Freezing survival by isolated malpighian tubules of the New Zealand alpine weta Hemideina maori

摘要

The ability of isolated Malpighian tubules from a freeze-tolerant insect, the New Zealand alpine weta (Hemideina maori), to withstand freezing was assessed by measuring post-freeze membrane potentials and rates of fluid secretion, The hemolymph of cold-acclimated Hemideina maori was found to contain relatively high concentrations of the cryoprotectants trehalose (>300 mmol l(-1)) and proline (41 mmol l(-1)). Survival of isolated Malpighian tubules was correspondingly high when a high concentration of trehalose was present in the bathing saline, Tubules allowed to recover for 20 min from a Ih freeze to -5 degrees C in saline containing 400 mmol l(-1) trehalose had a basolateral membrane potential of -53 mV compared with a potential of -63 mV in tubules not exposed to a freeze/thaw cycle, Fluid secretion in tubules that had experienced a freeze/thaw cycle in saline containing 400 mmol l(-1) trehalose was 9.9+/-2.6 nl h(-1) compared with 18.7+/-5.0 nl h(-1) (means +/- S.E.M., N=18) in tubules that had not been frozen, Tubules frozen in saline containing a lower concentration of trehalose (200 mmol l(-1)) or in glucose (400 mmol l(-1)) showed a similar ability to survive freezing to -5 degrees C, In contrast, freezing for Ih at -5 degrees C in saline containing 400 mmol l(-1) sucrose produced a 57 % decrease in membrane potential and an 88 % decrease in secretion rate, Tubules held in saline lacking high concentrations of sugars showed no survival after freezing to -5 degrees C for Ih, When frozen to -15 degrees C, tubules appeared to survive best in saline with the highest trehalose concentration (400 mmol l(-1)). Freezing damage was not simply the result of exposure to cold, since tubules chilled (unfrozen) to -5 degrees C for Ih were not compromised even when the bathing saline lacked a high sugar concentration, Exposure of tubules to a combination of low temperature and high osmolality mimicked damage caused by actual freezing: the membrane potential showed a 60 % recovery when the test was performed in saline containing trehalose, but showed no recovery in saline containing sucrose. [References: 43]