This method can help answer key questions. For example, how do oysters respond behaviorally to diel-cycling hypoxia and cyclical pH. How do they respond when diel-cycling hypoxia happens?
Do they close up? And after normoxia is restored, do they open back up? How do they respond?
The main advantage of this technique is that it measures the opening and the closing of the oysters in a very precise manner and is also a very affordable technique. Though this method can provide insight into the valve gape behavior of oysters in response to diel-cycling hypoxia, it can also be used for other species and it can be used to measure, for example, the adverse behavior of bivalves to contaminants or toxic algae. Plan to prepare 12 sensors;one for each channel on the Wheatstone bridge.
First, construct valve gape sensor cables. Twist two 32-gauge Teflon-coated wires of specific colors. Cut a length of twice the length of the desired gape sensor cable.
Make it twice the length from the aquarium to the SGM, plus a little extra. On one end, secure the wires to a power drill. Two people must hold either end of the wire which make the cable.
Now, the holders twist the cable using the drill on, while keeping the cable under tension. Next, cut the cable on the other end so there are four leads, two on each end. Then, strip two centimeters of insulation off each wire.
Next, tin the exposed wire. On one end, solder D-sub connector male crimp pins on the two leads. Then, insulate the leads with shrink tubing.
Next, solder the other two leads to the two contacts of a 1000-Ohm strain gauge that has 13.5 by 5.5 millimeter grids. Then, add a sensor identifying tag to the sensor cable. Now, seal the strain gauges using PCT two-way tape.
Use a 15 millimeter length of tape on the long ends and two millimeter lengths on the sides of the sensor. Next, apply Quick Gel to the sensor leads and to the attached open ends of the cables. Do not allow the Quick Gel to flow anywhere else.
Now, attach a second layer of PCT 2-A to the bottom and the top of the sensor. Periodically, while sealing the sensor, check the resistance to make certain it remains in range. To improve the seal, add thin strips of PCT 2-A to the edges, over the edges where the tape meets.
The strain gauges corrode quickly in sea water and thus must be very well sealed. The quality of the seal affects the useful life of the sensor. If any improvements can be made for this technique, it is how to better seal the strain gauges.
Generally, this experiment is performed on a dozen oysters. For demonstration purposes, only one will be manipulated. Begin with removing the oysters from the diel-cycling hypoxia aquaria before they enter the low plateau phase.
Then, clean and dry their shells. Next, use nail polish to write identifying numbers on each oyster. Then attach strain gauges to their valves using aquarium sealant.
Now, position the gauge sensor with the leads on the flat valve about midway along the shell and leave a loop of the strain gauge across the valves so as not to impede the opening of the valves. Next, glue the free end of the sensor onto the left valve. Now, check the resistance of the mounted sensors.
They must remain within the range of the readout electronics. Observe how the bending of a strain gauge caused by the oyster's opening and closing affects the voltage readings. Generally, 12 oysters are attached to 12 sensors for one experiment.
After 30 minutes, the aquarium sealant should be cured. Then, return the bivalves to an oxygenated holding bucket with flow-through water for at least five hours to allow the aquarium sealant to leach before submersion in the experimental aquaria. Later, transfer the oysters in their aquaria during the ramp-up phase to the super-saturation plateau.
Run all sensor cables to the SGM. Then, fill the 12 channels of the SGM with 12 sensor cables. Next, zero the gape signal for each individual sensor after using the potentiometer dials on the SGM.
At high solidities, plan to correct the drift in the offset of each sensor periodically so it stays in the range of the SGM and the Wheatstone bridge. When recovering the oysters from the aquaria to end the experiment, manually trigger each oyster to close. This produces a closing signal to know the direction and the magnitude of the closing signal.
During analysis, define the oyster as open if the sensor valve is larger than one half of the closed value and define the oyster as closed if the sensor value is smaller than one half of the closing value. Oysters exposed to uninterrupted normoxic estuarine water without hypoxia during the low plateau phase of the daily cycle were open most of the time. Most oysters exposed to severe hypoxia during the low plateau phase closed soon after their target-dissolved concentrations were reached.
Some closed even sooner. Being at a controlled or cyclical pH had no effect. Under mild hypoxia, closings often occurred late during the low plateau phase, instead of at the point when the target-dissolved oxygen was reached.
During the normoxia phase after the low plateau, oysters that experienced severe hypoxia during the low plateau were open most of the time and they often opened while dissolved oxygen levels were ramping up, even before the normoxia plateau had been reached. After watching this video, you should have a good idea how to make gape sensors and how to attach them to bivalves and how to measure behavioral responses of bivalves to diel-cycling hypoxia and cyclical pH. The apparatus is simple, low cost and easy to assemble.
After its development, this valve gape technique can also be used on other bivalve species and it can be used to measure the adverse behavior of bivalves to either contaminants or to toxic algae.
