Abstract
Cope’s gray treefrogs, Hyla chrysoscelis, accumulate glycerol during the period of cold acclimation that leads to the development of freeze tolerance. Glycerol must cross cell membranes in numerous processes during this time, including exit from hepatocytes where glycerol is synthesized and entry into other tissues, where glycerol is cryoprotective. Thus, we hypothesized that erythrocytes from H. chrysoscelis would be permeable to glycerol and that that permeability would be up-regulated during cold acclimation. Further, we hypothesized that glycerol permeability would be associated with the expression of aquaporins, particularly those from the glyceroporin sub-family. Erythrocytes from warm-acclimated treefrogs had high glycerol permeability at 20°C, as assessed by the time required for osmotic lysis following suspension in 0.2 M glycerol. That osmotic lysis, as well as uptake of radio-labeled glycerol, was inhibited by 0.3 mM HgCl3. Permeability assessed via osmotic lysis was markedly reduced at 5°C. These properties were similar in animals deriving from northern (Ohio) and southern (Alabama) populations, although suggestive (through statistical interactions) of greater glycerol permeability in northern animals. Erythrocytes expressed mRNA and protein for a previously described glyceroporin, HC-3. In cold-acclimated animals, HC-3 protein expression was up-regulated, but we could not detect a concomitant enhancement of glycerol permeability.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Agre P, Preston GM, Smith BL, Jung JS, Raina S, Moon C, Guggino WB, Nielsen S (1993) Aquaporin CHIP: the archetypal molecular water channel. Am J Physiol 265:F463–F476
Aldrich K, Saunders DK (2001) Comparison of erythrocyte osmotic fragility among ectotherms and endotherms at three temperatures. J Therm Biol 26:179–182
Carlsen A, Wieth JO (1976) Glycerol transport in human red cells. Acta Phys Scand 97:501–513
Costanzo JP, Lee RE (1991) Freeze-thaw injury in erythrocytes of the freeze-tolerant wood frog, Rana sylvatica. Am J Physiol 261:R1346–R1350
Costanzo JP, Wright MF, Lee RE (1992) Freeze tolerance as an overwintering adaptation in Cope’s grey treefrog (Hyla chrysoscelis). Copeia 1992:565–569
Cutler CP, Martinez A, Cramb G (2007) The role of aquaporin 3 in teleost fish. Comp Biochem Physiol 148A:82–91
Deane EE, Woo NYS (2006) Tissue distribution, effects of salinity acclimation, and ontogeny of aquaporin 3 in the marine teleost, silver sea bream (Sparus sarba). Marine Biotechnol (New York, NY) 8:663–671
DeGier J, Van Deenen LLM, Van Senden KG (1966) Glycerol permeability of erythrocytes. Experientia 22:20–21
Ferreira C, van Voorst F, Martins A, Neves L, Oliveira R, Kielland-Brandt MC, Lucas C, Brandt A (2005) A member of the sugar transporter family, Stl1p is the glycerol/H+ symporter in Saccharomyces cerevisiae. Mol Biol Cell 16:2068–2076
Hara-Chikuma M, Verkman AS (2005) Aquaporin-3 functions as a glycerol transporter in mammalian skin. Biol Cell 97:479–486
Hara-Chikuma M, Sohara E, Rai T, Ikawa M, Okabe M, Sasaki S, Uchida S, Verkman AS (2005) Progressive adipocyte hypertrophy in aquaporin-7-deficient mice: adipocyte glycerol permeability as a novel regulator of fat accumulation. J Biol Chem 280:15493–15496
Hite RK, Gonen T, Harrison SC, Walz T (2008) Interactions of lipids with aquaporin-0 and other membrane proteins. Pflug Arch 456:651–661
Irwin JT, Lee RE Jr (2003) Geographic variation in energy storage and physiological responses to freezing in the gray treefrogs Hyla versicolor and H. chrysoscelis. J Exp Biol 206:2859–2867
Lages FA, Lucas C (1995) Characterization of a glycerol/H+ symport in the halotolerant yeast Pichia sorbitophila. Yeast 11:111–119
Layne JR, Jones AL (2001) Freeze tolerance in the gray treefrog: cryoprotectant mobilization and organ dehydration. J Exp Zool 290:1–5
Layne JR, Stapleton MG (2009) Annual variation in glycerol mobilization and effect of freeze rigor on post-thaw locomotion in the freeze-tolerant frog Hyla versicolor. J Comp Physiol 179:215–221
Liu Y, Promeneur D, Rojek A, Kumar N, Frøkiær J, Nielsen S, King LS, Agre P, Carbrey JM (2007) Aquaporin 9 is the major pathway for glycerol uptake by mouse erythrocytes, with implications for malarial virulence. Proc Natl Acad Sci USA 104:12560–12564
Lucas C, da Costa M, van Uden N (1990) Osmoregulatory active sodium-glycerol co-transport in the halotolerant yeast Debaryomyces hansenii. Yeast 6:187–191
Mclean IW, Nakane PK (1974) Periodate-lysine-paraformaldehyde fixative. A new fixation for immunoelectron microscopy. J Histochem Cytochem 22:1077–1083
Mobasheri A, Wray S, Marples D (2005) Distribution of AQP2 and AQP3 water channels in human tissue microarrays. J Mol Hist 36:1–14
Mochida H, Nakakura T, Suzuki M, Hayashi H, Kikuyama S, Tanaka S (2008) Immunolocalization of a mammalian aquaporin 3 homolog in water-transporting epithelial cells in several organs of the clawed toad Xenopus laevis. Cell Tissue Res 333:297–309
Ohta K, Inoue K, Hayashi Y, Yuasa H (2006) Carrier-mediated transport of glycerol in the perfused rat small intestine. Biol Pharm Bull 29:785–789
Promeneur D, Liu Y, Maciel J, Agre P, King LS, Kumar N (2007) Aquaglyceroporin PbAQP during intraerythrocytic development of the malaria parasite Plasmodium berghei. Proc Natl Acad Sci USA 104:2211–2216
Rojek AM, Skowronski MT, Fuchtbauer E-M, Fuchtbauer AC, Fenton RA, Agre P, Rokiaer FJ, Nielsen S (2007) Defective glycerol metabolism in aquaporin 9 (AQP9) knockout mice. Proc Natl Acad Sci USA 104:3609–3614
Rojek A, Praetorius J, Frokiaer J, Nielsen S, Fenton RA (2008) A current view of the mammalian aquaglyceroporins. Ann Rev Phys 70:301–327
Roudier N, Verbavatz J-M, Maurel C, Ripoche P, Tacnet F (1998) Evidence for the presence of Aquaporin-3 in human red blood cells. J Biol Chem 273:8407–8412
Sauer A, Jurzion T, Meyerstein D, Meyersterin N (1991) Kinetics of hemolysis of normal and abnormal red blood cells in glycerol-containing media. Biochim Biophys Acta 1063:203–208
Schmid W (1982) Survival of frogs in low temperature. Science 215:697–698
Sohara E, Rai T, Miyazaki J, Verkman AS, Sasaki S, Uchida S (2005) Defective water and glycerol transport in the proximal tubules of AQP7 knockout mice. Am J Physiol 289:F1195–F1200
Storey JM, Storey KB (1985) Adaptations of metabolism for freeze tolerance in the gray tree frog, Hyla versicolor. Can J Zool 63:49–54
Tanaka S, Kurosumi K (1992) A certain step of proteolytic processing of proopiomelanocortin occurs during the transition between two distinct stages of secretory granules maturation in rat anterior pituitary corticotrophs. Endocrinology 131:779–786
Vom Dahl S, Haussinger D (1997) Evidence for a phloretin-sensitive glycerol transport mechanism in the perfused rat liver. Am J Physiol 272:G563–G574
Zimmerman SL, Frisbie J, Goldstein DL, West J, Rivera K, Krane CM (2007) Excretion and conservation of glycerol, and expression of aquaporins and glyceroporins, during cold acclimation in Cope’s gray tree frog, Hyla chrysoscelis. Am J Physiol 292:R544–R555
Zou C, Agar NS, Jones GL (2000) Haemolysis of human and sheep red blood cells in glycerol media: the effect of pH and the role of band 3. Comp Biochem Physiol 127A:347–353
Acknowledgments
We would like to thank Jim O’Boyle and Dr. Steven Secor for assistance with collecting frogs, and Venkateshwar Mutyam, Julie Carroll, and Matthew Armstrong for technical assistance. This research was supported in part by research Grant NSF IOB-0517301 to D.L.G. and C.M.K.
Author information
Authors and Affiliations
Corresponding author
Additional information
Communicated by H.V. Carey.
Rights and permissions
About this article
Cite this article
Goldstein, D.L., Frisbie, J., Diller, A. et al. Glycerol uptake by erythrocytes from warm- and cold-acclimated Cope’s gray treefrogs. J Comp Physiol B 180, 1257–1265 (2010). https://doi.org/10.1007/s00360-010-0496-4
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00360-010-0496-4