Thermal inactivation of volume-sensitive K+,Cl− cotransport and plasma membrane relief changes in human erythrocytes
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Previously, we reported that in mammalian erythrocytes irreversible annealing of spectrin heterodimers at 49–50 °C abolished cell volume-dependent regulation of ion carriers, thus suggesting an implication of a two-dimensional (2D) membrane carcass in volume sensing and/or signal transduction. To further examine this hypothesis, we employed atomic force microscopy. This method revealed folded membrane relief of fixed human erythrocytes with an average wave height of 3–5 nm covered by globular structures with a diameter of 40–50 nm and an average height of 1–2 nm. Erythrocyte swelling caused by reduction of medium osmolality decreased the height of membrane surface waves by 40 % and increased K+,Cl− cotransport by approximately sixfold. Both volume-sensitive changes of membrane relief and activity of K+,Cl− cotransporter were abolished by a 10-min preincubation at 50 °C. Our results strongly suggest that volume-dependent alterations of the human erythrocyte membrane relief are caused by reorganization of the 2D spectrin–actin network contributing to regulation of the activity of volume-sensitive ion transporters.
KeywordsCell volume Human erythrocytes K+,Cl− cotransport Membrane roughness Membrane relief oscillations
This work was supported by grants from the Natural Sciences and Engineering Research Council of Canada (to R.G. and S.N.O.), the Russian Foundation for Fundamental Research (12-04-00033-a to S.N.O; 10-04-00835-а to G.V.M. and E.Y.P.), and the Ministry of Education and Science of the Russian Federation (projects ##8477 and 8162). Manuscript editing by Ovid Da Silva and logistical services from the Research Support Office, CRCHUM are appreciated.
- 7.Francis LW, Lewis PD, Wright CJ, Conlan RS (2010) Atomic force microscopy comes of age. Biol Cell 102:133–143Google Scholar
- 8.Garay R, Nazaret C, Hannaert R, Cragoe E (1988) Demonstration of a K+, Cl− cotransport system in human red blood cells by its sensitivity to [(dihydroindenyl)oxy]alcanoic acid regulation of cell swelling and distinction from bumetanide-sensitive Na+, K+, Cl− cotransport system. Mol Pharmacol 33:696–701PubMedGoogle Scholar
- 9.Girasole M, Pompeo G, Cricenti A, Congiu-Castellano A, Andreola F, Serafino A, Frazer BH, Boumis G, Amiconi G (2007) Roughness of the plasma membrane as an independent morphological parameter to study RBCs: A quantitative atomic force microscopy investigation. Biochim Biophys Acta 1768:1268–1276PubMedCrossRefGoogle Scholar
- 17.Oberleithner H (2008) Nanophysiology: fact or fiction? Pfluger Arch.- Eur. J Physiol 456:1–2Google Scholar
- 21.Orlov SN, Kuznetsov SR, Kolosova IA, Aksentsev SL, Konev SV (1997) Volume-dependent regulation of ion transporters in human and rat erythrocytes: role of cytoskeleton and protein phosphorylation. Russ Physiol J 83(No5–6):119–147Google Scholar
- 25.Sachs JR (1998) How do red blood cells know how big they are? In: Okada Y (ed) Cell volume regulation: the molecular mechanism and volume sensing machinery. Elsevier Science, Tokyo, pp 3–13Google Scholar