Osmotic behaviour of human red blood cells: an interpretation in terms of negative intracellular fluid pressure.

1. The observation that human red blood cells do not shrink in hypertonic media as much as expected for ideal osmometers has previously been explained in terms of either a marked increase in the osmotic coefficient of the cell contents or an increase in the chloride content of the cells.2. Changes in suspension pH and haematocrit have been observed when the concentration of the unbuffered NaCl medium was doubled. The small increases in external pH, and the size of the volume decreases, are inconsistent with variations in the Cl content as a significant factor in the non-ideal osmotic responses.3. Membrane potentials of red cells in buffered media were followed using the fluorescent dye, diS-C(3)-(5). On shrinking at pH 7.4, the cells hyperpolarized ca. 5 mV as predicted if changes in the osmotic coefficient rather than in Cl content explained the osmotic behaviour.4. Regarding haemoglobin in concentrated solution as a solute with high osmotic coefficient is formally correct but is little help in understanding the properties of the solution. We have found it useful to consider separately haemoglobin and the rest of the contents of the cell. The haemoglobin then supports part of the total hydrostatic pressure on the cell leaving the crystalloid solution to experience a reduced fluid pressure. In greatly shrunken cells the contents act like a gel with the matrix of haemoglobin under compression and the fluid which fills the spaces within the matrix under tension.