Ionic currents in dispersed chemoreceptor cells of the mammalian carotid body

Ionic Cu en s in Dispe sed Chemo ecep o Cells o he Mammalian Ca o id Body j. UREI~/A, J. LOPEZ-L6PEZ, C. GONZALEZ, and j. LOPEZ-BARNEO F om he Depa amen o de Fisiologla y Bio isica, Facul ad de Medicina, Uni e sidad de Se illa, 41009 Se illa, Spain ABSTRACT Ionic cu en s o enzyma ically dispe sed ype I and ype II cells o he ca o id body ha e been s udied using he whole cell a ian o he pa ch-clamp echnique. Type II cells only ha e a iny, slowly ac i a ing ou wa d po assium cu - en . By con as , in e e y ype I chemo ecep o cell s udied we ound (a) sodium, (b) calcium, and (c) po assium cu en s. (a) The sodium cu en has a as ac i a ion ime cou se and an ac i a ion h eshold a ~-40 mY. A all ol ages inac i a ion ollows a single exponen ial ime cou se. The ime cons an o inac i a ion is 0.67 ms a 0 mV. Hal s eady s a e inac i a ion occu s a a memb ane po en ial o ~- 50 mV. (b) The calcium cu en is almos o ally abolished when mos o he ex e nal calcium is eplaced by magnesium. The ac i a ion h eshold o his cu - en is a ~-40 mV and a 0 mV i eaches a peak ampli ude in 6-8 ms. The calcium cu en inac i a es e y slowly and only dec eases o 27% o he maximal alue a he end o 300-ms pulses o 40 mV. The calcium cu en was abou wo imes la ge when ba ium ions we e used as cha ge ca ie s ins ead o calcium ions. Ba ium ions also shi ed 15-20 mV owa d nega i e ol ages he conduc ance s. ol age cu e. Deac i a ion kine ics o he calcium cu en ollows a biphasic ime cou se well i ed by he sum o wo exponen ials. A -80 mV he slow com- ponen has a ime cons an o 1.3 _+ 0.4 ms whe eas he as componen , wi h an ampli ude abou 20 imes la ge han he slow componen , has a ime cons an o 0.16 _+ 0.03 ms. These esul s sugges ha ype I cells ha e p edominan ly as deac i a ing calcium channels. The slow componen o he ails may ep esen he ac i i y o a small popula ion o slowly deac i a ing calcium channels, al hough o he possibili ies a e conside ed. (c) Po assium cu en seems o be mainly due o he ac i i y o ol age-dependen po assium channels, bu a small pe cen age o calcium-ac i a ed channels may also exis . This cu en ac i a es slowly, eaches a peak ampli ude in 5-10 ms, and he ea e slowly inac i a es. Inac i a ion is almos comple e in 250-300 ms. The po assium cu en is e e sibly blocked by e ae h- ylammonium. Unde cu en -clamp condi ions ype I cells can spon aneously i e la ge ac ion po en ials. These esul s indica e ha ype I cells a e exci able and Add ess ep in eques s o D . J. L6pez-Bameo, Depa amen o de Fisioiogla y Bio isica, Facul ad de Medicina, A enida de S/mchez Pizju~.n, 4, 41009 Se illa, Spain. D . J L6pez-L6pez and D . C. Gonz~lez' pe manen add ess is Depa amen o de Bioquimica y Biologia Molecula y Fisiologla, Facui ad de Medicina, Uni e sidad de Valladolid, C/Ram6n y Cajal 5, 47005 Valladolid, Spain. J. GgN. PHYSIOL. ~ The Rocke elle Uni e si y P ess 9 0022-1295/89/05/0979/21 $2.00 Volume 93 May 1989 979-999 979 on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 980 THE JOURNAL OF GENERAL PHYSIOLOGY" 9 VOLUME 93 9 1989 ha e a a ie y o ionic conduc ances. We sugges a possible pa icipa ion o hese conduc ances in chemo ecep ion. INTRODUCTION The mammalian ca o id bodies a e pai ed o gans ha sense he le el o oxygen en- sion (pO~) in a e ial blood. This senso y in o ma ion is sen o he cen al ne ous sys em whe e i induces an adequa e en ila o y esponse (De Cas o, 1928; Hey- mans e al., 1930; Fi zge ald and Lahi i, 1986). Among he di e en s uc u es o he ecep o complex (glomus o ype I cells, sub en acula o ype II cells, and ne e endings) ype I cells, he mos nume ous in he ca o id body, a e p esumably he p ima y chemo ecep o s. Type I cells a e ich in sec e o y g anules con aining dopamine, no epineph ine, and o he neu o ansmi e s and hey es ablish chemi- cal synapses wi h ne e endings. A dec ease in pO~ ul ima ely esul s in ansmi e elease by ype I cells and exci a ion o he a e en ibe s o he ca o id sinus ne e (see o e iews Eyzagui e and Zapa a, 1968; Fidone and Gonz~lez, 1986). The mechanism in ol ed in he ansduc ion o he hypoxic s imulus is unknown. I has been ecen ly shown ha ei he low pO~ o high ex e nal po assium can induce he elease o dopamine om ype I cells, which is dependen on ex e nal calcium and is inhibi ed by calcium channel blocke s (Fidone e al., 1982; Obeso e al., 1987). These da a sugges ha , in conco dance wi h he mechanism o s imulus- sec e ion coupling in many sec e o y sys ems, memb ane depola iza ion may play a pa in he esponse o ype I cells o hypoxia. The p esen esea ch wo k was unde aken o cha ac e ize he elec ical p ope - ies o he cellula elemen s o he ca o id body and o es he hypo hesis ha memb ane ionic conduc ances could be al e ed by changes in pO~. F om an elec o- physiological iewpoin ype I cells a e p ac ically unknown. Some p e ious elec o- physiological s udies ha e been pe o med using in aceUula eco ding mic oelec- odes, bu hey a e somewha con using and inconclusi e because hey we e p oba- bly done on cells damaged by he mic oelec ode impalemen . I has been epo ed ha ype I cells a e nonexci able and hei elec ical pa ame e s a e unal e ed by hypoxia (Eyzagui e e al., 1983; Acke and Pie uschka, 1977). These expe imen s we e, howe e , done on cells wi h an a e age es ing po en ial (-20 mV) ha is low enough o p oduce comple e inac i a ion o he ol age-dependen memb ane ionic conduc ances. We ha e pe o med expe imen s in acu ely dispe sed cells o he ca o id body subjec ed o whole-cell pa ch clamp, which is a echnique ha pe mi s accu a e elec- ical measu emen s in small cells (Hamill e al., 1981). Ou da a demons a e ha ype I cells ha e a a ie y o ol age-dependen ionic channels and ha hey a e able o gene a e la ge sodium- and calcium-dependen ac ion po en ials. Type II cells a e unexci able and on depola iza ion only gene a e a small ou wa d cu en . This a icle concen a es on he ull desc ip ion o he sodium, calcium, and po assium cu en s o ype I cells. The ollowing a icle illus a es ha in hese cells po assium channel ac i i y is e e sibly dec eased by hypoxia, which may be he memb ane mechanism esponsible o chemo ansduc ion. Some o his wo k has al eady appea ed in a sho epo (L6pez-Ba neo e al., 1988). on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 URENA ~ AL. Ionic Cu en s o Glomus Cells 981 METHODS Cell Dissocia ion and Cul u e Expe imen s we e pe o med on cells dissocia ed om abbi ca o id bodies. The p o ocol used o cell dissocia ion was wo ked ou a he Uni e si y o Valladolid in collabo a ion wi h D . Beni o He e os. Animals we e anes he ized and he whole egion o he ca o id a e y bi u ca ion was emo ed. Fou ca o id bodies we e dissec ed unde a mic oscope, cleaned, and hen each one o hem was cu in o wo o h ee small pieces. The issue was placed in a ial wi h 2 ml o a Ca ~+- and Mg~+- ee Ty ode solu ion o he ollowing composi ion, in millimola : 140 NaCI, 4.7 KCi, 5 Na py u a e, 3 suc ose, 5 glucose, and 10 HEPES. This solu ion also con ained ypsin (2 mg/ml), collagenase (2 mg/ml), and DNase (0.5 mg/ml). E e y 10 min he issue was i u a ed. A e 20 min a 37~ he ial was cen i uged a 800 g o 5 ain. The pelle was esuspended in ano he 2 ml o he same solu ion wi h collagenase (4 mg/ml), DNase (0.5 mg/ml), and albumin (5 mg/ml), and kep a 37~ o 20 min. The issue was i u a ed e e y 10 min. A he end o he incuba ion pe iod he p epa a ion was washed wice wi h he Ca ~+ and Mg2+- ee solu ion o emo e he enzymes, and he inal pelle was esuspended in 5 ml o minimum essen ial medium supplemen ed wi h giu amine (1%), penicillin-s ep omicin (2%), and e al cal se um (5%). Cells we e pla ed on sli e s o glass co e slips ea ed wi h poly-/-lysine and kep in a CO~ incuba o a 37~ un il use (4-48 h a e pla ing). A e he cells we e dissocia ed, mos o hem had a ound shape and wo dis inc popula ions could be easily dis inguished. Type I cells had a diame e be ween 9 and 13 #m and a ypical bi e in- gence appea ance unde he ligh mic oscope. A second popula ion o cells, classi ied as pu a i e ype II cells, had a smalle and mo e uni o m diame e (be ween 5 and 7 um). These wo cell ypes also had clea di e ences in hei elec ical p ope ies (see Resul s). A e 48 h in cul u e many cells began o show g owing p ocesses and acqui ed a bipola o an i egula shape. Solu ions Du ing he expe imen s a co e slip was ans e ed o a small chambe ha had a con inuous low o solu ion ha could be changed in ~15-20 s. The composi ion o he eco ding solu- ions is shown in Table I. In he ex and igu e legends solu ions a e indica ed as ex e nal/ in e nal. TI'X (500-1,000 nM) was added o he ex e nal solu ion o block Na channels and in mos expe imen s 3 mM Mg-ATP we e added o he in e nal solu ion o e a d wash-ou o Ca channels (Kos yuk, 1984; Fo scbe and Ox o d, 1985; Co a 1986). In some expe imen s designed o eco d K cu en s, he in e nal solu ion con ained a known concen a ion o ee Ca 2+, which was ob ained using Ca-EGTA bu e s. The concen a ions o Ca and EGTA used in hese expe imen s a e indica ed in he igu e legends. Unless o he wise no ed, he pH o he ex e nal and in e nal solu ions we e adjus ed o 7.4 and 7.2-7.3, espec i ely. Expe i- men s we e pe o med a oom empe a u e (20-25~ Reco ding Techniques Ionic cu en s o ype I cells we e eco ded using he whole-cell a ian o he pa ch-clamp echnique (Hamill e al., 1981). Pa ch elec odes we e usually ab ica ed om so hema oc i capilla ies (Hi schmann, Fede al Republic o Ge many) bu in se e al expe imen s bo osili- ca e glass (Kimax 51) was also used. In ou expe imen al condi ions, cu en eco ded wi h bo h ypes o elec odes we e indis inguishable. The elec ode ip was i e polished on a mic o o ge and elec ode esis ance a ied be ween 1 and 2 M L The pa ch-clamp ampli ie used in mos expe imen s was buil by us ollowing he s anda d design (Sigwo h, 1983; Ma - on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 982 THE JOURNAL OF GENERAL PHYSIOLOGY 9 VOLUME 93 9 1989 eson and A ms ong, 1984; A ms ong and L6pez-Ba neo, 1987). The cu en - o- ol age con e e was a Bu -B own OPA 111 (Tucson, AZ). F equency esponse was imp o ed by using low esis ance elec odes, elec onic compensa ion o se ies esis ance, and a ela i ely low eedback esis ance (100 M l). Cu en -clamp eco dings we e done in some cells using a Lis pa ch-clamp ampli ie (model LPC-7; Adams and Lis Associa es, L d., G ea Neck, NY). Da a Acquisi ion and Analysis An IBM-PC/AT compu e in e aced o he analog elec onics was used o pulse gene a ion and o acquisi ion, display, s o age, and analysis o he da a. The in e ace, ab ica ed in ou labo a o y, is buil on wo IBM p o o ype ca ds di ec ly connec ed o he expansion bus o he compu e (U e ia e ai., 1989). The pulse gene a o has an 8 bi digi al- o-analog con- e e (DCA 0808; Analog De ices, Inc., No wood, MA) and a p og ammable pe iphe al in e ace (PPI 8255A-5; NEC Mic ocompu e s, Inc.) wi h he po s p og ammed in ou pu TABLE I Composi ion o Solu ions Ex e nal NaC1 KCI CaCI~ MgCI2 BaCI~ CdCI~ TEACI HEPES 140 Na 140 2.7 2.5-10 .... 10 140 Na, 10 Ba 140 2.7 -- -- 10 -- -- 10 140 Na, 9 Mg 140 2.7 1 9 -- -- -- 10 140 Na, 0.5 Cd 140 2.7 1.5 -- -- 0.5 -- 10 100 Na, TEA 100 2.7 5 -- -- -- 40 10 In e nal NaCI KCI K-glu ama e KF CsCi CsFI MgCI~ HEPES EGTA 130 K -- 30 80 20 -- -- 2 10 5-10 130 Cs .... 110 20 2 10 5-10 90 Cs, 40 Na 40 -- -- -- 70 20 2 10 5-10 All alues a e gi en in millimola . mode (Pea man, 1977; Liu and Gibson, 1986). Cu en signals a e digi ized in a 12 bi ana- log- o-digi al con e e (ADC, AD578; Analog De ices, Inc.). The 12 lines o pa allel ou pu o he ADC a e ans e ed o he compu e RAM memo y by means o a PPI chip wi h he po s p og ammed in inpu mode (Pea man, 1977). The acquisi ion sampling in e al can be changed by he p og am o alues be ween 20 and 500/zs. Digi ized cu en signals a e dis- played in an oscilloscope a e being con e ed in o analog o m. In his sys em each cu en ace is de ined by 500 samples. In mos expe imen s linea ionic and capaci y cu en s we e sub ac ed using a P/4 p ocedu e (Bezanilla and A ms ong, 1977). Du ing analysis ail cu - en s we e i ed wi h one o he sum o wo exponen ials using a leas -squa es p ocedu e. RESULTS The da a p esen ed in his pape we e ob ained om o e 150 ype I chemo ecep- o cells subjec ed o ol age clamp. These cells had an a e age capaci ance o 7.27 +_ 2.26 pF (mean + SD, n = 50). The elec ical p ope ies o ype II cells, which gene a e only a iny ou wa d cu en , a e p esen ed a he end o he pape . on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 URENA ET AL. lO iC Cu en s o Glomus Cells 983 Ionic Cu en Componen s The majo componen s o ionic cu en eco ded in ype I cells a e shown in Fig. 1. Wi h he s anda d high K in e nal solu ion, depola iza ion o 30 mV om a holding po en ial o -70 mV elici ed a as inwa d cu en ollowed by a slowe ou wa d cu en (Fig. 1 A). A he epola iza ion o he 8-ms pulse he cu en changes sud- denly in di ec ion and a la ge inwa d ail cu en was eco ded. This esponse pa - e n was obse ed in e e y cell s udied al hough he ampli ude o he di e en cu - en componen s a ied om cell o cell. The ou wa d cu en was mainly ca ied by K ions since i disappea ed when all he K in he pipe e solu ion was eplaced by Cs. In his condi ion (Fig. 1 B) a as and sus ained inwa d cu en , ollowed by an inwa d ail, we e eco ded by a depo- A 140 Na.2.5Ca//130 K J / / .[ "i 140 Na,10Ca//130 Cs ! 140 Na,10 Ca,TTX//130 Cs ; ,i 1 2 ms -i nA 0.5 nA nA FIGURE 1. Majo cu en compo- nen s in ype I cells. (A) Cu en eco ded du ing a ol age s ep o 30 mV wi h e u n o he holding po en- ial (HP) o -70 mV a e 8 ms. (B) Blockade o he ou wa d po assium cu en by in e nal cesium. Pulse o 20 mV and HP o -80 inV. Calcium cu en is eco ded in isola ion by applica ion o he same ol age s ep in he p esence o TTX (C). Solu ion composi ion was as indica ed nex o each ace. Expe imen s EN1988J (A) and EN2188J (B and C). la iza ion o 20 mV om a holding po en ial o -80 inV. The addi ion o TTX o he ex e nal solu ion (Fig. 1 C) abolished he as componen o he inwa d cu en , which indica es ha i was due o he ac i i y o Na channels. Howe e , he slowly ac i a ing inwa d cu en and he ail emained unal e ed. These wo las cu en componen s we e he esul o he ac i i y o Ca channels since, as will be shown below, hey disappea ed a e eplacemen o ex e nal Ca by Mg, o when 0.5 mM Cd was added o he ex e nal solu ion. These esul s indica e he exis ence in ype I chemo ecep o cells o ol age-dependen Na, Ca, and K channels. P ope ies o he Sodium Cu en Cu en - ol age ela ions. Na cu en s we e eco ded in isola ion in cells whe e in e nal K ions we e eplaced by Cs (o a mix u e o Cs and Na), and Ca channels on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 984 THE JOURNAL OF GENERAL PHYSIOLOGY 9 VOLUME 93 9 1989 we e blocked by ex e nal Mg o Cd. A amily o Na cu en eco ds ob ained by depola iza ions o he indica ed memb ane po en ials is shown in Fig. 2 A. The hold- ing po en ial was -80 mV and he in e nal solu ion con ained 40 mM Na. Na cu - en had a ol age-dependen as ac i a ion ime cou se, a 0 mV i eached a peak in ~0.6-0.7 ms, and hen i inac i a ed comple ely. Cu en was inwa d a mem- b ane po en ials mo e nega i e ha l ENa and ou wa d wi h mo e posi i e memb ane po en ials. The cu en - ol age ela ion o his expe imen is shown in Fig. 2 B, whe e he peak cu en ampli ude is plo ed as a unc ion o he pulse memb ane po en ial (VM). Ac i a ion h eshold was ~- 40 mV, he peak o he I-V cu e occu ed a a VM A : . -40 o 60 nV 9 '-. / Io.s ~A 1ms -6O 1 -40 -20 FIGURE 2. Family o sodium cu - IM'nA ,/ en s eco ded du ing depola iza- -0.s / ions o -40, -20, 0, 20, 40, and 60 /Sp mV om a HP o -80 mV (A). Peak cu en ampli ude as a unc ion o -0.4 he memb ane po en ial is plo ed in B. The con inuous line was i ed by -0.2 eye. Solu ions (in millimola ): 140 20 Na, 0.5 Cd//90 Cs, and 40 Na. Expe imen MZ0288M. --0.2 o/ VM' nV 5/ o 0 o + 10 mV, and he e e sal po en ial was a +32 mV, which was only 1 mV apa om he ENa alue p edic ed by he Ne ns equa ion. Al hough inwa d Na cu en s we e obse ed in e e y cell s udied he ampli ude a ied om cell o cell. The maximal Na cu en ampli ude eco ded in cells wi hou in e nal Na was 0.46 _+ 0.2 nA (mean -+ SD, n = 6). Inac i a ion. In ype I cells inac i a ion o he Na cu en was, as in o he p ep- a a ions, a ol age-dependen p ocess. Fig. 3 A illus a es he ol age-dependence o s eady-s a e inac i a ion by measu ing he peak Na cu en elici ed by a depola iza- ion o 10 mV as a unc ion o V~ du ing a 50-ms condi ioning p epulse. In he o dina e, cu en ampli ude is no malized wi h espec o i s alue in he absence o on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 URENA ET AL. IOniC Cu aUs o Glomus Cells 985 p epulse. Hal s eady-s a e inac i a ion was a ~-50 mV, and a -25 mV he non- inac i a ed ac ion o IN, was < 10%. The inse in Fig. 2 B illus a es ha a 0 mV inac i a ion was comple e in <6 ms and ha i s ime cou se could be well i ed by a single exponen ial wi h a ime cons an o 0.67 ms. The plo in he same igu e shows ha he ime cons an o inac i a ion dec eased wi h memb ane depola iza- ion. Closing kine ics. The closing o Na channels was s udied in a ew cells by he applica ion o sho depola izing pulses (0.5-1 ms), which p e en ed he de elop- men o inac i a ion, and he eco ding o ail cu en s a he ins an o epola iza- ion. Na ails we e well i ed by single exponen ials and a -80 mV he ime con- A B LK "O O Z 0.5 VM,mV ' InA ; 2'0 VM,mV FIGURE 3. (A) S eady-s a e inac i a ion o sodium cu en s. The plo ep esen s no malized peak cu en ampli ude ob ained du ing a es s ep depola iza ion o 10 mV as a unc ion o he memb ane po en ial du ing a 50-ms condi ioning p epulse. (B) Time cou se o inac i a- ion. The inse is a sodium cu en eco ded by a ol age s ep o 0 inV. Inac i a ion is well i ed by an exponen ial wi h a ime cons an o 0.67 ms. The plo ep esen s he ime con- s an o inac i a ion (o dina e) measu ed in cu en s eco ded a a ious memb ane po en- ials (abscissa). Lines we e i ed by eye. HP, -80 mV. Solu ions (in millimola ): 140 Na and 0.5 Cd//130 Cs. Expe imen s EN2888J (A) and FE0388J (B). s an was be ween 60 and 70 #s. Thus, hese esul s indica e ha ype I chemo ecep- o cells ha e a p ominen Na cu en wi h p ope ies simila o hose ound in o he elec ically exci able cells. P ope ies o he Calcium Cu en Iden i ica ion o he cu en . Ca cu en s we e eco ded in cells dialyzed wi h he 130 Cs solu ion. Fig. 4 A shows a ace o inwa d cu en in his expe imen al con- di ion and wi h 10 mM Ca in he ex e nal solu ion. The pulse cu en was ollowed by a la ge inwa d all cu en . Exposu e o a di e en ex e nal solu ion wi h less Ca (9 mM Mg, 1 mM Ca) p oduced a ma ked educ ion o he s eady-s a e pulse cu - en and he ail, al hough a small componen o inac i a ing inwa d Na cu en on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 986 THE JOURNAL OF GENERAL PHYSIOLOGY. VOLUME 93 9 1989 FIGURE 4. Iden i ica mn o B he calcium cu en . (A) 140 Na//130 Cs 140 Na,TTX//130 Cs Inwa d cu en eco ded wi h ~,~ /~ ~~,.~a,z,~,,,~'~ ~ ~.9 ~,lCa he 140 mM Na, 10 mM Ca 9 Mg.1 Ca = ;;( ,, --~:-:: ex e nal solu ion du ing a ol - " :: ~~i" age s ep o 0 mV. On eplacing / . / calcium wi h magnesium (9 I 10 Ca : 10 Ca ~ 0.SnA 9 mM Mg, 1 mM Ca) he s eady ; inwa d cu en and he ail 2ms almos disappea bu a small componen o ansien sodium cu en emains. In he p esence o TFX (B) he isola ed calcium cu en eco ded in 10 mM Ca is almos abolished a e he in oduc ion o he 9 mM Mg, 1 mM Ca solu ion in he ba h. HP, -80 mV. Solu ions we e as indica ed in he igu e. Expe imen s EN2788J (A) and EN2588J (B). emained. A e Na channels we e blocked wi h TIX in ano he cell (Fig. 4 B), Ca cu en s could be eco ded in o al isola ion. This cu en ac i a es slowly and a 0 mV i eaches a maximum in ~8 ms. Fig. 4 B also illus a es he almos comple e disappea ance o he pulse cu en and he ail a e eplacemen o ex e nal Ca by Mg. In o he expe imen s wi h low ex e nal Ca (1.5 mM) he addi ion o 0.5 mM Cd o he ex e nal solu ion comple ely abolished he Ca cu en (see Fig. 2). Cu en - ol age ela ions. Fig. 5 (]e column, 10 Ca) shows calcium cu en s eco ded a a ious memb ane po en ials. Cu en gene a ed du ing he pulse, indi- ca ed be ween he a ows, ac i a es mo e apidly a mo e depola ized le els and is ollowed by a as ail whose ampli ude inc eases wi h he ampli ude o he depola - iza ion. These ails a e due o he low o Ca ions h ough he channels ha we e open du ing each pulse, and hei ime cou se e lec s ha o he closing o he Ca channels. Cu en ampli ude measu ed a he end o he pulse as a unc ion o pulse VM'mV 10 Ca -20 .... - ..... ~ F "! ":.: 13 nA 3ms lO Ba i/ ! i ./ 2 / i ? . . -. ~ - / .j FIGURE 5. Cu en h ough calcium channels ca ied by cal- cium and ba ium ions. Cu en s a e he esul o depola iza ions o he memb ane po en ials indi- ca ed nex o each ace wi h 10 mM Ca o 10 mM Ba in he ex e nal solu ion. The a ows indica e he onse and he end o he ol age s eps. All aces a e om he same cell. HP, -80 inV. Solu ions (in millimola ): 140 Na, 10 Ca (10 Ba), and T X//130 Cs. Expe imen MZ0288K. on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 URENA ET AL. Ionic Cu en s o Glomus Cells -60 -40 -20 0.4 0.6- 0.8- VM,mV 20 40 60 1 I , I IM,nA 987 FmURE 6. Calcium cu en - ol age ela ion. The cu en measu ed a he end o 8-ms pulses is plo ed as a unc ion o he pulse memb ane po en ial. The line was i ed by eye. HP, -80 mV. Solu ions (in millimo- la ): 140 Na, l0 Ca, and T X//130 Cs. Expe imen MZ0288K. memb ane po en ial is plo ed in Fig. 6. This I-V plo shows ha he ac i a ion h eshold was a --40 mV and ha he maximal inwa d cu en was ob ained a + ] 0 o + 20 mV. A hese I'M alues he a e age ampli ude o he Ca cu en mea- su ed wi h 10 mM ex e nal Ca was 0.4 .+ 0.2 nA (mean _+ SD, n = 8). The igh column o Fig. 5 shows eco dings ob ained in he same cell a e ex e - nal Ca was eplaced by Ba. The aces illus a e ha Ba ions lowed h ough Ca channels be e han Ca ions, as indica ed by he la ge size o he ails. In he ange be ween 0 and + 20 mV, o al Ba conduc ance was abou wo imes la ge han Ca conduc ance. Ba ions also a ec ed he conduc ance- ol age ela ion o Ca channels. Fig. 7 plo s he no malized ampli ude o ail cu en s eco ded wi h 10 mM Ca (do s) and 10 mMBa (squa es). A compa ison o bo h cu es indica es ha ex e nal Ba shi ed he ac i a ion cu e 15-20 mV in he nega i e di ec ion. Closing kine ics. I has been shown in p e ious igu es ha a he ins an o epola iza ion Ca cu en s we e ollowed by la ge ail cu en s. Examples o ails eco ded on epola iza ion o -80 mV a e a pulse o + 40 mV a e shown in Fig. 8 using 10 mM ex e nal Ca (A) o Ba (C) as cha ge ca ie s. The decay o he ail cu en had a clea biphasic ime cou se wi h a la ge as componen and a small slow componen . The slow componen was i ed by an exponen ial ex apola ed o @ N m E Z c g "0 c 0 0.5- -~o -go -2~ FIGURE 7. Conduc ance- ol age e- la ion o calcium channels wi h cal- cium and ba ium as cha ge ca ie s. Conduc ance, ep esen ed in he o dina e, was measu ed om he ampli ude o ail cu en s eco ded a he ins an o epola iza ion o 10- ms pulses. The pulse memb ane po en ial is ep esen ed on he abscissa. Reco dings a e om he same cell ba hed in 10 mM Ca (do s) and 10 mM Ba (squa es). Lines we e d awn by eye. HP, -80 mV. Solu- 2~0 4J0 6~ ions (in millimola ): 140 Na, l0 Ca VM,m (10 Ba), and TI'X//130 Cs. Expe i- men MZ0288K. on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 994 THE JOURNAL OF GENERAL PHYSIOLOGY 9 VOLUME 98 9 1989 a a ions ( o example Sah e al., 1988). The cu en has a as ac i a ion and inac i- a ion, is comple ely blocked by nanomola concen a ions o TTX, and is selec i ely ca ied by Na ions. Ac i a ion o 1Na in ype I cells is simila o he Na cu en eco ded in GHs cells (Ma eson and A ms ong, 1984) bu is somewha slowe han he cu en om he squid axon (Hodgkin and Huxley, 1952; Bezanilla and A m- s ong, 1977). The peak o he cu en - ol age cu e occu ed a 10 mV, which is simila o Na cu en s eco ded in GHs (Dubinsky and Ox o d, 1984; Ma eson and A ms ong, 1984) and ch oma in (Fenwick e al., 1982) cells, bu i is displaced owa d posi i e ol ages i compa ed wi h Na cu en s eco ded in neu ons (Sah e al., 1988) and neu oblas oma cells (Moolenaa and Spec o , 1978). These quan i a- i e di e ences we e also seen in he ol age-dependence o s eady-s a e inac i a- ion. In ou expe imen s hal o he Na channels we e inac i a ed a ~ -50 mV, a alue simila o ha measu ed in GH3 cells (Dubinsky and Ox o d, 1984; Ma eson and A ms ong, 1984), bu less nega i e han he one obse ed in cen al neu ons (-75 mV, Sahe al., 1988). Inac i a ion o Na cu en s om ype I cells ollows a single exponen ial ime cou se and is ela i ely as compa ed wi h ac i a ion. A sin- gle exponen ial decay has also been obse ed in GH3 cells (Vandenbe g and Ho n, 1984) al hough a biexponen ial ime cou se was equi ed o i Na inac i a ion in a and og ne e (Chiu, 1977; Neumcke and S amp li, 1982). Na cu en densi y in ype I cells, assuming ha he e is no memb ane in olding, was 0.15 mA/cm ~. This alue is wi hin he ange o he es ima ions done in GHs cells (Dubinsky and Ox o d, 1984; Ma eson and A ms ong, 1984) bu i is abou 10 imes smalle han in he squid axon. Thus, on quan i a i e g ounds he Na cu en o ype I cells esembles he INa o GH s and ch oma in cells, bu di e s in some aspec s om he cu en exis ing in neu ons and ne e cells. Despi e he exis ence o a ela i ely low densi y o Na channels, ype I cells, like o he sec e o y cells, can gene a e la ge ac ion po en ials. The ac ha in glomus cells s eady-s a e inac i a ion o IN, is displaced in he posi i e di ec ion may con ib- u e o a mo e e icien use o he Na channels a ailable (see also Ma eson and A m- s ong, 1984). Res ing po en ials measu ed in p e ious in acellula eco dings pe - o med in ype I cells we e in he neighbo hood o - 20 mV (Acke and Pie uschka, 1977; Eyzagui e e al., 1983). Ou esul s show ha >90% o gs, is inac i a ed a -25 mV, which explains he unexci abili y o he in acellula ly eco ded cells, which we e p obably damaged by he mic oelec odes. Calcium cu en . Blockade o Na channels by TTX e ealed in all ype I cells he exis ence o a Ca cu en . This cu en esembles/ca om o he e eb a e p epa a- ions in i s ime cou se, i s sensi i i y o ex e nal di alen ca ion block, and he abili y o Ba o subs i u e o Ca as cha ge ca ie (Fenwick e al., 1982; Hagiwa a and Ohmo i, 1982; Dubinsky and Ox o d, 1984; Ma eson and A ms ong, 1984). The peak o he cu en - ol age cu e was a 10-20 mV, which is close o he alues obse ed in o he cells. In equi alen expe imen al condi ions, he ampli ude o he Ca cu en is somewha la ge in ype I cells han in GHs (Hagiwa a and Ohmo i, 1982; Dubinsky and Ox o d, 1984; Ma eson and A ms ong, 1984), ch oma in (Fenwick e al., 1982), adenohypophysial (Co a, 1986), and panc ea ic be a (Hi ia and Ma eson, 1988) cells. Wi h 10 mM ex e nal Ca, he peak Ca cu en ampli ude was almos he same as he maximal Na cu en . on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 URENA ET AL. [Ol' ic Cu en s o Glomus Cells 995 Indica ions o he exis ence o mo e han one Ca channel ype ha e been obse ed in a numbe o sec e o y cells (Co a, 1986; Ma eson and A ms ong, 1986; Hi ia and Ma eson, 1988; Taba es e al., 1989), neu ons (Ca bone and Lux, 1984; Nowycky e al., 1985), and muscle (Bean, 1985). In hypophysial pa s in e media, panc ea ic be a cells, and ad enoco ical cells, FD and SD Ca channels a e dis inguished acco ding o hei di e en closing kine ics. FD channels also ha e a highe h eshold and inac i a e mo e slowly han SD channels. Ca ails in mos ype I cells ollow a clea biphasic ime cou se and a e well i ed by he sum o a small slow exponen ial and a as la ge exponen ial. The ime cons an s o he wo componen s a e compa able o hose measu ed in o he p epa a ions (Co a, 1986; Ma eson and A ms ong, 1986). Al hough he analysis o he deac i a ion kine ics o Ca cu en s sugges s he exis- ence in ype I cells o wo Ca channel ypes, a numbe o expe imen al obse a ions do no suppo his in e p e a ion. (a) Ca cu en o ype I cells inac i a e e y slowly wi h a ime cou se ha esembles ha o FD channels om o he p epa a- ions. (b) The ime cou se o ail cu en s eco ded a e sho and la ge depola iza- ions is simila and is no a ec ed by a 500-ms condi ioning p epulse. In o he cells he slow componen o he ails, which ep esen s he closing o SD channels, is ma kedly educed a e long (>50 ms) depola iza ions o by a condi ioning depola - izing p epulse. (c) The conduc ance- ol age ela ion wi h Ba ions as cha ge ca ie s is displaced 15-20 mV owa d nega i e ol ages. Ba also slows down he closing o he channels. These e ec s o Ba a e known o be media ed by hei in e ac ion wi h FD channels. Finally, (d) in he absence o exogenous in e nal Mg-ATP he Ca cu - en o giomus cells is e y labile and disappea s almos comple ely in 5-8 ain. This p ope y is ypical o FD channels whe eas SD channels seem o be esis an o dilu- ion o cy osolic componen s (Co a, 1986; Ma eson and A ms ong, 1986). All oge he his e idence indica es ha Ca cu en in ype I cells is mainly media ed by FD channels. A small popula ion o SD channels, esponsible o he slow componen o he ails, may exis bu unequi ocal p oo s o hei exis ence we e no ound. The possibili y s ill emains ha he slow componen o he ails is ei he a esul o he ac i i y o Ca channels ha a e di e en om SD channels, o ha i ep esen s a pa icula kine ic p ope y o FD channels in ype I cells. Po assium cu en . An ou wa d K cu en was eco ded in all ype I cells s ud- ied. This cu en was eco ded wi h a la ge ampli ude in cells wi h in e nal Ca con- cen a ions anging be ween 0.5 #M and <10 -1~ M, bu in a gi en cell i dec eased in ampli ude a e wash-ou o Ca channels. Thus, a pe cen age o his cu en may be due o he ac i i y o Ca-dependen K channels ac i a ed by he local ise in cy o- solic Ca ha ollows Ca in lux h ough unc ional Ca channels (Ma y and Nehe , 1985). Because o he apid disappea ance o Ca channels in ou p epa a ion, e en in he p esence o 3 mM in e nal Mg-ATP, we cen e ed on he s udy o he ol age- dependen K cu en a e wash-ou o Ca channels. The ol age-dependen K cu en o ype I cells has a ypical sigmoid ac i a ion ime cou se, a +40 mV i eaches hal maximal ampli ude in 3.5 ms, and he ea e i slowly inac i a es. The cu en is quali a i ely simila o delayed K cu en s om o he p epa a ions (Ad ian e al., 1970; Dubinsky and Ox o d, 1984; Ro sman and T ube, 1986; Ma eson and Ca melie , 1988). I is, howe e , slowe han K cu en s on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 996 TH~ JOURNAL OF GENERAL PHYSIOLOGY 9 VOLUME 93 9 1989 o he squid axon o neu ons dispe sed om he squid gian ibe lobe (Hodgkin and Huxley, 1952; A ms ong and L6pez-Ba neo, 1987), bu i s ac i a ion ime cou se is simila o cu en om e eb a e sec e o y cells (Dubinsky and Ox o d, 1984; Ro sman and T ube, 1986). The K cu en in ype I cells inac i a ed almos comple ely in 250 ms. This same beha io is ound in K cu en s om a numbe o p epa a ions including squid neu ons (Llano and Bookman, 1986), GHs cells (Ma - eson and Ca melie , 1988), and skele al muscle ibe s (Ad ian e al., 1970). The ol age-dependen K cu en o ype I cells was, as we e o he K cu en s, almos abolished by a la ge concen a ion o ex e nal TEA. In ou expe imen s he e e si- bili y o he TEA blockade was almos pe ec . Possible Pa icipa ion o Ionic Channels in he Physiology o Type I Cells Al hough ype I cells ha e been conside ed he bes candida es o being he p i- ma y chemo ecep o s since he ea ly wo k on he ca o id body, he basic mecha- nisms in ol ed in his p ocess ha e emained la gely unknown ( o a e iew see Bel- mon e and Gonz~lez, 1983). Recen in es iga ions ha e shown ha bo h hypoxia and high ex e nal po assium, which p esumably causes memb ane depola iza ion, p oduce sec e ion o dopamine in he ca o id body and ha his e ec is abolished by Ca channel an agonis s (Fidone e al., 1982; Alma az e al., 1986; Obeso e al., 1987). Thus, ype I cells may unc ion in a way simila o o he sec e o y sys ems in which Ca in lux h ough ol age-ga ed memb ane Ca channels is a c i ical e en leading o sec e ion. This idea could no be econciled wi h he ac ha ype I cells impaled wi h mic oelec odes we e ound o be unexci able (Eyzagui e e al., 1983), howe e , he esul s shown in his a icle demons a e ha hey ha e an app eciable densi y o ol age-dependen sodium and calcium channels and ha hey can epe i i ely gene a e ac ion po en ials. Na channels ha e, he e o e, an impo an ole in spike gene a ion and by p oducing a as depola iza ion con ib- u es o he opening o Ca channels, which a e well sui ed o as injec ion o Ca in o he cy osol. The Ca-ac i a ed componen o he K cu en may pa icipa e in spike epola iza ion, whe eas he slowly inac i a ing K channels a e mos p obably in ol ed in pacemaking. The indings epo ed he e led us o hypo hesize ha ionic channels in ype I cells migh be egula ed by en i onmen al 02 ension and ha hey migh he e o e be di ec ly implica ed in chemo ansduc ion. This hypo hesis was expe imen ally con- i med as illus a ed in he ollowing a icle. The au ho s wish o hank M s. Lola Gan o nina, Uni e si y o Se ille, o he aluable pa icipa- ion in some expe imen s. This esea ch was pa ially suppo ed by g an s PB86-250 and PB86-325 om Di eccion Gene al de In esdgaci6n Cien i ica y T6cnica. J. R. L6pez-L6pez is a ellow o Fondo de In es igaciones Sani a ias. O iginal e sion ecei ed 25 July 1988 and accep ed e sion ecei ed 2 Decembe 1988. REFERENCES Acke , H., and F. Pie uschka. 1977. Meaning o he ype I cell o he chemo ecep i e p ocess. An elec ophysiological s udy on cul u ed ype I cells o he ca o id body. In Chemo ecep ion in he on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 URENA El' AL. Ionic Cu en s o Glomus Cells 997 Ca o id Body. H. Acke , S. Fidone, D. Pallo , C. Eyzagui e, D. W. L ibbe s, and R. W. To - ance, edi o s. Sp inge -Ve lag, Be lin. 92-98. Ad ian, R. H., W. K. Chandle , and A. L. Hodgkin. 1970. Slow changes in po assium pe meabili y in skele al muscle. Jou ud o Physiology. 208:645-688. Alma az, L., C. Gon~lez, and A. Obeso. 1986. E ec s o high po assium on he elease o 3H- dopamine om he ca ca o id body in i o. Jou na/o Physiology. 379:293-307. A ms ong, C. M., and J. L pez-Ba neo. 1987. Ex e nal calcium ions a e equi ed o po assium channel ga ing in squid neu ons. &/ence. 236:712-714. Bean, B. P. 1985. Two kinds o calcium channels in canine a ial cells. Di e ences in kine ics, selec i i y, and pha macology. Jou nal o Gene al Physiology. 86:1-30. Belmon e, C., and C. Gonz~lez. 1983. Mechanisms o chemo ecep ion in he ca o id body: possible models. In Physiology o he Pe iphe al A e ial Chemo ecep o s. H. Acke and R. G. O'Reagan, edi o s. Else ie Science Publishing Co., Inc., Ams e dam. 197-220. Bezani a, F., and C. M. A ms ong. 1977. Inac i a ion o he sodium channel. I. Sodium cu en expe imen s. Jou nal o Gene al Physiology. 70:549-566. Ca bone, E., and H. D. Lux. 1984. A low ol age-ac i a ed, ully inac i a ing Ca channel in e e- b a e senso y neu ons. Na u e. 310:501-502. Chiu, S. Y. 1977. Inac i a ion o sodium channels: second o de kine ics in myelina ed ne e.Jou - nal o Physiology. 273:573-596. Co a, G. 1986. Calcium channel cu en s in pa s in e media cells o he a pi ui a y gland.Jou nal o Gene al Physiology. 88:83-105. De Cas o, F. 1928. Su la s uc u e e l'inne a ion du sinus ca o idien de l'homme e des mam- mi ~ es. Nou eaux ai s su inne a ion e la onc ion du glomus ca o icum. E udes ana omi- ques e physiologiques. T abajos del Labo a o io de In es igaciones Biolbgicas de la Uni e sidad de Mad /d. 25:331-380. Dubinsky, J. M., and G. S. Ox o d. 1984. Ionic cu en s in wo s ains o a an e io pi ui a y umo cells. Jou nal o Gene al Physiology. 83:309-339. Eyzagui e, C., L. Mon i-Bloch, Y. Hayashida, and M. Ba 6n. 1983. Biophysics o he ca o id body ecep o complex. In Physiology o he Pe iphe al A e ial Chemo ecep o s. H. Acke and R. G. O'Reagan edi o s. Else ie Science Publishing Co p., Inc., Ams e dam. 59-88. Eyzagui e, C., and P. Zapa a. 1968. A discussion o possible ansmi e o gene a o subs ances in ca o id body chemo ecep o s. In A e ial Chemo ecep o s. R. W. To ance edi o . Blackweli Scien i ic Publica ions, Inc., Ox o d. 213-251. Fenwick, E. M., A. Ma y, and E. Nehe . 1982. Sodium and calcium channels in bo ine ch oma in cells. Jou nal o Physiology. 331:599-635. Fidone, S. and C. Gonz~ilez. 1986. Ini ia ion and con ol o chemo ecep o ac i i y in he ca o id body. In Handbook o Physiology. The Respi a o y Sys em II. A. P. Fishman, edi o . Ame ican Physiological Socie y, Be hesda, MD. 247-312. Fidone, S., C. Gonz~lez, and K. Yoshizaki. 1982. E ec s o low oxygen on he elease o dopamine om he abbi ca o id body in i o.Jou nal o Physiology. 333:93-110. Fi zge ald, R. S., and S. Lahi i. 1986. Re lex esponses o chemo ecep o s imula ion. In Hand- book o Physiology. The Respi a o y Sys em II. A. P. Fishman, edi o . Ame ican Physiological Socie y, Be hesda, MD. 313-362. Fo sche , P., and G. S. Ox o d. 1985. Modula ion o calcium channels by no epineph ine in in e - nally dialyzed a ian senso y neu ons. Jou nal o Gene al Physiology. 85:743-763. Hagiwa a, S., and H. Ohmo i. 1982. S udies o calcium channels in a clonal pi ui a y cells. Jou - nal o Physiology. 336:649-661. Hamill, O. P., A. Ma y, E. Nehe , B. Sakmann, and F. S. Sigwo h. 1981. Imp o ed pa ch-clamp on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 998 THE JOURNAL OF GENERAL PHYSIOLOGY - VOLUME 93 9 1989 echniques o high- esolu ion cu en eco ding om cells and cell- ee memb ane pa ches. P liJge s A chi . 391:85-100. Heymans, C., J. J. Bouckae , and L. Dau ebande. 1930. Sinus ca o idien e ~ lexes espi a oi es. II. In luences espi a oi es ~ lexes de acid6se, de I'alcal6se, de i'anhyd ide ca bonique, de I'ion hyd og~ne e de l'anox~mie: sinus ca o idiens e 6changes espi a oi es dans les poumons e au del~ des poumons. A chi es Ime na iouales de Pha macodynamie e de Thk apie. 39:400--408. Hi ia , M., and D. R. Ma eson. 1988. Na channels and wo ypes o Ca channels in a panc ea ic B cells iden i ied wi h he e e se hemoly ic plaque assay. Jou nal o Gene al Physiology. 91:617- 639. Hodgkin, A. L., and A. F. Huxley. 1952. A quan i a i e desc ip ion o memb ane cu en and i s applica ion o conduc ion and exci a ion in ne e. Jou nal o Physiology. 117:500-544. Kondo, H., T. Iwanaga, and T. Nakajima. 1982. Immunocy ocbemical s udy on he localiza ion o neu on speci ic enolase and S-100 p o eins in he ca o id body o a s. Cell and Tissue Resea ch. 227:291-295. Kos yuk, P. G. 1984. Me abolic con ol o ionic channels in he neu onal memb ane. Neu oscience. 13:983-989. Liu, Y., and G. A. Gibson. 1986. Mic ocompu e Sys ems: he 8086/8088 Family. A chi ec u e, P og amming and Design. P en ice Hall, Inc., NJ. Llano, I., and R. S. Bookman. 1986. Ionic conduc ances o squid gian ibe lobe neu ons. Jou nal o Gene al Physiology. 88:543-569. L6pez-Ba neo, J., J. R. L6pez-L6pez, J. U e ia, and C. Gonz{dez. 1988. Chemo ansduc ion in he ca o id body: K cu en modula ed by pO~ in ype I chemo ecep o cells. Sc/ence. 241:580- 582. Ma eson, D. R., and C. M. A ms ong. 1984. Na and Ca channels in a ans o med line o an e io pi ui a y cells. Jou nal o Gene al Physiology. 83:371-394. Ma eson, D. R., and C. M. A ms ong. 1986. P ope ies o wo ypes o calcium channels in clonal pi ui a y cells. Jou nal o Gene al Physiology. 87:161-182. Ma eson, D. R., and P. Ca melie . 1988. Modi ica ion o K channel inac i a ion by papaln and N-b omoace amide. Biophysical Jou nal. 53:641-645. Ma y, A., and E. Nehe . 1985. Po assium channels in cul u ed bo ine ad enal ch oma in cells. Jou nal o Physiology. 367:117-141. Moolenaa , W. H., and 1. Spec o . 1978. Ionic cu en s in cul u ed mouse neu oblas oma cells unde ol age-clamp condi ions. Jou nal o Physiology. 278:265-286. Neumcke, B., and R. S amp li. 1982. Sodium cu en s and sodium cu en luc ua ions in a my- elina ed ne e ib es.J0u nal o Physiology. 329:163-184. Nowycky, M. C., A. P. Fox, and R. W. Tsien. 1985. Th ee ypes o neu onal calcium channel wi h di e en calcium agonis sensi i i y. Na u e. 316:440--443. Obeso, A., S. Fidone, and C. Gonz ilez. 1987. Pa hways o calcium en y in ype I cells o he ca o id body. Signi icance o he sec e o y esponse. In Chemo ecep o s in Respi a o y Con- ol. J. A. Ribei o and D. Pallo , edi o s. C oom Helm, London. 91-98. Pea man, J. B. 1977. Mic ocompu e -Based Design. McG aw-Hill Book Co., New Yo k. Ro sman, P., and G. T ube. 1986. Calcium and delayed po assium cu en s in mouse panc ea ic B-cells unde ol age-clamp condi ions. Jou nal o Physiology. 374:531-550. Sah, P., A. J. Gibb, and P. W. Gage. 1988. The sodium cu en unde lying ac ion po en ials in guinea pig hippocampal CA1 neu ons.Jou nal o ~l Physiology. 91:373-398. Sigwo h, F.J. 1983. Elec onic design o he pa ch-clamp. In Single Channel Reco ding. B. Sak- mann and E. Nehe , edi o s. Plenum Publislaing Co p., New Yo k. 3-35. on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989 URENA ET AL. Ionic Cu enla o Glomus Cells 999 Taba es, L., J. U e ia, and J. L6pez-Ba neo. 1989. P ope ies o calcium and po assium cu en s o clonal ad enoco ical cells. Jou nal o Gene al Physiology. 93:495-519. U e ia, J., J. c. Ma eos, and J. L6pez-Ba neo. 1989. Low-cos sys em o au oma ed acquisi ion, display, and analysis o ansmemb ane ionic cu en s. Medical and Biological Enginee ing and Compu ing. 27:94--100. Vandenbe g, C. A., and R. Ho n. 1984. Inac i a ion iewed h ough single sodium channels. Jou - nal o C, ene al Physiology. 84:535-564. on Sep embe 22, 2014jgp. up ess.o gDownloaded om Published May 1, 1989