hypothalamus-pituitary function, providing plausible explanation for frequent occurrence of sexual dysfunction and poor stress-response in PD patients.

키워드

참고문헌

  1. Advis JP, McCann SM, Negro-Vilar A (1980) Evidence that catecholaminergic and peptidergic (luteinizing hormonereleasing hormone) neurons in suprachiasmatic-medial preoptic, medial basal hypothalamus and median eminence are involved in estrogen-negative feedback. Endocrinology 107:892-901. https://doi.org/10.1210/endo-107-4-892
  2. Bellomo G, Santambrogio L, Fiacconi M, Scarponi AM, Ciuffetti G (1991) Plasma profiles of adrenocorticotropic hormone, cortisol, growth hormone and prolactin in patients with untreated Parkinson's disease. J Neurol 238:19-22. https://doi.org/10.1007/BF00319704
  3. Ben-Jonathan N, Hnasko R (2001) Dopamine as a prolactin (PRL) inhibitor. Endocr Rev 22:724-763. https://doi.org/10.1210/er.22.6.724
  4. Calne SM, Kumar A (2008) Young onset Parkinson's disease. Practical management of medical issues. Parkinsonism Relat Disord 14:133-142. https://doi.org/10.1016/j.parkreldis.2007.07.007
  5. Carrasco GA, Van de Kar LD (2003) Neuroendocrine pharmacology of stress. Eur J Pharmacol 463:235-272. https://doi.org/10.1016/S0014-2999(03)01285-8
  6. Chomzynski P, Sacchi N (1987) Single-step method of RNA isolation by RNA guanidium thiocyanate phenolchloroform extraction. Anal Biochem 162:156-159.
  7. Day HE, Vittoz NM, Oates MM, Badiani A, Watson SJ Jr, Robinson TE, Akil H (2002) A 6-hydroxydopamine lesion of the mesostriatal dopamine system decreases the expression of corticotropin releasing hormone and neurotensin mRNAs in the amygdala and bed nucleus of the stria terminalis. Brain Res 945:151-159. https://doi.org/10.1016/S0006-8993(02)02747-6
  8. Deumens R, Blokland A, Prickaerts J (2002) Modeling Parkinson's disease in rats: an evaluation of 6-OHDA lesions of the nigrostriatal pathway. Exp Neurol 175: 303-317. https://doi.org/10.1006/exnr.2002.7891
  9. Florio P, Zatelli MC, Reis FM, degli Uberti EC, Petraglia F (2007) Corticotropin releasing hormone: a diagnostic marker for behavioral and reproductive disorders- Front Biosci 12:551-560. https://doi.org/10.2741/2081
  10. Grunblatt E, Mandel S, Youdim MB (2000) Neuroprotective strategies in Parkinson's disease using the models of 6-hydroxydopamine and MPTP. Ann N Y Acad Sci 899:262-273.
  11. Kalra SP, Kalra PS (1985) Neural regulation of luteinizing hormone secretion in the rat. Endocr Rev 4:311-351. https://doi.org/10.1210/edrv-4-4-311
  12. Kan SF, Kau MM, Low-Tone Ho L, Wang PS (2003) Inhibitory effects of bromocriptine on corticosterone secretion in male rats. Eur J Pharmacol 468:141-149. https://doi.org/10.1016/S0014-2999(03)01663-7
  13. Kim K, Lim IS, Cho BN, Kang SS, Lee BJ, Choi KH, Chung CH, Lee CC, Cho WK, Wuttke W (1993) A partial blockade of catecholaminergic neurotransmission with 6-hydroxydopamine decreases mRNA level of gonadotropin releasing hormone in the male rat hypothalamus. Neuroendocrinology 58:146-152. https://doi.org/10.1159/000126524
  14. Kim K, Lee BJ, Cho BN, Kang SS, Choi WS, Park SD, Lee CC, Cho WK, Wuttke W (1994) Blockade of noradrenergic neurotransmission with diethyldithiocarbamic acid decreases the mRNA level of gonadotropinreleasing hormone in the hypothalamus of ovariectomized, steroid-treated prepubertal rats. Neuroendocrinology 59: 539-544. https://doi.org/10.1159/000126703
  15. Kostrzewa RM, Jacobowitz DM (1974) Pharmacological actions of 6-hydroxydopamine. Pharmacol Rev 26:199- 288.
  16. Mahesh VB, Brann DW (1992) Interaction between ovarian and adrenal steroids in the regulation of gonadotropin secretion. J Steroid Biochem Mol Biol 41:495-513. https://doi.org/10.1016/0960-0760(92)90375-S
  17. Martinovic JV, McCann SM (1977) Effect of lesions in the ventral noradrenergic tract produced by microinjection of 6-hydroxydopamine on gonadotropin release in the rat. Endocrinology 100:1206-1213. https://doi.org/10.1210/endo-100-4-1206
  18. Metz GA (2007) Stress as a modulator of motor system function and pathology. Rev Neurosci 18:209-222.
  19. Monda M, Viggiano A, Viggiano A, Viggiano E, Messina G, Tafuri D, De Luca V (2007) Sympathetic and hyperthermic reactions by orexin A: role of cerebral catecholaminergic neurons. Regul Pept 139:39-44. https://doi.org/10.1016/j.regpep.2006.10.002
  20. Negro-Vilar A, Advis JP, Ojeda SR, McCann SM (1982) Pulsatile luteinizing hormone (LH) patterns in ovariectomized rats: involvement of norepinephrine and dopamine in the release of LH-releasing hormone and LH. Endocrinology 111:932-938. https://doi.org/10.1210/endo-111-3-932
  21. Otake K, Oiso Y, Mitsuma T, Hirooka Y, Adachi K (1994) Hypothalamic dysfunction in Parkinson's disease patients. Acta Med Hung 50:3-513.
  22. Parmer RJ, Zinder O (2002) Catecholaminergic pathways, chromaffin cells, and human disease. Ann N Y Acad Sci 971:497-505. https://doi.org/10.1111/j.1749-6632.2002.tb04514.x
  23. Paxinos G, Watson C (1998) The Rat Brain in Stereotaxic Coordinates. Fourth edition. Academic Press Inc. San Diego.
  24. Ramaker C, van Hilten JJ (2000) Bromocriptine versus levodopa in early Parkinson's disease. Cochrane Database Syst Rev 2000(3):CD002258.
  25. Salgado-Pineda P, Delaveau P, Blin O, Nieoullon A (2005) Dopaminergic contribution to the regulation of emotional perception. Clin Neuropharmacol 28:228-237. https://doi.org/10.1097/01.wnf.0000185824.57690.f0
  26. Selvage DJ, Lee SY, Parsons LH, Seo DO, Rivier CL (2004) A hypothalamic-testicular neural pathway is influenced by brain catecholamines, but not testicular blood flow. Endocrinology 145:1750-1759. https://doi.org/10.1210/en.2003-1441
  27. Simpkins JW, Kalra SP (1979) Blockade of progesteroneinduced increase in hypothalamic luteinizing hormonereleasing hormone levels and serum gonadotropins by intrahypothalamic implantation of 6-hydroxydopamine. Brain Res 170:475-484. https://doi.org/10.1016/0006-8993(79)90965-X
  28. Stacy M, Galbreath A (2008) Optimizing long-term therapy for Parkinson disease: levodopa, dopamine agonists, and treatment-associated dyskinesia. Clin Neuropharmacol 31:51-56. https://doi.org/10.1097/WNF.0b013e318065b088
  29. Weidenfeld J, Feldman S, Itzik A, Van de Kar LD, Newman ME (2002) Evidence for a mutual interaction between noradrenergic and serotonergic agonists in stimulation of ACTH and corticosterone secretion in the rat. Brain Res 941:113-117. https://doi.org/10.1016/S0006-8993(02)02641-0