This method can help answer key questions in the environmental and ecotoxicalogical fields involving the relationship between cardiac and locomotor activities on the various acute and prolonged stressful conditions. The main advantages of this technique are that crayfish don't require complex prior manipulations or prolonged adaptation and are capable of bearing sensors for a few month between methods. The implications of this technique extend toward etiological, physiological, ecotoxicological, and industrial applications as crayfish aided noninvasive biomonitoring can be implemented in laboratories and water treatment and supply facilities.
Though this method can provide inside insight into crayfish cardiac, physiology, and behavior and water quality control, it can potentially be applied to other large freshwater and marine crustaceans. Visual demonstration of this method is critical for understanding the link between noninvasive sensor using near infrared light for evaluation in crayfish heart and movement detection modules based on image processing. A few days before beginning the procedure, select adult specimens with a carapace size of at least 30 millimeters.
And examine each specimen for the absence of disease and whether they lift both cheli when touched. Then allow the healthy crayfish to acclimate in a tank of settled tap water until the day of the procedure. As soon as the sensors have been prepared, open the sensor software on the computer and enter the number of crayfish to be fixed to the sensors and displayed on the screen.
Set the number of crayfish heart rates to be visualized. And use a paper towel to dry the dorsal carapace of the first animal. Wrap the cheli and abdomen of the crayfish in the paper towel to avoid damage and to eliminate additional stress to the animal.
And attach the censor to the dorsal carapace in a position that facilitates a maximal cardiac signal amplitude. Holding the crayfish with the sensor in one hand, use the other hand to add a drop of freshly mixed epoxy glue onto each of the four auxiliary wires located on the sensor. Then allow the glue to dry without moving the sensor for at least five minutes.
When the glue is no longer sticky to the touch, place the unwrapped crayfish and sensor into a box without water for a few more minutes until the glue is completely dry. Before moving the crayfish back into the tank, immerse the cephalothorax into the tank water for a few seconds several times to discharge the air that has accumulated in the gills. And leave the crayfish in the holding water tank for approximately one hour to remove any excess chemicals.
Then, release the crayfish into the experimental water tanks for one to two weeks of acclimation and under the appropriate experimental conditions depending on the observed physiological indices. As soon as the crayfish are placed in the experimental tanks, start the software. The video camera will automatically switch on.
Then select the movement detection option and locate each tank on the screen to start tracking the behavior and linking the behavior of the crayfish with the cardiac activity recordings. The recorded crayfish cardiac and behavioral activities can be save in a TXT format file. It is crucial to include the locomotion of the crayfish in the analysis as this activity can cause changes in the heart rate.
For example, in this experiment ten seconds after the start of the experiment a food odor was delivered into the tank via a peristaltic pump. At 14 seconds the crayfish recognized the stimulus and it's heart rate slightly decreased due to the so called orienting response. After 20 seconds, the heart rate increased resulting in a decrease in cardiac intervals.
At 26 seconds the crayfish moved toward the stimulus source and both the physiological excitation caused by the food odor and the locomotion initiation resulted in a substantial heart rate increase. At 37 seconds there was evidence of abrupt crayfish motion that could have substantially contributed to the heart rate growth during the reactions to the stimulus. Indeed, a disturbed crayfish typically demonstrates an increase in heart rate that is often associated with occasional locomotion.
However, even motionless crayfish can demonstrate a high heart rate that also indicates a pronounced stress. The heart rate of an undisturbed crayfish is characterized by a monotonic amplitude of the heart beat curve and by approximately equal cardiac intervals between each cardiac peak. While attempting this procedure it's important to remember that a quick and thorough sensor attachment will cause less stress to the experimental animals enabling the acquisition of precise physiological characteristics.
More advanced monitoring methods include the use of fully contactless crayfish monitoring which allows the heart beat frequency to be determined using only the combination of a near infrared sensor and a sensitive camera. After its development, this approach paved the way for researchers in the fields of behavior, physiology, reproduction, and androgyne of unrestricted crayfish and other large aquatic invertebrates to explore environmental and anthropogenic impact on bioindicator organisms. For the noninvasive biomonitoring, has a very practical application at the local variety in the Czech Republic as a real time water quality early monitoring system.
While the stability of the water state is continuously assessed to the dynamics of crayfish ecophysiological characteristics.
