A Strain Gauge Monitor (SGM) for Continuous Valve Gape Measurements in Bivalve Molluscs in Response to Laboratory Induced Diel-cycling Hypoxia and pH

Elka T Porter; Frederick S Porter

doi:10.3791/57404

JoVE Journal > Environment

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Ambiente

基于实验室诱导的昼夜循环缺氧和 pH 值的双壳软体动物连续气门目瞪口呆测量用应变计监测仪 (通用)

Published: August 01, 2018

doi:

10.3791/57404

Elka T Porter, Frederick S Porter

¹Yale Gordon College of Arts & Sciences,University of Baltimore, ²Goddard Space Flight Center,NASA

Summary

了解双壳悬浮馈线对环境变量 (如溶解氧) 的行为反应, 可以解释一些生态系统过程。我们开发了一个价格低廉, 以实验室为基础的应变计监测仪 (锦葵) 来测量牡蛎、牡蛎、昼夜循环缺氧和周期性 pH 值的气门打呵欠反应。

Abstract

研制了一种成本低廉、基于实验室的应变计阀, 用于监测双壳软体动物在昼夜循环缺氧时的气门打呵欠行为。一座惠斯通电桥连接到与牡蛎壳相连的应变计 (锦葵)。记录的信号允许的开放和关闭的双壳贝类将连续记录在两天期间的实验性昼夜循环缺氧和昼夜循环变化的 pH 值。在这里, 我们描述了一个用于开发廉价的应变计监视器的协议, 并在一个示例实验室实验中描述了如何使用它来测量东部牡蛎 (锦葵) 的气门打呵欠行为, 以应对昼夜循环缺氧和pH 值的周期性变化。对周期性严重缺氧 (0.6 毫克/升) 溶解氧条件下的牡蛎进行了气门打呵欠测量, pH 值、周期性轻度缺氧 (1.7 毫克/升) 条件和常氧 (7.3 毫克/升) 条件下无周期性变化。我们证明, 当牡蛎遇到重复的昼夜周期, 他们迅速关闭他们的壳, 以应对严重缺氧和关闭的时间滞后到轻度缺氧。当 normoxia 恢复时, 它们又迅速地打开了。牡蛎没有反应周期性的 pH 条件叠加在昼夜循环严重缺氧。在氧条件降低时, 三个以上的牡蛎同时关闭。我们证明, 牡蛎对昼夜循环缺氧的反应, 这必须考虑当评估的行为, 双壳贝类溶解氧。该阀可用于评估双壳软体动物对溶解氧或污染物的变化的反应。密封技术, 以更好地密封的阀门打哈欠应变计从海水需要进一步改进, 以提高传感器的寿命。

Introduction

缺氧,即溶解氧浓度足够低, 对生物和生态过程产生负面影响, 但通常功能上定义为 [做] < 2 毫克/升¹, 缺氧 (功能上定义为 0.0-0.2 毫克/升)在世界沿海水域、河口和深海²^、³发生频繁和严重的情况, 并经常因增加富营养化⁴^、⁵而加剧。随着缺氧和缺氧程度的增加, 动物对生境的影响和生境质量的丧失。预计气候变化会恶化缺氧和缺氧⁶。

在许多分层, 营养丰富的河口, 如美国切萨皮克湾, 季节性持续缺氧可以战胜和可能发生年复一年²年。此外, 昼夜的低氧循环经常出现在诸如切萨皮克湾和其他地点的河口, 并发生在夜间或清晨⁷^,⁸的凌晨时间。

大多数研究的重点是持续暴露的有机体对低 [做] 和对他们的耐受性缺氧和缺氧⁹^,¹⁰^,¹¹^,¹²^,¹³^,¹⁴.此外, 研究还观察了物种分布、丰度和物种组成的大规模转移, 以响应延长的低 [做]⁴^,¹⁵。通常对低 [做] 非常敏感的物种, 死于大众,¹⁶将剩下的物种转移到一个年轻的, 规模较小, 短命的动物群中, 例如, 在路易斯安那州-德克萨斯大陆架生态系统⁴上发现。

行为变化通常在社区崩溃^{之前 17}和研究报告了有机体的行为反应到延长的低 [做]⁴^,¹⁶^,¹⁷^,¹⁸^,¹⁹ ^,²⁰^,²¹^,²²^,²³^,²⁴^,²⁵。然而, 这些研究并不侧重于生物体对缺氧昼夜循环暴露的反应以及河口 [做] 可用性的波动性。

昼夜在浅水河口的循环缺氧已得到越来越多的认识, 因为研究监测 [做] 更频繁的过程中的天数与 sondes 在河口¹⁶^,²⁶。水可以保持缺氧几个小时, 在夜间或清晨小时, 当没有氧气的光合作用在夜间, 但高耗氧呼吸⁷^,¹⁶。研究还发现, 潮汐影响低条件下的昼夜循环, 当低潮与夜间²⁷的结束相吻合时, 观测到最极端的极小值。只有在几个小时的缺氧后才会回到 normoxia⁷^,¹⁶^,²⁸在每天的周期。

为了确定锦葵对昼夜循环缺氧和 ph 值的行为反应, 我们监测了暴露于实验室诱导的昼夜循环和周期性 pH 值的牡蛎瓣膜的开启和关闭。双壳贝类的打呵欠反应已被用来检测恶劣的环境条件。在反应污染物²⁹^、³⁰^、³¹、有毒藻类³²^、³³^、³⁴、热污染³⁵^、³⁶的情况下, 双壳贝类阀门关闭^,³⁷、食物数量减少³⁸^,³⁹^,⁴⁰, 哺养的政权³⁹,⁴¹, 再现³⁷^,⁴², 光周期⁴³^,⁴⁴、ph 值⁴⁵^、⁴⁶、ph 值和溶解氧的总和⁴⁷均已测定。例如, 打呵欠技术包括直接观测⁴⁸^,⁴⁹^,¹³, 连续测量使用簧片开关和磁铁 (Dreissena 显示器)⁵⁰, 或光纤传感器⁵¹需要清水。此外, 磁铁和磁场强度霍尔传感器已用于研究贻贝角⁵²^,⁵³^,⁵⁴^,⁵⁵和高频电磁感应系统这可以测量两个电线圈之间的不同距离, 被粘在阀门已使用⁵⁶^,⁵⁷^,⁵⁸^,⁵⁹。电磁感应系统需要高压源, 必须将电源输送到外壳⁵²的两侧。该系统还可作为 “MOSSELMONITOR” (http://mosselmonitor.nl/) 提供商业用途。

在紧张的研究预算, 我们建立了一个廉价的应变计监测 (通用), 以持续测量牡蛎目瞪口呆的实验室诱导昼夜循环的 [做] 和 pH 值, 在低能见度条件下。我们的系统也比竞争系统简单得多, 允许许多动物在实验中进行检测。我们想确定 C 的行为反应. 锦葵昼夜循环严重 ([做] = 0.6 毫克/升) 缺氧与控制 ph 值 (ph = 7.8) 和循环 ph 值 (ph = 7.8-7.0), 并对轻度 ([做] = 1.7 毫克/升) 缺氧的目瞪口呆反应。此外, 我们想确定牡蛎是否能够迅速响应在昼夜周期的变化, 以及他们如何反应时, normoxia 返回后, 缺氧事件。也许牡蛎是最佳地适应迅速地波动的环境在许多河口¹⁶^,²⁷在他们居住的地方被发现。虽然有更多复杂的阀门目瞪口呆显示器可用, 但通用提供了一种廉价的技术, 允许连续测量阀门在水中的目瞪口呆, 即使在低能见度条件。

图 1。用于气门打呵欠装置的惠斯通电桥.请单击此处查看此图的较大版本.

用于监测双壳类打呵欠的应变计传感器是聚酰亚胺支持下的一种曲折模式的电阻膜。少量的应变调节传感器的电阻。当呵欠导致传感器的电阻发生变化时, 双壳贝类会弯曲应变片。我们使用了一个零位, 平衡, 惠斯通电桥每双壳的通道, 如图 1所示, 以测量传感器电阻的变化。惠斯通电桥由电位器清零, 允许数据存储器使用相当高的增益。惠斯通电桥是一种标准方法, 用于准确测量未知电阻, 使用与已知电阻标准和电压表的比值。这个非常老技术的历史在 Ekelof (2001)⁶⁰被谈论。我们集成了12通道, 每一个都有自己的惠斯通电桥和零电位计, 进入应变仪显示器 (通用) 单元。

Protocol

1. 气门打呵欠装置的惠斯通电桥的施工注: 应变表名义上为1000ω, 所以要完全平衡和空桥, 所有组件应为1000Ω。如图 1所示, 焊锡两 1 kΩ精密电阻, 然后再到976ω电阻和100Ω10转电位器。典型的应变计范围是几个ω在1000ω有名无实, 因此只有这种抵抗需要被匹配。使用固定的976Ω电阻和可调电阻 (10 转电位器) 为此目的。将可变臂的一侧和 1 kΩ电…

Representative Results

暴露在不间断的常氧河口水中的牡蛎 (在日常周期的低高原阶段没有缺氧) 大部分时间都是开放的, 只是很少短暂关闭 (图 5)。当他们关闭从牡蛎到牡蛎不同。Loosanoff 和 Nomejko 194644和希金斯 198039也发现了这种模式。牡蛎也没有反应在黑暗和轻的阶段的区别。 <img alt="Figure…

Discussion

典型的研究集中在连续, 延长的时间期间低氧气条件和反应, 经常被测量作为生存, 动物。然而, 目前我们对动物对昼夜循环缺氧的行为反应的理解最小为⁶³。因此, 更多的研究应该集中在反应昼夜循环缺氧的有机体的行为, 这在夏天经常发生在许多河口⁷^,⁸。

在此, 我们提出了一种连续测量双壳贝类对昼夜循环缺氧和?…

Divulgazioni

The authors have nothing to disclose.

Acknowledgements

我们感谢梅林达 Forseth 拍摄牡蛎的照片和测量他们在 ImageJ 的打呵欠宽度。我们感谢丹尼斯 Breitburg 水族馆与昼夜循环缺氧和周期性 pH 条件的接触。我们感谢史密森环境研究中心, Edgewater, 马里兰, 为实验空间。低氧实验由国家海洋和大气管理中心资助, 为海岸海洋研究资助号。NA10NOS4780138 和史密森亨特登基金对丹尼斯 Breitburg。在缺氧实验过程中, 阀门的打呵欠测量是由华盛顿学院为埃尔卡 t. 波特提供的一项师资增强补助金资助的。

Materials

Campbell CR 10x data logger	Campbell Scientific, Logan, Utah		Or other data logger. At Campbell the CR 10X has been replaced with the CR 1000
Campbell CR 10x multiplexer	Campbell Scientific, Logan, Utah		Data logger needs to have space for 12 channels
Dsub connector male crimp pins	TE Connectivity	205089-1	pins for gape sensor leads
PCA tape	Micro Measurements Corp, NC		To seal the strain gauge
Duro Quick Gel	Ace Hardware		Superglue
SG13/1000-LY43 or LY41	Omega Engineering Inc., Stanford, CT		Strain gauges
32 AWG (7/40) teflon Alpha wires	AlphaWire, Elizabeth, NJ	2840/7	Sensor cables, different colors are available
1/16" heat shrink tubing	Qualtek	B01A3QKKO6	To seal the leads of the sensor cable
Weller WES51 Analog Soldering Station	Amazon		Lots of soldering, need a good soldering iron. https://www.amazon.com/Weller-WES51-Analog-Soldering-Station/dp/B000BRC2XU/ref=sr_1_23?s=hi&ie=UTF8&qid=1505654295 &sr=1-23&keywords=soldering+iron
Rosin Soldering Flux Paste	Amazon		Needed for soldering
60-40 Tin Lead Rosin Core Solder Wire	Amazon		Needed for soldering
Aquarium sealant	Home Depot		Attach sensors to bivalve
PC Laptop			Any old PC to run Campbell gape program
heat gun	Amazon		shrink shrink tubing
Drill	Hardware store, Amazon		for twisting wires to make sensor cables
AC to DC power module	Acopian	DB15-30	Wheatstone bridge power supply
Poteniometer	Clarostat	733A	Wheatsone bridge nulling
isolating BNC connector	Sterren Electronics	"200-148	Wheatstone bridge output for multimeter
Fused AC receptical panel module	Adam technologies	IEC-GS-1-200	Wheatstone bridge power supply connector
976 ohm 1% resistor	Vishay Dale	CMF50976R00FHEB	Wheatstone bridge resistor
1 kohm 1% resistor	Vishay Dale	CMF501K0000FHEB	Wheatstone bridge resistor
Potentiometer scale dial	Kilo International	462	10 turn dial for nulling potentiometer
DB25 male panel connector	TE connectivity	1757819-8	Data logger connector on Wheatstone bridge
DB25 female panel connector	TE connectivity	1757819-8	Sensor connector to Wheatstone bridge
perforated circuit board	Vector electronics	64P44WE	circuit board for mounting of bridge components
enclosure	Hammond Manufacturing	1444-29	Enclosure for sensor readout electronics

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Porter, E. T., Porter, F. S. A Strain Gauge Monitor (SGM) for Continuous Valve Gape Measurements in Bivalve Molluscs in Response to Laboratory Induced Diel-cycling Hypoxia and pH. J. Vis. Exp. (138), e57404, doi:10.3791/57404 (2018).