407

Biochimica et Biophysica Acta, 1071 (I 9~)[ ) 407-427 © 1991 Elsevier Science Publishers B.V. All rights reserved 0304-4157/91/$03.50

B B A R E V 85394

Review

Activation of ion transport pathways by changes in cell volume Balfizs Sarkadi ~ and John C. Parker 2 l National Institute of Haematology and Blood Transfusion, Budapest (Hungary) and 2 Department of Medicine, School of Medicine UNC-Chapel Hill, ('h.apel Hill, NC (U.S.A.) (Received 23 April 1991)

Contents I.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

407

II.

Condu,:tive ion transport pathways . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

409 409 411

A. Swelling activated K + channels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . B. Swelling activated CI - channels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . II1. Coupled transporters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . A. Swelling-activated transporters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1. [K + - C I - ] cotransport . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. K * / H + exchange . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3. C a 2 ~ / N a ÷ exchange . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . B. Shrinkage activated transporters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . t. N a + / H + exchange . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. [ N a + - K + - 2 C I - ] cotransport . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

412 412 412 413 413 415 415 416

IV. Some possible special volume serJsor and transduction mechanisms . . . . . . . . . . . . . . . . . . . . . . . A. Ionic strength and osmolality . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . B. p H or m e m b r a n e potential . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . C. Magnesidm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . D. Cytoskeleton . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . E. M e m b r a n e stretc~ . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . F. Concentration and dilution of regulatory enzymes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . G. Macromr~lecular crowding . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

41 / 418 419 419 419 420 420 420

Physiological role of cell volume activated ion transport pathways . . . . . . . . . . . . . . . . . . . . . . . .

421

V.

VI. S u m m a r y and conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

422

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

423

I, " ,troduction Animal cells regulate their volume preciseb'. Water is in thermodynamic equilibrium across the plasma membrane, and because ~,~jtoplasm contains a higher concentration of impermeant, charged solutes than extracellular fluid, a driving force exists that pulls small

Correspondence: B. Sarkadi, National Institute of Haematology and Blood T,:ansfusion, 1113 Budapest, Daroczi u. 24, Hungary.

ions and water into the cell. Since the anim~,l cell membrane is easily distended and cannot sustain a hydrostatic pressure gradient, there are of necessity mechanisms which oppose the tendency of cells to swell. According to a model formulated in a discussion between Dean and Davson in 1940 [55] and developed by Tosteson and Hoffman in 1960 [291], cell volume maintenance depends on the balanced functioning of pumps~ fueled by metabolic energy, moving solutes uphill against their electrochemical potential, and leaks, through which downhill solute movements occur.

408 TABLE I

TABLE I!

Ion transport pathways activated during cell volume regulation a

Intracellular signals that can trigger volume-activated ion transport pathways

Mechanism activated

Examples of cells or tissues

Regulatory volume decrease Conductive K + transport Conductive CI- transport [K + - C I - ] cotmnsport N a + / C a 2+ exchange K + / H + exchange

Lymphocytes, Ehrlich ascites cells, epithelial cells, platelets, granulocytes, CHO cells, hepatocytes, etc. Red cells of most species Carnivore red cells Amphiuma red cells

Regulatory volume increase N a + / H + exchange [ N a + - E + - 2 C I - ] cotrans-

port

Lymphocytes, CHO cells, red cells of some species, epithelial cells Ehflich ascites cells, red cells of some species, epithelial cells

a For reviews see Refs. 13, 33, 40, 53, 58, 88, 98, 128, 129, 147, 152, 159, 161, 176. 177, 193, 196, 227, 265, 274.

In the years since the publication of the pump-leak model, the leaks have been found to be as specific, complex and subject to regulation as the pumps. Electrochemically downhill ion movements are mediated either by channels or by tightly cgupled cotransport and countertransport mechanisms, both of which are specific for the transported species. Conductive channels and coupled transporters respond to a variety of stimuli, including hormones, growth factors, changes in membrane potential, and alterations in cytoplasmic free calcium. Some of these leak pathways are activated by changes in cell volume and conduct solute movements that contribute to cell volume regulation. Most animal cells, when placed into anisotonic * media, initially swell or shrink like more or less perfect osmometers. In response to osmotic swelling, most cells extrude cytoplasmic solute and water, resulting in regulatory volume decrease (RVD), while osmotic shrinkage activates systems that draw solute and water inward, resulting in regulatory volume increase (RVI). This review focuses on the events triggered by a perturbation in the volume of eukaryotic ceils. How cells detect a volume change and how that information is communicated to ion transporters in the plasma membrane is discussed. An important subject which is not covered in detail is the role of nonelectrolytes in volume regulation (reviewed in Ref. 40). The best studied volume-regulatory ion transport pathways are listed in Table I and have been ably reviewe0. [13,33,40,53,58,88,98,128,129,147,152,161, 176,177,193,196,227,265,274]. Much information on volume regulation has come from experiments in which * An anisotonic medium is one in which cells shrink or swell, while an anisoosmotic medium simply has a higher or lower osmotic concentration than the physiological value fro" th~ $iveu o~ganism. Accordingly, an isoosmotic medium can be anisotonic (e.g., isoosmotic urea induces swelling and |ysis in red cells of most spec.:es).

Signaling system

Transporters affected

Rise in cellular Ca 2+ concentration

K + channels a C I - channels b N a + / H + exchange b C a 2 + / N a + exchange b K + / H + exchange a

Phosphoinositides and lipid r~etabolites ( + protein kinase C)

N a + / H + exchange b C a 2 + / N a y exchange b [Na + - K +--2CI - ] cotransport b C I - channels ~

cAMP ( + cAMP-dependent

[K + - C I - ] cotransport b [Na +-K + - 2 CI- ] cotran.~,~ort b N a + / H + exchange b K + channels b C I - channels b

kinase)

Tyrosine kinase Unspecified Mg 2+ and/or ATP requiring reactions, probably involving kinase action GTP-binding proteins

N a + / H + exchange b [K + - C I - I cotransport b [ N a + - K + - 2 CI- ] cotransl~rt b Na + / H + exchange b K + channels b N a + / H + exchange ~

a Indicates that the signaling system is probably triggered by a volume stimulus. b Indicates that the transporter can be activated or inhibited by this signal but there is no evidence that the signaling system is triggered by a volume stimulus. (For detaiis and references see text.)

cells were caused to swell or shrink in anisotonic media. This approach employed cell sizing and flux methods and provided important descril~tive data. Recently, attention has been devoted ~.~ the regulation of membrane transport systems. Some of the findings are summarized in Table II. Whereas there are a number of intracellular signaling systems capable of modulating ion transport, only a few of them have been shown to be induced by a change in cell volume. This review must therefore reflect the limited state of knowledge in the field of volume regulation. Much is known about the behavior of the transporters, something is known about the signaling systems that can influence them, little is known aboat the response of these second messenger pathways to cell ,-plume, and almost nothing is known about how cell volume is perceived. Studies on the transporter molecules themselves make it clear that a given mode of transport may be carried out by different gene p'roducts. For example, two separate N a + / H + exchangers, one on the apical and one on the basolateral surface, with different amiloride affinities, have recently been identified [31]. Therefore, the designations '[K+-CI -] cotransport', ' N a ÷ / H + exchange', 'K + channels' or 'CI + channels' used in this review should be regarded as physiological

409 entities. Whenever possible, we will discuss the transporters as molecules. The reader will find this review emphasizes work in red cells. We acknowledge this bias and, although attempting to provide a general overview, apologize to those who deal with other cell types for possible errors of commission and omission.

11, Conductive ion transport pathways The word 'channel' in this review is used to denote all conductive transport pathways through which ions flow in a chemically uncoupled fashion. Thus, Ca2+/Na + exchange, which is electrogenie because of the stoichiometry of its coupling, is not regarded as a channel. II-A. Swelling-activated K ÷ channels

In several cell types hypoosmotic swelling activates a conductive K + transport pathway. This mechanism has been studied in detail in Ehrlich ascites cells (reviewed in Refs. 128, 129), lymphocytes [58,98], and various epithelial cells [53,144,193,247,265,274,309]. It has been shown to be present in plate!ets [168,287], granu|ocytes [145], cultured cells such as the C l i o [257] and osteosarcoma [318] lines as well as in hepatocytes [118,130]. In most tissues the activation of a swellingindt, ced K + conductance occurs simultaneously with that of an independent, conductive Cl- transport (see section II-B). The outward movemer~t of KCI, driven by the K + gradient, results in osmotically obliged water efflux and thus a regulatory volume decrease (RVD). Loss of cellular KCI in response to cell swelling was first described and analyzed in frog skirl epithelial cells by MacRobbie and Ussing [178] and recent data indicate that in certain tissues bicarbonate accompanies K + efflux [171,297,298]. The swelling-induced K + transport pathway in all the tissues mentioned above is highly selective for K + (and Rb +) over Na + and other cations, carries net charge, and does not require the presence or the cotransport of specific anions. The opening of the swelling-induced K ÷ channel does not depend on the direction of the existing K + gradient or on the membrane potential. This was shown by the experiments in which, in the presence of a high external K + concentration (which also causes membrane depolarization), the K + (and CI-) channels opened by a hypoosmotic shock allow a rapid KCi influx and thus further cell sa,,eliing [95,126,127,255,257]. The K+-channel opening in lyrnphocytes seems to be a graded response, corresponding to the extent of cell swelling [256]. A close relationship appears to exist between swelling-activated K + channels and calcium-activated K + channels, origin-qEy described in metabolically depleted red cells by

Gfirdos [83]. This highly selective, K+-conductance pathway ~: activated by submicromolar concentrations of intracellular free calcium and has been described in both eukaryotic and prokaryotic cells (for reviews see Refs. 144, 203, 220, 228, 259, 262, 295). Although calcium-dependent K + transport can be evoked independently of cell volume changes, Grinstein et al. [93] showed that swelling-activated K + transport in lymphocytes depends on cellular calcium, and this was subsequently found to apply to most cell types. In most epithelial cells, swelling-induced K + transpnrt and thus RVD require the presence of extracellular calcium and a net calcium influx [54,119,193,245, 246,313]. In lymphocytes, Ehrlich ascites and CHO ceil% on the other hand, external calcium is not required, but prolonged cellular calcium depletion eliminates the activation of K + channels by cell swelling [95,98,125,257]. Swelling-induced increases in cytoplasmic free calcium concentration have been shown to occur in various epithelial cell types [119,226,2452246, 313,314], and buffering of cytoplasmic calcium with chelators eliminated RVD [246]. In tracheal epithelial cells the swelling-induced release of calcium from internal stores was thought to promote volume regulatory K + fluxes [252]. The importance of calcium as a trigger for the activation of swelling-induced K + channels has not been appreciated in some studies with lymphocytes. For example, no swelling-induced calcium changes wet,: recorded in human peripheral blood lymphocytes employing the fluorescent dye Quin-2 as a reporter of cytosolic ionized calcium levels [237]. Moreover, Grinstein and Smith [106] reported swelling-induced activation of K + channels in human peripheral blood and rat thymic lymphocytes at normal or even subnormal cytoplasmic free calcium levels. On the other hand, in mouse thymocytes, hypoosmotic shock generaied a calcium signal and a consequent K + effiux [244]. Recent experiments in which the sensitive calcium indicator lndo-1 was used in human (Jurkat) lymphoblasts, showed a significant in~[ease in cytoplasmic calcium during hypoosmotic exposure and this calcium signal correlated with the activation of RVD (Tordai and Sarkadi, unpublished results). Further similarities between swelling-induced and calcium-induced K + channels include the actions of various drugs and toxins (Table liD. Quinine and other organic cations ha, e strong inhibitory effects in both cases. The mediatir~g role of calmodulin in both pathways is suggested by the effects of anti-ealmodulin drugs, although these compounds may have several non-specific effects on membrane functions. In addition, both swelling-induced and calcium-induced K + movements are significantly inhibited by intracellular acidification [51,58,98,144,275]. Thus, the observations cited above support the notion that the swelling-

410 TABLE llI Inhibitors of calcium-activated and swelling-activated K + channels, respectively Inhibitor

Calcium activated K + channels (references)

Swelling activated K + channels (references)

Quinine, quinidine Cetiedil Cationic dyes (e.g. diSC3 (5)) Calmodulin antagonists SH reagents Oligomycin (A) Baz +, La a * Charibdotoxin (CTX)

la 2a

lb 2b

3a 4a 5a 6a 7a 8a

3b 4b 5b 6h 7b 8b

la: (9, 232); lb: (94, 95, 126, 171); 2a: (15); 2h: (258); 3a: (268, 269); 3h: (258); 4a: (6, 148, 202, 284); 4b: (93-95, 127, 132, 258); 5a: (84); 5b: (258); 6a: (21,238): 6b: (258); 7a: (285, 316), To: (155, 305); 8a: (39, 108, 188, 312); 8b: (106).

activated and the calcium-activated K + channels are, if not identical, at least closely related. The responses of cells to a variety of stimuli are mediated by a sequence of events in which ligands bind to receptors, phospholipases are activat~d, inositol phospholipid derivatives are released, intracellular calcium is mobilized, and protein kinase C is activated by diacylglycerol [16]. In Ehrlich ascit~s cells, osmotic swelling was found to induce phosphoinositide breakdown and IPa liberation, and this was suggested to be causally related to calcium-dependent RVD [44]. Clearly, further studies are required to establish this mechanism in volume regulation. Activation of calcium-activated K + channels has been thought to be modulated by cAMP-dependent protein kinase [57,1'42,143,242,307] and G-proteins [19,290], but no detai!ed studies are yet available concerning the relationship of these observations to volume-dependent K + ti-ansport. The hypoosmotically-induced Rb + efflux in rabbit kidney epithelial cells was abolished after pertussis toxin preincubation, suggesting G protein involvement in this pathway [163]. A dependence of swelling-activated K + transport on cell and membrane structural components has also been suggested. Cytochalasin B and related drugs, which disrupt cytoskeletal elements, inhibited RVD in several epithelial cell types [72]. Further evidence that cell integrity is important for the activation of K + channels by cell swelling can be found in the report of Gfinstein ct al. [96] that although plasma membrane vesicles of volume-regulating thymocytes conducted ~welling-induced cation movements, there was no selectivity for K +, and neither calcium nor specific inhibitors showed the effects observed in intact cells. These observations are not surprising in view of the findings that membrane isolation and mild proteolysis

abolish the K ÷ selectivity and the calcium sensitivity of the K + channels in erythrocytes [286,314,315], and limited proteolysis removes the calcium dependence of isolated renal epithelium calcium-activated K + channels [142,143]. In the past few years considerable effort has been devoted to the isolation and reconstitution of swellingactivated and calcium-activated membrane K + channels. An affinity chromatography method, using quinine and furosemide as ligands, yielded a complex of membrane proteins which showed K + and Cl- channel activity [41]. The calcium-activated K + channel from kidney epithelial cells was isolated on a calmodulin affinity column [142], and was shown to be directly regulated by calmodulin and cAMP-dependent kinase [143]. This protein is a pH- and trypsin-sensitive hetero-dimer which closely corresponds to the calciumactivated 'maxi' K + channel (see below). The. role of this protein in volume-activated K + transport is at present unclear. Another promising technique for the isolation and characterization of swelling-activated K + channels may be the use of specific toxins for affinity purification [87,192]. Recent studies on the molecular biology of various K + channels indicate that they share a structurally similar region that constitutes the ion-selective, singlefile pore [115,280,321,322]. Moreover, in Drosophila the coding region for a pore-like structural component of the calcium-activated K + channel shows a high degree of homology to the pore domain of voltage-gated K + channels [241]. For a recent compilation of the moieodar models describing potassium channel gating and selectivity see Ref. 189. There is a large body of data concerning the biophysical characteristics of Ca2+-dependent K + channels. They are categorized according to their conductivity ('mini' and 'maxi' channels, with conductivities around 10-40 pS and 100-300 pS, respectively), drug sensitivity, voltage and internal cation dependence, as well as modulation by cAMP or G proteins [19,154, 276,296]. Unfortunately, relatively little information is available regarding the relationship between these K + channels and those activated by cell swelling. According to patch clamp studies, cell swelling appears to induce stretch-activated channels which in several cases were found to be non-selective (see below). Work by Sackin [250,251] indicates that in Nectutus kidney proximal tubules, cae+-insensitive, K +selective channels are directly opened by membrane stretch, and in a colon carcinoma (T84) cell line the calcium-independent, swelling-induced opening of a 20 pS K + channel was observed [288]. Also, stretchactivated, calcium-independent, selective K + channels were detected in auditory hair cells [167]. However, in most cases an indirect activation of K + channels during cell swelling has been suggested to occur via a

411 mechanism in which cell swelling stretches the membrane and opens Ca 2+ channels, which allow a calcium influx. The increased intracellular free calcium then activates K + channels [43,195,289,293]. The type of K ÷ channel ('maxi'/'mini' conductance, voltage dependence, etc.) activated under these conditions depends on the tissue investigated [193]. A similar, indirect mechanism was proposed with regard to the opening of swelling-induced, calcium-dependent K + channels in turgor-regulating plant cells [203].

II-B. Swelling activated Cl- channels In the RVD reaction, swelling-induced opening of selective K + channels frequently entails the independent activation of Ci- channels, allowing a rapid loss of KC! and osmotically obliged water. The induction of a conductive CI- transport under hypoosmotic conditions was first suggested in epithelial cells [178] and Ehrlich ascites cells [124]. Using specific inhibitors of swelling-activated K + transport, such as quinine or quinidine, it was demonstrated in lymphocytes that the opening of swelling-activated chloride channels occurs independently of the activation of K + channels. Tracer Cl- fluxes were increased either in the presence or absence of K + movement and when the ionophores valinomycin or gramicidin were added to bypass the quinine-blocked K + transport, volume changes were entirely dependent on the activation of Cl- conductance [93,95-97,255]. B-lymphocytes have rio native tendency to perform RVD, but it can nevertheless be shown in these cells that chloride channels are opened by cell swelling, aad if an artificial K + transport pathway is provided, RVD occurs [42,97]. The opening of the swelling-activated chloride channels showed a distinct threshold at a certain degree of cell swelling; it was an all-or-none response and a time-dependent closing of the CI- channels was observed, even when cell volume was held constantly above the activation threshold [256]. The independence of swelling-activated K + and Cl- channels was confirmed and extended in studies with Ehrlich ascites cells, where the time dependence of the closing reaction was also observed [125-127]. The interpretation af the data in Ehrlich ascites cells is somewhat compli~:ated by the simultaneous activation of electroneutral transport systems during cell swelling, as noted below. During the RVD reaction ;n lymphocytes, Ehrlich ascites, or CHO cells, the membrane permeability for Cl-, which is relatively low in the resting state, significantly surpasses K + conductance, and a membrane depolarization occurs [93,95,129,256,257]. The fue¢tioning of swelling.induced Cl- channels itself does not seem to be voltage-dependent [93A26,127,132,255, 256]. Swelling-activated chloride channels can trans-

port other small, monovalent anions, but not large organic anions such as gluconate [98,129]. A major question concerns the role of intracellular calcium in the activation of the volume-dependent Clchannels. In lymphocytes an increase in cellular C a 2+ mediated by the ionophore A23187 stimulated Cltransport [93], but to a much lesser degree than hypoosmotic shock [255,256]. Cellular calcium depletion, which abolished the opening of K + channels, did not inhibit the swelling-activated chloride channels [256]. On the other hand, Ehrlich ascites cells demonstrated a pronounced calcium dependence of swelling-activated chloride transport. Significant cell shrinkage was evoked by the calcium ionophore A23187; moreover, the time-dependent closing of swelling-activated chloride channels was abolished when cellular calcium was maintained at a high level [127]. Still, in these cells the effect of high cytoplasmic-free calcium on K + channels was greater than on Cl- channels, while the opposite was true with swelling, which caused a greater change in Cl- than K + conductance [151]. Calcium-activated Cl- channels are present in several other cell types [75] but their involvement in volume regulatory ion transport is questionable (see below). It has been suggested recently that the role of calcium in the activation of Cl- transport is an indirect one [20,150,151]. A Ca2+-dependent activation of leukotriene synthesis is postulated to be responsible for the activation of O - channels. Leukotriene D4 was shown to directly activate Ci- channels while inhibitors of leukotriene synthesis prevented swelling-induced Cl- transport [150]. The molecular nature of the Cl- channel(s) opened by cell swelling is still unknown. The pathway is inhibited by several drugs but usually at relatively high concentrations and in a non-selective manner. In iymphocytes, blockers of the red cell anion exchange pathway, SITS and DIDS, inhibited swelling-induced chloride transport, but only after preincubation at high concentrations (0.5-1 mM) and then incompletely [258]. Ehdich ascites cells were found to be utterly insensitive to the above inhibitors, but swelling-activated chloride channels in these cells were effectively blocked by diphenylamine carboxylate (DPC) and indacrinone (1), which inhibit some epithelial Cl- channels. In lymphocytes oligomycin C, oleic acid, and most calmodulin antagonists were found to inhibit sweUing-activated chloride transport [258]. These inhibitor studies offer little information as to the molecular nature of the channels. In patch clamp studies several anion transport pathways have been characterized [75,320]..These include volume-dependent, calcihm-activated, and various kinasa-activated chloride channels that coexist and have quite distinct features. Work in this area has been stimulated by recent discoveries regarding the genetic

412 basis of cystic fibrosis (CF), a disease in which there is a defect in the activation of cAMP-dependent Cichannels critical for fluid secretion in exocrine glands and airway epithelia [77,306]. Since the gene, its protein product (called CFTR), and the mutation(s) in the gene responsible for cystic fibrosis have been discovered [239,243] a vigorous effort has been underway to clarify the molecular mechanism of the disease. Recent data indicate that the expressed CFTR protein forms or activates DIDS-insensitive CI- channels which have a conductivity between 5 and 10 pS, have no rectifying properties, carry B r - > CI-> I- in this order of selectivity and are stimulated by cAMP [8,139,288]. Patch c~amp studies using both whole cell and excized patch recordings in iymphocytes (P. Gardner, personal communication) and epithelial cells [81,107,317] indicate that the volume-dependent CI- channels are different from both the cAMP activated and the Ca2++ calmodulin-activated CI- channels. The swellingactivated chloride channels are more DIDS-sensitive and have strong outward rectification properties with a conductivity in the range of 30-50 pS. Their anion selectivity order is I - > B r - > CI-, and they are rapidly inactivated by depolarizing pulses. Just how these channels are triggered by cell swelling, however, remains an open question.

Ill. Coupled transporters IliA. Swelling-activated transporters III-A.1. [K +-Cl-] cotransport [K+-C! -] cotransport is activated by an increase in volume of the red cells of many species, including fish, duck, sheep, rabbit, pig, dog, and human, plus some non-erythroid cell types including Ehrlich ascites cells [129], Necturus gallbladder epithelium [233], and kidney tubule [92]. The transport of K + and CI- through this route is rigorously selective for these two ions, electroneutral [28], ki~eticaUy asymmetrical [56,137], and subject to weak inhibition by loop diuretics [66,157], certaip stilbene disulfonates [158,173], and in low-K + sheep red cells by an antibody to the L antigen [66]. There is evidence that the regulation of this transporter by volume involves protein phosphorylation. The most compelling experiments are in rabbit red cells where an analysis of the "kinetics of activation and deactivation of the transporter led Jennings and AIRohil [134] to formulate a simple, unimolecular kinetic model: (phmphatase) Ki2 Resting state

~............ '

Active state

K21 (kinase - inhibited by swelling)

in which the equilibrium between resting and active

transporters is described by two rate constants, Kiz and K21. There was a 5-10 min delay in the activation of [K+-CI -] cotransport after cells were abruptly swollen, but there was hardly any delay in the deactivation of the pathway when swollen cells were returned to their normal volume. The activation delay was further prolonged by phosphatase inhibitors such as fluoride, vanadate, and okadaic acid [135]. The authors conclude that a phosphatase mediates the reaction associated with the turning on of the [K+-Ci -] cotransporter by swelling (KI2). The catalyst of the deactivation reaction (K2I) is a kinase. From the relative rates of activation/deactivation it was inferred that swe!iing causes a decrease in the activity of the kinase (K2~) rather than an increase in the activity of the phosphatase (K~2). Entirely similar results were obtained in experiments with dog red cells [222]. The model of Jennings and his colleagues [134,135] for regulation of [K+-Ci -] cotransport is consistent with several previous observations on the system, ineluding the gtimulation of [K+-CI -] cotlansport seen with ATP depletion of intact cells [65] and resealed ghosts [62]. In addition, the ~tlmulatory influences on [K+-CI -] cotransport of Mg 2+ depletion [160,221], and N-ethyl maleimide [156] might bc e"plained in terms of inhibition of the kinase (K21 in the model above) that returns the active transporter to its resting state. In apparent contradiction to this conclusion are the experiments of Sachs [249] and Kracke and Dunham [146] in which volume-activation of [K+-CI -] cotransport appears to require ATP. These viewpoints might be compatible if the mechanism for activation of [I~+CI-] eotransport were like some hormonal systems in having kinases/phosphatases that themselves are substrates subject to activation or deactivation by phosphorylation and/or dephosphorylation [46,261,300]. Although cyclic AMP can itself activate [K+-CI -] cotransport in the red cells of pigs [141] and rabbits [135], Jennings and Schulz [135] determined that neither protein kinase A nor C were likely mediators of the transporter's response to cell swelling. The mechanisms by which volume stimuli are transduced hzve been studied by several authors using [K +CI-] cotransport as an indication of response. Dunham and Logue [62] and Sachs [249] report that resealed ghosts, in which normal cell contents have been greatly diluted, continue to respond to osmotically-induced swellir~g with an increase in [K+-CI -] cotransport. This finding implies that the swelling stimulus becomes transduced in relation to a change in membrane shape, as from a flat disk to a plump disk, and that the dilution of cell contents is not part of the physiological message. The finding of Kracke and Dunham [146] that inside-out vesicles from sheep red eel.Is show an increased [K+-CI" ] cotransport with osmotic swelling is

413 particularly interesting in this regard, because such vesicles are likely to be deficient in many of the extrinsic proteins of the membrane skeleton [279], and the swelling of such vesicles might be expected to affect the radius of curvature of the membrane in a fashion just opposite to the swelling of intact cells or right,.sideout membrane preparations. Against any role for membrane shape or curvature, on the other hand, are the reports of Smith and Lauf [270] and Brugnara et ai. [27] ,'hat resealed ghosts show no swelling-induced K + transport and the observation of Jennings and Schulz [133] that changing red cell shape with various chemical agents has no effect on the volume response of K + flux. There is evidence that the link between cell swelling and [K+-C! -] cotransport may be influenced by the nature, and perhaps the concentration, of impermeant cell contents. Red cells from patients with hemoglobins S and C show an increased activity of [K+-CI -] cotransport relative to hemoglobin A cells at any given volume [30]. To some ex~.:,nt this difference may be related to the lower mean cell age of the red cell population in the hemoglobinopathic patients, because young red cells from people with homozygous hemoglobin A have an increased activity of [ K + - C I 1 cotransport relative to older ceils. But the sensitivity to cell swelling of red cells containing hemoglobins S and C is more than can be explained by a hemolytic state. Hemoglobin A has a glutamate on position 6 of the beta-globin chain. In hemoglobin S this position is filled by a valine, and in hemoglobin C by a lysine. Thus, the two mutant hemoglobins that affect [K+-Cl-] cotra~v,port are more positively charged than hemogIJbin A, and there is some evidence that their binding properties to the red cell membrane differ from those of hemoglobin A [26,29,30]. Another feature of red cells containing hemoglobins S and C that differentiates ~hem from hemoglobin A-containing cells is that they show activation of [K+-CI-] cotransport with mild acidification, an effect that cannot be explained by acid-induced cell swelling [30]. This property of sickle cells may be central to the pathogenesis of sickle cell crises and hemolytic anemia, because when the red cells encounter a moderately acid region of the circulation, e.g., in the spleen or the p!acenta, the [K+-C! -] cotransporter becomes stimulated and the cells lose salt and water, becoming rigid and unable to pass through small capillaries. It is not necessary to deoxygenate SS or CC cells to observe these changes [30]. Thus, the emphasis in the literature on deoxygenationinduced red cell transport may be irrelevant to the situation in vivo [22,26,29,30,166]. How these differences in globin structure influence membrane transport is not dear, but the effect of oku~aic acid in reducing the response of [ K " - C I - ] cotransport to swelling or to acidification in red cells with CC or SC

hemoglobins [204] suggests that the mutant hemoglobins may influence the equilibrium of a phosphatasekinase system like the one proposed by Jennings et al. [134,135] discussed above. III-A.2. K +/ H + exchange

As far as we are aware, the giant red cell of Araphiurea furnishes the only reported example of swellinginduced K + / H + exchange [32,34,35]. Reduction of the tonicity of the medium bathing these cells activates a coupled, electroneutral effiux of K + ions in exchange for protons. As the pH of the cytoplasm falls, an inward chemical gradient for bicarbonate develops. Bicarbonate enter,; the cell, and chloride leaves, both ions being shuttled by the band 3-mediated anion exchanger. The net result is an effiux of K ÷, chloride, and water and a restoration towards normal ce~l volume. There is evidence for coupling between K + / H ÷ exchange and N a + / H + exchange in these cells, suggesting that the same protein nmy carry out '-out.L types of transport, the former in response to swelling and the latter in response to shrinkage [34]. Activation of the K + / H + exchange mechanism can be accomplished by raising cytosolic free-calcium [33], and swelling of Amphiuma red cells causes a rise in cytosolic-free calcium, as determined by a null point method [35]. The affinity of the K + / H + exchanger for calcium was found to be increased by swelling the cells. Calmodulin antagonists inhibited the activation of K / H + exchange by cell swelling. PhorboI esters activated K + / H + exchange. Taken together, the information suggests a control mechanism for swelling-induced K + / H + exchange that is dependent on free calcium and protein kinase C and possibly involves a volume-induced change in specificity wherein a single transporter can conduct swelling-inducc,d K + / H + exchange and shrinkage-induced N a + / H + exchange [34,35]. An alternative explanation offered for the above experimental data by Lew and Bookehin [165] and disciassed by Sarkadi and Gfirdos [260] is the activation of a Ca2+-dependent K + channel by cell swelling and the presence of a relatively high H + conductance in the merr~brane of these cells. In this model the increased negativity of the membrane potential evoke0 by the swelling-induced K + conductance give.~ :[se ~.o a countertransport of protons, p~oducing the observed phenomenon of an apparent K + / H + exchange. III-A.3. Ca 2 +/Na + exchange

Red cells ~3f dogs [269], cats [209], ferrets [1871, and bears [310,311] all carnivores - begin their life in the marrow like other ceils, wJt~, a high potassium content and a N a + / K ~ pump. But when carnivore red cells ripen past the reticulocyte stage, the N a * / K + pump ceases to function [179]. As a consequence, carnivore =d cells in their maturity have ,:3"toplasmic concentra-

414 tions of Na t and K + that are not greatly different from plasma. Potassium cannot serve a volume-regulatory function in these cells because its transmembrahe gradient is too small. Although swelling-activated [K+-CI -] cotransport can be shown in some carnivore red cells [221,311], its physiological role is unclear. To protect themselves against swelling, carnivore red cells extrude Na t , their major internal cation, by performing Ca2+/Na + exchange in what has come to be called the 'reverse' mode [60]. Calcium enters from the plasma, down its electrochemical gradient, in exchange for Na t , which is extruded from the cell against its gradient; the Ca 2+ is the~ pumped out via the ATP-dependent Ca 2+ pump [4,25,78,187,206-208,210, 310,311]. When assayed by Ca 2t influx or Na + efflux measurements, Ca2+/Na + exchange in dog red cells is activated by cell swelling [210], although when measured as Ca 2t eftlux, Ca2t-Na t exchange is reported not to be volume-dependent [4]. Ca2+/Na + exchange, first described in heart muscle and aerve [12,235], is found in many cell types including photorecevtors, smooth muscle, epithelia, sperm,

TABLE IV Some modulators of Ca2+/Na + exchange Modulator

Effect

Tissue or cell

Reference

C~osolic Ca2+

Stimulation of Ca2+ influx Stimulation (deregulation) Stimulation of Ca2+ influx Increase of affinity for Cai2+ and Nap+ Stimulation Stimulation Stimulation Inhibition

Heart Nerve Heart

120 60, 50 120

Heart

120

Heart

230

Heart Heart Heart Heart

225 225 225 37

Stimulation Blocks adenosine stimulation Stimulation

Heart Heart

24 24

Heart RBC RBC

47, 140, 231 216 217 140 271

Stimulation

Heart Smooth muscle Smooth muscle RBC

Stimulation

Nerve

59

Cytosolic chymotrypsin C~osolic MgZ+-ATP

Phospholipases Phosphatidic acid Proteases Calmodulin inhibitors Adenosine Pertussis toxin Oxidants Hemolysisr:s,~aling Insulin Angiotensin11 External Mg2+ Replacement of chloride

Stimulation Stimulation Stimulates Ca 2+ influx Inhibition

271 212

by SCNVanadate

and platelets [2,138]. As far as we are aware, the only red cells that are capable of Ca2+/Na + exchange are those of carnivores and perhaps a mouse erythroleukemia cell [272]. It is interesting that in a rare breed of dogs whose mature red cells retain the N a + / K + pump and have a high K + content [179], the Ca2+/Na + exchanger is still present [78]. Unlike other coupled transporters discussed in this review, the Ca2+/Na + exchanger.conducts a current. There are several likely isoforms of the exchanger [236]. One isoform in heart sarcolemma has a stoichiometry of I C a 2 t for 3 Na t ions (Ca2+/3 Na+). In retinal rod outer segments, however, the transporter may function as a calcium-potassium cotransporter/ sodium exchanger ([K+-Ca2+]/4 Na t ) carrying one positive charge in a direction opposite to the movement of one calcium [149]. There is a long list of weak inhibitors of C a 2 t / N a t exchange that includes quinidine [210], anthracyclines [114], harmaline, amiloride analogues, dichlorobenzamil, some calcium-entry blockers, and some local anesthetics [116,138,229]. Amrinone, a cardiotonic drug, stimulates C a 2 t / N a t exchange in dog red cells [211]. None of the compounds investigated to date has either the specificity or the affinity to be useful its a selective Ca2+/Na t exchange reagent. Complementary DNA for one isofo~m of the Ca 2 F/Na t exchanger has been cloned from cardiac sarcolemma. The transporter has a calculated l~olypeptide molecular mass of 108 kDa, but with attached carbohydrates its molecular mass is nearer to 160 kDa [199]. There are many modulators of Ca2+/Na t exchange (Table IV). The amino acid sequence of the exchanger suggests several potential modulation sites, including a cationic region that is likely to be a calmodulin-binding domain and a serine residue on the cytoplasmic portion of the protein that might be a phosphorylation site [199]. Certain unphysiologic stimuli cause a substantial increase in the activity of the transporter. These include the addition of proteolytie enzymes to the cytoplasmic face of the membrane, exposure to strong oxidizing agents, treatment with phospholipases, replacing chloride with thiocyanate, and hemolysis and resealing [120,212,2!6,217,225,231]. Apparently, cells have the capability to exchange Ca 2+ for Na + at a much faster rate than is seen under normal conditions, and therefore the system must be under tight regulation. The exchanger can operate either as a Ca 2+ effiux pathway or in a mode that promotes Na + extrusion and Ca 2+ entry. Among the influences that govern the direction in which it transports are the internal calcium concentration. Paradoxically, internal C a 2+ is required for the transporter to function as a calcium influx-sodium extrusion pathway, and this mode of

415 operation also appears to require ATP [60,6L120]. Further evidence for a modulatory role of internal calcium has recently been published by Niggli and Lederer [200]. Although a role for protein phosphorylation in the regulation of Ca2+/Na + exchange has been proposed [37], and although vanadate, a protein phosphatase inhibitor, appears to stimulate Na÷-de pendent Ca 2+ efflux [59] no direct evidence for such modulation is known to us. Thece is general agreement that Ca2+/Na + exchange is responsible for Na + extrusion and volume regulation in carnivore red cells (see above), but it is not universally observed that cell swelling turns on Ca2+/Na + exchange. In dog red cells there is a clearcut stimulation of Ca 2+ influx via the Ca2+/Na + exchanger with cell swelling [210,217], but the transporter in ferret red cells show only a modest degree of swelling activation [187]. When measured as Ca 2+ efflux dependent on external Na + there is reportedly no volume effect in dog red cells [4]. An explanation for this discrepancy may be that in the experiments failing to show swelling-induced activation of C a / N a exchange the cells were depleted of ATP a n d / o r Mg 2+, or they were treated with vanadate-all measures designed to inhibit the Ca 2÷ pump [4,187]. If activation of Ca/Na exchange by cell swelling involves a kinase/phosphatase system, then metabolic perturbations of the sort mentioned might well obscure the normal physiologic process.

III-B. Shrinkage-activatea transporters III-B. 1. Na + / H + exchange An electrically silent, 1:1 N a ÷ / H + exchange is found in many cell types, including epithelia, excitable tissue, germ cells, immunocytes, and red cells. The transporter is activated by a variety of stimuli, including hormones, growth factors, thrombin, and fertdization (see Ref. 100). Amiloride and its analogues are among the most potent inhibitors of N a + / H + exchange, but amiloride itself can inhibit other processes in the cell [17,105]. A eDNA for at least one isoform of the transporter has been cloned and sequenced [253]. The transport protein has a glycosylated M r of 110 kDa with an estimated 10 transmembrane segments [254]. It is likely that there are multiple, genetically distinct N a + / H ~ exchangers with different amiloride sensitivities, subcellular localization, and regulatory responses [31]. Activation of N a + / H + exchange by cell shrinkage was first shown by Siebens and Kregenow [264,266] and Cala [32] in Amphiuma red cells, and has been found subsequently in other species of red cells and other cell types (see Refs. 67, 100). In lymphocytes, Ehrlich ascites cells and in some epithelia a preliminary swelling must be imposed on the cells before shrinkage-activate,J

7.5

A

0 ~

~'O

c E

5.0

i

| x

®.

\

a

-¢,

Oa

6.0

6~

A

!

|

u

I

I

I

6.4

8.6

6.8

?.0

7.2

7.4

Fig. 1. Effect of cell shrinkage on the activation of H + efflux through the N a + / H + exchanger in rat thymic lymphocytes. The initial rate of proton efflux is plotted as a function of cytoplasmic pH. Cells suspended in isotonic media (285 mOsm, solid symbols) show a steep dependence of efflux rate with falling internal pH, but at physiological pH (7.1) the efflux rate is zero. Wh,en the same assay is performed in shrunken cells (550 mOsm, open symbols) the whole curve is shifted to the right so that at physiological pH the N a + / H + exchanger is activated. (Reproduced from Ref. 99, with the permission of the authors and the publishers.)

N a + / H + exchange can be observed [96,97,129,193]. In conjanction with chloride-bicarbonate exchange, N a + / H + exchange mediates the entry of sodium, chloride, and osmotically obliged water into shrunken cells, thereby adjusting their volume upward towards normal [361. Among the most important intracellular inducers of N a + / H + exchange is cytosolic acidification [10], which has an effect that transcends the role of protons as transported substrates and has led to the notion that there is a proton modifier site on the transporter [100]. Many of the stimuli that activate N a + / H + exchange exert their influence via an increase in the affinity of the cytoplasmic face of the transporter for protons, so that the mechanism functions at intracellular pH values that in the unstimulated cell would be above the threshold for activation [100]. This is true of the volume stimulus: shrinking the cell shifts the pH activation curve as shown in Fig. 1 [99]. N a + / H + exchange is subject to stimulation by a number of well-recognized second messengers, including cyclic AMP [23,198], analogues of diacyiglycerol [101,102], and cytosolic free calcium, although the latter effect may be an indirect one, secondary to a calcium-induced loss of potassium and therefore of cell volume [103]. Extracellular agents that bind to membrane recept¢rs can turn on N a + / H + exchange without the mediation or a2y recognized second-messenger pathway. For example, in ~:,~me tissues stimulation of the transporter by b,~ta-adrenergie agents can occur

416 without measurable changes in cAMP and without any effect of agents known to inhibit GTP-binding regulatory proteins [80]. In platelets, some evidence for G protein involvement in the downqegulation of Na + / H + exchange was surmised oi~ the basis of fluoride effects [267]. There is little evidence that the above-mentioned signaling systems, all of which can modulate N a + / H + exchange, are involved in the activation of this transporter by cell shrinkage, although a recent report claims that shrinkage activation of the N a + / H + exchanger in perfused barnacle muscle involves G proteins and can be simulated by cholera toxin [52]. Several observations suggest that protein phosphorylation might be an intermediate event in the activation of Na+/H ~ exchange. ATP depletion of cultured human epithelial cells decreases the affinity of the N a + / H + exchanger for internal protons, and this effect is reversible [38]. Weinman et al. [304] present evidence that N a + / H + exchange can be inhibited or down-regulated by cAMP- :luced phosphorylation of a putative regulatory polypeptide of 42 kDa extracted from rabbit kidney brush border membranes. Recently Sardet et al. [254], transfected hamster lung fibroblasts (which lack N a + / H + exchange) with cDNA from the human Na+/H + exchanger and demonstrated that when the transfectant cells were treated with epidermal growth factor, thrombin, phorbol ester, or fetal bovine serum, the transporter itself became phosphorjlated over a time course that paralleled its physiological activation as monitored by alkalinization of the cytoplasm. Whether the phosphorylation caused the activation of N a + / H + exchange or was a consequence of the cytoplasmic pH change was not considered. Information about volume-dependent activation of N a + / H + exchange remains meagre. Grinstein et al. [101] found in lymphocytes that osmotic shrinkage was associated with phosphorylation of a 60 kD membrane polypeptide [100,101,103], although it is not clear whether this is related to the activation of N a + / H + exchange. Okadaic acid, .a serine-threonine protein phosphatase inhibitor that readily gets into cells, was recently found to increase the sensitivity of the dog red cell Na+/H + exchanger to shrinkage, so that it became functional even in slightly swollen cells. The agent also slowed the rate at which the N a + / H + exchanger turned off following abrupt cell swelling [18,222].

III-B.2. [Na +-K ÷-2 Cl -] cotransport Coupled, electroneutral mow,ment of Na +, K +, and CI- occurs in nearly all tissue types studied, including epithelia, excitable tissues, and red cells [86,113]. Much of the literature about volume regulation in Ehrlich ascites cells refers to a shrinkage-activated [Ha+-Cl -] cotransporter, but r,~cent 'sork suggests that this important pathway is in r¢ality a [Na+-K+-2CI -] cotransporter [136,164]. A!though there has been some

LN

OUT

I N°°

,

Eo

N'a~!o Cl NO Eo

Ei

Cli

C|-Ei

IF C|'K[i

Ki

Ch KCl No Eo

CI K CI NO'Eo

CI.KCl F i

Ir--

Nai

CIKCINaEi

Fig. 2. A glide-symmetry model for the [Na+-K+-2 C I - ] cotransport, as proposed by Lytle and McManus [172] for the functioning of the transporter in duck red cells. The sequential binding and release of the transported ions and the exclusive transmembrane movement of the empty and fully loaded transporters explains the experimental

findings for the net and exchangemovementsof ions, respectively. (Reproduced from Ref. 220, with the permissionof the publishers.) dispute about its stoichiometry, it is now agreed that one full turnover results in the net translocation of one Na ÷, one K +, and 2 Cl- ions, as originally proposed by Geck et al. [86]. Disagreements about the stoichiometry were resolved when it was realized that the system could conduct Na*-Na + and K+-K + exchange but that no net transport could occur without all four ions crossing the membrane. An elegant glide symmetry model has been proposed to account for the data on net and exchange movements of ions via this pathway (Fig. 2, Refs. 63, 131, !72, 185, 191). The [Na÷-K+-2C! -] cotransporter is inhibited by diuretics such as furosemide and bumetanide. A derivative of the latter drug has been used to identify and purify the transport protein from a number of tissues. There is now general agreement that it is a 150-200 kDa glycoprotein [113]. A eDNA for a 200 kDa isoform of the transporter has recently been clotted from shark rectal gland (Lytle and Forbush, personal communication, with permission). The [Na+-K+-2C! - ] cotransporter is subject to regulation by many influences including beta-adrenergie catecholamines, atrial naturetic factor, thrombin, epidermal growth factor, vasopressin, and insulin. In some tissues these agents are stimulatory and in others they inhibit transport. In duck red cells, up-regulation of [Na+-K+-2CI - ] cotransport in response to a variety of stimuli correlates with an increase in b:~metanide bindi~g sites on the outer cell surface [112], suggesting either that activation of the transporter ilivolves its insertion into the membrane or that only functioning transporters bind bumetanide. There is evidence that the [Na+-K+-2CI -] cotransporter is regulated by phosphorylation. In various tissues it can be activated or inactivated by cAMP, phorbol esters, and cytosolic-free calcium [113]. In perfused

417 squid giant axon the transporter is inactivated by ATP-depleting solutions ~t the cytoplasmic face of the membrane, and the rate of inactivation is slowed when vanadate or fluoride :are infused. Similar results were found in ferret red cells [71]. The results are interpreted as indicating that the [Na+-K÷-2CI -] cotransporter or its regulatory system must be phosphorylated to be active and that mitigation of the effects of ATP depletion by vanadate and fluoride is due to the activity of these agents as phosphatase inhibitors [5]. In turkey red cells, c A M P or adrenergic stimulation of the [Na +-K+-2CI-] cotransporter is associated with phosphorylation of a 230 kDa membrane protein called goblin, but goblin is not phosphorylated when the pathway is activated by cell shrinkage [3]. Compelling evidence for phosphorylation of the [Na÷-K+-2C! -] cotransporter itself is provided by the report of Pewitt et al. [224]. In duck red cells kinase inhibitors like K-252a and H-9 block the activation of [Na+-K+-2CI-] cotransport both by agents thought to act via cAMPdependent protein kinase and also by cAMP-independent stir~uli, such as cell shrinkage. Okadaic acid, a protein serine/threonine phosphatase inhibitor, stimulated [Na+-K+-2C! -] cotransport and bumetanide binding and increased the level of phosphorylation of many membrane proteins, not including the cAMP-dependent phosphorylatio.,~ sites in goblin. A 150 kDa membrane protein from these cells that binds bumetanide - presumably the [Na+-K+-2Ci -] c transporter itself - was found to be a phosphoprotein, but there were no studies reported to indicate t~. ~ t~ activity as a transporter was affected by its le, =l "* phosphorylation. Recently Lytle and FcrDush [1'/4] hay : shown that a 200 kDa bumetanide-binding membrane protein from shark rectal gland is phosphorylated in response to activation by forskolin and also by changes in cell volume. A role for cytosolic-free magnesium as a modulator of [Na+-K+-2CI -] cotransport was suggested by Fiatman [70] in ferret red cells and confirmed by Starke and McManus [278] in red cells of ducks. This finding may explain the str.~king differences in activity of this pathway in the red cells of various healthy human donors [64,281]. Mairbaurl and Hoffman [180-182] report that people with high levels of red cell [Na+-K ÷2C!-] cotransport have high levels of free cytosolic Mg 2+, which in turn correlates with low levels of 2,3-diphosphoglycerate [110]. It may be that the Mg 2+ modulation of [Na÷-K+-2CI -] cotransport is closely related to phosphorytatior: of the transporter. Iv'. Some possible special volume sensor and tran,~luction mechanisms

The possible roles of known second messengers in activating volume-responsive transporters have been

discussed above. In this section those mechanisms which could serve as special sensors or transducers of the volume stimulus are focused upon. One way to approach the problem of volume perception is to measure the 'set point' for volumeactivated transporters: in a plot of volume-activated transport flux versus cell volume, the set point is the volume above or below which the flux is activated or inactivated. Perturbations of the cell that alter the set point for transporters might exert their action on the volume receptor. For example, changing the anion of red cells from chlor?de to thiocyanate - a manoeuver that has profound effec~ on the isoelectric point of the cell's lmpermeant solutes [223] - moves the set point for shrinkage-.activated N a + / H + exchange [109,213], sweUing-activatcd Ca2+/Na + exchange [212] and swelling-activated [ K ' - C I - ] cotransport [162] all down to a lower cell volume, so that the cell behaves as if it were swollen and seeks a lower steady-state volume * The same effect can be achieved by depleting cells of cytosoiic magnesium [22i]. The set point can be made to move in the other direction, thus raising the volume threshold at which the swelling-activated fluxes are triggered, by loading the cells with magnesium or lithium ions or pretreating them with the protein phosphatase inhibitor, okadaic acid [222]. With the exception c f stretch-activated channels, it is unlikely that the transport proteins themselves act as volume sensors. There is evidence, for example, that the voib~c rests..,_,, i~J some cells can be disabled, ~:~ "":~e "ic tra=~porters still functional. In rat thyn~o~,,, ~~".32] and deg :ed cells [215] low concentrations of inaleimides inhibit Na ~/H + exchange in response to cell shrinkage without impairing the response of the exchanger to cytoplasmic acidification. Magnesium depletion renders the N a + / H + exchanger of dog red ceils utterly unresponsive to cell shrinkage, but N a + / H + exchange can still be activated in magnesium-depleted cells by lowering cytoplasmic pH [219]. In intact dog red cells the N a + / H + exchanger can be made more sensitive to ceil shrinkage by raising the concentration in the cytosol of lithium or protons [214]. However, in resealed dog ghosts only pr¢,tons [104], not Li +, will stimulate the N a + / H + exchanger and change its set point for volume activation [221]. Presumably, Li + acts on a cell mechanism that detects or transduces volume signals, possibly a G-protein [11], and is disrupted by the process of hemolysis and resealingo * Stimulation of [K+-Ci - ] cotransport by SCN-, a non-halide, requires some ek'planation.Lauf [162] found that pre-incubationof red cells with SCN-, then transfer of cells back to CI - increased [K+-CI - ] cotransport. Under the circumstancesof these experiments virtuallyall the cell anion would be equilibrated with that of the medium; therefore, th~ sthnulatory effect of SCN- did not require that a high concentrationof that anion be present during the flux measurement.

418 Further evidence for a mechanism in the cell that responds to volume signals and communicates with membrane transporters is the finding that there is co-ordination between shrinkage-induced and swelling-induced responses. This was first shown in duck red cells, where [Na +-K+-2 CI-] cotransport (activated by cell shrinkage, catech¢lamines, hypoxia, and a rise in cytosolic magnesium) and [K+-CI -] cotransport (activated by cell swelling and cell magnesium depletion) are apparently orchestrated so that they never function simultaneously or at cross purposes [111,278]. This apparent co-ordination of transporters in the duck red cell could come about as a result of a change in specificity of a single transport protein from [Na+-K +2 CI-] to [K+-CI-]. However, in the dog red cell there are two systems, swelling-induced [K+-CI -] cotrans. port and shrinkage-induced Na+/H + exchange, that carry no ions in common and yet sho~ reciprocal responses to a variety of perturbations. C a ~ / N a + exchange in these cells appears to respond in a manner parallel to [K+-CI -] cotransport (Table V). TABLE V Reciprocal responses of volume-activated transporters in dog red cells Perturbation

Na + / H + exchange

[K + -CI- ] cotransport

Ca 2+/Na + exchange

Swelling

Inhibits (slow) Activates (fast) Activates Inhibits Activ,tes Activates Inhibits

Activates (slow) Inhibits (fast) Inhibits Activates Inhibits Inhibits Activates

Activates NT Inhibits NT NT Activates NT NT Activates

Shrinkage Mg2 +-loading Mg2+-depletion Okadaic acid Lithium loading SCN- loading

NT, not tested. For details and references see text.

There is reciprocity also in the kinetics of activation and deactivation by volume perturbation. Like rabbit red cells [1341, dog red cells show a 5-10 min delay in the activation of swelling-induced [K+-CI -] cotransport. In addition, the swelling-induced deactivation of N a + / H + exchange is a slow process. Cell shrinkage, on the other hand, effects a prompt activation of N a + / H + exchange and deactivation of [K+-CI -] cotransport [222]. These observations point to the existence of some volume-sensitive mechanism that acts as a go-between or switch between the two transporters. The effects of okadaic acid have led to the proposal that a kinase-phosphatase system governs the activation state, not only of [K+-CI -] cotransport, as discussed above [134,135], but also of N a + / H + exchange (Fig. 3, Ref. 222). Finally, volume-perturbed cells can be treated with fixatives, so that their transporters behave as if the cell

[ K - e l l OFF

[K-CI] ON

SLOW SH

HI M g " HIU"

~

A

SWELL

'

Na/H ON

8

LO Mg'"

Na/H OFF

Fig. 3. Model for the coordinated activation and inactivation of [K+-CI - ] cotransport and Na+/H + exchange pathways in dog red cells. Cell shrinkage, high Mg 2+, high Li + concentrations and phosphatase inhihitors, e.g., okadaic acid, shift the system into state A (Na + / H + exchange activated, [K +-CI- ] cotransport inactivated), while cell swelling, low cellular Mg 2+, or exposure of the cells to SCN- (see footnote, section IV) promote a shift into state B (Na+/H + exchange inac'Jvated, [K+-CI - ] colransport activated). Transformation from state A to B is a slow process and requires the action of protein phosphatase(s), while a shift from state B to state A is rapid and requires ATP-dependent kinase action.

had the volume at which the fixative was applied [218]. This has been done with glutaraldehyde and N-phenylmaleimide for shrinkage-activated Na+/H + exchange in dog red cells [197,215,222]. The latter compound can fix swelling-induced [K+-CI -] cotransport as well (Parker and Colclasure, unpublished observati s ) . What is not precisely ~!ear ,~! these experiments is whether the fixatives exert their effect on the transport mechanisms themselves or on a regulator ef transport. In the case of N-phenylmaleimide, fixation of the N a + / H + exchanger in the 'off' configuration does not preclude its activation by cYtosolic acidification [215]. If there is a volume-sensing mechanism separate from the transporters, then h~3w does it work? Even if known second messengers, e.g., cellular calt:~um, cAMP or phosphoinositides convey the signal, how do ceils perceive that they are shrunken or swollen? How is the volume perturbation registered? Is it by virtue of a change in shape with an attendant deformation of the cell skeleton? Is it due to a change in concentration of some key cytoplasmic substance? Is it related to the tension on the membrane? In the following sections some of these alternatives will be discussed.

IV-A. Ionic strength and osmolality An obvious candidate for a volume transducer in cells exposed to anisotonic medic, might be the change in ionic strength or osmolality itself. But the replacement of a large fraction of ions by non-permeating

419 nonelectrolytes, e.g. sucrose in the bathing media, has no significant effect on cell volume regulation in most cell types [9~;~1.It has also been shown in a number of preparations that when cell volume is changed isoosmotically by ~dding or subtracting ~olute from the cell, the same response occurs as when cells are swollen or shrunken osmotically (see for example Ref. 213). Activation of the Na +/H + exchanger by hypertonic shrinkage in several cell types requires a previous hypoosmotic shock and a regulatory volume decrease (see sectien III-B.I). The loss of cellular solute during this pretreatment may be responsible for this phenomenon, thus the activation of the Na+/H + exchanger may be modulated by cell ion a n d / o r osmotic content. Recently Foskett and Spring [72i showed with an intracellular chloride-sensitive dye, SPQ, that shrinkage of salivary acinar cells ,during muscarinic stimulation is associated, not only with a reduction of chloride content, but also of chloride concentration [72,73]. This occurs because of a selective loss of chloride in excess of other anions. It is conceivable that cells having lost chloride during RVD might, upon replacement in an isotonic medium, have a low chloride concentration and that this, perhaps by mediation of a chloride-bicarbonate exchanger, would lower the cytoplasmic pH, thus stimulating Na +/H + exchange.

IV-B. pH or membrane potenti,ff It is plausible that as cells shrink or swell the concentration and dilution of charged, impermeant cell contents might result in a change in the distribution of protons or other permeant ions that would be re~e':ted .;-. a change in cytoplasmic pH [32] or even of membrane potential [122]. Indeed, it was proposed that in red cells the pK of hemoglobin might be altered by the changes in its concentration that attend cell volume perturbation [85], but so far as we are aware, this notion has not found experimental support, and the likelihood that cytoplasmic pH or membrane potential are general volume transducers seems remote [76,122, 273]. Still, cellular pH may significantly modulate volume-dependent ion movements, e.g., cytoplasmic alkalinization activates swelling-induced Ca2+/Na + exchange (see section II-A.3.), and cellular acidification strongly inhibits swelling-activated K + channels (see section II-A). The role of cell pH as a ohysiological regulator of the volume-induced potassium channel in lymphocytes has been suggested by Deu:sch end Lee [58]. As discussed earlier, in sickle cell disease an acidification-induced activation of red cell [K+-CI -] cotransport and consequent cell shrinkage may be relevant to the pathogenesis of vase-occlusive crises. Intracellular pH changes resulting from the functioning of a given volume-induced transport pathway may result in the activation of other pH-sensitive eel-

ume-reguh~tory pathways. An interesting example is the activation of N a + / H + exchange (causing cell swelling) in cells which undergo a regulatory volume decrease by the opening of K + channels, which produces H + uptake and intracellular acidification [168,169]. Changes in membrane potential often follow volume perturbation, but they are probably not induced by the cell swelling per s e More likely, they reflect alterations in membrane cond~uctance induced by the volume-regulatory response [98,227]. As noted earlier, when conductive K + and Cl- pathways are activated during RVD reaction, the Cl- conductance usually surpasses that of K +, and membrane depolarization occurs. Such an effect, may in turn modulate the activity of the K + pathway: membrane depolarization was shown to increase the calcium sensitivity of a Ca2+-activated K + channel [68].

IV-C. Magnesium Cytosolic magnesium modulates swelling- and shrinkage-activvted transporters in the red cells of ferrets [70], ducks [278] and dogs [221]. Magnesium is a relatively impermeam intfaceUular ion [69], and might plausibly be expected to change its intracellular concentra~ion with swelling or shrinkage. Cellular magnesium concentration has been shown to affect the calcium sensiti,,'ty of calcium-activated K + channels [89], and this ,. ze~ may be relevant in the modulation of volome-act~, :1 K + conductance. Give,, the evidence presented above for modulation of transporters by kina~es and phosphatases and the catalytic role for magnesium in most kinase and some phosphatase reactions, the candidacy of this cation as a volume transducer would seem strong indeed. Although some evidence supports a role for magnesium as a volume transducer [277], it has yet to he shown that changes in cell volume alt¢: free magnesium. Changes m cell volume ought to affect the concentration, not only of magnesium cation, but also of substances in the cell like ATP that bind ~lagnesium. The role of magnesium as a voiume transduce.r is ihetefore as yet unproven.

IV:D. Cytoskelewn When cells change volume they change shape, and there must be associated movements of the membrane, the membrane skeleton, and the cytoskeleton that could possibly play a role in volume transduction. Evidence in favor of the role of the membrane skeleton derives from experiments in which volume regtl|ation by coils was either inhibited or stimulated by microfilament disruptive agents like the cytocha!asins [53,227]. The cytoskeletonomediated fusion of cytoplasmic membrane vesicles with the plasma membrane was suggested to

420 insert new, volume-regulatory ion channels [72]. The difficulty with these experiments is that they show a role for microfilaments in cell integri~j, but they do r~ot prove that filament stretch or deformation is the volume signal. Drastic changes in the radius of curvat,~re of red cells induced by agents that either cause an evagination or an invaginatien of the membrane exert no influence on the activation of swelling-induced [K~--CI - ] cotransport [133]. A cogent discussion of the role of mechanical deformation in generating biochemical sig,,als that might influence membrane transporters and involve certain known second messenger systems can be found in Watson's recent review [303].

IV-E. Membrane stretch Volume regulation and volume perception in several prokar~otic and eukariotic cell types are postulated to involve the opening of stretch-activated ion channels, in most cases with a broad specificity for both cations and anions (for recent reviews see Refs. 75, 193, 196. 248). This notion originated with the observation that when negative pressure was exerted on membrane patches, channels were activated. As discussed above (section II-A), although some reports indicate the occurrence of stretch-activated selective K + channels, the triggering of swelling-induced K + channels in several types of cell membranes is indirect: it entails the inflow of calcium that follows the opening of stretchactivated calcium channels. In the case of swellingactivated CI- channels, a more direct role of membrane stretch has been suggested (see section II-B). A low conductance (about 20 pS), mechanosensitive, non-selective ion channnel in opossum kidney cells [292] was shown to open under hypoosmotic conditions. Since this channel carries both Na +, K +, and Cl- ions, its role in a volume regulatory reaction is questionable. Several reports indicate the presence of stretch activated cation channels, carrying both monovalent and divalent cations in skeletal muscle membranes [247,248], and gadolinium ions were reported to specifically inhibit such transport pathways [319]. Another example of mechanosensitive ion channels is the 'maxi' CI- channel observed in Escherichia coil [184]. Negative hydrostatic pressure, corresponding to a small change in medium osmolality, significantly activates this probably non-selective anion channel, which has a conductance of nearly 1000 pS. In its most general form, this concept presumes that membrane skeletal deformation is the volume transducer, because there are putative connections between the membrane skeleton and the stretch-activated channels [248]. Mediation of membrane stretch via adenyl cyclase has been proposed [302,303]. Two difficulties with the idea that membrane stretch is a general mechanism for volume signaling are that it does not explain

shrinkage-activated tran.~ort, and it is hard to reconcile with the geometry of many cells, particularly red cells, which can accommodate a~ much as a 60% increase in volume without a cha~£¢ in surface area [123]. Mechanical tension ('stretch') along the plane of the membrane could only be exerted at the point where the cell reaches its iy¢ic volume, and yet degrees of swelling much less than this are sufficient to activate [K+-CI -] cotransport [221].

IV-F. Concentration and ddution of regulatory enzymes Jennings and Schulz [133] pre~:eneed a model in which a hypothetical kinase-phosphatase regulatory system might report changes in cell volume based on a differential effect of concentration/dilution on the enzymes responsible for phosphorylation and dephosphorylation. Although the scheme is plausible, the authors did not claim they had evidex~ce to support it.

IV-G. Macromolecular crowding Recently it was proposed that the primary volu.~,ae signal in dog red cells is generated by the concentra. tion and dilution of cytosolic proteins [49]. This notion is based on the observation that resealed ghosts display shrinkage-activated N a + / H + exchange only when their cytosolic protein concentration exceeds 30 g / d l (.~,imilar to native red cells). This effect is totally independent of the actual volume of the resealed ghosts. Partial substitution of albumin for hemoglobin in the ghosts showed that Na+/H + exchange was activated at the same critical cytosolic protein concentration. A similar set of observations has been reported for [K-Cl] cotransport in resealed dog ghosts [48]. High concentrations of inert macromolecules can greatly influence the specific activity of certain enzymes, a phenomenon termed 'macromolecular crowding' [132,190,263,323]. The kinetics and the equilibria of reactions may be different in the crowded cytoplasm [79] versus the dilute solutions in which enzymes are usually assayed [190]. Log differences in specific activity of a glycolytic enzyme, glyc,eraldehyde-3-phosphate dehydrogenase, in response to small changes in the ctmcentration of a variety of ambient proteins, have been reported by Minton [190]. l[ macromolecular crowding can affect the activity of em~ymes, and if volume activation of transport involves 'gnzymes such as phosphatases and kinases, as recently, suggested [134,135,174,222], then it seems plausible., that changes in the activities of cytoplasmic or memb,,ane-associated phosphatases a n d / o r kinases might ar!se from concentration and dilution of hemoglob/n with the shrinking and swelling of red ceils. Given the crowded state of the cytoplasm in most

42i cell types [79], these observat&o~s may apply to other volume-sensitive responses.

V. Physiological role of,cell ~olume-acfivated ion t~nsport pathwa3,s The volume regulatory' apparatus in animal cells is extremely sensitive: changes as small as 2-5% of the original cell volume, ~c~:urring even in the most gradual way, may result in the activation of specific transport pathways [40A70]. Perturbations of cell volume could conceivably arise from two circumstances: the cell might be subjected to an aaisotonic environment, or the solute content of the cell might change. In higher animals the tonicity of extracellular fluid is nearly constant, and most cells are not subjected to severe osmotic stress. However, in the renal medulla, in lumenal surface epithelia, and in the portal venous system significant changes in extracellular osmolarity occur even under physiological circumstances. An example ,d' volume-regulatory behavior in t,iuo, in response to an anisotonic extracellular medium, is furnished by the mammalian red cell, which in traveling through the capillaries of the renal medulla encounters a hypertonic environment. The red cell is protected from shrinking by :ts ability to equilibrate urea across its membrane very rapidly, via a specific transporter [175]. Renal tubular cells subjected to enduring hyper° tonicity control their volume mainly by adjusting their content of nonelectrolytes (see Ref. 14). The transcription of the aldose reductase gene in kidney cells is activated by cell shrinkage, with the result that sorbito! accumulates within the cells and protects them from dehydration [82]. Similarly, hypertonicity stimulates transcription of a message for the synthesis of a glyeine/betaine transporter in renal cells, which are thereby enabled to accumulate protective solute [240]. In the mucous membranes of the airway and the gastrointestinal tract, cells are intermittently exposed to anisotonic media. The volume-regulatory ion flux pathways reviewed here protect the epithelia from these effects by stabilizing cell water content during the processes of secretion and absorption. The beststudied tissues in this respect are the skin, urinary bladder and gall bladder of amphibia, but the recent literature reports investigations on mammalian tissues as well [153,193,2941. Large amounts of salt and water flow through the cells in 'high resistance' epithelia, utilizing some of the transport pathways that are activated by cell volume changes. Models of the deployment of channels and electroneutral transporters in various epithelia are given in a recent comprehensive review by O'Grady et al. [20!1. Transeellular ion and water flow is fueled primarily by the electrochemical gradients generated by the N a + / K + pump on the basolateral membrane

but requires the synchronized work of conductive Na*, K +, and CI- channels plus [Na+-K+-2 CI-] cotransporters which may exist on e;ther surface of the epithelium (see Refs. 40, 53, 88, 193, 234, 265, 282, 299). Models describing the functioning of such systems [153,283] suggest that cell volume change~ may play a role in eo6rdinating the communication among transporters on opposite :urfaces of the cell ('tran~cellular cross-laik'). For example, when hormonal stimulation operas Na ~ ei~annels in the apical membrane, the cells swell, and swelling-activated Ci- and K + channels on the baso!ateral membrane become activated, making possible transceUular movement of salt and water in the absorption process. Conversely, when apical C!channels are activated (e.g., by cAMP-dependent stimuli), increased secretion may entail a sequence which mvolv,:~ chloride loss, potassium loss, cell shrinkage and then the shrinkage-induced activation of the basolateral [Na ÷-K+-2 Ci-] cotransporter, thus promoting net movements of fluid from serosa to mucosa. Cell volume change can be part of a hormonal response in an epithelium. Muscarinic stimulation of salivary gland cells, for example, causes a rise in cytosolic calcium followed by a shrinkage of the cell due to loss of KC1 and water. A volume-recovery phase (RVI) ensues, and the interaction of the systems can give rise to oscillations of cytoplasmic calcium as well as of cell volume (Fig. 4, Refs. 73, i97). Muscarinic agonists can also effect rapid activation of N a + / H + exchange in salivary gland cells [183]. A pathological circumoance in which cell volume can become perturbed has been revealed by studies on ischemic cardiac cells. Hypoxia or acidosis, in addition to affecting the supply of ATP for the N a + / K ÷ pump, may cause inappropriate stimulation of the N a + / H ÷ exchanger and therefore lead to a cell volume increase,



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Carbachol Fig. 4. Oscillations of cell volume (triangles, solid line) and cytoplasmic free calcium (open circles, dotted line) following application of carbachol to a single rat salivary gland acinar cell. Note that an increase in calcium is associated with volume loss, and vice versa. (Reproduced from Ref. 73, with the permission of the authors and the publishers.)

~2 triggering the activation of volume regulatory pathways [7]. It i~as recently been postulated that cell volume changes may themselves be part of a second messenger signaling system [118,301]. The opening of hormoneand neuroh'ansmitter-gated Cl- channels (e.g., the GABA and glycine receptors in central nervous system inhibitory synapses, see Ref. 75) may lead to changes in both membrane potential and in cell volume. Cellular signaling that involves an increase in cytoplasmic calcium may also induce volume changes by opening K + a n d / o r CI- channels (see Refs. 72, 73). Recent studies suggest a regulatory role of cell volume in the function of liver ceils [90,117,118]. Physiological increases in amino acid (e.g., glutamine or glycine) concentrations in portal venous blood lead to a Na+-dependent amino acid uptake by liver ceils, which causes them to swell and to activate volume regulatory K + (and probably CI-) efflux pathways [308]. Moreover, cell swelling, induced either by amino acid uptake or by a hypoosmotic shock may itself play a second me",:cnger role, resulting in profound changes in hepati~: metabolism: glycogenolysis, glycotysis, and proteolysis are inhibited, while glycogen and protein synthesis are stimulated [ 117,118]. Mediator roles for membrane stretch-activated calcium influx a n d / o r mitochondrial swelling have been suggested in connection with these observations [118]. Volume regulatory ion fluxes apparently also have a crucial role in several plant species. In marine algae and terrestrial non-lignified plants, the maintenance of a turgor pressure, balancing the osmotic gradient which acts on the plasma membrane, provides a mechanical force to resist gravity and deformation. In Mimosa and in carnivorous plants leaf movement is also driven by the changes in the turgor pressure in the motor cells. The molecular basis of these phenomena seems to be the opening of swelling-induced a n d / o r mechanosensitive K ~ and CI- channels. In most plant tissues, as indicated earlier for the animal cells, stretch-activated opening of Ca -'+ channels with a consequent increased cytoplasmic calcium seems to be responsible for the activation of the K ÷ and CI- channels [194,203]. Other physiologic volume-regulatory functions might include the modulation of cell water content associated with growth and maturation: witness the decrease in water content and volume of red cells as they develop. In some species this process involves the conversior~ from a high-potassium to a low-potassium cell [220]. Interference with the normal maturation of red cells, e.g., by treatment of the animal with hydroxyurea, can lead to disturbances in cell water content [205]. Expression of the oncogene Ha-ras in growth-arrested cultured fibroblasts stimulates mitogenesis and cellular replication. These events are associated with a stimulation of two shrinkage-activated transporters, N a + / H +

exchange and [Na+-K+--2 CI-] cotransport, with the result that the cells rapidly accumulate salt and water. Whether the changes in solute and cell volume are part of a signaling system or the result of stimuli leading to cell growth and replication is unknown [186]. A topic about which little information could be found is the cellular mechanism by which the osmotic concentration in the animal body is perceived and regulated. Osmoreceptors in the brain detect small changes in body fluid tonicity and generate such responses as thirst and the control of antidiuretic hormone levels. Hypotonic swelling of cardiac atrial cells induces the release of atrial natriuretic factor (ANF), which produces natriuresis in the kidney [45,91]. It seems plausible that these physiologic receptor functions involve volume perception, but the mechanisms are not understood on a cellular basis. The principal function of the volume regulatory apparatus in animal cells appears to be the stabilization of cell water content in the face of the forces mentioned h~ the introduction that tend to disperse the cell corten~s towards a state of equilibrium w~th the surrounding ~j~cL..~. Atl advantage of belonging to the animal kingdom is that one can move about in the atmosphere a,,wl flex ~r~e's cells. Without carefu!!y balanced pumps and leaks man would live like plants, encased in rigid cellulose walls, or would be obliged to live in an osmotically congenial aquatic environment so composed that there would be no driving force moving water across cell membranes. VI. Summary and conclusions Swelling-activated K + and CI- channels, which mediate RVD, are found in most cell types. Prominent exceptions to this rule include red cells, which together with some types of epithelia, utilize eleetroneutral [K+-CI -] cotransport for down-regulation of volume. Shrinkage-activated Na + / H + exchange and [Na+-K +2 CI-] cotransport mediate RVI in many cell types, although the activation of these systems may require special conditions, such as previous RVD. Swellingactivated K + / H + exchange and Ca2+/Na+ exchange seem to be restricted to certain species of red cells. Swelling-activated calcium channels, although not carrying sufficient ion flux to contribute to volume changes may play an important role in the activation of transport pathways. In this review of volume-activated ion transport pathways we have concentrated on regulatory phenomena. We have listed known secondary messenger pathways that modulate volume-activated transporters, although the evidence that volume signals are transduced via these systems is preliminary. We have focused on several mechanisms that might function as volume sensors. In our view, the most important candi-

423

dates for this role are the structures which detect deformation or stretching of the membrane and the skeletal filaments attached to it, and the cxtraordinary effects that small changes in concentration of cytoplasmic macromolecules may exert on the activities of cytoplasmic and membrane enzymes (macromolecular crowding). It is noteworthy that volume-activated ion transporters are intercalated into the cellular signaling network as receptors, messengers and effectors. Stretchactivated ion channels may serve as receptors for cell volume itself. Cell swelling or shrinkage may serve a messenger function in the communication between opposing surfaces of epithelia, or in the regulation of metabolic pathways in the liver. Finally, these transporters may act as effector systems when they perform regulatory volume increase or decrease. This review discusses several examples in which relatively simple methods of examining volume regulation led to the discovery of transporters ultimately found to play key roles in the transmission of information within the cell. So, why volume? Because h's functionally important, it's relatively cheap (if you happened to have everything else, you only need some distilled water or cencentrated salt solution), and since it involves ma~y disciplines of experimental 0iology, it's fun to do. References 1 Aabin, B. and Hoffmann, E.K. (1986) Acta Physiol. Scand. 128, 42A. 2 Allen, T.J.P,., Noble, D. and Reuter, H. (1989) (eds.) in Sodium-Calcium Exchange (Oxford University Press, Oxford, UK). 3 Alper, S.L., Beam, K.G. and Greengard, P. (1980) J. Biol. Chem. 255, 4864-4871. 4 Altamirano, A.A. and Beauge, L. (1985)Cell Calcium 6, 503-525. 5 Altamirano, A.A., Breitwieser, G.E. and Russell, J.M. (1988) Am. J. Physiol. 254, C582-C586. 6 Alvarez, J., Garcia-Saacho, J. and Herreros, B. (1986) Biochim. Biophys. Acta 857, 2911-294. 7 Anderson, S.E., Murphy, E., Steenbergen, C., London, R.E. and Cala, P.M. (1990) Am. J. Physiol. 259, C940-C948. 8 Ande:son, M.P., Rich, D.P., Gregory, R.J., Smith, A.E. and Welsh, M.J. (1991) Science 251,679-682. 9 Armando-Hardy, M., EIIory, J.C.. Ferreira, H.G., Fleminger, S. and Lew, V.L. (1975) J. Physiol. (London) 250. 32-33P. 10 Aronson, P., Nee, J. and Suhm, M.A. (1982) Nature 299, 161163. 11 Avissar, S., Schreiber, G.. Danon, A. and Belmaker, R.H. (1988) Nature 331, 440-442. 12 Baker, P.F., Blaustein, M.P., Hodgkin, A.L. and Steinhardt, R.A. (1969) J. Physiol. (London) 200, 431-458. 13 Ballanyi, K. and Grafe, P. (1988)Renal Physiol. Biochem. I1, 142- 157. 14 Beck, F., Dorge, A. and Thurau, K. (1988) Renal Physiol. Biochem. 11,174-186. 15 Berkowitz, L.R. and Orringer, E.P. (1982) Blood Cells 8, 283287. 16 Berridge, M.J. (1987) Annu. Rev. Bioehem. 56, 159-193.

17 Besterman, J.M., May, W.S., LeVine, H., Cragoe, E.J. and Cuatrecasas, P. (1985) J. Biol. Chem. 260, 1155-1159. 18 Bianchini, L., Smith, J.D., Woodside, M., Takai, A. and Grinstein, S. (1991) FASEB J. 5, A669. 19 Birnbaumer, L., Abramowitz, J. a,'.:l Brown, A. (1990) Biochim. Biophys. Acta 1031, 163-224. 20 Bjerregaard, H.F. (1989) Pfluegers Arch. 414, 193-199. 21 Blum, R.M. and Hoffman, J.F. (1971) J. Membr. Biol. 6, 315-328. 22 Bookehin, R.M., Ortiz, O.E. and Lew, V . L (1991) J. Clin. Invest. 87,113-124. 23 Borgese, F., Garcia-Romeau, F. and Morals, R. (1987) J. Physiol. (Lorx;on) 382, 123-144. 24 Brechler, V. Pavoine, C., Lotersztajn, S., Garbarz~ E. and Pecker, F. (1990) J. Biol. Chem. 265, 16851-16855. 25 Brown, A.M. and Wood, M.A. (1987) J. Physiol. (London) 382, 142P. 26 Brugnara, C., Kopin, A.S., Bunn, H.F. and Tosteson, D.C. (1985) 1. Clin. Invest. 75, 1608-1617. 27 Brugnara, C., Van Ha, T. and Tosteson, D.C. (1988) A~I. J. Physiol. 255, C346-C356. 28 Brugnara, C., Van Ha, T. and Tosteson, D.C. (1989) Am. J. Physiol. 256, C994-C1003. 29 Brugnara, C., Van Ha, T. and Tosteson, D.C. (1989) Blood 74. 487--495. 31) Brugnara, C. (1989) J. Membr. Biol. 111, 69-81. 31 Burns, K.D., Homma, T. and Harris, R.C. (1990) J. Am. Soc. Nephrol. i, 714 (abstract). 32 Cala, P.M. (1980) I. Gen. Physiol. 76, 683-708. 33 Cala, P.M. (1983) Mol. Physiol. 4, 33-52. 34 Cala, P.M. (1986) Curr. Topics Membr. TransT. 26, 79-99. 35 Cala, P.M., Mandel, L.J. and Murphy, E. (1986) Am. J. Physiol. 250, C423-C429. 36 Cala, P. and Grinstein, S. (1988) in Na/H Exchange (Grinstein, S., ed.), 201-208, CRC Press, Boca Raton, FL. 37 Caroni, P. and Carafoli, E. (1983) Eur. J. Biochem. 132, 451-460. 38 Cassel, D., Katz, M. and Rc,tman, M. (!986) J. Biol. Chem. 261. 5460-5466. 39 t ,,,~tle, N.A. and Strong. P.N. (1986) FEBS Lett. 20,9, II7-121. 40 Chan~berlin M.E. and Strange, K. (1989) Am. J. Physiol. 257, C159-C173 41 Cher,,'.,~a, f~ ~.}.and Zeuthe, T. (1987) Aeta Physiol. Stand. 129, ~37-138,. 42 Cheung, R.K., Grinstein, S., Dosch, H.-M. and Gelfand. E.W. (1982) J. Ce, Physiol. 112, 189-196. 43 Christensen, O. (1987) Nature 330, 66-68. 44 Christensen, S., Hoffmann, E.K., Saermark, T.S. and Simonsen, L.O. (1988) J. Physiol. (London) 403, 109P. 45 Clemo, H.F. and Baumguricn, C.M. Am. J. Physiol. 260. C681C690. 46 Cohen, P. (1988) Proc. Roy. St~c. Lond. Ser. B 234, 115-144. 47 Cole!asure, G.C. and Parker, J.C. (19901 Comp. Biochem. Physiol. 96A, 147-150. 48 Colcl~sure, G.C. and Parker, J.C. (1991) J. Den. Fh:~ic~l. Abstracts, December, 1991. 49 Colclasure, G.C. and Parker, J.C. (1991) J. Gen. Physiol., in press. 50 Condrescu, M., Gerardi, A. and DiPolo, R. (1088) Biochim. Biophys. Acta 946, 289-298. 51 Cornejo, M., Guggino, S.E. and Guggino, W.B. (1989) J. Membr. Biol. 1i 0, 49-55. 52 Davis, B.A., Hogan, E.M. and Boron, W.E. (1991) Am. J. Physiol. (in press). 53 Davis, C.W. and Finn, A.L. (1987) Fed. Proc. Fed. Am. Soc. Exp. Biol. 44, 2520-2525. 54 Davis, C.W. and Finn, A.L. (1987) J. Gen. Physiol. 89, 687-702. 55 Davson, H. (1940) Cold Spring Harbor Symp. Qvant. Biol. 8, 255-268.

424 56 Delpire, E. and Lauf, P.K. (1991) J. Gen. Physiol. 97, 173-193. 57 De Peyer, .i.E., Cachilin, A.B., Levitan, I.B. and Reuter, H. (1982) Proc. Natl. Acad. Sci. USA 79, 4207-4211. 58 Deutsch, C. and Lee, S.C. (1988) Renal Physiol. Biochem. I1, 260-276. 59 DiPoio. R. and Beauge, L. (1981) Biochim. Biophys. Acta 645, 229-236. 60 DiPoio. R. and Beauge, L. (1986) Biochim. Biophys. Acta 854, 298-306. 61 DiPolo, R. and Beauge, L. (1987) J. Gen. Physiol. 90, 505-525. 62 Dunham, P.B. and Logue, P.J. (1986) Am. J. Physiol. 250, C578-C583. 63 Duhm, J. (1987) J. Membr. Biol. 98, 15-32. 64 Duhm, J, and Gobel, B.O. (1984) J. Membr. Biol. 77, 243-254. 65 Ellory, J.C, a,:d Dunham, P.B. (1980) in Membrane Transport in Erythrocytes (Lassen, U.V., Ussing, H.H. and Wieth, J.O., eds.) pp. 409-423, Munksgaard, Copenhagen. 66 EIIory, J.C. and Hall, A.C. (1988) Comp. Biochem. Physiol. 90A. 533-537. 67 Ericson, A.-C. and Spring, K.R. (1982) Am. J. Physiol. 243, C146-C150. 68 Findlay, I. (1984) J. Physiol. (London) 350, 179-195. 69 Flatman, P.W. (1984) J. Membr. Biol. 80, 1-14. 70 Platman, P.W. (1988) J. Physiol. (London) 397, 471-487. 71 Flatman, P.W. (1991) J. Physiol. (London) 437, 495-510. 72 Foskett, J.K. and Spring, K.R. (1985) Am, J. Physiol. 248, C27-C36. 73 Foskett, J.K. and Melvin, J.E. (1989) Science 244, 1582-1585. 74 Fc~kett, J.K. (1990) Am. J. Physiol. 259, C998-C1004. 75 Franciolini, F. and Petris, A. (1990) Biocbim. Biophys. Acta 1031, 247-259. 76 Freedman, J.C. and Hoffman, J.F. (1979) J. Gen. Physiol. 74, 157-185. 77 Frizzell, R.A., Rechkemmer, G. and Shoemaker, R.L. (1986) Science 233, 558-560. 78 Pujise, H., Yamada, I., Masuda, M., Miyazawa, Y., Ogawa, E. and Takahashi, R. (1991) Am. J. Physiol. 260, C589-C597. 79 Fulton, A.B. (1982) Cell 30, 345-347. 80 Ganz, M.B., Pachter, JA. and Barber, D,L. (1990) J'. Biol. Chem, 265, 8989-8992. 81 Garcia-Diaz, J.F. (1991) Biophys. J. 59, 646a. 82 Gar¢ia-Perez, A., Martin, B., Murphy, H.R., Uchida, S., Murer, H., Cowley, B.D., Handler, J.S. and Burg, M.B. (1989) J. Biol. Chem. 264, 16815-16821. G~,rdos, G. (1958) Biochim. Biophys. Aeta 30, 653-654. 84 G~rdos, G., Sz:~sz, I. and Sarkadi, B. (1975) FEBS Proc. 35, 167-180, 85 Ga~-Bobo, C.M. and Solomon, A.K. (1968) J. Gen. Physiol. 52, 825-853. 86 Geek, P., Pietrzyk, C., Burckhardt, B.-C., Pfeiffer, B. and Heinz, E. (1980) Biochim. Biophys. Acta 600, 432-447. 87 Giangiacomo, K.M., Gareia-Calvo, M., Garcia, M.L. and McManus, O.B. (1991) Biophys. J. 59, 214a. 88 Gil!es, R. (1988) Renal Physiol. Biochem. 11,277-288. 89 Golowasch, J., Kirk'wood, H. and Miller, C. (1986) J. Exp. Biol. 124, 5.-13. 90 Graf, J., Haddad, P., Haeussinger, D. and Lang, F. (1988) Renal Physiol. Biochem. 11, 202--220. 91 Greenwald, J.E., Apkon, M., Hruska, K.A. and Needleman, P. (1989) J. Clin. Invest. 83, 1061-1065. 92 Gregor, R. and Sehlatter, E. (1983) Pflueger's Arch. 396, 325334. 93 Grinstein, S., Clarke, C.A. and Rothstein, A. (1982) Phil. Trans. Roy. Soc. Lond. Ser. B, 299, 509-518. 94 Grinstein, S., Dupre, A. and Rothstein, A. (1982) J. Gen. Physiol. 79, 849-868.

95 Grinstein, S., Clarke, C.A., Dupre, L al~d Rothstein, A. (1982) J. Gen. Physiol. 80, 801-823. 96 Grinstein, S., Cohen, S., Sarkadi, B. and Rothstein, A. (1983) J. Cell. Physiol. 116, 352-362. 97 Grirstein, S., Clarke, C.A., Rothstein, A. and Gelfand, E.W. (1983) Am. J. Physiol. 245, C160-C163. 98 Grinstein, S., Rothstein, A., Sarkadi, B. and Gelfand, E.W. (1984) Am. J. Physiol. 246, C204-C215. 99 Grinstein, S., Rothstein, A. and Cohen, S.J. (1985) J. Gen. Physiol. 85, 765-788. 100 Grinstein, S. and Rothstein, A. (1986) J. Membr. Biol. 90, 1-12. 101 Grinstein, S.~ Goetz-Smith, J.D., Stewar~, D., Beresford, B.J. and Mellors, A. (1986) J. Biol. Chem. 261, 8009-8016. 102 Grinstein, S., Cohen, S., Goetz, J.D., Rothstein, A., Mellors, A. and Gelfand, E.W. (1986) Curr. Topics Mcmbr. Transp. 26, 115-134. 103 Grinstein, S. and Cehen, S. (1987) J. Gen. Physiol. 89, 185-213. i9~ Grinstein, S. and Smith, J.D. (1987) J. Biol. Chem. 262, 90889092. 105 Grinstein, S. (1988) (ed.) N a / H Exchange, pp. 1-347, CRC Press, Boca Raton, FL. 106 Grinstein, S. and Smith, J. (1990) J. Gen. Physiol. 95, 97-120. 107 Grygorczyk, R. (1991) Biophys. J. 59, 646a. 108 Guggino, S.E., Guggino, W.B., Green, N. and Sacktor, B. 0987) Am. J. Physiol. 252, C128-C137. 109 Guizouarn, H., Scheuring, U., Borgese, F., Motais, R. and Garcia-Romeu, F. (1990) J. Physiol. (London) 428, 79-94. 110 Gupta, R.K., Benkovic, J.L. and Rose, Z.B. (1978) J. Biol. Chem. 253, 6172-6176. 111 Haas, M. and McManu~ T.J. (1985) J. Gen. Physiol. 85, 649-667. 117 Haas, M. and Forbusb B. (1986)J. Biol. Chem. 261, 8434-8441. 113 Haas, M. (1989) Annu. Rev. Physiol. 51,443-457. 114 Harper, J.R., Orringer, E.P. and Parker, J.C. (1979) Res. Commun. Chem. Pathol. Pharmacol. 26, 277-284. 115 Hartmann, H.A., Kirseh, G.E., Drewe, J.A., Tagliatela, M., Joho, R.H. and Brown, A.M. (1991) Science 251,942-944. 116 Hata, T., Makino, N., Nakanishi, H. and Yanaga, T. (1988) Mol. Cell. Biochem. 84, 65-76. 117 Haussinger., D., Lang, F., Bauers, K. and Gerok, W. (1990) Eur. J. Biochem. 188, 689-695. 118 Haussinger, D. and Lang, F. (1991) Biochem. Cell. Biol 69, 1-4. 119 Hazama, A. and Okada, Y. (1988) J. Physiol. (London) 402, 687-702. 120 Hilgeman, D.W. (1990) Nature 344, 242-244. 121 Hladky, S.B. and Rink, T.J. (1977) in Membrane Transport in Red Cells, (EIIory, J.C. and Lew, V.L. eds.) pp. 115-136, Academic Press, London. 122 Hladky, S.B. and Rink, T.J. (1978) J. Physiol. (London) 274, 437-446. 123 Hoffman, J.F., Eden, M., Barr, J.S. and Bedell, H.S. (1958) J. Cell. Comp. Physiol. 51,405-414. 124 Hoffmann, E.K. (1978) in Osmotic and Volume Regulation, (Jorgensen, C.B. and Skadhauge, E., eds.) pp. 397-417, Munksgaard, Copenhagen. 125 Hoffmann, E.K., Simonsen, L.O. and Lambert, I.H. (1984) J. Membr. Biol. 78, 211-222. 126 Hoffmann, E.K. (1985) Fed. Proc. Fed. Am. Soc. E~p Riol..44, 2513-2519. 127 Hoffmann, E.K., Lambert, I.H. and Simonsen, L.O. (1986) J. Membr. Biol. 91,227-244. 128 Hoffmann, E.K., Lambert, I.H. and Simonsen, L.O. (1988) Renal Physiol. Biochem. 11, 221-247. 129 Hoffmann, E.K. and Simonsen, L.O. (1989) Physiol. Rev. 69, 315-382. 130 Howard, L.D. and Wondergem, R. (1987) J. Membr. Biol. 100, 53-61.

425 131 lkehara, T., Yamaguchi, H., Hosokawa, K. and Miyamoto, H. (1990) Am. J. Physiol. 258, C599-C609. 132 Jarvis, T.C., Ring, D.M., Daube, S.S. and yon Hippel, P.H. (1990) J. Biol. Chem. 265, 15160-15167. 133 Jennings, M.L. and Schulz, R.K. (1990) Am. J. Physiol. 259, C960-C967. 134 Jennings, M.L. and AI-Rohil, N. (1990) J. Gen. Fhysiol. 95, 1021-1040. 135 Jennings, M.L. and Schulz, R.K. (1991) J. Gen. Physiol. 97, 799-817. 136 Jensen, B.S. and Hoffmann, E.K. (1991) Acta Physiol. Scand., in p~ess. 137 Kaji, D. (1989) Am. J. Physiol. 256, CI214-C.!223. 138 Kaczorowski, GJ., Slaugh'.er, R.S., King, V.F.K. and Garcia, M.L. (1989) Biochim. Biophy~ ~cta 988, 287-302. 139 Kantner, N., Hanrahan, J.W, Jensen, T.J., Nalsmith, A.L., Sun, S., Ackerley, C.A, Reyes, E.F., Tsui, L.-T., Rommono, J.M., Bear, C.E. and Riordan, J.R. (1991) Cell 64, 681-691. 140 Kato, M. and Kato, K.J. (1988) Mol. Cell. Biochem. 83, 15-25. 141 Kim, H.D., Sergeant, S., Forte, L.R., Sohn, D.H. and Ira, J.H. (1989) Am. J. Physiol. 256, C772-C778. 142 Klaerke, D.A., Petersen, J. and Jorgensen, P.L. (1987) FEBS Lett. 216, 211-216. 143 Klaerke, D.A., Karlish, S.J.D. and Jorgensen, P.L. (1987) J. Membr. Biol. 95, 105-.112. 144 Klaerke, D.A. and Jergensen, P.L. (1988) Comp. Biochem. Physiol. 90A, 757-765. 145 Kov;ics,T., Sz~isz, 1., Sarkadi, B., Brezanoczi, F. and Gfirdos, G. (1989) Acta Biochem. Biophys. Acad. Sci. Hung. 24, 83-99. 146 Kracke, G.R. and Dunham, P.B. (1990) Proc. Natl. Aead. Sci. USA 87, 8575-8579. 147 Kregenow, F.M. (1981) Annu. Rev. Physiol. 43, 493-505. 148 Lackington, I. and Orrego, F. (1981) FEBS Left. 133, 103-106. 149 Lagnado, L. and MeNaughton, P.A. (1990) J. Membr. Biol. 113, 177-t91. 150 Lambert, I.H., Hoffmann, E.K. and Christensen, P. (1987) J. Membr. Biol. 98, 247-256. 151 Lambert, I.H., Hoffmann, E.K. and Jorgensen, F. (1989) J. Membi. Biol. 111, 113-132. 152 Lang, F., Messner, G. and Rehwald, W. (1986) Am. J. Physiol. 250, F953-F962. 153 Larsen, E.H. (1991) Physiol. Rev. 71,235-283. 154 Latorre, A. and Miller, C. (1983) J. Membr. Biol. 71, 11-30. 155 Lau, K.R., Hudson, R.L. and Schultz, S.G. (1984) Proc. Natl. Acad. Sci. USA 81, 3591-3594. 156 Lauf, P.K. and Theg, B.E. (lqg0) Biochem. Biophys. Res. Commun. 92, 1422-1428. 157 Lauf, P.K. (1982) J. Comp. Physiol. 146, 9-16. 158 Lauf, P.K. (1984) J. Membr. Biol. 77, 57-62. 159 Lauf, P.K. (1985) J. Membr. Biol. 88, !-13. 160 Lauf, P.K. (1985) Am. J. Physiol. 249, C271-C278. 161 Iauf, P.K. (1988) Renal Physiol. Biochem. 11,248-259. 162 Lauf, P.K. (1991) Am. J. Physiol. 260, C503-C512. 163 Le Maout, S., Tauc, M., Koeehlin, N. and Poujeol, P. (1990) Biochim. Biophys. Acta, 1025, 29-39. 164 Levinson, C. (1991) FASEB J. 5, A670. 165 Lew, X~.L. and Bookchin, R.M. (1986) J. Membr. Biol. 92. 5%74. 166 Lew, V.L., Freeman~ C.J., Ortiz, O.E. and Bookchin, R.M. (1991) J. Clin. Invest. 87, 100-112~ 167 Li, M.J. and Iwasa, K.H. (1991) Biophys. J. 59, 456a. 168 Livne, A., Grinstein, S. and Rothstein, A. (1987) J. Cell. Physiol. 131, 354-363. 169 Livne, A. and Hoffmann, E.K. (1990) J. Membr. Biol. 114, 153-157. 170 Lohr, J.W. and Grantham, J.J. (1986) J. Clin. Invest. 78, 11651172.

171 Lopes, A. and Guggino, W. (1987) J. Membr. Biol. 97, 117-125. 172 Lytle, C. and MtManus, T.J. (1986) J. Gen. Physiol. 88, 36a. 173 Lytle, C.Y. and McManus, T.J. 0987) J. Gen. Physiol. 90, 28a-29a. 174 Lyric, C. and Forbush, B. 0990)J. Cell Biol. I l l , 312a. 175 Macey, R.I. (1984) Am. J. Physiol. 246, C195-C203. 176 MacKnight,/'~.D.C. and Leaf, A. (1977) Physiol. Rev. 57, 510573. 177 MacKnight, A.D. (1988) Renal Physiol. Biochem. 11, 114-141. 178 MacRobbie, E.A.C. and Ussing, H.H. (1961)Acta Physiol. Stand. 53, 348-365. ~'~o.. ~:,.~..~:, v. and Inaba, M. (1985)J. Biol. Chem. 260, 3337-3343. 180 Mairbaurl, H. and Hoffman, J.F. (1989) FASEB J. 3, 2056 (A579). 181 Mairbaurl, H. and Hoffman, J.F. (1990) Biophys. J. 57, 94a. 182 Mairbaurl, H. and Hoffman, J.F. (1991) Pflueger's Arch., in press. 183 Manganel, M. and Turner, R.J. (1991) J. Biol. Chem. 266, 10182-10188. 184 Martinac, B., Buechner, M., Delcour, A.H., Adler, J. and Kung, C. (1987) Proe. Natl. Acad. Sei. USA 84, 2297-2301. 185 McManus, T.J. (1987) Fed. Proc. Fed. Am. Soc. Exp. Biol. 46, 2278-2381. 186 Meyer, M., Maly, K., Uberall, F., Hoflacher, J. and Grunicke, H. (1991) J. Biol. Chem. 266, 8230-8235. 187 Milanick, M.A. (1989) Am. J. Physiol. 256, C390-C398. 188 Miller, C., Moczydlowski, E., Latorre, A. and Phillips, M. (1985) Nature 313, 316-318. 189 Miller, C. (1991) Science, 252, 1092-1096. 190 Minton, A.P. (1983) Mol. Cell. Biochem 55, 119-140. 191 Miyamolo, H., Ikehara, T., Yamaguchi, H., Hosokawa, K., Yonezu, T. and Masuya, T. (1986) J. M~mbr. Biol. 92, 135-150. 192 Moczydlowski, E., Lucchesi, K. and Ravindram, A. 0988) J. Membr. Biol. ! 05, 95-111. 193 Montrose-Rafizadeh, C. and Guggino, W.B. (1990) Annu. Rev. Physiol. 52, 761-772. 194 Moran, N. (1991) Biophys. J. 59, 456a. 195 Morris, C.E. and Sigurdson, W.J. (1989) Science 243, 807-809. 196 Morris, C.E. (1990) J. Membr. Biol. 113, 93-107. 197 Motais, R., Borgese, F., Scheuring, U. and Gareia-Romeu, F. (1989) J. Gen. Physiol. 94, 385-400. 198 Motais, R., Scheuring, U., Borgese, F. and Garcia-Romeu, F. (1990) in Progress in Cell Research, Vol. I, (Ritchie, J.M., Magistreui, P.J. and l~olis, L., eds.) pp. 179-193, Elsevier, Amsterdam. 199 Nicoll, D.A., Longoni, S. and Philipson, K.D. (1990) Science 250, 562-565. 200 Niggli, E. and Lederer, W.J. (1991) Nature 349, 621-624. 201 O'Grady, S.M., Palfrey, H.C. and Field, M. (1987) Am. J. Physiol. 253, C177-C!92. 202 Okada, Y., Yada, T., Ohno-Shosaku, T. and Oiki, S. (1987) J. Membr. Biol. 96, 121-128. 203 Okazaka, Y. and Tazawa, M. (1990)J. Membr. Biol. 114, 189194. 204 Orringer, E.P., Broekenbrough, J.S., Whitney, J.A., Glosson, P.S and Parker, J.C. (1991) Am. J. Physiol., in press. 205 Orringer, E.P., Blythe, D.S.B., Johnson, A., Phillips, G., Dover, G.J. and Parker, LC. (1991) Blood 78, 212-216. 206 Ortiz, O.E. and Sjodin, R.A. (1984) J. Physiol. (London) 354, 287-301. 207 Parker, J.C. (1973) J. Gen. Physiol. 62, 147-156. 208 Parker, J.C., Gitelman~ H.J., Glosson, P.S. and Leonard, D.L. (1975) J. Gen. Physiol. 65, 84-96. 209 Parker, J.C. (1977) in Membrane Transport in Red Cells, (EIIory, J.C. and Lew, V.L., eds.) pp. 427-465, Academic Press, London. 210 Parker, J.C. 0978) J. Gen. Physiol. 71, 1-17.

426 211 212 213 214 215

Parker, J.C. and Harper, J.R. (1980) J. Clin. Invest. 66, 254-259. Parker, J.C. (1983) Am. J. Physiol. 244, C318-C323. Parker, J.C. (1983) Am. J. Physiol. 244, C324-C330. Parker, J.C. (1986) J. Gen. Physiol. ~,7, 189-200. Parker, J.C. and GIosson, P.S. (1987) Am. J. Pbvsol. 253, C60-C65. 216 Parker, J.C. (1987) Am. J. Physiol. 253, C580-C587. 217 Parker, J.C. (1988) Biochim. Biophys. Acta 943, 463-470. 218 Parker, J.C., Glosson, P.S. and Walstad, D.L. (1988) Mol. Cell. Biochem. 82, 91-95. 219 Parker, J.C., Gitelman, H.J. and McManus, T.J. (1989) Am. J. Physiol. 257, C1038-C1041. 220 Parker, J.C. and Dunham, P.B. (1989) in Red Blood Cell Membranes (Agre, P. and Parker, J.C., eds.) pp. 507-561, Marcel Dekker. 221 Parker, J.C,, MclVlanus, T.J., Starke, L.C. and Gitelman, H.J. (1990) J. Gen. Physiol. 96, 1141-1152. 222 Parker, J.C., Colclasure, G.C. and McManus, T.J. (1991) J. Gen. Physiol., in press. 223 Paync, J.A., Lytle, C.Y. and McManus, T.J. (1990) Am. J. Physiol. 259, C819-C827. 224 Pewitt, E.B., Hegde, R.S., Haas, M. and Palfrey, H.C. (1990) J. Biol. Chem, 265, 20747-20756. 225 Philipson, K.D. (1985) Annu. Rev. Physiol. 47, 561-571. 226 Pierce, S.K., Politis, A.D., Smith, L.H. and Rowland, L.M. (1988) Cell Calcium 9, 120-140. 227 Pierce, S.K. and Politis, A.D. (1990) Annu. Rev. Physiol. 52, 27-42. 228 Preston, R.R., Walleq-Friedman, M.A., Saimi, Y. and Kung, C. (1990) J. Membr. Biol. 115, 51-60. 229 Ravens, U. and Wettwer, E. (1989) J. Cardiovasc. Pharmacol. 14 (Suppl. 3), $30-$35. 230 Reeves, J.P. (1985) Curt. Topics Membr. Transp. 2". 77-127. 231 Reeves, J.P., Bailey, C.A. and Hale, C.C. (1986) J. Biol. Chem. 261, 4948-4955. 232 Reichstein, E. and Rothstein, A. (1981) J. Membr. Biol. 59, 57-63. 233 Reuss, L. (1983) Nature 305, 723-726. 234 Reuss, L. (1988) Renal Physiol. Biochem. 11, 187-201. 235 Reuter, H. and Seitz, N. (1968) J. Physiol. (London) 195, 451470. 236 Reuter, H. (1991) Nature 349, 567-568. 237 Rink, TJ., Sanchez, A., Grinstein, S., Rothstein, A. (1983) Biochim. Biophys. Acta 762, 593-596. 238 Riordan, J,R. and Passow, H. (1971) Biochim. Biophys. Acta 249, 601-605. 239 Riordan, LR., Rommens, LM., Kerem, B., Alon, N., Rozmahel, R., Grzelczak, Z., Zielenski, J., Lok, S., Plavsic, N., Chou, J.-L., Drumm, M.L, Iannuzzi, M.C., Collins, F.S. and Tsui~ t,.-C. (1989) Science 245, 1066-1073. 240 Robey, A.B., Kwon, H.M., Handler, J.S., Garcia-Perez, A. and Burg, M.B. (1991) J. Biol. Chem. 266, 10400-10405. 241 Robertson, G.A., Atkinson, N.S. and Ganetzky, B. (1991) Binphys. J. 59, 196a. 242 Romero, P.J., Ortiz, C.E. and Melitto, C. (1990) J. Membr. Biol. 116, 19-29. 243 Rommens, J.M., Iannuzzi, M.C., Kerem, B., Drumm, M.G., Melmer, G., Dean, M., Rozmahel, R., Cole, J.L., Kennedy, D., Hikada, N., Zsiga, M., Buchwald, M., Riordan, J.R., Tsui, L.-C. and Collins, F.S. (1989) Science 245, 1059-1065. 244 Ross, P.E. (1991) Biophys. J. 59, 598a. 245 Rothstein, A. and Mack, E. (1990) Am. J. Physiol. 258, C827C834. 246 Rothstein, A. and Mack, E. (1991) Am. J. Physiol., in press. 247 Roy, G. and Sauve, R. (1987) J. Membrane Biol. 100, 83-96. 248 Sachs, F. (1987) Fed. Proc. Fed. Am. Soc. Exp. Biol. 46, 12-16. 249 Sachs, J.R. (1988) J. Gen. Physiol. 92, 685-711.

250 251 252 253 254 255 256 257 258 259

260

261 262 263 264 265

266 267 268 269 270 271 272 273 274 275 276 277 278 279 280 281 282 283 284 285 286 287

Sackin, H. (1987) Am. J. Physiol. 253, FI253-FI262. Sackin, H. (1989) Proc. Nail Acad. Sci. USA 86, 1731-1735. Sanderson, MJ. and Dirksen, E.R. (1991) Biophys. J. 59, 650a. SardeL C., Franchi, A. and Pouyssegur, J. (1989) Cell 56, 271280. Sardet, C., Counillon, L., Franchi, A. and Pouysseguer, J. (1990) Science 247, 723-726. Sarkadi, B., Mack E and Rothstcin, A. (1983) J. Gen. Physiol. 83, 497-512. Sarkadi, B., Mack, E. and Rothstein, A. (1983) J. Gen. Physiol. 83, 513-527. Sarkadi, B., Attisano, L., Grinstein, S., Buchwald, M. and Rothstein, A. (1984) Biochim. Biophys. Acta 774, 159-164. Sarkadi, B., Cheung, A., Mack, E., Grinstein, S., Gelfand, E.W. and Rothstein, A. (1985) Am. J. Ph::siol. 248, C480-C487. Sarkadi, B. and Gfirdos, G. (1985) ir~ Enzymes of Biological Membranes, 2nd edn. Vol. 3 (Martonosi, A., ed.) pp. 193-234, Plenum Press, New York. Sarkadi, B. and Gfirdos, G. (1989) in The Red Cell Membrane (Raess, B.U. and Tunnicliff, G., eds) pp. 369-396, Humana Press, Clifton, NJ. Shenolikar, S. and Nairn, A.C. (!991) Adv. Second Messenger Phosphoprotein Res. 23, 1-121. Schwartz, W. and Passow, H. (1983) Annu. Rev. Physiol. 45, 359-374. Sherwin, D.E. and Windsor, D.J. (1990) Eur. J. Biochem. 190, 523-529. Siebens, A.W. arid Kregenow, F.M. (1978) Physiologist 21, 110. Siebens, A.W. (1985) in The Kidney: Physiology and Pharmacology (Seldin, D.W. and Giebisch, G., eds.) pp. 91-115, Raven Press, New York. Siebens, A.W. and Kregenow, F.M. (1985) J. Gen. Physiol. 86, 527-564. Siffert, W., .!ak~s, K.H. and Akkerman, J.W. (1990) J. Biol. Chem. 265, 15441-15448. Simons, T.J.B. (1976) Nature 267, 467-469. Simon~, T.J.B. (1979) J. Physiol. (Londn,) 2S8. ~t81-507. Smith, D.K. and Lauf, P.K. (19•5) Biochim. Biophys. Acta 818, 251-259. Smith, JB., Cragoe, E.J..~nd Smith, L. (1987) J. Biol. Chem. 262 11988-11994. Smith, R.L., Macara, I.G., Levenson, R., Housman, D. and Cantley, L. (1982) J. Biul. Chem. 257, 773-780. Solomon, A.K., Toon, M.R. and Dix, J.A. (1986) J. Membr. Biol. 91,259-273. Spring, K.R. and Ericson, A.-C. (1982) J. Membr. Biol. 69, 167-176. Stampe, P. and Vestergaard-Bogind, B. (1985) Biochim. Biephys. Acta, 815, 313-321. Stampe, P. and Vestergaard-Bogind, B. (1989) J. Membr. Biol. 112, 9-14. Starke, L.C. (1989) Ph.D. Thesis, Duke University Der}artment of Physiology. Starke, L.C. and McManus, T.J. (1990) FASEB J. 4, A8i8. Steck, T.L. (1974) J. Cell Biol. 62, 1-19. Stevens, C.F. (1991) Nature 349, 657-658. Stewart, G.W. (1988) J. Physiol. (London) 401, 1-16. Strange, K. (1991) Am. J. Physiol. 260, F225-F234. Strieter, J., Stephenson, J.L., Palmer, L.G. and Weinstein, A.M. (1990) J. Gen. Physiol. 96, 319-344. Szfisz, I. and Gfirdos, G. (1974) FEBS Lett. 44, 213-216. Szfisz, I., Sarkadi, B. and Gfirdo~, G. (1978) Acta Biochim. Biophys. Acad. Sci. Hung. 13, 133-141. Sze, H. and Solomon, A.K. (1979) Biochim. Biophys. Acta 554, 180-194. Szirmai, M., Sarkadi, B., Szfisz, I. and Gfirdos, G. 0988) Haematologia 21, 33-40.

427 288 Tabeharani, J.A. and Hanrahan, J.W. (1991) Biophys. J. 59, 6a. 289 Taniguchi, J. and Guggino, W.3. (1989) Am. J. Physiol. 257, F347-352. 290 Toro, L., Ramos-Franco, J. and Stefani, E. (1990) J. Gen. Physiol. 96, 373-394. 291 Tosteson, D.C. and Hoffman, J.F. (1960) J. Gen. Physiol. 44, 169-194. 292 Ubl, J., Murer, H. and Kolb, H.~. (1988) J. Membr. Biol. 104, 223-232. 293 Ubl, J., Murer, H. and Kolb, H.A. (1988) Pflueger's Arch. 412, 551-553. 294 Ussing, H.H. (19t)0) in Cell Volume Regulation. Comparative Physiology, Vol. 4, (Beyenbach, K.V.. ed.), pp. 87-113, Karger, Basel. 295 Vanselow, K.H. and Hansen, U.P. (1989) J. Membr. Biol. 110, 175-187. 296 Vestergaard-Bogind, B., Stamp'_, P.and Christophersen, P. (1987) J. Membr. Biol. 95, 121-i30. 297 Volkl, H. and Lang, F. (1988) Pflueger's Arch. 411,514-519. 298 Volkl, H. and Lang, F. (1988) Pflueger's Arch. 412, 1-6. 299 Volkl, H., Paulmichl, M. and Lang, F. (1988) Renal Physiol. Biochem. I 1,158-173. 300 Walsh, D.A., Newshohne, P., Cawley, K.C., Van Pattefi, S.M. and Angelos, K.L. (1991) Physiol. Rev. 71,285-304. 301 Waterford, M. (1990) Trends Biochem. Sci. 15, 329-330. 302 Watson, P. (1990) J. Biol. Chem. 265, 6569-6575. 303 Watson, P.A. (1991) FASEB J. 5, 2013-2019. 304 Weinman, E.J., Steplock, D., Bui, G., Yuan, N. and Shenolikar, S. (1990) Am. J. Physiol. 258, FI254-FI258. 305 Welling, P.A., Linshaw, M.A. and Sullivan, L.P. (1985) Am. J. Physiol. 249, F10-F27.

306 Welsh, M.J. (1986) Science 232, 1648-1650. 307 Wen, Yo, Famulsky, K.S. and Carafgli. E. (1984) Biochem. Biophys. Res. Commun. 122, 319-324. 308 Wettstein, M., Dahl, S., Lang, F., Gerok, W. arid Haussinger, D. (1990) Biol. Chem. Hoppe-Seyler 371,493-501 309 Wiener, H., Klaerke, D.A. and JOrgensen, P.L (1990) J. Membr. Biol. i17, 275-283. 310 Willis, J.S., Nelson, R.A., Gordon, C., Vilaro, P. and Zhao, Z. (1990) Comp. Biochem. Physiol. 96A, 91-96. 311 Willis, J.S., Nelson, R.A., Livingston, B. and Marjanovic, M. (1990) Comp. Biochem. Physiol. 96A, 97-105. 312 Wolff, D., Cecchi, X., Spalvins, A. and Canessa, M. (1988) J. Membr. Biol. 106, 243-252. 313 Wong, S.M.E. and Chase, H. (1986) Am. J. Physiol. 250, C841C852. 314 Wong, S.M.E., DeBeil, M.C. and Chase, H.S. (1990) Am. J. Physiol. 258, F292-F296. 315 Wood, P.G. and Mueller, H. (1984) Eur. J. Biochem. 141, 91-95. 316 Wood, P.G. and Mueller, H. (1985) Eur. J. Biochem. 146, 65-69. 317 Worrell, R.T. and Frizzell, R.A. (1991) Biophys. J. 59, 27a. 318 Yamaguchi, D.T., Green, J., Kleeman, V.R. and Muallem, S. (1989) J. Biol. Chem. 264, 4383-4390. 319 Y~ng. X.C. and Sachs, F. (1989) Science, 243, 1068-1071. 320 Yellen, G (1987) Annu. Rev. Biophys. Chem. 16, 227-246. 321 Yellen, G., Jurman, M.E., Abramson, T., MacKinnon, R. (1991) Science 251,939-942. 322 Yool, A.J. and Schwarz, T. (1991) Nature (London) 349, 700-704. 323 Zimmerman, S.B. and Harrison, B. (1987) J. Biol. Chem. 84, 1871-1875.

Activation of ion transport pathways by changes in cell volume.

Swelling-activated K+ and Cl- channels, which mediate RVD, are found in most cell types. Prominent exceptions to this rule include red cells, which to...
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