当鱼类处于高盐碱环境下时，体内HCO3-浓度升高会导致代谢性碱中毒，同时渗透压升高而引发渗透失衡。传统鱼类生理学认为HCO3-的分泌和排泄是由鳃上皮细胞的离子交换和肾脏的重吸收而完成的。前期研究发现，高盐碱环境下鱼体内HCO3-并不通过鳃和肾排出，而肠道可能起着关键作用。本研究以盐碱水域土著鱼类青海湖裸鲤为研究对象，观测饥饿状态下青海湖裸鲤由淡水过渡到高盐碱水后，饮水率变化规律；研究活体条件下青海湖裸鲤肠道HCO3-的分泌和排泄规律；研究高盐碱环境下青海湖裸鲤调控肠道HCO3-分泌的关键基因表达规律，以此探索鱼类在盐碱环境下肠道HCO3-分泌和排泄规律。本研究结果将解释青海湖裸鲤应对盐碱升高的适应策略，为研究鱼类对高盐碱环境的适应机制提供新证据，同时为青海湖裸鲤保护预警机制及保护措施的制定提供依据。

When fish are transferred to saline-alkaline water, there is a transitory metabolic alkalosis and osmotic imbalance with increased HCO3- and Na+,Cl- concentrations in the plasma. In the view of traditional fish physiology, the excretion of HCO3- is controlled by ion exchange at the gill epithelia and reabsorption in tubular cells. Gymnocypris przewalskii is endemic to the austere environment of Lake Qinghai. This lake waters are saline-alkaline with pH value as 9.4, carbonate alkalinity as 29 mmol/L and salinity as 15.3. Living in lake water poses a significant challenge for water and salt balance in G. przewalskii. This fish may use special pathways to secret and excrete HCO3- when adapting to saline-alkaline environmental conditions. Previous studies had shown that, when G.przewalskii acclimated to saline-alkaline water, internal excess HCO3- cannot completely be excreted by the gills or the kidney. In contrast it appears that intestinal HCO3- secretion and excretion may play an important role in the acid-base balance and osmoregulation. This proposal will examine acid-base and osmolality homeostasis behavior through testing the variation of drinking rate of naked carp exposed to saline-alkaline lake water; Investigating the HCO3- secretion and excretion characteristic with vitro methods; Find the key genes participated in the intestinal HCO3- secretion .The results of this study will reveal the intestinal HCO3- secretion and excretion mechanism of the fish species lived in saline-alkaline water environments,and provide evidence to the study of saline-alkaline tolerance mechanisms of fish and protection of the G. przewalskii.

本课题选择盐碱水域土著鱼类青海湖裸鲤为研究对象，对盐碱环境下鱼类饮水率特征、肠道HCO3-排泄规律以及SLC家族基因表达定量进行了探索研究。结果发现：青海湖裸鲤在盐碱水中的饮水率显著高于对照组，其中湖水组约为对照组的2倍，饮水率最高的1.1倍湖水组饮水率为对照组的10倍；盐度组和湖水组裸鲤在胁迫24 h后肠道排泄物中HCO3-浓度显著性升高，其中湖水组上升最为明显，约为对照组的1.7倍；分析肠液中离子组成，发现盐碱水中青海湖裸鲤肠道对K+、Ca2+、Mg2+以及HCO3-发挥分泌功能；对Na+、Cl-发挥吸收功能，肠道沉淀以MgCO3为主。碱度组、盐度组和湖水组青海湖裸鲤中肠slc4a1、slc4a2、slc4a4和slc26a6基因的表达量在胁迫4天过程中均呈现出先升高后回落的现象，其中湖水组裸鲤中肠SLC4、SLC26家族基因表达量上调最为明显，尤其是slc26a6基因表达量最高上升为对照组的4.9倍，推测盐碱环境下青海湖裸鲤通过肠道Cl–/ HCO3-交换子 (slc4a1、slc4a2、slc26a6)、Na+- HCO3-联合转运子分泌和排泄机体内积累的碱，这一调节途径有助于青海湖裸鲤补偿因水环境中盐碱度升高而造成的渗透及酸碱失衡；在注射SLC蛋白抑制剂DIDS后，裸鲤血液pH继续上升，血液HCO3-浓度也显著升高，同时肠道排泄HCO3-浓度明显下降，说明注射DIDS后，由SLC基因表达合成的Cl–/ HCO3-交换子、Na+- HCO3-联合转运子蛋白活性受到抑制，导致HCO3-、CO32–不能及时排出而在体内累积，使得机体碱中毒症状加剧。胞内碳酸酐酶基因及活性实验结果表明青海湖裸鲤鳃肾通过降低碳酸酐酶基因表达水平来补偿盐碱胁迫造成的呼吸性碱中毒并参与渗透调节过程。本项目研究结果为盐碱环境下鱼类肠道酸碱调节机制的揭示奠定了基础。