Browsing by Author "Tseng, Yung-Che"

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Brain functioning under acute hypothermic stress supported by dynamic monocarboxylate utilization and transport in ectothermic fish
(2014-08-08) Tseng, Yung-Che; Liu, Sian-Tai; Hu, Marian Y; Chen, Ruo-Dong; Lee, Jay-Ron; Hwang, Pung-Pung
Abstract Background The vertebrate brain is a highly energy consuming organ that requires continuous energy provision. Energy metabolism of ectothermic organisms is directly affected by environmental temperature changes and has been demonstrated to affect brain energy balance in fish. Fish were hypothesized to metabolize lactate as an additional energy substrate during acute exposure to energy demanding environmental abiotic fluctuations to support brain functionality. However, to date the pathways of lactate mobilization and transport in the fish brain are not well understood, and may represent a critical physiological feature in ectotherms during acclimation to low temperature. Results We found depressed routine metabolic rates in zebrafish during acute exposure to hypothermic (18°C) conditions accompanied by decreased lactate concentrations in brain tissues. No changes in brain glucose content were observed. Acute cold stress increased protein concentrations of lactate dehydrogenase 1 (LDH1) and citrate synthase (CS) in brain by 1.8- and- 2.5-fold, paralleled by an increased pyruvate to acetyl-CoA transformation. To test the involvement of monocarboxylate transporters (MCTs) under acute cold stress in zebrafish, we cloned and sequenced seven MCT1-4 homologues in zebrafish. All drMCT1-4 are expressed in brain tissues and in response to cold stress drmct2a and drmct4a transcripts were up-regulated 5- and 3-fold, respectively. On the contrary, mRNA levels of drmct1a, -1b and -4b in zebrafish brain responded with a down regulation in response to cold stress. By expressing drMCTs in Xenopus oocytes we could provide functional evidence that hypothermic stress leads to a 2-fold increase in lactate transport in drMCT4b expressing oocytes. Lactate transport of other paralogues expressed in oocytes was unaffected, or even decreased during cold stress. Conclusion The present work provides evidence that lactate utilization and transport pathways represent an important energy homeostatic feature to maintain vital functions of brain cells during acute cold stress in ectotherms.
Branchial NH4+-dependent acid–base transport mechanisms and energy metabolism of squid (Sepioteuthis lessoniana) affected by seawater acidification
(2014-08-06) Hu, Marian Y; Guh, Ying-Jey; Stumpp, Meike; Lee, Jay-Ron; Chen, Ruo-Dong; Sung, Po-Hsuan; Chen, Yu-Chi; Hwang, Pung-Pung; Tseng, Yung-Che
Abstract Background Cephalopods have evolved strong acid–base regulatory abilities to cope with CO2 induced pH fluctuations in their extracellular compartments to protect gas transport via highly pH sensitive hemocyanins. To date, the mechanistic basis of branchial acid–base regulation in cephalopods is still poorly understood, and associated energetic limitations may represent a critical factor in high power squids during prolonged exposure to seawater acidification. Results The present work used adult squid Sepioteuthis lessoniana to investigate the effects of short-term (few hours) to medium-term (up to 168 h) seawater acidification on pelagic squids. Routine metabolic rates, NH4+ excretion, extracellular acid–base balance were monitored during exposure to control (pH 8.1) and acidified conditions of pH 7.7 and 7.3 along a period of 168 h. Metabolic rates were significantly depressed by 40% after exposure to pH 7.3 conditions for 168 h. Animals fully restored extracellular pH accompanied by an increase in blood HCO3− levels within 20 hours. This compensation reaction was accompanied by increased transcript abundance of branchial acid–base transporters including V-type H+-ATPase (VHA), Rhesus protein (RhP), Na+/HCO3− cotransporter (NBC) and cytosolic carbonic anhydrase (CAc). Immunocytochemistry demonstrated the sub-cellular localization of Na+/K+-ATPase (NKA), VHA in basolateral and Na+/H+-exchanger 3 (NHE3) and RhP in apical membranes of the ion-transporting branchial epithelium. Branchial VHA and RhP responded with increased mRNA and protein levels in response to acidified conditions indicating the importance of active NH4+ transport to mediate acid–base balance in cephalopods. Conclusion The present work demonstrated that cephalopods have a well developed branchial acid–base regulatory machinery. However, pelagic squids that evolved a lifestyle at the edge of energetic limits are probably more sensitive to prolonged exposure to acidified conditions compared to their more sluggish relatives including cuttlefish and octopods.
Development in a naturally acidified environment: Na+/H+-exchanger 3-based proton secretion leads to CO2 tolerance in cephalopod embryos
(2013) Hu, Marian; Lee, Jay-Ron; Lin, Li-Yih; Shih, Tin-Han; Stumpp, Meike; Lee, Mong-Fong; Hwang, Pung-Pung; Tseng, Yung-Che
吳郭魚在鹽度轉移過程中肌肉內麩胺酸/麩醯胺酸與糖解作用相關代謝之研究
(2016) 王宏原; Wang, Hong-Yuan
臺灣養殖吳郭魚在出貨前多會進行海水轉移步驟以清除肉質的不良風味。本研究運用廣鹽性莫三比克吳郭魚(Oreochromis mossambicus)探討在不同鹽度處理下，肌肉內麩胺酸(glutamate)/麩醯胺酸(glutamine) 以及糖解反應相關代謝機制之作用。本論文研究結果顯示在轉移至10 ‰海水處理後，肌肉中麩醯胺酸合成酶 (GLUL)與麩胺酸脫氫酶 (GLUD)的基因表現量有顯著提升，同時麩醯胺酸與其他胺基酸含量亦有上升；而轉移至20 ‰海水處理的結果顯示肝醣磷酸化酶 (PYG)基因表現量有顯著上升，六碳糖激酶 (HK)與甘油醛3-磷酸去氫酶 (GPD)基因表現量則有明顯下降的趨勢，此外葡萄糖亦有累積的結果。而在各鹽度處理下，肌肉中氨含量並無顯著變化。因此我們推測：在10 ‰海水處理後會使吳郭魚肌肉傾向將麩胺酸與銨離子結合生成麩醯胺酸，並累積於肌肉中；而轉移至20 ‰海水處理後由於糖解作用受阻致使葡萄糖逐漸累積於肌肉中。由以上莫三比克吳郭魚適應高滲透度緊迫環境所採用的生理策略，導致肌肉組織中胺基酸與葡萄糖的累積，可能是造成人類在食用味覺上產生「鮮味」的原因。
廣鹽性魚類適應鹽度變異中腸道與肝臟麩胺酸的運輸和代謝機轉
(2016) 彭慧文; Peng, Hui-Wen
環境鹽度的變異已知會刺激廣鹽性魚類腸道中鈉鉀幫浦進行主動運輸以調節離子及水分的恆定；而肝臟則會進行代謝作用，以提供魚體各組織充足的能量需求。非必需胺基酸中的麩胺酸(glutamate)是生物體中含量最多的胺基酸，過去在魚類代謝機制相關的研究中，麩胺酸的運用已知與適應環境鹽度變異有關，然而麩胺酸在廣鹽性魚類腸道及肝臟的代謝調節機轉仍不明確。　　本研究運用日本種青鱂魚(Oryzias latipes)作為廣鹽性魚類實驗物種。在鹽度轉移過程中，腸道和肝臟中麩醯胺酸運輸蛋白(SAT1)及麩胺酸和麩醯胺酸相關的合成酵素(GLS, GLUL)基因表現亦隨著環境鹽度增加而有顯著變化，且上述隨鹽度變化波動的運輸蛋白和酵素皆被發現表現在腸道和肝臟細胞中。此外，鹽度刺激會使麩胺酸代謝產生毒性氨，因此本研究發現與尿素合成相關的基因在腸道(CPS, OTC)和肝臟(ASL, ARG, AGMAT)中會被誘發以進行尿素生合成反應。綜合以上的結果推測：鹽度刺激會促進廣鹽性魚類腸道和肝臟中麩胺酸與麩醯胺酸的代謝和運輸，因代謝產出的含氮廢物會藉由尿素循環機制合成尿素。此適應性代謝轉換機制不僅有利於個體能量的適時供給，並能有助於維持組織細胞內的滲透壓恆定。
海洋酸化對黑點青鱂魚生理恆定及表觀遺傳修飾之影響評估
(2017) 劉姿延; Liu, Tzu-Yen
工業革命後人類的現代化活動造成大氣中二氧化碳濃度逐漸上升，當二氧化碳溶入海水會解離出碳酸根離子與氫離子，造成海水的 pH 值下降，這個現象稱為「海洋酸化」。許多研究指出：海洋酸化會影響海洋生物的存活率、多樣性及生理恆定，因此被認為是個全球性的環境議題。本實驗選用海水青鱂魚(Oryzias melastigma)作為長期高碳酸馴養試驗的模式魚種，實驗結果有別於過去在廣鹽性日本青鱂魚的發現，其胚胎並無發育遲緩的現象，而酸化處理後的胚胎個體及成魚的鰓與腸中之酸鹼離子調控蛋白群(AE1a、 NBCa 及 NHE2)基因表現量均會顯著上升。此外，在酸化海水發育成熟的個體卵巢中， AE1a 的啟動子甲基化程度會顯著提升。而酸化個體產下之 F1 子代酸鹼離子調控蛋白群的啟動子並未有顯著甲基化的趨勢，其基因表現量與排氫離子能力仍保持顯著提升。綜合以上結果我們推論：硬骨魚母體可能透過表觀遺傳的機制將生理適應訊息傳遞給子代，子代為了適應酸化環境會進而修飾啟動子的甲基化程度，並且持續增強酸鹼調節的能力；此外， AE1a、 NBCa 和 NHE2 為硬骨魚類調控酸鹼平衡之主要蛋白，並且能夠作為觀察表觀遺傳現象之有效生物標記。
神經內分泌胜肽趨同調控虎斑烏賊體液酸鹼恆定之探討
(2016) 陳玉奇; Chen, Yu-Chi
頭足類動物與硬骨魚類的胚胎對於調控體液酸鹼平衡的模式具有相似性，然而在頭足類動物上，這種生理現象於演化發育上的分子調控機制所知仍有限。本研究運用虎斑烏賊(Sepia pharaonis)的胚胎進行高碳酸的環境刺激，探測催產素同源基因sepiatocin、pro-sepiatocin及其受器sepiatocin-related receptor (spr)的表現，並檢測虎斑烏賊的耐酸能力。研究結果發現：虎斑烏賊的胚胎於不同發育時期能忍受不同程度的碳酸壓力，最低可至pH7.0。而藉由原位雜合分析亦發現spr會表現在虎斑烏賊胚胎的表皮及鰓上，sepiatocin及pro-sepiatocin和zn12標記的神經細胞有共定位的表現，顯示頭足類由神經細胞分泌sepiatocin及pro-sepiatocin。此外，高碳酸的壓力一方面會於不同的時間點顯著刺激pro-sepiatocin、sepiatocin與spr的表現，同時也影響虎斑烏賊胚胎表皮的離子調節蛋白(vha、nbc、nhe3、rhp、nhe3)及p63的基因表現，顯示與sepiatocin相關的激素可能參與體液酸鹼恆定，而頭足類胚胎與硬骨魚胚胎的酸鹼調控機制與表皮調控路徑的相似性應為在生存競爭下，兩者生理機制趨向趨同演化的證據之一。
系統性研究魚類面臨溫度波動的能量調適策略
(2021) 王敏真; Min-Chen Wang
none
缺氧誘導因子與廣鹽性硬骨魚類滲透壓生理調節之關聯
(2017) 趙仕堯; Chao, Shih-Yao
穩定體液滲透度是廣鹽性魚類在遭遇環境鹽度變異時重要的生理適應策略。而眾多非生物性環境擾動均會影響生物缺氧誘導因子(HIFs)蛋白功能的穩定性，進而影響細胞運作。然而目前對於魚類面臨滲透壓波動挑戰時，細胞內HIFs相關訊息網絡與滲透壓恆定的研究仍非常缺乏。本研究選用India medaka (Oryzias melastigma)作為暴露高鹽環境的廣鹽性硬骨魚實驗物種，實驗結果發現其仔魚在面臨鹽度上升刺激時，HIF-1α的基因和蛋白質表現量皆會顯著上升，而部分HIF-1α的核酸訊號亦會表現在仔魚表皮的離子細胞。此外，HIF-1α蛋白的弱化處理，除了會顯著地影響仔魚鈉鉀幫浦的蛋白表現量，亦會降低其在高鹽處理時表皮對於鈉與氯離子的排放；然而，仔魚表皮上離子細胞的表現密度，並未受到HIF-1α蛋白的弱化影響。我們據此推論：HIF-1α於表皮細胞的表現，有助於廣鹽性魚類在面臨環境鹽度挑戰時進行體液調節機制。
魚類表皮細胞麩胺酸-麩醯胺酸循環調控滲透恆定之研究
(2015) 黃珮甄; Huang, Pei-Chen
對廣鹽性硬骨魚類而言，維持魚體內滲透壓平衡以適應環境鹽度變化是相當重要的生理恆定機制，而最主要負責調節魚體滲透壓及離子恆定為魚鰓表皮的離子細胞，其可經由活化不同離子通道(例如：鈉鉀幫浦，Na+-K+-ATPase)或相關酵素，進行主動運輸模式進行生理調整，也因此需要消耗大量的能量。故適時的能量代謝啟動與即時的滲透度調節對魚類適應鹽度過程是極為必要的。從先前研究已得知，當環境鹽度變動時，廣鹽性硬骨魚類體內有麩胺酸累積和相關的代謝反應，然而其基本的細胞分子機制仍不明確。本研究運用廣鹽性青鱂魚作為實驗物種，證實了魚類表皮細胞中麩胺酸代謝及其運輸機制對於鹽度適應過程中之重要性。青鱂魚鰓中「麩胺酸家族」成員胺基酸含量會因環境鹽度波動有明顯上升；除此之外，魚鰓中麩胺酸及麩醯胺酸相關的運輸通道蛋白(EAATs, SAT1)及相對應合成酵素(GLS, GLUL)的基因表現相亦隨著環境鹽度增加而有顯著提升，且上述通道蛋白與酵素皆會表現在胚胎的上皮膜細胞。而鹽度刺激促使麩胺酸代謝產生的毒性氨(NH3)，將進而促使尿素循環的速率決定酵素(CPS, OTC)的誘發反應，以合成尿素參與滲透調節。綜觀上述，環境鹽度改變可促進廣鹽性魚類鰓表皮中麩胺酸及麩醯胺酸進行代謝及運輸，代謝過程中產出的含氮廢物會藉由尿素循環啟動轉合成尿素。上述代謝途徑的誘發不僅有利於提供足夠的能量供給，且助於維持表皮細胞的滲透恆定。
龜山島淺海熱泉怪方蟹的硫化氫轉換與後續運輸機制研究
(2018) 周佩璇; Chou, Pei-Hsuan
海底熱泉旁通常具有高濃度硫化氫釋放，而硫化氫會抑制生物的呼吸代謝，進而限制生物的生存。在此環境生存的生物，諸如深海熱泉中的螃蟹、貽貝、鬚腕類生物已知具備特殊的細胞解硫機制。然而，龜山島周遭的淺海熱泉組成，有別於典型的海底熱泉系統，在此區域棲息的硫磺怪方蟹(Xenograpsus testudinatus)對硫化物的耐受與適應機制，亦多屬推測並缺乏系統性的探討。本論文研究目的旨在瞭解硫磺怪方蟹在其原生高硫環境是如何有效地進行硫化氫相關的解毒機制。實驗結果發現：怪方蟹血液具有亞磺酸(hypotaurine)與硫代牛磺酸(thiotaurine)的存在；在高硫環境中，怪方蟹第三、五對鰓的牛磺酸運輸蛋白(taurine transporter, TAUT，可運輸taurine、hypotaurine、thiotaurine)之表現量明顯高於無硫環境馴養的個體。以上結果皆暗示怪方蟹可能具備將S2-與hypotaurine代謝生合成thiotaurine，以降低硫毒性再進行運輸的適應能力。此外，本研究亦發現在原生高硫環境生存的怪方蟹鰓中，與硫運輸相關的表皮膜蛋白solute carrier family 26 member 11 (SLC26A11)基因與蛋白表現量均顯著高於無硫馴養的實驗個體。此結果暗示硫化氫氧化後形成毒性較低的硫酸根產物，可藉由鰓表皮細胞底膜的SLC26A11將硫酸根離子運輸至血液。根據以上結果我們推論：硫磺怪方蟹具有特殊的硫轉換機制，並在其鰓表皮上具有專一的硫相關運輸蛋白，可有效地降低生存在高硫環境中持續存在的毒性緊迫，以利在熱泉環境中生存。