ADAPTATIONS OF PLASMA-MEMBRANE GLUCOSE-TRANSPORT FACILITATE CRYOPROTECTANT DISTRIBUTION IN FREEZE-TOLERANT FROGS

Natural freeze tolerance in several anuran species involves the accummulation of high concentrations of glucose as a cryoprotectant in body fluids and tissues. The present study identifies an important new molecular mechanism supporting freeze tolerance, an adaptive increase in the capacity for facilitated transport of cryoprotectant across plasma membranes by increasing the numbers and/or activity of plasma membrane glucose transporters. Glucose transport by membranes isolated from liver and skeletal muscle was analyzed in two species, the freeze-tolerant wood frog Rana sylvatica and the freeze-intolerant leopard frog Rana pipiens. Membranes from both liver and muscle of R. sylvatica displayed much higher rates of carrier-mediated glucose transport, measured by a rapid filtration technique, compared with corresponding rates for R. pipiens membranes. For the liver V(max) values for glucose transport by membrane vesicles were 69 +/- 18 and 8.4 +/- 2.3 nmol . mg protein-1 . s-1 at 10-degrees-C for R. sylvatica and R. pipiens, respectively. This difference was due primarily to a greater number of glucose transporters in R. sylvatica liver membranes; the total number of transporter sites, determined by cytochalasin B binding, was 4.7-fold higher in the freeze-tolerant species. For muscle membranes, the V(max) for glucose transport was 4.9 +/- 1 and 0.6 +/- 0.16 nmol . mg-1 . s- 1 at 22-degrees-C for R. sylvatica and R. pipiens, respectively. However, in muscle there were no differences in the number of membrane transporters between species. We conclude that the greater capacity for plasma membrane glucose transport in R. sylvatica is a key adaptation that facilitates the rapid export of glucose cryoprotectant from liver and its uptake into other organs during the early hours of freezing exposure.