Kidney International, Vol. 51 (1997), pp. 454—461 ION CHANNELS AND HEME PROTEINS AS SENSORS

Oxygen sensing by ion channels

JoSE LOPEZ-BARNEO, PATRICIA ORTEGA-SAENZ, ANTONIO MOLINA, ALFREDO FRANCO-OBREGON, JUAN UREA, and ANTONIO CASTELLANO

Departamento de Fisiologla Médica y Biofisica, Universidad de Sevilla, Facultad de Medicina, Sevilla, Spain

Oxygen is required for the survival of all higher life forms due 02-sensitive ion channels and chemotransduction in arterial to its role in mitochondrial respiration permitting ATP synthesis chemoreceptors by oxidative phosphorylation. Oxygen is also necessary for many The carotid bodies, located in the bifurcation of the carotid other cellular functions, including the synthesis of steroid hor-artery, are the main arterial oxygen chemoreceptors in mammals. mones or the generation of reactive oxygen species in the macro-The mechanisms underlying the basic transductional process have phage defense response. In mammals, 02 diffuses into the bloodremained obscure for decades, but most authors have traditionally in the , is concentrated within the erythrocytes by its bindingascribed a central role to the glomus, or type I, cells, which are of to hemoglobin and is distributed throughout the body by theneuroectodermal origin. These cells contain numerous cytosolic . Arterial and tissular 02 tension (P02) are keptgranules rich in and other and within restricted limits by short- and long-term adaptative mech-peptides, establishing synapses with afferent sensory fibers of the anisms set in motion by changes in 07 availability due tosinus nerve. Hence, it was widely accepted that glomus cells act as environmental or physiological circumstances. An immediatethe primary 02 sensors releasing transmitters that stimulate the response to a hypoxic stress is the reflex increase in respiratoryafferent fibers of the sinus nerve in response to low P02 [1, 21. rate resulting from the activation of arterial 02 chemoreceptors.However, direct proofs for the chemoreceptive properties of In these organs, decrements in arterial P02 are translated intoglomus cells have come from patch-clamp experiments demon- afferent nerve discharges stimulating the brain stem respiratorystrating that these cells, which were originally thought to he non-excitable, can generate Na and Ca2-dependent action centers to produce hyperventilation [1, 2]. Acute regional hypoxiapotentials [ii, 12] and contain voltage-gated K' channels whose also produces vasorelaxation of most systemic arteries in an effort activity is inhibited by lowering 02 tension [12—15]. More recently, to increase 02 delivery to hypoxic tissues, whereas it constrictsthe simultaneous recording of membrane ionic currents with pulmonary resistance arteries, thereby diverting blood toward the patch clamp techniques and of cytosolic [Ca2 '] by microfluorim- better ventilated alveoli [3, 4]. Besides the rapid functionaletry, in combination with amperometric detection of dopamine in adaptations to , protracted alterations in 02 availabilitysingle glomus cells, has allowed a detailed characterization of also lead to changes in the expression of genes encoding forseveral 02-dependent responses at the cellular level [16—1 81. various hormones, enzymes, and growth factors [5—8]. One of the Themajor electrophysiological properties of rabbit glomus cells best known examples is the enhancement of the synthesis andand their dependence on 02 tension are summarized in Figure 1. secretion of erythropoietin by the kidney and liver in response toOne of the most relevant features of glomus cells is that with chronic , which increases the 02-carrying capacity ofdepolarizations to positive membrane potentials hypoxia selec- the blood by augmenting the production of red blood cells [9]. tively reduces K current amplitude (Fig. IA). In rabbit glomus Despite the paramount importance of the physiologic functionscells, this is due to the interaction of 02 with a specific, voltage- regulated by 02 tension the molecular and cellular processesdependent, delayed rectifier K channel of single channel con- underlying the oxygen-dependent control systems are largelyductance ---20 pS in normal K concentrations [191. As illustrated unknown. With the exception of the recently identified hemopro-in Figure 1B, low P02 decreases the channel open probability tein involved in bacteria N2 fixation [10], the nature of the 02without altering the single channel conductance. The inhibition of sensors is still an enigma. In the past few years it has beenthe K current by low P02 is manifested as an increase of cellular demonstrated that ion channels represent novel molecular entitiesexcitability which, as shown in Figure 1C, results in an augmen- participating in 02 chemoreception. It has been shown that thetation of the action potential frequency. Together, these experi- activity of various kinds of voltage-gated ion channels can bemental observations give support to the view that glomus cells act as presynaptic-like elements where the increase in action potential modulated by 02 and that these channels might play a key role in firing in response to hypoxia leads to Ca2 influx and transmitter several cellular responses to hypoxia. release [16—18, 20]. The role of ion channels in oxygen chemotransduction sug- gested by the early patch clamp experiments was questioned by some investigators who proposed the mitochondria as the location for the 02 sensor and the major source of the Ca2 required for © 1997 by the International Society of Nephrology the hypoxia-induced secretion in glomus cells [21, 22]. However,

454 López-Barneo et al: Oxygen sensing by ion channels 455

A B C

Normoxia (control) Ca2 K Normoxia Hypoxia +20 open closed 1 Normoxia 1_-80 pr 'K 41• iU11 0.5 nA '-iL. I 2pA

Hypoxia 0.8 0. 'Ca 8 ms !flsr,j04 Fig. 1. Major 02-dependent electrophysiological properties of glomus cells. A. Macroscopic inward and outward currents of glomus cells and reversible inhibition of the outward current by hypoxia (P02 ——20 mm Hg). Control and recovery traces in normoxia (P02 =150mm Hg) are shown superimposed. In this experiment TTXwasadded to block the Na conductance. B. Single-channel recordings from an excised membrane patch containing at most one open 02-sensitive K channel. Depolarizing applied from —80 to +20 mV. Ensemble averages indicating the single-channel open probability in normoxia and hypoxia are from 15 and 22successiverecordings in the two experimental conditions. C. Effect of hypoxia on the firing frequency of a glomus cell during the first 150 ms after switching from voltage- to current-clamp (indicated by the vertical arrow). Note that low P02 evokes an increase in the slope of the pacemaker potentials. (Modified from (18, 20, 381.) recent work done on isolated glomus cells loaded with fluorescent In summary, the experiments on single, dispersed glomus cells dyes has firmly established that the increase of cytosolic Ca2strongly suggest that 02-sensitive ion channels have a major role elicited by low P02 depends almost exclusively on Ca2 entryin the sensory function of the . Ion channels appear to through plasmalemmal voltage-gated ion channels [16—18]. Un-transduce changes in P02 into a cytosolic [Ca2] signal which der normoxic conditions (P02 —150 mm Hg), the average restingtriggers the release of the transmitters that subsequently activate cytosolic [Ca2] in glomus cells is -—-60 nM and rises to 100 to 800 afferent fibers of the sinus nerve. Glomus cells are known to flM upon acute exposure to environmental low P02 (between 50contain catecholamines as well as numerous neuropeptides, how- and 10 mm Hg) [16]. The parallel time courses of these twoever, the putative transmitter that activates the afferent nerve variables (P02 and cytosolic [Ca2]) are illustrated in Figure 2A,fibers is still unknown. The participation of 02-sensitive ion which also includes the signal from an oxygen-sensing electrodechannels in the physiology of the glomus cell is compatible with placed near the glomus cell. The elevation of cytosolic [Ca2] inother mechanisms, for example, the 02-sensitivity of mitochon- response to repeated exposure to hypoxia is very reversible and, indna! functions [22], also possibly contributing to chemotransduc- a given cell, highly reproducible (Fig. 2B). Since hypoxia favorstion in situations of protracted exposure to very low P02 levels. action potential firing in rabbit glomus cells and these cells contain a large population of voltage-gated Ca2 channels, it can Oxygen sensing by ion channels in vascular smooth muscle be expected that those maneuvers that abolish the activity of cells voltage-gated Ca2 channels would also eliminate the response to Blood oxygen tension is known to be an important factor in the low P02. In cells subjected to whole cell voltage-clamp, thelocal regulation of circulation [3, 4, 23].Inmost systemic arteries current through voltage-gated Ca2 channels reversibly disap-a decrease of P02 induces favoring the of pears when Ca2 is removed from the external solution (Fig. 2C)02-deprived tissues. This same behavior is also apparent in the or when 0.2 m Cd2 (a powerful Ca2 tchannel blocker) is addedmain trunk of the . By contrast, hypoxia pro- (Fig. 2D). Thus we also examined the effect of hypoxia in glomusduces of the small distal branches of the pulmo- cells before, during and after exposure to external solutionsnary arteries. Hypoxic pulmonary vasoconstriction is critical for lacking Ca21 or with 0.2 ma Cd2 '.Thesetreatments completelyfetal survival because it helps to maintain the high pulmonary abolished the increase of cytosolic [Ca2J induced by low P02vascular resistance necessary to divert blood through the ductus (Fig. 2 E, F). Brief removal of external Ca21 (Fig. 2E) or additionarteriosus. In the adult, hypoxic pulmonary vasoconstriction of 0.2 mrvt Cd2 to the external solution (Fig. 2F) did not causeseems to contribute to the proper regional distribution of blood any deleterious action on the cells since the responses to hypoxiaflow in the , favoring the irrigation of the better ventilated appeared to be normal when the standard external solution wasalveoli [24]. Despite their physiological importance, the mecha- reintroduced in the chamber. nisms underlying the vasomotor responses to hypoxia are largely 456 LOpez-Barneoet a!: Oxygen sensing by ion channels

A B

P02

145 200 H H H H H

E E 20 Cytosolic [Ca2] 100 200 i- 0

0L 0 2 mm

C D t 0 Ca2 0.2 Cd2

Control Recovery Control 1 nA Recovery 1 ms

E F 450 H H H H H H

0 Ca2 200 0.2 Cd2 300

+ (5 150 100

0 0 2 mm

Fig. 2. Hypoxia promotes voltage-dependent Ca2 influx and elevation of cytosolic Ca2 in glomus cells. A. Parallel changes of 02 tension in the chamber and the modification of cytosolic Ca2 in a fura 2-loaded glomus cell. B. Changes of cytosolic Ca2 during repeated exposure of a glomus cell tohypoxia (H; P02 =30mm Hg). C andD. Calciumcurrents recorded in two patch clamped glomus cells during 8-ms depolarizations to +20 mV from a holding potential of —80 mV. The 0 mvt Ca2 and 0.2 mvi Cd2 external solutions reversibly abolished the current. E andF.Changes of cytosolic Ca2 in response to hypoxia (H; P02 =30mm Jig) in two unclamped cells bathed in a solution of standard ionic composition or after introduction of 0 mvi Ca2 (E) or 0.2 mvt Cd2 (F) solutions to block the voltage-gated Ca2 channels. (Modified from [16]; see this reference for details). López-Barneo et al: Orygen sensing by ion channels 457

A Hypoxia Control Recovery

100 pA

5 ms

B C 60 K 300 1.0 S.. • 0.8 200

'(H> — 0.6 + '(C) 0 100 0.4 .4'

0.2 0 160 120 80 40 0 Hypoxia P02, mm Hg 1 mm

Fig.3. Inhibition of voltage-gated Ca2 channels by low P02 in arterial myocytes. A. Macroscopic calcium currents recorded from a myocyte dispersed from the celiac artery during 15-ms step depolarizations to +10 mV from a holding potential of —80 mV. Exposure to hypoxia (switching from an external solution equilibrated with P02 —150 mm Hg to another with P02 —20 mm Hg) induces an inhibition in current amplitude. Reversibility is illustrated by the recovery trace. B. Calcium current amplitude as a function of P02. Current amplitude normalized to the amplitude of the current in normoxia (ordinate) as a function of P02 (abscissa). Whole-cell currents were elicited with step depolarizations to +10 mV from holding potentials of either —70 or —80 mV during the development of hypoxia in the bath. Note that the effect of hypoxia is more pronounced just below —80mm Hg. The data have been pooled from experiments performed on 3 celiac and 2 main trunk pulmonary myocytes. C. Simultaneous effects of membrane depolarization and low P02 on cytosolic Ca2 concentration in a coronary myocyte loaded with fura-2. Low P02 reversibly counteracted the rise in cytosolic Ca2 induced during the exposure to 60 mi external K. The shaded regions indicate the time during which the external solutions were switched. (Modified from [30, 31]).

unknown. It has been proposed that hypoxia may cause theP02 might cause vasodilation due to the decrease of the cytosolic release of vasoactive substances from either the endothelium or[Ca2] resulting from the inhibition of L-type Ca2 channel neighboring hypoxic tissues; however, a considerable amount ofactivity [30, 31]. This response to hypoxia is illustrated in Figure work suggests that arterial smooth muscle contractility might be3A with recordings of calcium currents obtained from a celiac also under direct control of blood P0, [23—26]. myocyte during depolarizing pulses to + 10 mV. The recordings It has been postulated that hypoxic vasorelaxation may be duedemonstrate an —40% reversible reduction in current amplitude to the opening of ATP regulated channels in the smooth muscleupon exposure to low PU2. The current amplitudc—P02 relation- cell membrane [271. In coronary arterial myocytes, exposure toship is shown in Figure 3B where normalized current amplitude anoxia or to P02 < 40 mm Hg for several minutes leads to the(ordinate) is plotted as a function of °2tensionin the chamber opening of ATP-regulated K (KAT?) channels producing mem-(abscissa). Current inhibition becomes most apparent as the P02 brane hyperpolarization and the closure of voltage-dependentlevel in the bath falls just below 100 mm Hg. Therefore, the Ca2 Ca2 channels [28]. Although KA-ip channels are surely importantchannels in our preparations are modulated within PU2 limits for hypoxic vasorelaxation during protracted exposures to ex-comparable to those previously described to cause vasodilation. tremely low P02 levels, the existence of other 02-dependentThe consequence of a reduction in Ca2 channel activity is the membrane mechanisms is suggested because the sensitivity ofreduction in voltage-dependent Ca2 influx and, hence, relax- most arteries to hypoxia occurs without a compromise of energyation. This type of response is illustrated at the single cell level in metabolism and over a physiologic range of P02 values [29]. a fura-2 loaded coronary myocyte (Fig. 3C). As in most excitable Recent work on dispersed arterial myocytes has indicated that lowcells, exposure to high extracellular K leads to the elevation of 458 López-Barneoet al: Oxygen sensing by ion channels

Normoxia Hypoxia Channel and sensor •1 Channel Independent sensor

Fig. 4. Schematic representation of various mechanisms proposed to explain the modulation of ion channels by oxygen tension. See text for explanation.

Mediator

Channel Associated sensor (channel subunit?)

cytosolic [Ca2 I due to membrane depolarization and subsequentinflux [34 —361, a sequence of events similar to that involved in 02 opening of voltage-gated Ca2 channels. This rise of Ca2 couldchemotransduction in carotid body glomus cells [20]. Moreover, be reversibly inhibited within seconds of lowering P02. Sincemyocytes dispersed from pulmonary resistance arteries seem to these observations are seen even in the presence of glibenclamidecontain a higher density of delayed rectifier K channels, which (5 IM; a blocker of KAT,channels),they reinforce the view thatare those exhibiting sensitivity to P02, than myocytes dispersed low P02 modulates voltage-dependent Ca2 influx through anfrom conduit vessels [36]. Recent data have also demonstrated a effect on Ca2 channel activity [31]. differential distribution and 02 sensitivity of Ca2 channels in In contrast with the hypoxic relaxation of systemic vessels, lowmyocytes dispersed from distinct locations along the pulmonary P02 provokes constriction of the pulmonary resistance arteries [4,arterial tree [37]. In proximal conduit myocytes, Ca2 channels 24, 32, 33]. Recently, several groups have shown that hypoxiagenerally behave as in sytemic arteries, exhibiting inhibition by inhibits the activity of voltage-dependent K channels in pulmo-low P02, whereas an opposite sensitivity (increase in calcium nary myocytes and, hence, proposed that hypoxic pulmonarycurrent amplitude during exposure to hypoxia) is seen in distal vasoconstriction could be partially due to the inhibition of the Kresistance myocytes. Therefore, it seems highly plausible that conductance, which in turn might lead to depolarization and Ca2 hypoxic-induced changes in the K and Ca2 conductances in López-Barneo et al: Oxygen sensing by ion channels 459

A ______+40mV B L —80 mV Vm Control/recovery 0 mV

mi Untransfected CHO cells

Control/recovery

Transfected with Sh K cDNA

m

2 nA 1 nA

2 ms 5 ms Fig. 5. Modulation of Shaker K'- currents by changes of P02. A. Transfection of CHO cells with Shaker B eDNA leads to the expression oflarge macroscopic K'- currents which are recorded with the whole-cell configuration of the patch clamp technique. B. Hypoxia (P02 —20 mm Hg) produces a reversible decrease in the amplitude of the K'- current recorded during depolarizations to two different membrane potentials. distal myocytes act concomitantly to enhance the pressor responsehave little effect on channel closing or inactivation [30, 31, 40]. of resistance pulmonary arteries to low P02, thus contributing toTherefore, it seems that in all these channel classes common acute hypoxic pulmonary vasoconstriction. 02-sensing domains exist that similarly influence their gating properties. The various mechanisms that can be proposed to Is oxygen sensitivity an intrinsic property of ion channels? explain how 02 tension influences ion channel function are Although the modulation of ion channel gating by 02 tensionsummarized in Figure 4. One possibility (depicted in Fig. 4, panel was initially thought to be rather specific for a K* channel class1) is that 02 could directly interact with a specific region of the a present in glomus cells, it seems now that °2 modulates thesubunits or domains forming their basic structure. For example, activity of several types of voltage-dependent ion channels located02 could bind reversibly to coordination complexes formed by in different tissues and involved in various cellular functions.metal-protein sites as it occurs in other molecules [411;however, There are several possibilities to explain how low P02 couldthe structural motifs allowing this chemistry to occur have not influence ion channel activity, though the available data stronglybeen identified thus far. An alternative scenario is that 02 tension suggest that the 02-ion channel interaction occurs through acould be detected by a membrane-associated sensor close to the sensor intrinsic to the plasma membrane that either forms part ofion channel. Such a sensor might be independent (panel 2) or the channels or is a molecule closely associated with them.associated with the channel molecule (panel 3). The case in which Reversible modifications of ionic currents by hypoxia are recordedthe 02 sensor-K channel interaction is direct or occurs in the in dialyzed cells after blockade of G proteins with GTP-y-S andphase of the membrane is the most likely because, as indicated with highly buffered cytosolic pH and soluble mediators, such asabove, water-soluble mediators, washed-out in excised patches, (10 ATP or Ca2 [18, 38]. In addition, reversible hypoxic inhibition ofnot seem to be required for the 02-ion channel interaction. It has K channel activity is observed in excised membrane patches ofbeen proposed that a membrane-bound NADPH-oxidase could glomus cells and central neurons [38, 39]. Furthermore, changesact as putative °2 sensor that could be coexpressed with the in °2 tension produce specific alterations in the kinetic parame-channels, and depending on the P02 levels, modify the redox ters of the channels. In 02-sensitivc ion channels low P02 reducesstatus of specific residues in the channel protein [421. In CHO current amplitude and slows down the activation time course, butcells transfected with eDNA of a recombinant K' channel of the 460 LOpez-Barneoet al: Oxygen sensing by ion channels

Shaker family (Fig. 5A), we have observed that P02 appears to 9. GOLDBERG MA, DUNNINGSP,BUNN HF: Regulation of the erythro- regulate the macroscopic K currents in a similar manner to that poietin gene: Evidence that the oxygen sensor is a heme protein. occurring in carotid body glomus cells (Fig. 5B). Since it is known Science242:1412—1415, 1988 10.GILLES-GONZALEZMA,DITFA S, HELINSKI OR:Ahaemprotein with that CHO cells do not express NADPH oxidase [43], these results kinase activity encoded by the oxygen sensor of Rhizobium meliloti. might indicate that voltage-gated, delayed rectifier, K channels Nature 350:170—172, 1991 possess intrinsic 02 sensitivity. Although these data must be 11. DUCHEN MR, CADDY KWT, KIRBY GC, PA1-ITERSON DL, PONTE J, considered as very preliminary, they open avenues for future BIsC0E TJ: Biophysical studies of the cellular elements of the rabbit carotid body. Neuroscience 26:291—311, 1988 experimental work. 12. LOPEZ-BARNEO J, LOPEZ-LOPEZ JR, URE$A J, GONZALEZ C: Chemo- transduction in the carotid body: K current modulated by PU2 in Concluding remarks type I chemoreceptor cells. Science 242:580—582, 1988 02-sensitive ion channels are more broadly distributed than 13. DELPIANO MA, HESCHELER J: Evidence for a P02-sensitive K channel in the type I cell of the carotid body. FEBS Lett 249:195—198, previously thought and they seem to participate in several °2 1989 dependent cellular functions. Here we discuss some of the phys-14. PEERS C: Hypoxic supression of K currents in type I carotid body iologic functions of 02-sensitive ion channels studied in recent cells: Selective effect on the Ca2-activated K current. Neurosc Left years; however, it is possible that these are only representative 119:253—256, 1990 examples of other processes where 02-sensitive channels are 15. STEA A, NURSECA:Whole-cell and perforated patch recordings from 02-sensitive rat carotid body cells grown in short- and long-term involved. In fact, we speculate that 02-sensitivity might be an culture. Pfiugers Arch 418:93—101, 1991 intrinsic characteristic of several ion channel families sharing16. UREAJ,FERNANDEZ-CHACONR,BENOTAR,ALVAREZ DE TOLEDO common 02-sensing domains. The identification of the structural G, LOPEZ-BARNEO J: Hypoxia induces voltage-dependent Ca2 entry motifs involved in 02 sensing by ion channels may help to and quantal dopamine secretion in carotid body glomus cells. Proc understand how cells sense the changes in 02 leading to the NatlAcad Sci USA 91:10208—10211, 1994 17. BUCKLER KJ, VAUGHAN-JONES RD: Effects of hypoxia on membrane hypoxic regulation of gene expression. It also may lead to the potential and intracellular calcium in rat neonatal carotid body type I design of pharmacological tools for a broad spectrum of patho- cells. J Physiol (Lond) 476:423—428, 1994 physiological conditions. 18. MONTORO Ri, UREIA J, FERNANDEZ-CHACON R, ALVAREZ DC To- LEDO G, LOPEZ-BARNEO J: Oxygen sensing by ion channels and Acknowledgments chemoreception in single glomus cell. J Gen Physiol 107:133—143, 1996 19. GANFORNINA MD, LOPEZ-BARNEO J: Single K channels in mem- This study was supported by grants from DGICYT of the Spanish brane patches of arterial chcmoreceptor cells are modulated by 02 Ministry of Science and Education, the European Community, and the tension. Proc Nat Acad Sci USA 88:2927—2930, 1991 Andalusian Government. P.0-S. and A.M. have a predoctoral fellowship 20. LOPEZ-BARNEO J, BENOT AR, URE8A J: Oxygen sensing and the of the Spanish Ministry of Science and Education. A.F-0. is recipient of electrophysiology of arterial chemoreceptor cells. News Physiol Set a postdoctoral international fellowship from NATO. A.C.'s present ad- 8:191—195, 1993 dress is: Departamento de Ciencias Fisiologicas Humanas y de Ia Nutri- 21. BIscoE TJ, DUCHEN MR: Responses of type I cells dissociated from cion, Universidad de Barcelona, Avenida Diagonal 643, E-08028, Barce- the rabbit carotid body to hypoxia. J Physiol (Lond) 428:39—5 9, 1990 lona, Spain. 22. LAHIRI 5: Chromophores in 02 chemoreception: The carotid body model. News Physiol Sci 9:161—165, 1994 Reprint requests to José López-Banzeo, M.D., Ph.D., Departamento de 23. SPARKS H: Effect of local metabolic factors on vascular smooth FisiologIa Médica y Biojisica, Universidad de Sevilla, Facultad de Medicina, muscle, in Handbook of Physiology. The Cardiovascular System. II. Avenida Sanchez Pizjudn 4, E-41009, Sevilla, Spain. Vascular Smooth Muscle, edited by BOHR DF, SOMLYO AP, SPARKS HV, Bethesda, American Physiological Society, 1980, pp 475—5 13 References 24. WEIR EK, ARCHER SL: The mechanism of acute hypoxic pulmonary vasoconstriction: The tale of two channels. FASEB J 9:183—189, 1995 1. Fio SJ, GONZALEZ C: Initiation and control of chemoreceptor 25. FISHMAN AP: Hypoxia on the pulmonary circulation: How and where activity in the carotid body, in Handbook of Physiology. The Respiratory it acts. Circ Res 38:221—231, 1976 System (vol II), edited by FISHMAN AP, Bethesda, Maryland, Ameri- 26. HALES CA: The site and mechanism of oxygen sensing for the can Physiological Society, 1986, pp 247—312 pulmonary vessels. Chest 88:235S—240S, 1985 2. Mc DONALD DM: Peripheral chemoreceptors: Structure-function 27. DAUT J, MAIER-RUDOLPH WN, VON BECKERATH N, MEHRKE G, relationships in the carotid body, in Regulation of Breathing, edited by GUNTHE K, GOEDEL-MEINEN L: Hypoxic dilatation of coronary arter- HORNBEINTF,New York, Dekker, 1981, pp 105—320 ies is mediated by ATP-sensitive potassium channels. Science 247: 3. DAWSON CA: Role of pulmonary vasomotion in physiology of the 1341—1344, 1990 lung. Physiol Rev 64:544—616, 1984 28. DART C, STANDEN NB: Activation of ATP-dependent K channels by 4. WADSWORTH RM: Vasoconstriction and vasodilator effects of hyp- hypoxia in smooth muscle cells isolated from the pig coronary artery. oxia. TIPS15:47—53,1994 J Physiol (Lond) 483:29—39, 1995 5. SHWEIKI D, ITIN A, SOFFER D, KESHET E: Vascular endothelial growth 29. DETAR R: Mechanism of physiological hypoxia-induced depression of factor induced by hypoxia may mediate hypoxia-initiated angiogene- vascular smooth muscle contraction. Am J Physiol 238:H761—H769, sis. Nature 359:843—845, 1992 1980 6. KOUREMBANAS S, HANNAN RL, FALLER D: Oxygen tension regulates 30. FRANCO-OBREGON A, URErA J, LOPEZ-BARNEO J: Oxygen-sensitive the expression of the platelet-derived growth factor-B chain gene in calcium channels in vascular smooth muscle and their possible role in human endothclial cells. J Clin Invest 86:670—674, 1990 hypoxic arterial relaxation. Proc NatI Acad Sci USA 92:4715—4719, 7. CZYZYK-KRESKAMZ,DOMINSKI Z, KOLE R, MII.LHORN DE: Hypoxia 1995 stimulates the binding of a cytoplasmic protein to a pyrimidinc-rich 31. FRANCO-OBREGON A, LOPEZ-BARNEO J: Low PU2 inhibits Ca2 sequence in the 3'-untranslatcd region of rat tyrosine hydroxylase channel activity in arterial smooth muscle cells. Am I Physiol (in press) mRNA. J Biol Chem 269:9940—9945, 1994 32. HARDER DR, MADDEN JA, DAWSON C: A membrane electrical 8. GOLDBERG MA, SCHNEIDERTJ:Similarities between the oxygen- mechanism for hypoxic vasoconstriction of small pulmonary arteries sensing mechanism regulating the expression of vascular endothelial from cat. Chest 88:233S—235S, 1985 growth factor and erythropoietin. I Biol (hem 269:4355—4359, 1994 33. MADDEN JA, VADUI.A MS, KURUI' VP: Effects of hypoxia and other López-Barneo et al: Oxygen sensing by ion channels 461

vasoactive agents on pulmonary and cerebral artery smooth muscle38. GANFORNINA D, LOPEZ-BARNEO J: Potassium channel types in arterial cells. Am J Physiol 263:L384—L393, 1992 chemoreceptor cells and their selective modulation by oxygen. J Gen 34. POST JM, HUME JR, ARCHER SL, WEIR EK: Direct role for potassium Physiol 100:401—426, 1992 channel inhibition in hypoxic pulmonary vasoconstriction. Am J 39. JIANGC,HADDADGH:A direct mechanism for sensing low oxygen Physiol 262:C882—C890, 1992 levels in central neurons. Proc NatlAcad Sci USA 91:7198—7201, 1994 35.YUANX,GOLDMANWF, T0D ML, RUBINLi,BLAIJSTEIN MP: 40. GANFORNINA D, LOPEZ-BARNEO J: Gating of 02-sensitive K chan- Hypoxia reduces potassium currents in rat pulmonary but not mesen- nels of arterial chemoreceptor cells and kinetic modifications induced teric arterial myocytes. Am J Physiol 264:L116—L123, 1993 by low P02. J Gen Physiol 100:427—455, 1992 36. ARCHER SL, HUANG JMC, REEVE HL, HAMPL V, TOLAROVA S, 41. KARLIN KD: Metalloenzymes, structural motifs, and inorganic models. MICHELAKIS E, WEIR K: Differential distribution of electrophysi- ologically distinct myocytes in conduit and resistance arteries Science 261:701—708, 1993 determines their response to nitric oxide and hypoxia. Cir Res 42. CRoss AR, HENDERSON L, JONES OTG, DELPIANO MA, HENTSCHEL J, 78:431—442, 1996 ACKER H: Involvement of an NAD(P)H oxidase as a P02 sensor 37. FRANCO-OBREGONA,LOPEZ-BARNEO J: Differential oxygen sensi- protein in the rat carotid body. Biochem J 272:743—747, 1990 tivity of calcium channels in rabbit smooth muscle cells of conduit 43. HENDERSON LM, BANTING G, CHAPPELL JB: The arachidonate acti- and resistance pulmonary arteries. J Physiol (Lond) 491:511—518, vable NADPH oxidase-associated H channels. J Biol Chem 270: 1996 5905—5916, 1995