Cardiac Ionic Currents and Acute Ischemia: From Channels to Arrhythmias

101 Pages·1999·1.12 MB·English

PHYSIOLOGICALREVIEWS
Vol.79,No.3,July1999
PrintedinU.S.A.
Cardiac Ionic Currents and Acute Ischemia:
From Channels to Arrhythmias
EDWARD CARMELIET
CentreforExperimentalSurgeryandAnesthesiology,UniversityofLeuven,Leuven,Belgium
I. Introduction 917
A. Aimandgeneraloutline 917
B. Ischemiamodels 918
C. Generalbiochemicalchangesduringischemiaandreperfusion 918
II. IonChannelsandTransporters 920
A. Ionchannelsandtransportersintheplasmamembrane 920
B. Ionchannelsinintracellularorganelles 942
C. Gapjunctionchannels 947
III. IschemiaSyndromes 949
A. Changesinionconcentrations 950
B. Amphiphilesandfattyacids 963
C. Radicals 966
D. Catecholamines 969
E. ExtracellularATP,adenosine,andACh 974
F. Stretchandvolumechanges 977
IV. ElectricalChangesandArrhythmiasDuringIschemiaandUponReperfusion 978
A. Electrophysiologicalchangesatthecellularandmulticellularlevels 978
B. Arrhythmias 982
V. ConcludingRemarksandPerspectives 987
Carmeliet,Edward.CardiacIonicCurrentsandAcuteIschemia:FromChannelstoArrhythmias.Physiol.Rev.79:
917–1017,1999.—Theaimofthisreviewistoprovidebasicinformationontheelectrophysiologicalchangesduring
acuteischemiaandreperfusionfromthelevelofionchannelsuptothelevelofmulticellularpreparations.Afteran
introduction,sectionIIprovidesageneraldescriptionoftheionchannelsandelectrogenictransporterspresentin
theheart,morespecificallyintheplasmamembrane,inintracellularorganellesofthesarcoplasmicreticulumand
mitochondria,andinthegapjunctions.Thedescriptionisrestrictedtoactivationandpermeationcharacterisitics,
whilemodulationisincorporatedinsectionIII.Thissection(ischemicsyndromes)describesthebiochemical(lipids,
radicals,hormones,neurotransmitters,metabolites)andionconcentrationchanges,themechanismsinvolved,and
theeffectonchannelsandcells.SectionIV(electricalchangesandarrhythmias)issubdividedintwoparts,withfirst
adescriptionoftheelectricalchangesatthecellularandmulticellularlevel,followedbyananalysisofarrhythmias
during ischemia and reperfusion. The last short section suggests possible developments in the study of ischemia-
relatedphenomena.
I. INTRODUCTION calpropertiesatthecellularandmulticellularlevelandto
analyzetheir roleinthegenesisof acutecardiacarrhyth-
A. Aim and General Outline mias (sect. IV).
Thereadermainlyinterestedintheelectrophysiological
Theaimofthepresentreviewistoprovideadescrip- changesduringischemiaandarrhythmiasmaydirectlystart
tion of ionic channels and electrogenic transporters withsectionIIIandreturnlater,whennecessaryforabetter
presentintheheart(sect.II),todescribethebiochemical understandingofcertainprocessesandnotions,tosectionII.
andionconcentrationchangesduringacuteischemiaand The reader with a main interest in the biophysics of ion
earlyreperfusionandtheeffectsonchannels(sect.III),to channelsisdirectedfirsttosection II.
translate these findings in terms of modulation of electri- Section II provides a description of the activation-
0031-9333/99$15.00Copyright©1999theAmericanPhysiologicalSociety 917
918 EDWARD CARMELIET Volume79
inactivation kinetics and the permeation processes of farction but does not allow a direct analysis of the
channels and carriers; their role in the genesis of electri- changes in the ionic currents involved. For this pur-
cal activity is briefly discussed in this section. Section II pose, voltage-clamp measurements have been applied
does not include information on modulation or pharma- to multicellular preparations and single cells subjected
cology. It is subdivided in sections describing channels to hypoxia, uncouplers of the mitochondrial oxidative
and transporters in the plasma membrane, in the gap chain, inhibitors of glycolysis, superfusion with a solu-
junction,andinintracellularorganelles(sarcoplasmicre- tioncontainingahighconcentrationofK(cid:49)andH(cid:49),and
ticulumandmitochondrion).Althoughinformationabout deficient in glucose. In other models, the complex na-
eventualchangesduringischemiaislackingforanumber ture of the ischemic process has been dissected in
ofchannelsandtransporters,itishopedthatasystematic different facets of specific biochemical changes that
description and analysis will be helpful to understand occur during ischemia: amphiphiles, radicals, cat-
more fully the actual state of knowledge, to put this echolamines, adenosine, ACh, and stretch.
information into perspective, and eventually to help to
plan adequate experiments.
C. General Biochemical Changes During Ischemia
SectionIIIdescribeshowchannelsandtransporters
and Reperfusion
are modulated. The modulatory processes are not de-
scribed for each channel separately but are grouped
into “syndromes” related to ischemia, such as changes Underaerobicconditions,NADHandFADH ,formed
2
in ion concentration ([K(cid:49)] , [Na(cid:49)], [Ca2(cid:49)], and duringglycolysisandinthecitricacidcycle,transfertheir
o i i
[Mg2(cid:49)], where the subscripts o and i refer to extracel- electronstoO throughtheelectrontransportchain.This
i 2
lularandintracellular,respectively);disturbanceoflip- provides the energy to build up the chemiosmotic gradi-
ids resulting into the accumulation of long-chain acyl- entthatdrivesthesynthesisofATP.Oxidativephosphor-
carnitines, lysophosphoglycerides, fatty acids, and ylation is coupled to the demands of the cell. A feedback
arachidonic acid; production of radicals; secretion of isgeneratedbythebreakdownproductsofATP(772)and
neurotransmitters, hormones, and metabolites, with theriseinmitochondrialCa2(cid:49)thatmodulatestheactivity
concomitant stimulation of adrenergic, purinergic, and of the mitochondrial dehydrogenases (692) and of the
muscarinic receptors; and the genesis of stretch. The ATP synthase (199).
analysisofeachsyndromeincludesadescriptionofthe The turnover rate of high-energy phosphates is
changes, the mechanisms involved, the effect on chan- 30–40 mol(cid:122)g wet wt(cid:50)1(cid:122)min(cid:50)1, while the storage is
nels and transporters, and the final outcome at the only15mol/gwetwt.Thismeansthatthetimelimitfor
electrophysiological and arrhythmia levels. exhaustion is short and only 15–30 s (339). When oxy-
Section IV contains two major subdivisions: the elec- gen falls below a critical level in the cytoplasm, the
trophysiological changes at the cellular and multicellular electrontransportandtheprocessofH(cid:49)ejectioninthe
levelandthegenesisofarrhythmias.SectionIVAcontains mitochondrion stops. The energy stored in the proton
a description of the changes in resting and action poten- electrochemical gradient becomes insufficient to syn-
tial, in excitability, refractoriness, and conduction and is thesizeATPinappropriatequantity.Someoftheenergy
followedbyadiscussionofpossiblemechanisms.Section in ATP may be used to maintain the mitochodrion
IVB starts with an analysis of the general processes in- membrane potential (Em) and to inhibit irreversible
volved in the genesis of arrhythmias, followed by a de- reactionsleadingtocelldeath(772).Asaconsequence,
scription of the type of arrhythmias encountered during [ATP] may be expected to fall (Fig. 1) and [ADP] to
ischemiaandreperfusion,andfinallyadiscussionofpos- increase. The cell, however, has efficient means to
sible mechanisms. maintain the ATP level: 1) energy demand falls very
rapidly during the first 30 s of ischemia as a conse-
quence of contraction failure, 2) an important amount
B. Ischemia Models
of phosphocreatine (PCr) is continuously used to re-
store ATP from ADP with concomitant increase in P,
i
Cardiac ischemia is characterized by a deficient and 3) anaerobic glycolysis starts and intensifies.
energetic input as well as a deficient waste removal. The fall in the ATP/ADP stimulates the glycolytic
The result is failure of contraction, deterioration of pathway by activation of three important enzymes: 1)
electrical behavior, and eventual death of the cell. At hexokinase responsible for the formation of glucose-6-
theorganismlevel,theendpointmaybelethalarrhyth- phosphate; 2) phosphofructokinase, responsible for the
mias or mechanical pump failure. To study ischemia, transformation of fructose-6-phosphate to fructose-1,6-
different experimental models have been used. Coro- biphosphate; and 3) pyruvate kinase for the formation of
nary artery ligation or obstruction by a local thrombus pyruvate. The reactions leading to pyruvate are stimu-
mimics closely the clinical settings of myocardial in- latedbutglucosemetabolismstopsatthepyruvate-lactate
July1999 CARDIAC IONIC CURRENTS AND ACUTE ISCHEMIA 919
comestoopronounced,glycolysisinturnisinhibited,and
ATP synthesis is further reduced.
WheneverthefreeenergyofATPhydrolysisexceeds
theenergystoredintheprotonelectrochemicalgradient,
netATPhydrolysismayoccur;theATPsynthasethenacts
asanATPhydrolase,andtheenergyisusedtoupdatethe
electrical gradient in the mitochondrion (224) (Fig. 2).
Although the arrest of electron flow is expected to cause
depolarizationofthemitochondrialmembraneduringan-
oxia, such depolarization only occurs after a delay (244).
The delay corresponds to the time needed to block gly-
colysis to cause serious ATP deficiency and rise in
[Ca2(cid:49)].BlockingATPhydrolysisbyoligomycinslowsthe
i
fallin[ATP](808)and delaysactivation ofthemitochon-
drial mega-channel. When [ATP] drops to levels (cid:44)1 mM
andatthesametimetheconcentrationofH(cid:49),[Ca2(cid:49)] ((cid:46)1
i
(cid:109)M), PO3(cid:50) ((cid:46)10 mM), and long-chain acylcarnitines
4
(LCAC) increase (772), the mitochondrial membrane be-
comes abnormally leaky through activation of the mega-
channel or transition pore in the inner membrane of the
mitochondrion. Efflux of ATP through the plasma mem-
brane is an additional reason for a fall of intracellular
ATP. In the extracellular medium, this ATP is further
brokendowntoadenosineandmayactivatepurinergicP
1
and P receptors.
2
Block of oxidative metabolism and fall in ATP/ADP
FIG.1.Metabolicconsequencesofischemiaonintramitochondrial will have consequences on lipid metabolism, the genera-
levelsofATP,Pi,andphosphocreatine(PCr)aspercentofcontrol[ATP] tion of oxidative stress, the release of catecholamines,
obtainedfrom31P-NMRspectrainperfusedferrethearts.[FromMarban
and ion concentrations.
etal.(640).Copyright1990AmericanHeartAssociation.]
Uponreperfusionafter5–30minofischemia,oxygen
consumption rapidly recovers. The NADH/NAD(cid:49) quickly
stage,duetoinhibitionofpyruvatedehydrogenase.Pyru- decreases,althoughthelevelmayremainhigherthanthe
vate is transformed in L-lactate under simultaneous oxi- control for some time. After brief (5–10 min) ischemic
dation of NADH to NAD(cid:49). At the same time, phosphory- bouts, PCr concentration quickly returns to normal, but
lase a is transformed in its active form, facilitated by recoveryofATPisslow.Afterlongerperiodsofischemia,
accompanyingincreasesinAMP,P,Ca2(cid:49),andcAMPand PCr concentration still recovers within 5 min, but ATP,
i
increasesthebreakdownofglycogen.Highglycogencon- whichmayhavedroppedto50%,staysatthislowlevelfor
tent of cells postpones the development of contracture 30 min or more (641).
during ischemia (190). Finally, glucose transport through The preischemic pattern of substrate utilization is
the cell membrane is stimulated, secondary to a translo- restored, i.e., oxidation of fatty acids (FA), is the main
cation of glucose transporters from an intracellular pool contributortoATPsynthesis;thelevelofglycolysis,how-
to the plasma membrane (1155) and increases the flow ever, remains elevated in the early period of reperfusion
down to pyruvate. Glycolytic enzymes are especially and is important for ATP generation. This situation may
denseinthesubsarcolemmalspace;stimulationofanaer- explain why block of glycolysis at this time is disadvan-
obicglycolysisatthislevelisimportantfortheregulation tageous and results in aggravation of Ca2(cid:49) overload and
of intracellular ion concentrations and channels (KATP) release of intracellular enzymes. Such a block occurs
(1082). when FA are present in high concentration (621). A high
Stimulation of the glycolytic pathway explains why rateofFAoxidationgeneratesNADHwhichinhibitspyru-
[ATP] stays remarkably constant during the initial 10–15 vate dehydrogenase. Glycolysis which stops at the pyru-
min of ischemia. However, the free energy change upon vate level is accompanied by an overproduction of pro-
hydrolysisofATPfallsimmediatelybecauseoftherisein tonsandmaynegativelyaffecttherecoveryof[Na(cid:49)] and
i
[ADP] (281). This fall has important consequences for a [Ca2(cid:49)] (612). High levels of lactate are deleterious for
i
number of ATP-driven transporters such the Na(cid:49)-K(cid:49)- recovery.
ATPase and the Ca2(cid:49)-ATPase. The anaerobic glycolytic The high level of oxygen consumption early after
process is self-inhibiting however. When acidosis be- reperfusionstaysincontrasttothecontinuingdeficiency
920 EDWARD CARMELIET Volume79
FIG.2.Ionicchannelsandcarriersinouterandinner
mitochondrial membranes. Under aerobic conditions,
electron-carrying chain ejects protons and generates an
importantelectricalgradientoverinnermembrane.Proton
inflow through synthase provides necessary energy for
ATP synthesis. In anaerobic conditions, mitochondrion
should depolarize; synthase however may now act as an
ATPaseandguaranteepersistenceofelectricalaswellas
proton gradient. Ca2(cid:49) homeostasis in mitochondrion is
dependent on this electrical and proton gradient. MCC,
multiple conductance channel, mega-channel, or perme-
ability transition pore; mCS, mitochondrial cento/pico-
siemenschannel;KATP,ATP-sensitiveK(cid:49)channel;ANC,
adenine nucleotide carrier; VDAC, voltage-dependent an-
ionchannel.
of the contractile machinery or “stunning.” This uncou- substancesarelost.Onthemicroscopiclevel,cellsappear
pling between oxygen consumption and contractility is necrotic, and later important fibrosis develops.
not due to impairment of the respiratory chain flux and
insufficient ATP synthesis. Energy is sufficiently present
II. ION CHANNELS AND TRANSPORTERS
butapparentlynotused.Asapossibleexplanationforthe
high rate of oxygen consumption, the existence of futile
cycles has been proposed, such as the cycling of Ca2(cid:49)
A. Ion Channels and Transporters in the
between the cytoplasm and mitochondria (189). The ab-
Plasma Membrane
sorption of Ca2(cid:49) by the mitochondria as well as its re-
movalviatheNa(cid:49)/Ca2(cid:49)exchangerdependsontheenergy
1. Na(cid:49) channels
flux in the electron chain. The ejection of protons gener-
ates the negative matrix potential that drives the absorp- In most excitable cells, the Na(cid:49) current is responsi-
tion of Ca2(cid:49) and creates the proton gradient that is re- ble for the upstroke and the conduction of the action
quired for removing Na(cid:49) from the mitochondrial matrix potential. In heart, the density of the channel is low in
viatheNa(cid:49)/H(cid:49)exchangerandsubsequentlyCa2(cid:49)viathe sinoatrial node (SAN) and atrioventricular node (AVN)
Na(cid:49)/Ca2(cid:49) exchanger. Adenosine 5(cid:57)-triphosphate, which cells and highest in Purkinje cells. The Na(cid:49) channel is
foritssynthesisisalsodependentontheprotongradient, voltageoperatedandshowsactivationandinactivation.It
is used by the plasma membrane Na(cid:49)-K(cid:49) pump to keep is specifically blocked by tetrodotoxin (TTX) (for a gen-
cytosolic [Na(cid:49)] low (Fig. 2). Under normal conditions, eral review, see Ref. 291); modulation of channels is
this cycle only requires little energy, but in the case of described in section III.
Ca2(cid:49) overload, the expenditure may become excessive. A) ACTIVATION AND INACTIVATION. When a cardiac cell is
The situation is aggravated by an eventual intermittent depolarized, an inward current is generated that rises
openingofthemega-channelactivatedbyelevatedmatrix rapidly and decreases afterward on a much slower time
[Ca2(cid:49)] (91). The leak created causes breakdown of the course. The rise is sigmoidal (567) or exponential (685).
mitochondrialelectricalgradient.Activationofthemega- Thetimetopeakshortenswithdepolarization(634).The
channelmaypropagatefrommitochondriontomitochon- decline in macroscopic current or inactivation is best
drion, creating a Ca2(cid:49) wave in the cytosol (449). described by a sum of two exponentials. The time con-
Reperfusion after more than 30 min leads still to an stant of the first component becomes shorter with depo-
important but only transient recovery of oxygen con- larization.
sumption,whilecontractilefunctionremainsseverelyand Peakcurrent-voltagerelationcanbetranslatedintoa
persistently depressed. Intracellular enzymes and other conductance-voltage relation from which an activation
July1999 CARDIAC IONIC CURRENTS AND ACUTE ISCHEMIA 921
curve can be constructed. Peak current is voltage depen-
dent and increases for more negative holding potentials.
Therelationhasbeenexplainedasachangeinavailability
of the channel to become activated. Both relations, acti-
vationandinactivation,canbedescribedbyaBoltzmann
distribution,withslopesof(cid:59)6mV.Midpointsofvoltages
in multicellular preparations are (cid:50)30 mV for the activa-
tion process and (cid:50)85 mV for the inactivation process.
The time course and the steady-state properties of
the current have been explained in terms of two voltage-
dependentprocesses,activationandinactivationindepen-
dent from each other (425). The opening and closing of
the channel depends on the movement of charged gates,
andthehypothesisthuspredictstheexistenceofagating
current. In heart cells, such a gating current has been
measuredfortheactivationprocess(50,350,372,486)but
not for the inactivation process, as if inactivation is volt-
age independent. The activation gating current, however,
becomes smaller during the development of inactivation.
Thegatingchargebecomesimmobilized,suggestingsome
coupling between the two processes (372, 486). Depend-
ing on the type of cell, coupling between activation and
inactivation is variable. Coupling is strong in neuroblas-
tomacells(9) inwhichactivationshowsa variableonset
andinactivationfollowsrapidly.Incardiaccells,thecou-
pling is weak; openings always occur at the beginning of
the depolarizing pulse (activation is fast) and their dura-
tion is variable (inactivation is slower and variable) (69,
90, 567, 587, 856).
Recovery from inactivation is normally very fast
(1–10 ms), with rates increasing upon hyperpolarization.
After long depolarizations, recovery can become very
slow (order of seconds). The process has been called
slow inactivation (125, 160, 825, 876). The existence of
this phenomenon may be important to understand
changes occurring during and after ischemia, where the
cells are subjected to long depolarizations.
IstheremorethanonetypeofNa(cid:49)current?Inheart
cells, decay of the Na(cid:49) current is normally very rapid. In stepsFfIrGo.m3.(cid:50)H1y2p0oxtoia(cid:50)in5c0rmeaVsewsepreersaipsptelinetdNtoa(cid:49)acchealln-antetalcahcetidviptya.tcVholftraogme
a limited range of potentials, where activation and inac- aratventricularmyocyte.A:control.B:duringhypoxia,differentcells.
Bottom trace in each series is average current of 50 traces. [Adapted
tivation overlap, a small noninactivating current or win-
fromJuetal.(489).]
dowcurrentcanberecorded,whichisduetothecycling
ofchannelsbetweentherested,activated,andinactivated
state (27, 325, 755). This is not due to any abnormal in the hyperpolarized direction. This may cause more
behavior of the channel, but in pathological conditions, pronounced overlapping of activation and inactivation,
the overlapping may increase, resulting in an enhance- generatinginthiswayaconstantsteady-statecomponent.
ment of the current. Thecurrentdeactivatesimmediatelyuponhyperpolariza-
A slowly inactivating Na(cid:49) current, which represents tion. At the single-channel level, it is characterized by a
a small percentage of the total Na(cid:49) current, can be re- bursting behavior, i.e., clusters of repetitive short open-
cordedinrabbit(125)andcanine(325)Purkinjefibers,in ings (5 ms) sometimes alternating with long openings
rat ventricular cells (826), and in expressed human Na(cid:49) (order of 200 ms) (1187). An activity characterized by
channels(801)overabroadrangeofpotentials.Intherat, short openings ((cid:44)0.5 ms), becoming shorter with depo-
it becomes more pronounced during hypoxia (489) (Fig. larization, has been described as “background” current
3). Part of it does not inactivate completely. Compared because it carries current at the resting potential (1187).
withthefastNa(cid:49)currentcomponent,activationisshifted Thecurrentundergoesinactivation,however,andisvery
922 EDWARD CARMELIET Volume79
selectiveforNa(cid:49).AccordingtoBo¨hleandBenndorf(90), The short intracellular segment between domains III and
one and the same channel can show many modes of IV has been identified as the fast inactivation gate. Spon-
gating behavior; this channel may thus not be different taneous deletion of the KPQ segment in the junctional
from the slowly inactivating one. The slowly inactivating part gives rise to the LQT3 syndrome, characterized by a
current has been described as being more sensitive to slowingoftheinactivationprocessandgenerationoflong
blockbyTTX(174),afindingwhichhasbeenadvancedin action potentials. The absence or weak voltage depen-
favor of the existence of a different isoform; the result, dency of the inactivation process is consistent with the
however,canalsobeexplainedbytheslowkineticsofthe location of this inactivation gate outside the electrical
TTX block (126). fieldofthemembrane.Slowinactivationinvolvesconfor-
After exposure of membrane patches to lysophos- mational changes in the external pore (C-type inactiva-
phatidylcholine (108, 1002), a persistent current can be tion) (1058). The presence of sialic acid at the external
recorded over a voltage range of (cid:50)150 to 0 mV. It is surface shifts the activation and inactivation in the nega-
probablyduetoamodulationofthefastcomponent.The tivedirection(64).Interactionwiththecytoskeletonmod-
currentdoesnotdeactivateuponhyperpolarizationandis ulatestheinactivationandactivationvoltagedependence;
substantial at the resting potential. It may play an impor- F-actin disruption and microtubule stabilization acceler-
tantroleasaninwardleakcurrent,causingK(cid:49)lossduring ate the shift in the negative direction that occurs during
ischemia. whole cell recording (636). The two (cid:98)-subunits exert a
A Na(cid:49)-dependent, TTX-insensitive background cur- modulatory role, speed up inactivation, and decrease
rent is induced by high concentrations of ACh in guinea block by local anesthetics (633).
pig ventricular myocytes (659). The current reverses at The permeation process is dependent on the hydro-
(cid:50)25mVinnormalTyrodesolution,anditssingle-channel philic part of the (cid:97)-subunit between transmembrane seg-
conductance has been estimated by noise analysis to be ments 5 and 6. The selectivity for Na(cid:49) is due to specific
(cid:59)2pS(897).Becauseofitsdifferentpharmacologicaland amino acids in this region, and mutation of only two
single-channel characteristics, it can be regarded as a criticalaminoacidresiduesissufficienttoconferperme-
different isoform. ability properties similar to the Ca2(cid:49) channel. A cysteine
B) ION PERMEATION. In the presence of 150 mM [Na(cid:49)]o, in domain I is responsible for the high affinity of the
single-channel conductance is 20–25 pS; it is dependent channel for Zn2(cid:49) and Cd2(cid:49) and low sensitivity to TTX of
on the concentration of external Na(cid:49) with a dissociation the cardiac isoform.
constant (K ) of 300–400 mM (885) and shows different
d
substates (554, 727, 856). The channel is permeable to 2. Ca2(cid:49) channels
Na(cid:49)andLi(cid:49)andmuchlesstoK(cid:49)(9.5%)andCs(cid:49)(2%)or
tetramethylammonium((cid:44)1%)(885).AsmallCa2(cid:49)perme- Different types of Ca2(cid:49)-permeable channels have
ability exists (3) and may increase in the presence of beendescribedintheplasmamembraneofheartcells:the
ouabain and upon (cid:98)-receptor activation (840). The cur- L- and T-type channels, both voltage activated, and a
rent-voltage relation is linear in the absence of bivalent backgroundchannel(seeRef.673).Theycanbedifferen-
ions;deviationfromthislinearbehaviorinnormalbathing tiatedonthebasisoftheirelectrophysiologicalandphar-
solutions is due to a voltage-dependent block by [Ca2(cid:49)] macological characteristics. The density of the T- and
o
and[Mg2(cid:49)] ;electricaldistanceis0.3–0.4(884).Theblock L-type channels differs in different sections of the heart.
o
by [Ca2(cid:49)] and [Mg2(cid:49)] is characterized by a reduction in The ratio of T-type over L-type channel is highest in
o o
apparentsingle-channelconductance,suggestingaveryfast Purkinje and sinoatrial cells where it approaches the
block.Otherbivalentionsefficientlybutmoreslowlyblock valueof0.2–0.6,anditislessinatrialandventricularcells
thecurrent:Cd2(cid:49)(cid:46)Mn2(cid:49)(cid:46)Co2(cid:49)(cid:46)Ca2(cid:49)(cid:46)Mg2(cid:49)(cid:46)Ba2(cid:49). where the ratio only attains 0.015–0.025 (48).
OfimportancetonoteisthehighlysensitiveblockbyZn2(cid:49) A) BACKGROUND CA2(cid:49) CHANNELS. Calcium-permeable chan-
andCd2(cid:49)(297,1042);theseionsaremuchlesseffectivein nels are seen following incorporation of plasma mem-
neuronal and skeletal muscle cells, whereas the opposite braneproteinfractionsinbilayers(806).Novoltagesteps
sensitivityexistsforTTX.Internal[Mg2(cid:49)]blocksoutward are required for activation, and spontaneous single-chan-
current through the channel (786). The block is slightly nel activity with long openings of (cid:46)100 ms can be re-
voltagedependentwithanelectricaldistanceof0.18from corded at negative E values of (cid:50)90 mV. The conduc-
m
inside. tance is 7 pS in isotonic Ba2(cid:49); the channels are not
At the molecular level, the channel consists of an blocked by dihydropyridines (DHP) and show no run-
(cid:97)-subunit (260 kDa) and two (cid:98)-subunits (36 and 33 kDa down. Their selectivity is not pronounced with a perme-
each)(291).The(cid:97)-subunit,whichissufficientforchannel ability ratio P /P of 10.
Ba Cs
activity, is composed of six transmembrane segments A similar channel but with higher conductance (22,
repeatedfourtimesinatetramericstructure.Thevoltage 45,and78pSinBa2(cid:49))hasbeendescribedinneonatalrat
sensor for activation has been related to the S4 segment. hearts (187). It is not blocked by Cd2(cid:49) or nifedipine;
July1999 CARDIAC IONIC CURRENTS AND ACUTE ISCHEMIA 923
instead, activity is rather increased in the presence of conclusion that inactivation is dependent on voltage as
DHPbutsuppressedbyprotamine.Inratventricularmyo- well as on Ca2(cid:49) influx. The latter or Ca2(cid:49)-induced inacti-
cytes, the channel is induced by exposure of the inside- vation is the faster process, whereas the voltage-induced
out patches to phenothiazines (593). Activity is also in- inactivation is rather slow. Activation and inactivation
creased by exposure to oxygen free radicals and show a remarkable overlapping (window current) (414,
metabolic inhibition (1056). The channel may be respon- 900).
sible for the Na(cid:49)-independent Ca2(cid:49) entry pathway de- B) Intracellular Ca2(cid:49) or current-dependent inactiva-
scribed for rat trabeculae (583). tion. Evidence for intracellular Ca2(cid:49)-dependent inactiva-
B)L-TYPECA2(cid:49)CHANNEL.CalciuminfluxthroughtheL-type tionatthewholecelllevelisbasedonthechangeintime
Ca2(cid:49)channelisresponsiblefortheupstrokeoftheaction courseofcurrentdecaywithchangesin[Ca2(cid:49)] (seeRef.
i
potentialintheSANandAVNandplaysanimportantrole 673) (Fig. 4, A and B). The decay is faster the larger the
in determining the plateau and eventual spike-dome ap- Ca2(cid:49)current,anditisslowedinthepresenceofintracel-
pearanceoftheactionpotentialinothercardiaccells.Itis lular Ca2(cid:49) buffers. At the single-channel level (cell-at-
further responsible for the coupling between excitation tached patches), Ca2(cid:49) permeation through the channel
and contraction, induces release of Ca2(cid:49) from the sarco- reducestheopenprobabilityofsubsequentreopeningsof
plasmic reticulum, and regulates intracellular Ca2(cid:49) load. the channel and shifts the gating mode toward a mode
In this way it determines activity of a number of mito- with long-lived closed states (454). In excised patches,
chondrial and cytoplasmatic Ca2(cid:49)-sensitive enzymes. with Ba2(cid:49) as the charge carrier, steady-state elevation of
I) Kinetics. A) Activation and inactivation. Thresh- Ca2(cid:49) in the range of micromolar concentration or flash
old for activation is around (cid:50)25 mV, and half-maximum photolysis of Ca2(cid:49) reduces the open probability of the
activationisattainedatabout(cid:50)15mVformostcells(see Ca2(cid:49) channels. It is especially the Ca2(cid:49) originating from
Ref. 673) and at more positive potentials ((cid:50)3 mV) in the the sarcoplasmic reticulum (SR) that is responsible for
AV node (373). The rise in current follows a sigmoidal theinactivationprocessduringthefirst50msofdepolar-
timecourse,suggestingamultistepprocessastheunder- ization (949, 1154); at later times also Ca2(cid:49) permeating
lying mechanism. Activation is preceded by a gating cur- through the Ca2(cid:49) channel contributes to the inactivation
rent (50, 352, 486, 898). The density of the channels de- (949). The increase in cytosolic Ca2(cid:49) by the release from
rivedfromthegatingcurrentismuchlargerthantheionic theSRisindeed10-foldgreaterthantheCa2(cid:49)enteringthe
current density, suggesting that some of the channels cell via the L-type Ca2(cid:49) channel (915). In favor of this
although gating are not carrying current. explanation is the observation that inactivation is much
At the single-channel level, three modes of activity slower after depletion of the SR by caffeine or in the
havebeendistinguished(137,403).Inmode1,thechannel presenceofryanodine.Itisimportanttonotethat[Ca2(cid:49)]
shows repetitive short ((cid:44)1 ms) openings and closures seen by the channel may importantly deviate from the
(0.2and2ms),formingaburstofactivityseparatedfrom cytosolic[Ca2(cid:49)]asmeasuredbyfluorescencetechniques.
otherburstsbylongerclosures.Anumberofconsecutive Especially during the first 50 ms of a depolarizing pulse
bursts may be grouped in a cluster. A variable waiting the concentration seen by the channel may be much
time precedes the openings; it decreases at more depo- higher. Such a difference may explain why the relative
larizedlevelsandcorrespondstothefasteractivationand inhibition of the current estimated from the Ca2(cid:49) tran-
shorter time to peak values of the Ca2(cid:49) current. Mode 2 sient is greater during this initial period than later (611).
occurs in the presence of DHP agonists (403) or after The existence of an intact cytoskeleton is important
(cid:98)-receptor stimulation (137, 767) and is characterized by in determining the local [Ca2(cid:49)]. The cytoskeleton nor-
much longer open times. Mode 3 is characterized by the mally keeps the channels separated from each other and
complete absence of openings or presence of only rare limits the rise in local [Ca2(cid:49)]. In ischemia, the cytoskele-
short openings. The frequency of this latter mode in- ton may become disturbed. Disruption of the cytoskele-
creases with preceding depolarizations and corresponds ton structure by colchicine or cytochalasin, with conse-
to the occurrence of steady-state inactivation. quent clustering of the channels, favors inactivation,
The L-type Ca2(cid:49) current inactivates in two ways: a whereas substances such as taxol and phalloidin that
voltage-dependent and a current-dependent way. The ex- stabilize the skeleton remove inactivation and improve
istenceoftwotypesofinactivationexplainsthecomplex reopening (taxol and colchicine act on microtubules;
timecourseofcurrentdecayandthepresenceofadipin phalloidin on F-actin) (311).
the inactivation curve. Half-maximum steady-state inacti- Differentmechanismshavebeenproposedtoexplain
vation occurs at (cid:50)20 to (cid:50)30 mV. The curve shows a [Ca2(cid:49)]-dependent inactivation (719). 1) A fall in driving
i
minimum at (cid:59)0 mV and increases again at more positive forceisimprobable.Incell-attachedpatches,single-chan-
potentials. When intracellular Ca2(cid:49) is well buffered, this nel conductance does not change, whereas open proba-
turningupisabsentandthedecayofthecurrentduringa bilityismarkedlyreduced.2)Dephosphorylationbyphos-
pulseismuchslower.Theseobservationshaveledtothe phatase (e.g., calcineurin) and proteolysis by Ca2(cid:49)-
924 EDWARD CARMELIET Volume79
FIG.4.A:existenceoffacilitationduringrecoveryof
Ca2(cid:49)currentfrompreviousinactivationindogventric-
ularmyocytes.Paireddepolarizingpulsesof100-msdu-
rationwereappliedfromaholdingpotentialof(cid:50)80mV
to(cid:50)10mVwithadelayvariablebetween50and3,000
ms. Ca2(cid:49) current recovers rapidly within 30 ms to its
control value; afterward, current transiently becomes
larger.FacilitatoryeffectrequirespresenceofCa2(cid:49)in-
fluxandisabsentwhenexternalsolutioncontainsBa2(cid:49).
[Adapted from Tseng (991).] B: Ca2(cid:49)-induced inactiva-
tion. Ca2(cid:49) currents (top traces) and Ca2(cid:49) transients
(bottom traces) in guinea pig ventricular myocytes in
presence of 2 different external Ca2(cid:49) concentrations
(Ca ). Superimposed tracings for control (a) and after
o
Ca2(cid:49) loading (b) are shown. Note marked, rapid Ca2(cid:49)-
inducedinactivationinpresenceofCa2(cid:49)overload,but
also partial recovery during pulse. I, current; [Ca2(cid:49)],
i
intracellular Ca2(cid:49) concentration. [From Sipido et al.
(911).Copyright1995AmericanHeartAssociation.]
stimulatedproteases(calpain)(351)aremechanismsthat carried by Ba2(cid:49) or Sr2(cid:49)), the current decays with time,
mayberesponsibleforlong-termchangesinCa2(cid:49)channel and the rate of decay is faster the higher the depolariza-
behavior but are too slow to explain the time course of tion.2)Inactivationdevelopswhenprepulsesareapplied
changes during a single depolarization. 3) Calcium binds that do not result in Ca2(cid:49) inward current. 3) Inactivation
to the channel protein and induces a change in configu- is present in channels incorporated in lipid bilayers with
ration.Intheclonedchannel,aCa2(cid:49)bindingmotif(anEF Ca2(cid:49) buffered (806). 4) Inactivation is slowed by trypsin
hand) exists at the COOH terminal of the (cid:97) -subunit; treatment, but the intracellular Ca2(cid:49)-dependent inactiva-
1C
deletion eliminates Ca2(cid:49)-induced inactivation. The site is tionisnotaffected(862).Nochargemovementoccurson
located near the inner mouth but outside the electrical inactivation, but as inactivation of the current proceeds,
field(205,719).ThereductionofCa2(cid:49)currentby[Mg2(cid:49)] the charge movement that accompanies activation be-
i
also has been explained by a direct binding to this site, comes smaller (351, 898).
reducing the number of functional channels (1132). Be- D) Recovery from inactivation: Ca2(cid:49)-induced facili-
cause Ca2(cid:49) binding is a fast process, it provides an ex- tation.Uponhyperpolarization,theCa2(cid:49)currentrecovers
planation for the time course of inactivation and the from inactivation induced by a previous depolarization.
observation that Ca2(cid:49) oscillations are accompanied by Because inactivation is voltage and intracellular Ca2(cid:49)
equivalent changes in current (911). Photolysis of Ca2(cid:49) dependent,itislogicaltoexpectreprimingalsotodepend
resultsinarapidinactivation,within20ms(351)to75ms on these two parameters. For the voltage-induced inacti-
(46);theprocessisnotaccompaniedbyachangeingating vation, the rate and the degree of recovery is greater the
current, confirming that intracellular Ca2(cid:49)-induced inac- more hyperpolarized the membrane, with time constants
tivation and voltage-dependent inactivation are two dis- in the order of 300 ms at (cid:50)50 mV and 100 ms or shorter
tinct phenomena (273). at (cid:50)80 mV. A much slower component (seconds) is
C) Voltage-dependent inactivation. The evidence for present after long depolarizations, indicating the occur-
voltage-dependent inactivation is based on the following renceofslowinactivation(94,868).Thisslowinactivation
observations (see Ref. 673). 1) Under conditions where mayplayaroleinoverdrivesuppression(1074).Recovery
intracellular Ca2(cid:49)-dependent inactivation is excluded from Ca2(cid:49)-induced inactivation as such is voltage inde-
(e.g., current carried by monovalent cations or current pendent (911, 912) but indirectly it is modulated by volt-
July1999 CARDIAC IONIC CURRENTS AND ACUTE ISCHEMIA 925
age, since the fall in [Ca2(cid:49)] is dependent in part on the theporemouthofthechannel,whichattractbivalentions
i
Na(cid:49)/Ca2(cid:49) exchange that is faster the more negative the and increase their local concentration (331), is probably
E . thereasonforthisbehavior.InthepresenceofbothCa2(cid:49)
m
At negative holding potentials, recovery from inactiva- and Ba2(cid:49), the current across the channel is smaller than
tion may show an overshoot, i.e., the Ca2(cid:49) current tran- in the presence of either Ca2(cid:49) or Ba2(cid:49) alone (997). This
siently becomes larger than in steady state. The original kindofbehaviorhasbeencalledanomalousmolefraction
findingofanovershootwasmadeinPurkinjefiberstreated behavior and suggests a multi-ion channel. Different ions
with digitalis (505) but can also be observed without Ca2(cid:49) thusinteractinsuchawaythatthefluxofonespeciesis
overload(seereferencesinRefs.606,770).Thepotentiated hamperedbythepresenceoftheotherspecies.Multi-ion
Ca2(cid:49)currentischaracterizedbyalargerpeakandaslower occupancyalsoexplainsthehighconductanceandatthe
timecourseofdecay(991,1159,1193)(Fig.4). same time the high selectivity. High selectivity is condi-
At the single-channel level, facilitation is character- tionedbyhighaffinity;highconductancebythepresence
ized by an increase in open probability (Po) with a larger ofmorethanoneioninthechannelandrepulsionofone
proportion of long openings (413) (mode 2) and an in- ionbytheother.Themulti-ionnatureofthechannelalso
crease in number of functional channels (1132). In all
explainstheflickeryblockbehaviorofthechannelinthe
these approaches, it is clear that a moderate increase in
presence of elevated proton concentration (784).
Ca2(cid:49)isrequired(991,1159,1193);theovershootinrecov-
The channel can be blocked by a number of extra-
ery is inhibited by rising intracellular Ca2(cid:49) buffering, or cellular bi- and trivalent ions such as Mg2(cid:49), Mn2(cid:49), Co2(cid:49),
byryanodineorusingBa2(cid:49)asthecurrentcarrier.Exces- Ni2(cid:49), Cd2(cid:49), Zn2(cid:49), and La3(cid:49) (997). The ions Cd2(cid:49), Zn2(cid:49),
sive rises in Ca2(cid:49) lead to inhibition by Ca2(cid:49)-induced and La3(cid:49) block the channel in a voltage-dependent way
inactivation (351). with an apparent electrical distance of 0.15 for Cd2(cid:49) and
HowelevatedCa2(cid:49)causesfacilitationremainsamat- Zn2(cid:49) and 0.60 for La3(cid:49). Intracellular Mg2(cid:49) is needed to
terofdebate.Phosphorylationofthechannelproteinisa
activateenzymesthatphosphorylatethechannelbutmay,
possibility. Flash photolysis induces facilitation with a
on the other hand, reduce the current that has been
delay, and the effect is counteracted by inhibitors of
augmented by isoproterenol or BAY K 8644 by a direct
protein kinases (PK) (18, 1159). Other groups, however,
blocking action and by activating phosphatases (1090,
did not find an effect of PK inhibitors (46, 1132), and
1132).ThecAMP-dependentphosphorylationreducesthe
facilitation still occurred with nonhydrolyzable ATP ana-
sensitivity to [Mg2(cid:49)] block (1131).
logs (1132) or even improved (46). The conclusion of i
The cardiac Ca2(cid:49) channel molecular structure con-
these authors is that a Ca2(cid:49)-nucleotide complex directly
potentiatesCa2(cid:49)current(I )throughaphosphorylation- sistsoffoursubunits:two(cid:97)-subunits,(cid:97)1and(cid:97)2,a(cid:98)-sub-
Ca unit,anda(cid:100)-subunit(a(cid:103)-subunitisexclusivelyexpressed
independent mechanism. The existence of two phases in
inskeletalmuscle).The(cid:97)-subunitissufficienttoexpress
the recovery (decrease followed by an overshoot) has
channel activity. It resembles the Na(cid:49) channel with four
practical consequences in determining down- or upregu-
times six transmembrane segments, a highly charged S4
lation of I as a function of frequency and diastolic E .
Ca m segment that probably acts as the voltage sensor for
II) Permeation and selectivity. The channel is 500–
1,000timesmorepermeabletobivalentionssuchasCa2(cid:49) activation, and an intracellular link between domain II
and Ba2(cid:49) than to monovalent ions. The exclusion of and III responsible for inactivation. Trypsin treatment
removes voltage-dependent inactivation but not internal
monovalent ions depends on the presence of a minimum
Ca2(cid:49)-dependentinactivation(862).BindingofCa2(cid:49)tothe
concentrationofbivalentions.Intheabsenceofbivalent
COOHterminalisapossiblemechanismforCa2(cid:49)-induced
ions, the channel becomes highly permeable to monova-
lent ions (651). Although restricted, the permeability for inactivation (205).
K(cid:49) is responsible for a substantial current during the The highly conserved glutamate residues located in
action potential, the reason being that the concentration the pore region of all four repeats are involved in high-
of intracellular K(cid:49) is quite high compared with the nano- affinitybivalentbinding(1146).AmutationfromEtoQin
molar free Ca2(cid:49) concentration. The K(cid:49) contribution also domain III has shown that this group is the strongest
explains why the reversal potential (E ) of the Ca2(cid:49) determinant of Ca2(cid:49) binding (331).
rev
current is much less positive than expected for the equi- Thefunctionofthe(cid:97)-subunitismarkedlymodulated
librium potential for Ca2(cid:49). The bivalent ion current by the (cid:98)-subunit. Coexpression of the two results in a
throughthechannelincreaseswiththeconcentrationand fourfold increase of peak current, which is not due to a
showssaturation.Comparedwiththesingle-channelcon- change in single-channel conductance (331) but to a
ductanceinisotonicBa2(cid:49)(8–10pSand15–25pS,seeRef. marked increase in density of functional channels (488,
673),thesingle-channelconductanceatthephysiological 718, 759). The gating current remains unchanged, but
concentration of 1 mMCa2(cid:49) was found to be surprisingly coupling between conductance and gating is improved
high at 7 pS (1162). The presence of negative charges at (759).
926 EDWARD CARMELIET Volume79
The L-type Ca2(cid:49) channels is highly regulated; this I , I , and I ), and a current (the inward rectifier
KATP KNa KAA
aspect is analyzed in section III. IK1) that apparently does not gate and can be called a
C)T-TYPECA2(cid:49)CHANNEL.ACa2(cid:49)currentofshortduration background current. Under physiological circumstances
isactivatedatpotentialsmorenegativethanthethreshold thevoltage-activatedK(cid:49)currents,I amongtheligand-
KACh
for the L-type Ca2(cid:49) current (48, 726; see review in Ref. activated and I , play an important role in shaping the
K1
1026).ThecurrentiswellrepresentedinSANcells,atrial normal action potential. Under ischemic conditions, li-
cells, Purkinje cells, and nodal cells. In the embryonic gand-activated currents, especially I and I , be-
KATP KAA
chick ventricle, it is the major Ca2(cid:49) current. The current come primordial, whereas some of the “physiological”
isnotfoundinhumanatrium(258,606)orhumanventri- currents are inhibited. Voltage-activated K(cid:49) currents
cle(76).Thechannelhasbeenproposedtoplayarolein showactivationandinactivationupondepolarization;the
pacemaking.Itmayalsointerferewithsteroidgenesis,cell rates of these two processes can vary from fast to ultra-
proliferation, and cardiac growth (400). slow. Ligands can bind to receptors, which then activate
I) Activation, inactivation, and repriming. Thresh- the channel via a G protein, or can interact directly with
old foractivationisaround (cid:50)70to(cid:50)50mVandmaximum an intracellular site of the channel.
activationisseenat(cid:50)30to(cid:50)10mV(seeRef.1026).Inacti- The amino acid composition of most K(cid:49) channels is
vationisrapidandcomplete,withtimeconstantsof30msat known(201),andrecently,themolecularstructureofthe
(cid:50)50mV,becomingshorteratmoredepolarizedlevels.This porehasbeenelucidated fromX-ray analysisofthecrys-
behavior is opposite to the L-type channel, where inactiva- tallized molecule (234). The channels show a remarkable
tion is decelerated at positive potentials. Steady-state inac- homology with the Na(cid:49) and Ca2(cid:49) channels. However,
tivation extends from (cid:50)85 to (cid:50)40 mV with half maximum whereas Na(cid:49) and Ca2(cid:49) channels consist of tandemly
around (cid:50)60 mV and slope of 5.5 mV (14, 36, 412). An linkedfourdomainsofsixtransmembranesegmentsthat
increase in [Ca2(cid:49)] does not induce inactivation but rather are connected in one long polypeptide, only one domain
i
facilitatestheT-typecurrent(13,994).Atthesingle-channel with six transmembrane segments is found for the K(cid:49)
level, this increase in current is characterized by a shift to channel. Segment 4 shows a high density of positive
longopenings(mode2behavior). charges and acts as potential sensor. Two types of inac-
Repriming is voltage dependent and becomes faster tivationhavebeendescribedinexpressedchannels(794):
with hyperpolarization: 250 ms at (cid:50)70 mV and 100 ms at N-type inactivation in which the negatively charged NH
2
(cid:50)90 mV (145). It is slower the longer the preceding de- terminal acts as a ball and blocks the open channel and
polarization,suggestingtheexistenceofslowinactivation C-type inactivation (COOH terminal) in which conforma-
(412). tional changes on the extracellular side close to the pore
II)Permeation.In100mM[Ca2(cid:49)],thesingle-channel result in some kind of constriction. C-type inactivation is
conductanceis8pS,comparedwith20pSfortheLtype. sensitivetodrugbindingandextracellularK(cid:49).Theligand-
Contrary to the behavior of the L-type channel, perme- activated and background channels have a simpler struc-
ability for Ca2(cid:49) and Ba2(cid:49) ion is the same (48, 900). The ture and contain only two transmembrane segments. Re-
channelispermeabletoSr2(cid:49),blockedbyNi2(cid:49),butmuch cently, a new family of K(cid:49) channels with two pore
less sensitive to Cd2(cid:49). Extracellular protons inhibit the segments in tandem and four transmembrane segments
channel with greater efficiency than the L-type current, have been expressed. They act as background channels
whereas intracellular protons have no effect (1000). Ex- (530);someofthemareactivatedbyarachidonicacidand
tracellular Mg2(cid:49) reduce the current and shift the activa- polyunsaturated fatty acids (280). A tetrameric structure
tion and inactivation curves in the positive direction for all K(cid:49) channels is highly likely. The pore in K(cid:49) chan-
(1113). nels is formed by a stretch of 19 amino acids in the link
Recently two isoforms, the G and H isoform of the betweenS5andS6inthefourrepeats.ThemotifGYG(or
(cid:97)-subunit, have been cloned (760). Sialic acid probably ofGFG)intheP-regionisthesignatureofK(cid:49)selectivity,
formsanimportantcomponentoftheextracellularpartof but other residues also participate in determining K(cid:49)
the channel (1150). selectivity.
A) K(cid:49) OUTWARD CHANNELS WITH FAST ACTIVATION.Upon depo-
3. K(cid:49) channels larization, three different K(cid:49) currents are rapidly acti-
vated.Theycanbedistinguishedbytheirrateofinactiva-
Because the equilibrium potential of K(cid:49) is rather tion, which is relatively fast for I , slow to ultraslow for
to
negative, all cardiac K(cid:49) channels when activated will I , and nonexistent for I , also called background
Kur Kss
carry outward current, repolarize the membrane during current. On the latter current, no detailed information is
the action potential, or stabilize the membrane at a hy- available at the present time.
perpolarizedlevel.AmongthemanyK(cid:49)currents,distinc- I)Thefasttransientoutwardcurrent.Thefasttran-
tioncanbemadebetweenvoltage-activatedcurrents(I , sient outward current (I ) is a transient outward K(cid:49)
to to
I , I , I , and I ), ligand-activated currents (I , current that is rapidly activated and inactivated and
Kur Kss Kr Ks KACh