Diagram of the RBC transport systems that participate in Ca²⁺-induced dehydration. The top part of the figure shows the transporters of Ca²⁺, Cl^-, and H⁺ responsible for the net gain of CaCl₂ by RBCs when exposed to the ionophore A23187. With RBCs suspended in a Ca²⁺-containing plasmalike medium, addition of the ionophore A23187 triggers an electroneutral entry of Ca²⁺ in exchange for protons with a Ca²⁺:2H⁺ stoichiometry. With CO₂ concentrations at equilibrium across the RBC membrane, the anion exchanger (AE) and CO₂ shunt operate jointly like an electroneutral Cl^-:H⁺ cotransport,²³  known as the Jacob-Stewart mechanism (JS).^24-26  Together, the ionophore and JS mediate net CaCl₂ transport since the proton fluxes cancel out. Ca²⁺ transport is rate limited by the ionophore concentration because the high constitutive expression of the AE ensures nonlimiting speed in the co- and counter-ion transfers. The cytoplasmic Ca²⁺-buffering behavior of RBCs (B, CaB) may be approximated by [Ca²⁺]_i = α[Ca_T]_i, where α is approximately 0.3 over a wide range of [Ca²⁺]_i values and [Ca_T]_i is the total calcium content of the cells.^27,28  Thus, normal RBC [Ca_T]_i levels are hardly detectable, among the lowest of any cell in nature.^29-31  At sufficiently high ionophore concentrations (usually > 10 μM in 10% hematocrit RBC suspensions) and appropriate external Ca²⁺ concentrations ([Ca²⁺]_o > 100 μM), the induced Ca²⁺ influx exceeds the V_max of the PMCA in all the RBCs causing their [Ca²⁺]_i levels to approach equilibrium with [Ca²⁺]_o in a uniform manner.²⁰  The ionophore-induced equilibrium sets [Ca²⁺]_i /[Ca²⁺]_o = ([H⁺]_i/[H⁺]_o).^2,32  Addition of Co²⁺ in excess of [Ca²⁺]_o instantly blocks Ca²⁺ transport by the ionophore allowing the PMCA to extrude the induced Ca²⁺ load.^6,33  Co²⁺ is itself transported by the ionophore but at normal [Mg²⁺]_i levels it has no effect on PMCA-mediated Ca²⁺ fluxes.³⁴  The bottom part of the figure shows the transport systems that participate in the Ca²⁺-induced rapid dehydration response. Elevated [Ca²⁺]_i triggers the dehydration process by activating the Gardos channels. Gardos channel activation hyperpolarizes the cell (E, membrane potential) driving Cl^- out, resulting in the net loss of KCl and water. With full Gardos activation, dehydration is rate limited by the Cl^- permeability.³⁵  Replacement of approximately 10 mM Cl^- by the more permeable anion SCN^- maximizes the dehydration rate because rapid SCN^-:Cl^- exchange via AE, or SCN^- diffusion across the membrane (dashed line), continually replenish the intracellular SCN^- supply.¹⁸  Gardos-mediated dehydration has a very low temperature coefficient. With nonlimiting anion movement, maximal dehydration of RBCs may be attained within 10 to 30 minutes at 0°C to 4°C.¹⁸