The ability of cells to readjust their volume after swelling, a phenomenon known as regulatory volume decrease (RVD), is a fundamental biological achievement guaranteeing survival and function of cells under osmotic stress. This article reviews the mechanisms of RVD in mammalian cells with special emphasis on the activation of ion channels during RVD.

1 Stimulation of chemotaxis of human polymorphonuclear leucocytes (PMNs) with the chemoattractive peptide fMLP (N-formyl-Met-Leu-Phe) is paralleled by profound morphological and metabolic alterations like changes of intracellular pH (pH i ) and cell shape. The present study was performed to investigate the interrelation of cell volume (CV) regulatory ion transport, pH i and migration of fMLP stimulated PMNs. 2 Addition of fMLP to PMNs stimulated directed migration in Boyden chamber assays and was accompanied by rapid initial intracellular acidi®cation and cell swelling. 3 Inhibition of the Na + /H + exchanger suppressed fMLP stimulated cell migration, accelerated the intracellular acidi®cation and inhibited the fMLP-induced cell swelling. 4Step omission of extracellular Na + caused intracellular acidi®cation, which was accelerated by subsequent addition of gastric H + /K + ATPase inhibitor SCH 28080, or by omission of extracellular K + ions. In addition Na + removal caused cell swelling, which was further enhanced by fMLP. 5 H + /K + ATPase inhibitors omeprazole and SCH 28080 inhibited stimulated migration and blunted the fMLP-induced increase in CV. 6 Increasing extracellular osmolarity by addition of mannitol to the extracellular solution caused cell shrinkage followed by regulatory volume increase, partially due to activation of the Na + /H + exchanger. In fMLP-stimulated cells the CV increase was counteracted by simultaneous addition of mannitol. Under these conditions the fMLP stimulated migration was inhibited. 7 The antibacterial activity of PMNs was not modi®ed by Hoe 694 or omeprazole. 8 Western analysis with a monoclonal anti gastric H + /K + ATPase b-subunit antibody detected a glycosylated 35 kD core protein in lysates of mouse and human gastric mucosa as well as in human PMNs. 9 The results indicate that fMLP leads to cell swelling of PMNs due to activation of the Na + /H + exchanger and a K + -dependent H + -extruding mechanism, presumably an H + /K + ATPase. Inhibition of these ion transporters suppresses the increase in CV and precludes PMNs from stimulated migration.

Cell volume regulation is an essential feature of most cells. After swelling in hypotonic media, the simultaneous activation of potassium and chloride channels is believed to be the initial, time-determining step in cell volume regulation. The activation of both pathways is functionally linked and enables the cells to lose ions and water, subsequently leading to cell shrinkage and readjustment of the initial volume. NIH 3T3 fibroblasts efficiently regulate their volume after swelling and bear chloride channels that are activated by decreasing extracellular osmolarity. The chloride current elicited in these cells after swelling is reminiscent of the current found in oocytes expressing an outwardly rectifying chloride current termed ICln. Introduction of antisense oligodeoxynucleotides complementary to the first 30 nucleotides of the coding region of the ICln channel into NIH 3T3 fibroblasts suppresses the activation of the swelling-induced chloride current. The experiments directly demonstrate an unambiguous link between a volume-activated chloride current and a cloned protein involved in chloride transport.

The Na⁺/H⁺ exchangers (NHEs) are among the major ion transporters involved in cell volume regulation. NHE activation leads to a cellular influx of Na⁺ ions and extrusion of H⁺ ions, which are readily replenished from intracellular buffers. This will result in a net import of Na⁺. In many systems NHE operates in parallel to Cl^–-/ HCO3₃^– exchange, resulting in cellular uptake of NaCl. The influx of osmotically obliged water will consequently lead to cell swelling. This makes NHEs suitable to serve as powerful mechanisms for increasing cell volume (CV). The low volume threshold for NHE activation enables the cells to respond to very minute reductions of the CV. By the coupling to the export of H⁺ ions cell volume regulatory NHE activation may lead to changes in intracellular pH. On the other hand NHEs are activated by a broad variety of ligands and by intracellular acidosis, which, in turn, may consequently lead to cell swelling. In addition, NHEs are linked to other intracellular proteins and structures, like e.g. the cytoskeleton, which themelves are involved in the regulation of numerous cellular processes. Therefore NHEs link CV regulation to a diversity of cellular functions, both in physiological and pathophysiological conditions. Six isoforms of the Na⁺/H⁺ exchanger, termed NHE1 - 6, have been cloned so far. NHE 1 - 5 are located in the plasma membrane, whereas NHE6 is sorted to the mitochondrial membrane. NHE1 and NHE6 are the ubiquitously expressed isoforms. The expression of the isoforms NHE2 to NHE5 is restricted to specific tissues and the pattern of their expression, as well as their subcellular localization indicate that they fulfill specialized functions. Cell shrinkage induced activation has been shown for NHE1,2 and 4. In contrast, NHE3 is inhibited by cell shrinkage. In many cells several isoforms are present and assigned to specific membrane domains where they may serve a functional crosstalk between the different ion transporters.

Expression of the Ha-ras oncogene has been reported to stimulate the dimethylamiloride sensitive Na+/H+ exchanger and Na+,K+,2Cl- cotransport, both transport systems which are involved in cell volume regulation. The present study has been performed to test for an influence of ras oncogene expression on cell volume regulation in NIH 3T3 fibroblasts expressing the Ha-ras oncogene (+ ras). As controls served NIH 3T3 fibroblasts not expressing the ras oncogene (-ras). In isotonic extracellular fluid, the cell volume of + ras cells (2.70 +/- 0.08 pl) is significantly greater than the cell volume of -ras cells (2.04 +/- 0.10 pl). Both, + ras and -ras cells exhibit a regulatory cell volume increase in hypertonic extracellular fluid and a regulatory cell volume decrease in hypotonic extracellular fluid. The regulatory cell volume decrease is inhibited by 1 mmol/l quinidine and barium, the regulatory cell volume increase is inhibited in -ras and +ras cells by dimethyl-amiloride (100 mumol/l) and, only in +ras cells, by furosemide (100 mumol/l) and bumetanide (10 mumol/l). In conclusion, expression of the ras oncogene leads to a shift of the set point for cell volume regulation to greater cell volumes, which may contribute to the activation of the Na+/H+ exchanger and Na+,K+,2Cl- cotransport.