龙眼体胚发生相关未知蛋白DlUP-3基因的克隆与表达分析

在龙眼体胚发生早期的蛋白质组学研究中,发现1个体胚发生相关未知蛋白DlUP-3,通过简并引物结合RACE技术进行其基因全长序列克隆。结果显示:(1)克隆到的龙眼体胚发生相关未知蛋白基因DlUP-3的全长cDNA序列为1 681bp,开放阅读框由1 017个核苷酸组成,编码338个氨基酸(GenBank登录号为GQ167202)。(2)生物信息学分析发现,该基因推导蛋白分子量为36 854.2Da,pI为9.05;该蛋白为Ras蛋白质家族成员,具有ATP/GTP-binding site motif A(P-loop)结合位点和1个典型的Ras_like_GTPase superfamily组件,无典型信号肽结构,但有跨膜螺旋的亲水性蛋白;不规则卷曲是其最大量的结构元件,散布于整个蛋白质中。(3)实时荧光定量PCR分析显示,该基因在龙眼体胚发生过程中均有表达,其中以胚性愈伤组织阶段表达量最低,而球形胚阶段最高。研究表明,DlUP-3基因在龙眼体胚发生过程尤其是球形胚阶段有重要的作用,为进一步研究该基因在龙眼体胚发生过程中的功能奠定了基础。     

龙眼胚性愈伤组织DRM1基因的克隆及其定位与表达分析

该研究以龙眼胚性愈伤组织的转录组数据为基础,对龙眼胚性愈伤组织DlDRM1基因进行克隆和生物学信息分析,并检测其在体胚发生过程中不同发育阶段、不同浓度的外源激素(2,4-D、IAA、KT)处理下及不同组织部位的表达,以揭示DlDRM1基因在龙眼体胚发生过程中的功能。结果表明:(1)从龙眼转录组unigene序列筛选获得龙眼结构域重排甲基化酶1基因(命名为DlDRM1)全长序列,并利用RT-PCR法从‘红核子’龙眼胚性愈伤组织中克隆获得DlDRM1基因的cDNA全长序列(GenBank登录号为KY990493);DlDRM1基因cDNA全长2 574bp,包括494bp的5′UTR,184bp的3′UTR,可编码包含631个氨基酸的蛋白质。(2)生物信息学分析显示,DlDRM1是一个不稳定的亲水蛋白,不含信号肽,不存在跨膜结构域,其分子式为C_(3104)H_(4839)N_(851)O_(984)S_(28);序列比对和系统进化分析表明,龙眼DlDRM1与脐橙DRM相似度最高(76.85%),二者亲缘关系也最为接近。(3)实时荧光定量PCR分析发现,DlDRM1在龙眼各组织器官中均有表达,且在果肉中表达量最高,其次是花蕾,在叶中的表达量最低;DlDRM1基因在非胚性愈伤组织中表达量最高,而且在非胚性愈伤向胚性愈伤转变过程中DlDRM1基因的表达量呈逐步下降趋势,说明DlDRM1基因与体胚胚性呈负相关关系,在龙眼体胚发生过程中可能发挥着重要的作用;一定浓度的IAA和2,4-D能够促进DlDRM1基因的表达,而KT则抑制DlDRM1的表达。(4)亚细胞定位结果表明,DlDRM1定位于细胞核和细胞膜上。     

龙眼组蛋白去乙酰化酶基因DlHDT1的克隆与特性分析

组蛋白去乙酰化是植物表观遗传调控的重要组成部分,对染色体结构修饰和基因表达调控发挥着重要的作用。为深入探究组蛋白去乙酰化酶基因(histone deacetylase 1,HDT1)在龙眼体胚发生过程中的功能,该研究结合龙眼基因组数据,采用RT-PCR方法克隆得到龙眼组蛋白去乙酰化酶基因(DlHDT1),对其进行生物信息学分析及亚细胞定位观察,同时结合转录组数据分析DlHDT1在体胚发生过程中的FPKM值,并利用qRT-PCR技术检测PEG6000和NaCl处理下DlHDT1的表达模式。结果表明:(1)DlHDT1基因CDS序列全长918 bp,编码305个氨基酸,该蛋白为不稳定亲水性蛋白,不含信号肽和跨膜结构,共含43个磷酸化位点,相对分子量为32 585.54 Da,等电点为4.65;进化树分析显示龙眼DlHDT1与漾濞槭亲缘关系最近(78.76%)。(2)亚细胞定位显示,DlHDT1蛋白定位于细胞核中;顺式作用元件分析发现DlHDT1基因含有大量光响应元件和脱落酸、茉莉酸甲酯等激素及逆境胁迫响应元件;转录组数据显示,DlHDT1在龙眼体胚发生不同时期均有表达,在胚性愈伤组织(EC)阶段表达最低,在球形胚(GE)阶段表达最高。(3)qRT-PCR显示,在PEG6000和NaCl处理下DlHDT1基因,呈下调表达趋势,推测DlHDT1可能参与调控龙眼对干旱及盐胁迫的响应,并存在负调控关系。研究认为,DlHDT1为核定位基因,可能参与龙眼体胚形态建成并在龙眼响应非生物逆境胁迫过程中发挥重要作用。     

龙眼WOX家族基因鉴定及在体胚发生早期的表达分析北大核心CSCD

为探究龙眼WUSCHEL相关的同源异型盒(WUSCHEL-related homeobox,WOX)家族基因的生物学功能与表达模式,该研究基于龙眼全基因组数据库对DlWOX家族成员进行鉴定与生物信息学分析,并利用实时荧光定量PCR(qRT-PCR)技术检测验证其在龙眼体胚发生早期三个阶段以及在不同激素处理下的表达模式。结果表明:(1)共筛选出13个龙眼DlWOX家族成员,均为不稳定蛋白;亚细胞定位预测显示DlWOX定位于细胞核与细胞骨架上;进化树分析发现,DlWOX家族分为远古支、中间支和WUS(WUSCHEL基因是WOX家族中最先发现的基因)支。(2)基因结构分析发现,DlWOX内含子数在0~19个之间,其大部分的编码蛋白都含有基序motif1与motif2,部分成员含有特异的基序;DlWOX启动子顺式作用元件包含大量光与激素响应元件。(3)对DlWOX在龙眼不同组织部位及体胚发生早期的表达模式分析发现,该家族部分成员在龙眼叶片中高表达,DlWOX14.1、DlWOX14.2和DlWOX9A在胚性愈伤组织阶段(EC)高表达;qRT-PCR分析显示,大部分龙眼DlWOX家族成员响应茉莉酸甲酯(MeJA)和赤霉素(GA)的调控,除DlWOX6外,其余成员在GA与MeJA处理下均上调表达,其中DlWOX9A在GA和MEJA处理下表达量显著上调。研究发现,龙眼DlWOX9A转录组测序结果与qRT-PCR结果的表达量存在差异并且趋势也不完全相同,推测WOX家族在龙眼的整个体胚发生早期起着重要的作用,尤其是在GE阶段;龙眼DlWOX基因在进化过程中存在高度的保守性,部分DlWOX家族成员可能通过响应GA与MeJA激素在龙眼体胚发生过程中发挥作用。     

龙眼体胚发生过程中DCL基因的分子特性及表达分析

从龙眼转录组unigene序列筛选获得龙眼DCL基因(命名为DlDCL)全长序列,并结合生物信息学及实时荧光定量等方法,对龙眼DCL基因进行研究,以明确DlDCLs在龙眼体胚、不同生长组织部位中的表达规律及其对激素和光质应答反应,为进一步研究龙眼体胚过程中DlDCLs基因的调控研究奠定基础。结果表明:(1)龙眼转录组数据存在DlDCL1、DlDCL2、DlDCL3和DlDCL4四个DCL家族成员,且基于Unigene的FPKM值发现不同基因在体胚发生阶段具有差异表达。(2)生物信息学分析发现,DlDCLs成员间基本理化性质较为类似,均为亲水性不稳定蛋白、不含信号肽、可进行跨膜运动,但也存在一定差异,如DlDCL2为碱性蛋白,而其他3个成员为酸性蛋白;亚细胞定位预测显示,DlDCLs均定位于细胞核中,但DlDCL2也存在定位于叶绿体中;对DCL蛋白结构域预测显示,DCL是高度保守的蛋白。(3)系统进化树分析显示,不同物种的DCL分为4个分支,同源的DCL蛋白都聚为一类,且DlDCLs与柑橘DCL亲缘关系更为接近。(4)实时荧光定量PCR分析表明,DlDCLs在龙眼非胚性愈伤组织和体胚发生过程中的表达模式差异较大,但DlDCLs在愈伤组织阶段均有较高的表达量,推测DlDCLs在体胚发生过程可能具有功能的独立和协作。DlDCL1和DlDCL2在叶片、花器官等组织部位中的相对表达量较高,暗示DlDCL1和DlDCL2可能参与到光合作用和花器官的发育;DlDCLs还受2,4-D、MeJA、SA激素和光质诱导,表明DlDCLs可能参与激素和光质调控。     

龙眼体胚CDC48基因的克隆、原核表达及其在体胚发生过程中的表达分析

为探讨龙眼(Dimocarpus longan)体胚CDC48基因的表达方式,采用RT-PCR和RACE方法,从龙眼胚性愈伤组织中克隆得到1条长度为2620 bp、含有完整开放阅读框的DlCDC48基因cDNA序列(GenBank登录号:EU606206)和长度为2418 bp的DNA序列(GenBank登录号:FJ590953)。DlCDC48编码1个含有805个氨基酸的蛋白质。DlCDC48基因不含内含子。生物信息学分析表明:DlCDC48蛋白为不具跨膜结构域的亲水性胞质蛋白,不具有信号肽,定位在细胞核;与其他植物的CDC48有较高的同源性。将DlCDC48基因构建成原核表达载体,经IPTG诱导表达了1个分子量约为89 kD的蛋白。利用实时荧光定量PCR(qPCR)技术,DlCDC48在龙眼体胚发育过程中的各个阶段均有表达,其中球形胚时的表达量最低,胚性紧实球形结构阶段的表达量最高。这为进一步研究CDC48基因在植物体胚发生中的作用奠定基础。     

龙眼UGD6基因克隆及其表达特性分析

该试验采用RT-PCR和RACE技术,对龙眼多糖合成的关键基因尿苷二磷酸-葡萄糖6-脱氢酶基因(DlUGD6)进行分离克隆、生物信息学分析和亚细胞定位研究,并采用qRT-PCR技术,对其在龙眼体细胞胚胎发生、合子胚发育及不同组织器官中的表达模式进行分析。结果表明:(1)DlUGD6基因的cDNA序列全长1 860bp,包含开放阅读框1 443bp,编码480个氨基酸(GenBank登录号KU198438);生物信息学分析显示,DlUGD6属于稳定的酸性亲水蛋白,不含信号肽,具有跨膜结构和3个典型的保守结构域,属于UDP-葡萄糖/GDP-甘露糖脱氢酶家族;进化树分析表明,DlUGD6与柑橘亲缘关系较近。(2)洋葱内表皮GFP荧光定位观察发现,DlUGD6定位于细胞质;qRT-PCR结果显示,DlUGD6在龙眼非胚性愈伤组织中表达量相对较高,且在其他体胚发育阶段也均有稳定表达;在合子胚发育中子叶胚形成后第8天(S3)和第24天(S7)时表达量最高,整体呈"W"型;在不同组织器官中,DlUGD6在花药和茎中的表达量最高,且整体上生殖器官中的表达水平高于营养器官。研究认为,DlUGD6基因可能参与龙眼生长发育各个阶段中细胞壁多糖合成。     

龙眼DlPPO1基因的克隆及其表达调控分析

该研究根据同源克隆技术,利用RT-PCR和RACE技术,以‘四季蜜’龙眼叶片cDNA为模板,获得龙眼多酚氧化酶基因(polyphenol oxidase,PPO)的3个转录本DlPPO1-a、DlPPO1-b和DlPPO1-c的cDNA全长序列(KM387405、KM516087和KM516088)和1条DNA序列DlPPO1(KU837229)。DlPPO1-a、DlPPO1-b和DlPPO1-c的全长分别为1 969、1 960和1 920bp,包含相同的完整开放阅读框1 800bp并编码599个氨基酸;该基因与荔枝、橄榄和枣等物种的PPO基因同源性较高。生物信息学分析表明,DlPPO1保守结构域具有多酚氧化酶的典型结构域特征。利用实时荧光定量PCR技术检测DlPPO1表达结果表明,在龙眼体胚发生过程中,DlPPO1从心形胚时期开始上调表达至子叶胚时期达到最高,推测其在龙眼体胚发生中后期可能发挥重要作用;DlPPO1在龙眼叶片中表达量最高,其次是花芽,而在其他组织部位表达量较低。激素和非生物胁迫处理下的表达分析表明,水杨酸(SA)、低浓度茉莉酸甲酯(MeJA)、NaCl、甘露醇及PEG可诱导DlPPO1基因上调表达,这些表达模式暗示其可能参与多种非生物胁迫应答过程。     

龙眼ERF家族成员鉴定及其在体胚发生早期的表达

为了解龙眼ERF家族的基本性质以及在体胚发生早期的表达规律,该研究对龙眼基因组鉴定出108个ERF家族成员进行基本理化性质分析与进化树构建,并结合龙眼转录组及miRNA、lncRNA数据库对DlERF家族成员与lncRNA、miRNA之间的关系预测与表达模式分析。结果显示：(1)理化性质及进化树分析表明,AP2/ERF结构域在龙眼中相对最为保守,DlERF对龙眼的抗胁迫能力以及对病菌的防御能力可能起着重要作用。(2)RNA Seq中的表达量分析可知,95个ERF基因在转录组中检测到表达,在愈伤组织(EC)、不完全胚性紧实结构(ICpEC)与球形胚(GE)阶段中分别存在31、11与53个ERF基因高表达。(3)qRT PCR结果显示,在龙眼体胚发生早期显著表达5个ERF基因中,Dlo_008317.1ERF15 2、Dlo_022310.1ERF1 1与Dlo_009939.1ERF98在GE阶段表达量显著高于EC与ICpEC阶段,Dlo_009070.1ERF106 3与Dlo_022634.1ERF22 1则分别在EC与ICpEC阶段高表达。(4)DlERF家族成员与相关lncRNA、miRNA表达量分析显示,LTCONS_00013739对靶基因Dlo_008317.1ERF15 2为正调控关系;Dlo_miR413与Dlo_miR1510a共同靶向Dlo_009070.1ERF106 3,从EC到GE阶段均表现出显著负调控关系,而Dlo_miR413与Dlo_008317.1ERF15 2的表达量表现出正相关,推测相比于Dlo_008317.1ERF15 2,Dlo_miR413更倾向于调控Dlo_009070.1ERF106 3;调控Dlo_009939.1ERF98相关的Dlo_miR408与Dlo_miR774b,从表达趋势来看,Dlo_miR774b在龙眼体胚发生早期过程能够负调控Dlo_009939.1ERF98;Dlo_miR399c与Dlo_022310.1ERF1 1在EC到GE阶段可能存在负调控关系。(5)不同梯度浓度的乙烯处理使得DlERF基因表达显著下调。这些发现提供了有关龙眼体胚发生早期DlERF的重要见解,从而为将来对龙眼体胚发生早期过程中DlERF的功能分析奠定了基础。     

龙眼体细胞胚胎发生过程中DlWUS的克隆与表达分析

以龙眼‘红核子’LC2悬浮细胞系诱导的胚性愈伤组织为基本材料,按照龙眼体细胞胚胎同步化方法诱导获得龙眼体胚不同阶段材料,并以龙眼体细胞胚胎发生不同阶段混合材料作为试验材料,采用RT-PCR结合RACE技术分离并克隆龙眼中编码同源异型结构域蛋白的转录因子WUSCHEL(简称DlWUS)的cDNA全长及DNA序列,并进行序列分析与表达分析。结果表明:DlWUS的cDNA全长1 110bp,开放阅读框(ORF)858bp,共编码285个氨基酸(GenBank登录号为KM017506),DlWUS的DNA包含2个内含子。序列分析表明,DlWUS是一个不稳定的亲水蛋白,不含信号肽,亚细胞定位于细胞核,具跨膜结构和Homeodomain超级家族的保守结构域以及WUS转录因子家族特有的WUS box和EAR-like结构域,推测该目的基因确实为WUS转录因子。系统进化分析显示,龙眼DlWUS与脐橙WUS归为一个分支,亲缘关系较近。实时荧光定量PCR分析结果表明,在龙眼体细胞胚胎发生整个过程中,DlWUS均有表达,但仅在球形胚时期表达量较高,说明DlWUS可能主要在球形胚阶段发挥作用,并且在一定浓度范围内,外源施加IAA和GA3能够促进DlWUS基因的表达,而外源施加SA则抑制DlWUS基因的表达。     

龙眼果实采后失水果皮褐变与活性氧及酚类代谢的关系

研究了(10±1)℃和50%相对湿度贮藏条件下"福眼"龙眼果实果皮褐变与活性氧和酚类代谢的关系.结果表明,采后失水导致龙眼果实果皮褐变,果皮活性氧清除酶SOD、CAT、APX、GR活性和内源抗氧化物质AsA、GSH含量下降,O-2产生速率和MDA含量增加,细胞膜透性迅速增大;PPO和POD活性增加,总酚和类黄酮含量明显下降.据此认为,果皮褐变可能是细胞的活性氧代谢失调,细胞膜结构破坏,使PPO、POD与酚类物质(含类黄酮)接触、酚类物质氧化的结果.     

龙眼核提取液的降血糖作用

以常规降血糖药(格列苯脲)为阳性对照,通过对正常小鼠和糖尿病小鼠进行降血糖治疗试验,研究了龙眼核提取液的降血糖作用。结果表明,龙眼核提取液能有效地缓解经四氧嘧啶诱发的糖尿病小鼠体内的高血糖症状,降血糖率达77.4%,具有良好的降血糖效果。     

龙眼生理落果中的纤维素酶、果胶酶活性及脱落酸含量

水涨龙眼脱落幼果的纤维素酶活性与落果比率之间具显著相关性（r=0．886）。在生理落果期间,脱落幼果纤维素酶活性高出正常幼果37％。果胶酶活性对生理落果的影响不甚明显,仅在落果中期表现出较强酶活性。在整个生理落果期间,脱落幼果脱落酸含量始终处在高于正常幼果的状态,其含量比正常幼果的脱落酸含量高167倍。幼果中纤维素酶活性强和脱落酸含量高是促进水涨龙眼生理落果的重要因素。     

琯溪蜜柚汁胞在发育与粒化过程中APX活性变化及其同工酶分析

对不同发育时期的琯溪蜜柚(Citrus grandis'Guanximiyou')汁胞进行APX活性测定及同工酶分析.结果表明,随着汁胞的发育,APX活性增大;而柚子衰老腐烂时,APX活性迅速降低.APX同工酶随着汁胞发育而发生变化,花后150 d的APX同工酶增加了1个组分(迁移率0.66),且随着汁胞的成熟,其表达量增加.花后230 d时柚子开始衰老腐烂,花后242 d已难以看到大部分APX同工酶酶谱.粒化汁胞的APX活性比正常汁胞大,同工酶酶谱亮度和清晰度也大,推测琯溪蜜柚汁胞在粒化过程中APX可清除活性氧61由基,抵抗氧化损伤.     

Characterization of ascorbate peroxidase in soybean under flooding and drought stresses

Flooding and drought are the two different forms of water stress that adversely affect the growth and development of soybean plant in particular at early stage. Ascorbate peroxidase (APX) is a known antioxidant enzyme that plays key role in abiotic stresses. To investigate the changes in APX in soybean under drought and flooding stresses, western blotting, enzyme activity assay and biophoton emission techniques were used. Flooding stress was imposed by adding excess amount of water in the sand and drought by withholding water supply. Under flooding stress, a decrease in APX was detected with time. Completely opposite trend was evident in hypocotyl and root of plants exposed to drought. Western blotting and APX activity results are complementary to each other. Biophoton emissions further confirmed the increasing and decreasing trend of APX under drought and flooding stress, respectively.     

Mitochondrial and peroxisomal ascorbate peroxidase of pea leaves

The isoenzyme pattern and the substrate specificity of the membrane-bound mitochondrial and peroxisomal ascorbate peroxidases (APX; EC 1.11.1.11) from pea leaves are studied. The substrate specificity of both APXs was assayed using the electron donors ascorbate and pyrogallol, whereas o-dianisidine, hydroquinone, tetramethylbenzidine and 4-methoxy-α-naphthol were also assayed with mitochondrial APX (mitAPX). In leaf mitochondria, the specific activity of APX was similar with pyrogallol and ascorbate, the activity being inhibited by p-CMS. mitAPX showed low activity with the guaiacol peroxidase (GPX)-type substrates, tetramethylbenzidine and 4-methoxy-α-naphthol. Activity of mitAPX with hydroquinone suggest a potential role of mitAPX in the drainage of electrons from the mitochondrial electron chain at the level of ubiquinone. In peroxisomes, the APX (perAPX) specific activity was much higher with pyrogallol than with ascorbate. This perAPX was more sensitive to incubation with Triton X-100 than the mitAPX. By native PAGE the mitAPX was resolved in 6 isoenzyme bands, and the activity of the 3 main bands (mitAPX III, III′ and IV) was inhibited by p-CMS. These 3 major isozymes were also present in mitochondrial membrane fractions. Staining for GPX activity with 4-methoxy-α-naphthol revealed that the APX detected in mitochondria did not have the capacity to oxidize 4-MN, and therefore cannot be considered as true GPX. When intact peroxisomes and peroxisomal membranes were subjected to native PAGE, no APX activity could be detected and this was probably due to the inactivation of perAPX. Results obtained suggest that pea mitochondrial APX (mitAPX) represent a distinct and novel isozyme different from those APXs of chloroplast and cytosolic origin previously reported. The peroxisomal APX (perAPX), however, appears to ressemble the chloroplast APXs as regards its sensitivity to Triton X-100.     

Regulation of ascorbate peroxidase activity in dark-grown radish cotyledons by a catalase inhibitor, 3-Amino-1,2,4-Triazole

The present study was performed to see the physiological role of cytosolic ascorbate peroxidase (APX) and its relationship to other enzymes involved in the H₂O₂ scavenging metabolism, and also to elucidate the regulation of APX expression in dark-grown radish (Raphanus sativus L. cv Taiwang) cotyledons. To do so, 3-amino-l,2,4-triazole (aminotriazole), a known specific inhibitor of catalase, was used to simulate a catalase-deficient phenomenon in cotyledons. Aminotriazole, in very low concetration (10^-4 M), inhibited remarkably the development of catalase activity in cotyledons during dark germination. This inhibition of catalase by aminotriazole, however, did not result in any significant changes in the growth response and the H₂O₂ level of developing cotyledons. In addition, the development of guaiacol peroxidase (GPX) activity was also not significantly affected. Unlike GPX, cytosolic APX activity was induced rapidly and reached a 1.7-fold increase in aminotriazole treated cotyledons at day 7 after germination. However,in vitro incubation of cytosolic APX preparation from cotyledons with aminotriazole did not result in any significant change in activity. One cytosolic APX isozyme (APXa) band involved in this APX activation was predominantly intensified in a native polyacrylamide gel by activity staining assay. This means that this APXa isozyme seems to play a key role in the expression of cytosolic APX activity. On the other hand, 2-day-old control seedlings treated with exogenous 1 mM H₂O₂ for 1 h showed a significant increase of cytosolic APX acitivity even in the absence of aminotriazole. Also, 2 μM cycloheximide treatment substantially inhibited the increase of APX activity due to aminotriazole. Based on these results, we suggest that a radish cytosolic APX could probably be substituted for catalase in H₂O₂ removal and that the expression of APX seems to be regulated by a change of endogenous H₂O₂ level which couples to APX protein synthesis in a translation stage in cotyledons.     

Comparative studies of H2O2 detoxifying enzymes in green and greening barley seedlings under cadmium stress

Cadmium(Cd) stress induced alterations in the activities of several representatives of the enzymatic antioxidant defense system such as guiacol peroxidase (POD), catalase (CAT) and ascorbate peroxidase (APX) were comparatively studied in green and greening barley seedlings that represent two different stages of plant development. Although roots were the main site of Cd accumulation, 1.5–3% of Cd was translocated into leaves and it induced oxidative damage which was indicated by the reduced chlorophyll and increased malondialdehyde content of the leaves. In roots of both types of seedlings exposed to various Cd concentrations, the APX activity was enhanced without any increase in the activity of POD. In leaves, however, elevated activities of both POD and APX could be observed. In roots of green seedlings at high concentration of Cd, the APX activity was reduced on the fourth day of culture but no inhibition was found in the POD activity. Leaf CAT which mainly represented the peroxisomal enzyme activity did not display any changes under Cd stress. Our results show that at both developmental stages barley seedlings exhibit a well-defined activity of the enzymatic antioxidant system, which operates differentially in roots and shoots subjected to Cd stress.     

The Effect of the Removal of the Area Postrema on Insulin and IGF-1-Induced Cardiovascular and Sympathetic Nervous Responses

Previous studies have demonstrated that insulin and IGF-1 both increase lumbar sympathetic nerve activity (LSNA) and decrease mean arterial pressure (MAP). We hypothesized that the peripheral responses to insulin and IGF-1 are mediated, at least in part, via the central nervous system. In this study we determined the effects of the peripheral administration of both insulin and IGF-1 on cardiovascular dynamics and LSNA following removal of the area postrema (APX), a major site of blood-brain communication. Insulin infusion in normal rats decreased MAP but increased HR and LSNA. When insulin was infused in APX rats it also decreased the MAP but the MAP recovered rapidly and plateaued at a level equivalent to normals after 40 min. Insulin significantly increased the HR and LSNA in the APX rats compared to normals. However, when hypoglycemia was prevented by glucose infusion, the HR and LSNA responses to insulin in the APX rats were similar to normals. IGF-1 also decreased MAP and to a greater extent in the APX rats compared to normals but the increased LSNA in APX rats was equivalent to normals. The APX rats when compared to normals had a greater sensitivity to insulin-induced hypoglycemia while IGF-1 decreased the plasma glucose to a lesser degree in APX rats. We conclude that insulin and IGF-1 entry into the CNS at least via the area postrema does not contribute significantly to the hypotensive response and that the greater depressor response to IGF-1 is likely due to enhanced vascular sensitivity in APX rats. The increased HR and LSNA following insulin were likely mediated by an increased reflexive response to hypoglycemia.