Hemolymph Glucose Levels as a Measure of Crayfish Stress: A Methodology Using a Human Glucometer

The objective of this study was to develop a methodology for measuring hemolymph glucose levels in crayfish of the genus Orconectes using a human glucometer. A secondary objective was to confirm through the use of this methodology that subjecting crayfish to acute stress in the form of a short‐term exposure to a basic solution or to chronic stress in the form of prolonged exposure to a nitrate‐containing solution, would increase hemolymph glucose levels. To measure the effects of acute exposure to a basic solution on crayfish hemolymph glucose levels, crayfish were placed in a beaker containing a basic solution (Ringer's buffer, pH=8) or a control solution (Ringer's buffer, pH=7) for a period of 10 seconds, after which time they were removed from the beakers and hemolymph was collected with a syringe. A ReliOn Prime glucometer (Wal‐Mart Stores, Inc.) was used to measured glucose levels (mg/dl) directly from the hemolymph. In order for the glucose levels to be within the sensitivity of the glucometer, hemolymph was mixed with high glucose Dulbecco's Modified Eagle Medium (DMEM; Gibco) in a 3:1 ratio. Glucose levels were determined by subtracting the levels of glucose measured in the DMEM solution from the glucose levels measured in the DMEM+hemolymph solution and correcting for the dilution factor. Crayfish acutely stressed through immersion in a basic solution had significantly higher hemolymph glucose levels compared to control crayfish (45.8 ± 5.9 vs 12.4 ± 3.8 mg/dl, two‐tailed t‐test; p<0.0001). To measure the effects of chronic exposure to a solution containing nitrate on crayfish hemolymph glucose levels, crayfish were housed in control tanks (no nitrate) or tanks containing 14 mg/L nitrate for one week, after which time hemolymph was drawn and glucose levels measured using the methodology described above. Crayfish chronically stressed through exposure to a nitrate‐containing solution had significantly higher hemolymph glucose levels compared to control crayfish (45 ± 14.3 vs 28.4 ± 8.8 mg/dl, two‐tailed t‐test; p=0.045). These results show that implementing this methodology human glucometers can be used to determine crayfish hemolymph glucose levels, making them an easy, inexpensive, and reliable alternative for this type of measurement. We were also able to show that hemolymph glucose levels are increased in both chronically and acutely stressed crayfish compared to control crayfish. These experiments can be used in physiology classrooms to demonstrate and discuss the role of the autonomic nervous system in an animal's response to stressful environmental conditions. Support or Funding Information This research was funded by the Denison University Department of Biology. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .