Protocols of cardiac ion channels INaL Ito1 IKur IKs IK1

Eunjung ParkFDACDERONDDCRP

herg 1

CIPA project-2CIPA project-2

Notes

bull The references are listed in the last slide or if necessary in the bottom of the slide

bull The references in the pulse sequence table are hyperlinked to Pubmed

bull The percentages of channel sources and temperatures in each ion channel were based on selected literatures in this presentation That is not intended to show all relevant published data

Late Sodium Channel Current

bull INaL inhibition Suppress ventricular arrhythmiasndash reduction of intracellular calcium loadingndash decrease of intracellular sodium

bull Potentially selective late INa Blockers at relative low concentrationndash Ranolazine selectivity 9 to 45 fold strong in blocking late vs

peak INa

ndash Amiodarone TTX lidocaine quinidine phenytoin etc

Peak Na(1-2 ms duration)

Phase 2 and early phase 3(100 - 750 ms duration)

Ref Cardiovasc Drugs Ther (2013) 27 79-89

Late sodium channel protocolsRef ID Year Drug Holding (mV) Late Na (ms) Late Na (mV) Peak Na

(ms mV)Frequency

(Hz)

173 1997 -120 100 -20 same 02

100 1998 -150 720 -20 24 -90 to 30

174 1998 R56865 -100 350 -20 No data 02

177 2000 Flecainide -150 Mean (20 and 235) -20 same 5

179 2000 BDF 9198 -140 250 -30 No info

178 2001 Amiodarone -160 200-220 -30 No time info 01

180 2002 -140 750 -40 25 -30

186 2006 -140 2000 na No info 01

205 2009 Ranolazine -140 Mean (46 and 48) -20 24 -20

199 2012 -120 300 -20 No info

160 2013 Ranolazine -130 50 -10 20 -10

195 2013 GS967 -120 220 -20 20 -20

Ex) Ref ID 195peak late

-120 mV complete removal of inactivation

-20 mV

INa enhancer

Pulse20 vs 220 ms

Notes

bull The references are listed in the last slide or if necessary in the bottom of the slide

bull The references in the pulse sequence table are hyperlinked to Pubmed

bull The percentages of channel sources and temperatures in each ion channel were based on selected literatures in this presentation That is not intended to show all relevant published data

Late Sodium Channel Current

bull INaL inhibition Suppress ventricular arrhythmiasndash reduction of intracellular calcium loadingndash decrease of intracellular sodium

bull Potentially selective late INa Blockers at relative low concentrationndash Ranolazine selectivity 9 to 45 fold strong in blocking late vs

peak INa

ndash Amiodarone TTX lidocaine quinidine phenytoin etc

Peak Na(1-2 ms duration)

Phase 2 and early phase 3(100 - 750 ms duration)

Ref Cardiovasc Drugs Ther (2013) 27 79-89

Late sodium channel protocolsRef ID Year Drug Holding (mV) Late Na (ms) Late Na (mV) Peak Na

(ms mV)Frequency

(Hz)

173 1997 -120 100 -20 same 02

100 1998 -150 720 -20 24 -90 to 30

174 1998 R56865 -100 350 -20 No data 02

177 2000 Flecainide -150 Mean (20 and 235) -20 same 5

179 2000 BDF 9198 -140 250 -30 No info

178 2001 Amiodarone -160 200-220 -30 No time info 01

180 2002 -140 750 -40 25 -30

186 2006 -140 2000 na No info 01

205 2009 Ranolazine -140 Mean (46 and 48) -20 24 -20

199 2012 -120 300 -20 No info

160 2013 Ranolazine -130 50 -10 20 -10

195 2013 GS967 -120 220 -20 20 -20

Ex) Ref ID 195peak late

-120 mV complete removal of inactivation

-20 mV

INa enhancer

Pulse20 vs 220 ms

Late Sodium Channel Current

bull INaL inhibition Suppress ventricular arrhythmiasndash reduction of intracellular calcium loadingndash decrease of intracellular sodium

bull Potentially selective late INa Blockers at relative low concentrationndash Ranolazine selectivity 9 to 45 fold strong in blocking late vs

peak INa

ndash Amiodarone TTX lidocaine quinidine phenytoin etc

Peak Na(1-2 ms duration)

Phase 2 and early phase 3(100 - 750 ms duration)

Ref Cardiovasc Drugs Ther (2013) 27 79-89

Late sodium channel protocolsRef ID Year Drug Holding (mV) Late Na (ms) Late Na (mV) Peak Na

(ms mV)Frequency

(Hz)

173 1997 -120 100 -20 same 02

100 1998 -150 720 -20 24 -90 to 30

174 1998 R56865 -100 350 -20 No data 02

177 2000 Flecainide -150 Mean (20 and 235) -20 same 5

179 2000 BDF 9198 -140 250 -30 No info

178 2001 Amiodarone -160 200-220 -30 No time info 01

180 2002 -140 750 -40 25 -30

186 2006 -140 2000 na No info 01

205 2009 Ranolazine -140 Mean (46 and 48) -20 24 -20

199 2012 -120 300 -20 No info

160 2013 Ranolazine -130 50 -10 20 -10

195 2013 GS967 -120 220 -20 20 -20

Ex) Ref ID 195peak late

-120 mV complete removal of inactivation

-20 mV

INa enhancer

Pulse20 vs 220 ms

Late sodium channel protocolsRef ID Year Drug Holding (mV) Late Na (ms) Late Na (mV) Peak Na

(ms mV)Frequency

(Hz)

173 1997 -120 100 -20 same 02

100 1998 -150 720 -20 24 -90 to 30

174 1998 R56865 -100 350 -20 No data 02

177 2000 Flecainide -150 Mean (20 and 235) -20 same 5

179 2000 BDF 9198 -140 250 -30 No info

178 2001 Amiodarone -160 200-220 -30 No time info 01

180 2002 -140 750 -40 25 -30

186 2006 -140 2000 na No info 01

205 2009 Ranolazine -140 Mean (46 and 48) -20 24 -20

199 2012 -120 300 -20 No info

160 2013 Ranolazine -130 50 -10 20 -10

195 2013 GS967 -120 220 -20 20 -20

Ex) Ref ID 195peak late

-120 mV complete removal of inactivation

-20 mV

INa enhancer

Pulse20 vs 220 ms

Cardiac potassium channels

ID248 name clone Channel blocker role note

Ito1 Transient outward Kv43Kv14 4-AP (closed state)bullEarly rapid repolarization (Phase I)bullDetermine the height of early plateau

Ito2 is not found in cardiac cells

IKur Ultra rapid delayed rectifier Kv15 4-AP (selective) bullAtrial repolarization

IKs Slow delayed rectifier KCNQ1(KvLQT1) KCNE1 (hminK) Chromanol (293B)

bullAtrial and ventricular repolarizationbullHeart rate dependent

Activation ~0 mV

IKr rapid delayed rectifier Kv111 (hERG) E-4031bullRapid activation and inactivation strong inward rectificationbullCardiac APD and refractoriness

Activation ~-30mV[K+] dependent

IK1 Inward rectifier Kir21 Ba2+ (IC50= 20 uM)

bullMaintain the resting membrane potentialbullControl excitabilitybullInitiate the final repolarization phase

Nav15

Kv43 (Ito f) Kv15 (IKur)

KvLQT1minK (IKs)

Cav12 (ICaL) hERG (IKr)

Cav32 (IcaT)

Kir 21 (IK1)

Summary of Ito1 protocolsbull Distribution Atrial PF epicardial M cells gt endocardialbull Pharmacology A common finding in cardiac disease is reduction in Ito1 density with a loss

of heterogeneity of Ito expression and associated action potential prolongation

bull Channel sources1 Transfected cell lines CHO L cell COS-7 cell HEK 293 and Xenopus Oocyte (818 = 44)2 Myocyte (56) human AM (3) rat VM (4) Canine VM 92) and rabbit VM (1)

Guinea Pig (0 very low expression of Ito)

bull Temperature ndash RT = 1215 (80)ndash near or real physiological 315 (20)

1 Pulse sequencesndash Holding -90 to -40 (mode -80 mV) ndash Activation 0 to 75 mV (mode 50 mV)ndash Duration of activation 150 ndash 500 ms (mode 300 ms)

bull Ito measurement peak current or after 70ms of activation

Pulse sequences Ito

Ito (Kir 43)

ID ChannelCurrent

Speciescells CPD Temp Vh(mV)

Prepulse(ms mV)

Activation(ms mV)

Voltage -Dependence(ms mV)

Intervalfrequency

47 hKv43hKChIP22 CHO Methadone RT -90 300 40

49 rKv43 MouseL cell Vanoxerine RT -80 300 20 15s

50 hKv43 HEK293 Alfuzosin RT -80 300 20 15s

206 Ito RatVM Zacopride 24 -40 na 500 -40 to 80 (10)

210 Ito HumanAM Cyamemazine

32-34 -50 500 60

214 Kv43 MonkeyCOS7 Pentamidine na -80 na 500 -40 to 60

215 Ito CanineVM Pioglitazone na -80 5ms -40 na 50

220 Ito RatVM Ajmaline RT -75 300 60

221 rKv42 rKv43 HEK293 RSD1235 25 -80 Brief -50 250 60 1Hz

222 Ito RatVM FK-506 22-24 -80 na 50 1000 -50 to 70 01Hz

224 hKv43 HEK293 Allitridi 22-23 -80 300 60 300 -80 to 60

245 Ito HumanAM DPO RT -80 150 40

256 Ito RabbitVM Carvedilol 34 -60 300 40 300 -50 to 60

263 Kv43 CHO (transient) Spironolactone canrenoic acid

RT -80 250 50 250 -90 to 50 10s

267 Ito ratVM Ebastine na -90 na 75 na -30 to 75

273 Ito CanineVM Thymol 37 -80 5ms- 40 400 50 400 -10 to 60

281 rKv14 Xenopus Oocyte Haloperidol 22 -80 300 0

283 Ito HumanAM Clotrimaxole 21-22 -50 300 50 300 -40 to 60 02Hz

VM= ventricular myocyte AM= Atrial myocyte

Summary of Ito1 protocolsbull Distribution Atrial PF epicardial M cells gt endocardialbull Pharmacology A common finding in cardiac disease is reduction in Ito1 density with a loss

of heterogeneity of Ito expression and associated action potential prolongation

bull Channel sources1 Transfected cell lines CHO L cell COS-7 cell HEK 293 and Xenopus Oocyte (818 = 44)2 Myocyte (56) human AM (3) rat VM (4) Canine VM 92) and rabbit VM (1)

Guinea Pig (0 very low expression of Ito)

bull Temperature ndash RT = 1215 (80)ndash near or real physiological 315 (20)

1 Pulse sequencesndash Holding -90 to -40 (mode -80 mV) ndash Activation 0 to 75 mV (mode 50 mV)ndash Duration of activation 150 ndash 500 ms (mode 300 ms)

bull Ito measurement peak current or after 70ms of activation

Pulse sequences Ito

Ito (Kir 43)

ID ChannelCurrent

Speciescells CPD Temp Vh(mV)

Prepulse(ms mV)

Activation(ms mV)

Voltage -Dependence(ms mV)

Intervalfrequency

47 hKv43hKChIP22 CHO Methadone RT -90 300 40

49 rKv43 MouseL cell Vanoxerine RT -80 300 20 15s

50 hKv43 HEK293 Alfuzosin RT -80 300 20 15s

206 Ito RatVM Zacopride 24 -40 na 500 -40 to 80 (10)

210 Ito HumanAM Cyamemazine

32-34 -50 500 60

214 Kv43 MonkeyCOS7 Pentamidine na -80 na 500 -40 to 60

215 Ito CanineVM Pioglitazone na -80 5ms -40 na 50

220 Ito RatVM Ajmaline RT -75 300 60

221 rKv42 rKv43 HEK293 RSD1235 25 -80 Brief -50 250 60 1Hz

222 Ito RatVM FK-506 22-24 -80 na 50 1000 -50 to 70 01Hz

224 hKv43 HEK293 Allitridi 22-23 -80 300 60 300 -80 to 60

245 Ito HumanAM DPO RT -80 150 40

256 Ito RabbitVM Carvedilol 34 -60 300 40 300 -50 to 60

263 Kv43 CHO (transient) Spironolactone canrenoic acid

RT -80 250 50 250 -90 to 50 10s

267 Ito ratVM Ebastine na -90 na 75 na -30 to 75

273 Ito CanineVM Thymol 37 -80 5ms- 40 400 50 400 -10 to 60

281 rKv14 Xenopus Oocyte Haloperidol 22 -80 300 0

283 Ito HumanAM Clotrimaxole 21-22 -50 300 50 300 -40 to 60 02Hz

VM= ventricular myocyte AM= Atrial myocyte

Pulse sequences Ito

Ito (Kir 43)

ID ChannelCurrent

Speciescells CPD Temp Vh(mV)

Prepulse(ms mV)

Activation(ms mV)

Voltage -Dependence(ms mV)

Intervalfrequency

47 hKv43hKChIP22 CHO Methadone RT -90 300 40

49 rKv43 MouseL cell Vanoxerine RT -80 300 20 15s

50 hKv43 HEK293 Alfuzosin RT -80 300 20 15s

206 Ito RatVM Zacopride 24 -40 na 500 -40 to 80 (10)

210 Ito HumanAM Cyamemazine

32-34 -50 500 60

214 Kv43 MonkeyCOS7 Pentamidine na -80 na 500 -40 to 60

215 Ito CanineVM Pioglitazone na -80 5ms -40 na 50

220 Ito RatVM Ajmaline RT -75 300 60

221 rKv42 rKv43 HEK293 RSD1235 25 -80 Brief -50 250 60 1Hz

222 Ito RatVM FK-506 22-24 -80 na 50 1000 -50 to 70 01Hz

224 hKv43 HEK293 Allitridi 22-23 -80 300 60 300 -80 to 60

245 Ito HumanAM DPO RT -80 150 40

256 Ito RabbitVM Carvedilol 34 -60 300 40 300 -50 to 60

263 Kv43 CHO (transient) Spironolactone canrenoic acid

RT -80 250 50 250 -90 to 50 10s

267 Ito ratVM Ebastine na -90 na 75 na -30 to 75

273 Ito CanineVM Thymol 37 -80 5ms- 40 400 50 400 -10 to 60

281 rKv14 Xenopus Oocyte Haloperidol 22 -80 300 0

283 Ito HumanAM Clotrimaxole 21-22 -50 300 50 300 -40 to 60 02Hz

VM= ventricular myocyte AM= Atrial myocyte

Kinetics Pulse Sequences Ito

ID channel cells kinetics Vh (mV)

1st (ms mv)

2nd (ms mV)

3rd (ms mv)

4th (ms mV)

224 hKv43 HEK293 Inactivation -80 -10 to 60

Channel availability -80 na -100 to 40 300 50

Activation conductance -80 8 na na

Recovery time from inactivation -80 30s 50 t -80 300 50 -80

Close state inactivation -90 300 50 3000 -90 t -50 300 50

283 Ito HumanAM Voltage dependence of activation and inactivation

-80 250 -100 to 30 (10) na 50

Recovery time from inactivation -80 30 -40 300 50 t -80 300 50

Extracellular buffer solution of ItoID NaCl NMDG KCl CaCl2 MgCl2 HEPES glucose dextrose NaH2PO4 Na-

acetate Blocker pH pH buffer

47 49 50 137 4 18 1 10 10 74 NaOH

206 135 54 18 1 10 20 033 CdCl2 (01 CaL) BaCl2 (02 Ik1) 74 NaOH

210 137 4 18 12 10 11 CdCl2 (02) 74 CdCl2

214 140 4 2 1 10 74 NaOH

215 144 56 25 12 5 11 74 na

220 135 54 09 09 10 10 033 CoCl2 (2) 74 NaOH

221 135 5 1 1 10 28 74 NaOH

222 135 4 18 1 2 11 TTX (001)Co++ (2) 74 NaOH

224 140 54 18 1 10 10 033 73 NaOH

245 132 4 12 10 111 Nisoldipine (04 uM) na na

256 143 54 18 05 5 56 025 Nisoldipine (0003) or CoCl2 (2) and TTX (001) 74 NaOH

263-1 130 4 1 1 10 10 74 NaOH

263-2 140 4 1 1 10 10 TTX (003) CoCl2 (2) 74 NaOH

267 137 54 2 1 10 10 74 NaOH

273 140 5 18 1 6 6 Nifedipine (0005) 74 NaOH

283 140 5 1 5 10 033 BaCl2 (01) CdCl2 (02) 74 NaOH

NMDG N-methyl-D-glucamine 263-1 CHO cells 263-2 mouse VM

Pipette solution of ItoID KCl K-

aspartate MgCl2 EGTA ATP GTP HEPESNa2-

phospho creatinne

NaCl CaCl2 KOH KH2PO4 pH pH buffer

47 49 50

130 5 5 4 10 72 KOH

206 150 1 5 3 5 73 KOH

210 130 1 5 5 5 7 72 KOH

214 110 142 10 4 10 517 72 KOH

215 45 100 1 10 na 5 72

220 25 130 1 1 5 01 5 3 725 KOH

221 130 1 5 4 01 10 72 KOH

222 130 05 3 04 25 72 KOH

224 20 110 1 5 5 01 10 5 72 KOH

245 20 110 5 5 10 72

256 80 40 10 5 5 5 065 60 72

263 42 80 5 5 5 3 10 72 KOH

267 135 5 5 5-10 10 19 72 KOH

273 45 100 1 10 3 5 72 KOH

283 20 110 1 5 5 01 10 5 72 KOH

Summary of IKur (Kv1531) protocolsbull Distribution a major repolarizing current in human atria but seems to play no

role in the ventricle bull Pharmacology selective IKur blockers to restore and maintain sinus rhythm

without a risk of ventricular pro-arrhythmia

bull Channel sources1 Transfected cell lines CHO L cells HEK 293 and Xenopus Oocyte (710 =

70)2 Myocyte (30) species ndash human AM and mouse VM

bull Temperature RT = 100bull Pulse sequences

ndash Holding range -80 to -50 mV (mode -80 mV)ndash Activation 20 - 60 mV (mode 50 mV)ndash Duration of activation 150 ndash 1000 ms (mode 300 ms)

1 IKur measurement steady state current at the end of depolarization and tail currents

Pulse sequences IKurID Channel

CurrentSpeciescells CPD Temp Vh

(mV)Prepulse Activation

(ms mV)Tail current

Voltage -Dependence(ms mV)

Interval (s)

49 hKv15 MouseL cell vanoxerine RT -80 300 20 10s

221 Kv15 HEK293 RSD1235 25 -80 200 60

224 hKv15 HEK293 Allitridi 22-23 -80 300 50 300 -40 to 60

245-1 hKv15 CHO DPO RT -80 150 40 na -30

245-2 Ikur HumanAM DPO RT -50 300 40 na -30

245-3 hKv15 XO DPO na -80 na 40

262 hKv15 CHO SSR149744C RT -70 1000 30 20s

263 hKv15 Mouse LtK- Spironolactonecanrenoic acid

RT -80 500 60 500 -40 500 -80 to 60

263 Ikur MouseVM Spironolactonecanrenoic acid

RT -80 200 40 500 50 10s

283 Ikur HumanAM clotrimazole 21-22 -50 100 40 150 50 na -30 150 -40 to 50

Ikur (Kv15)

Pipette solution of ItoID KCl K-

aspartate MgCl2 EGTA ATP GTP HEPESNa2-

phospho creatinne

NaCl CaCl2 KOH KH2PO4 pH pH buffer

47 49 50

130 5 5 4 10 72 KOH

206 150 1 5 3 5 73 KOH

210 130 1 5 5 5 7 72 KOH

214 110 142 10 4 10 517 72 KOH

215 45 100 1 10 na 5 72

220 25 130 1 1 5 01 5 3 725 KOH

221 130 1 5 4 01 10 72 KOH

222 130 05 3 04 25 72 KOH

224 20 110 1 5 5 01 10 5 72 KOH

245 20 110 5 5 10 72

256 80 40 10 5 5 5 065 60 72

263 42 80 5 5 5 3 10 72 KOH

267 135 5 5 5-10 10 19 72 KOH

273 45 100 1 10 3 5 72 KOH

283 20 110 1 5 5 01 10 5 72 KOH

Summary of IKur (Kv1531) protocolsbull Distribution a major repolarizing current in human atria but seems to play no

role in the ventricle bull Pharmacology selective IKur blockers to restore and maintain sinus rhythm

without a risk of ventricular pro-arrhythmia

bull Channel sources1 Transfected cell lines CHO L cells HEK 293 and Xenopus Oocyte (710 =

70)2 Myocyte (30) species ndash human AM and mouse VM

bull Temperature RT = 100bull Pulse sequences

ndash Holding range -80 to -50 mV (mode -80 mV)ndash Activation 20 - 60 mV (mode 50 mV)ndash Duration of activation 150 ndash 1000 ms (mode 300 ms)

1 IKur measurement steady state current at the end of depolarization and tail currents

Pulse sequences IKurID Channel

CurrentSpeciescells CPD Temp Vh

(mV)Prepulse Activation

(ms mV)Tail current

Voltage -Dependence(ms mV)

Interval (s)

49 hKv15 MouseL cell vanoxerine RT -80 300 20 10s

221 Kv15 HEK293 RSD1235 25 -80 200 60

224 hKv15 HEK293 Allitridi 22-23 -80 300 50 300 -40 to 60

245-1 hKv15 CHO DPO RT -80 150 40 na -30

245-2 Ikur HumanAM DPO RT -50 300 40 na -30

245-3 hKv15 XO DPO na -80 na 40

262 hKv15 CHO SSR149744C RT -70 1000 30 20s

263 hKv15 Mouse LtK- Spironolactonecanrenoic acid

RT -80 500 60 500 -40 500 -80 to 60

263 Ikur MouseVM Spironolactonecanrenoic acid

RT -80 200 40 500 50 10s

283 Ikur HumanAM clotrimazole 21-22 -50 100 40 150 50 na -30 150 -40 to 50

Ikur (Kv15)

Summary of IKur (Kv1531) protocolsbull Distribution a major repolarizing current in human atria but seems to play no

role in the ventricle bull Pharmacology selective IKur blockers to restore and maintain sinus rhythm

without a risk of ventricular pro-arrhythmia

bull Channel sources1 Transfected cell lines CHO L cells HEK 293 and Xenopus Oocyte (710 =

70)2 Myocyte (30) species ndash human AM and mouse VM

bull Temperature RT = 100bull Pulse sequences

ndash Holding range -80 to -50 mV (mode -80 mV)ndash Activation 20 - 60 mV (mode 50 mV)ndash Duration of activation 150 ndash 1000 ms (mode 300 ms)

1 IKur measurement steady state current at the end of depolarization and tail currents

Pulse sequences IKurID Channel

CurrentSpeciescells CPD Temp Vh

(mV)Prepulse Activation

(ms mV)Tail current

Voltage -Dependence(ms mV)

Interval (s)

49 hKv15 MouseL cell vanoxerine RT -80 300 20 10s

221 Kv15 HEK293 RSD1235 25 -80 200 60

224 hKv15 HEK293 Allitridi 22-23 -80 300 50 300 -40 to 60

245-1 hKv15 CHO DPO RT -80 150 40 na -30

245-2 Ikur HumanAM DPO RT -50 300 40 na -30

245-3 hKv15 XO DPO na -80 na 40

262 hKv15 CHO SSR149744C RT -70 1000 30 20s

263 hKv15 Mouse LtK- Spironolactonecanrenoic acid

RT -80 500 60 500 -40 500 -80 to 60

263 Ikur MouseVM Spironolactonecanrenoic acid

RT -80 200 40 500 50 10s

283 Ikur HumanAM clotrimazole 21-22 -50 100 40 150 50 na -30 150 -40 to 50

Ikur (Kv15)

Pulse sequences IKurID Channel

CurrentSpeciescells CPD Temp Vh

(mV)Prepulse Activation

(ms mV)Tail current

Voltage -Dependence(ms mV)

Interval (s)

49 hKv15 MouseL cell vanoxerine RT -80 300 20 10s

221 Kv15 HEK293 RSD1235 25 -80 200 60

224 hKv15 HEK293 Allitridi 22-23 -80 300 50 300 -40 to 60

245-1 hKv15 CHO DPO RT -80 150 40 na -30

245-2 Ikur HumanAM DPO RT -50 300 40 na -30

245-3 hKv15 XO DPO na -80 na 40

262 hKv15 CHO SSR149744C RT -70 1000 30 20s

263 hKv15 Mouse LtK- Spironolactonecanrenoic acid

RT -80 500 60 500 -40 500 -80 to 60

263 Ikur MouseVM Spironolactonecanrenoic acid

RT -80 200 40 500 50 10s

283 Ikur HumanAM clotrimazole 21-22 -50 100 40 150 50 na -30 150 -40 to 50

Ikur (Kv15)

Extracellular buffer solution of IKurID NaCl NMCG K-

gluconate KCl CaCl2 MgCl2 HEPES glucose dextrose NaH2PO4 Na-acetate Blocker pH pH

buffer

49 137 4 18 1 10 10 74 NaOH

206 135 54 18 1 10 10 033 CdCl2 (01) BaCl2 (02) 74 NaOH

210 137 4 18 12 10 11 CdCl2 (02) 74 NaOH

221 135 5 1 1 10 28 74 NaOH

224 140 54 18 1 10 10 033 73 NaOH

245-1 132 4 18 12 10 111 72 na

245-2 132 4 12 10 111 nisoldipine (04)

245-3 116 4 10 725

262 140 54 2 2 15 10 74 NaOH

263 130 4 1 1 10 10 74 NaOH

281 96 2 18 1 5 74 NaOH

283 140 5 1 1 5 10 033BaCl2 (02 Ik1) CdCl2 (Ica)

74 NaOH

245-1 CHO 245-2 human AM 245-3 XO (inside out VC)

Pipette solution of IKurID KCl K-

aspartate MgCl2 EGTA ATP GTP HEPESNa2-

phosphocreatinne

NaCl CaCl2 K-gluconate KH2PO4 pH pH

buffer

49 130 5 5 4 10 72 KOH

206 150 1 5 3 5 73 KOH

210 130 1 5 5 5 7 72 KOH

221 130 1 5 4 01 10 72 KOH

224 20 110 1 5 5 01 10 5 72 KOH

245-1 15 32 5 5 5 119 735

245-2 20 110 5 5 10 72

245-3 4 10 003 116 725

262 140 2 11 10 2 73 KOH

263 42 80 5 5 5 3 10 72 KOH

283 20 110 1 5 5 01 10 5 72 KOH

245-1 CHO 245-2 human AM 245-3 XO (inside out VC)

Summary of Iks protocolsbull Distribution all cell types but less in midmyocardial myocytes

bull Pharmacology Atrial and ventricular repolarizationHeart rate dependentbull subunit KCNQ1 (channel forming) Kv71 KvLQT1bull subunit KCNE1 (accessary unit activation of KCNQ1) minK (minimum potassium

subunit)

bull Channel sources1 transfected cell lines CHO HEK 293 Cos-7 cells and Xenopus Oocyte (2136 = 58)2 Myocyte (42) species ndash Canine VM Guinea pig VM rabbit VM and mouse VM (mode

GP)

bull Temperature RT = 1629 55 physiological temperature = 45

bull Pulse sequencesndash Holding potential range = -90 to -30 (mode -80)ndash Depolarization potential 20-60 mV ndash Duration of depolarization 50 ms ndash 10s (mode 2s)ndash Repolarization potential -80 to -10 mV (mode -40 mV)ndash Duration of repolarization 05 ndash 5s (mode 2s)

bull IKs measurement steady state current at the end of depolarization and tail currents

Pulse sequences IKsID Channel

CurrentSpeciescells Temp Vh

(mV)Prepulse (ms mV)

Activation (s mV)

deactivation(s mV)

Interval (s)

47 hKvLQT1hminK CHO RT -80 2 40 01 Hz

49 hKv711hKCNE1 CHO RT -80 2 20 05 -40 15s

50 hKv711hKCNE1 HEk293 RT -80 2 20 05 -40 15s

184 IKs canineepiardial cells 37 -50 2 40 na -30 na

214 Kv711minK HEK293 na -80 5 -60 to 60 5s -50 na

215 IKs caninemyocyte na -40 5 50

224 hKCNQ1hKCNE1 HEK293 22-23 -80 3 -60 to 60 (20) na -50 na

229 hCNE1-hKCNQ1 Transgenic mousemyocyte RT -75 20 -40(Na block) 2 60 2 -40 006 Hz

230 KCNQ1KCNE1 XenopusOocyte 20-22 -80 5 30 na -70

232 KCNQ1KCNE1 HEK293 36 -80 2 -80 to 110 (10) 2 -60 6s

233 KCNQ1KCNE1 CHO na -80 80 -90 3 20 1 -50 005Hz

237 KCNQ1 CHO na -80 na -80 to 60 1 -40

239 KCNQ1KCNE1 HEK293 RT -70 1 -70 to 60 na -40

241 KCNQ1 andor minK MouseCOS-7 22-25 -80 3 -120 to 60 (20) 2 -30

242 hKvLQT1hminK HEK293 19-36 -80 3 -50 to 60 (10) 5 -50

242 hKvLQT1hminK HEK293 19-36 -50 1 or 2 50 na -50 10s

244 KCNQ1 andor KCNE1 CHO na -90 3 -70 to 70 (10) na -60

245 IKs GPMyocyte 35 -50 1 50

249 KCNQ1KCNE1 HEK293 33-34 -40 1 -40 to 80 (10) na -40

Iks (Kv71minK kvLQT1 KCNQ1KCNE1)

Pipette solution of IKurID KCl K-

aspartate MgCl2 EGTA ATP GTP HEPESNa2-

phosphocreatinne

NaCl CaCl2 K-gluconate KH2PO4 pH pH

buffer

49 130 5 5 4 10 72 KOH

206 150 1 5 3 5 73 KOH

210 130 1 5 5 5 7 72 KOH

221 130 1 5 4 01 10 72 KOH

224 20 110 1 5 5 01 10 5 72 KOH

245-1 15 32 5 5 5 119 735

245-2 20 110 5 5 10 72

245-3 4 10 003 116 725

262 140 2 11 10 2 73 KOH

263 42 80 5 5 5 3 10 72 KOH

283 20 110 1 5 5 01 10 5 72 KOH

245-1 CHO 245-2 human AM 245-3 XO (inside out VC)

Summary of Iks protocolsbull Distribution all cell types but less in midmyocardial myocytes

bull Pharmacology Atrial and ventricular repolarizationHeart rate dependentbull subunit KCNQ1 (channel forming) Kv71 KvLQT1bull subunit KCNE1 (accessary unit activation of KCNQ1) minK (minimum potassium

subunit)

bull Channel sources1 transfected cell lines CHO HEK 293 Cos-7 cells and Xenopus Oocyte (2136 = 58)2 Myocyte (42) species ndash Canine VM Guinea pig VM rabbit VM and mouse VM (mode

GP)

bull Temperature RT = 1629 55 physiological temperature = 45

bull Pulse sequencesndash Holding potential range = -90 to -30 (mode -80)ndash Depolarization potential 20-60 mV ndash Duration of depolarization 50 ms ndash 10s (mode 2s)ndash Repolarization potential -80 to -10 mV (mode -40 mV)ndash Duration of repolarization 05 ndash 5s (mode 2s)

bull IKs measurement steady state current at the end of depolarization and tail currents

Pulse sequences IKsID Channel

CurrentSpeciescells Temp Vh

(mV)Prepulse (ms mV)

Activation (s mV)

deactivation(s mV)

Interval (s)

47 hKvLQT1hminK CHO RT -80 2 40 01 Hz

49 hKv711hKCNE1 CHO RT -80 2 20 05 -40 15s

50 hKv711hKCNE1 HEk293 RT -80 2 20 05 -40 15s

184 IKs canineepiardial cells 37 -50 2 40 na -30 na

214 Kv711minK HEK293 na -80 5 -60 to 60 5s -50 na

215 IKs caninemyocyte na -40 5 50

224 hKCNQ1hKCNE1 HEK293 22-23 -80 3 -60 to 60 (20) na -50 na

229 hCNE1-hKCNQ1 Transgenic mousemyocyte RT -75 20 -40(Na block) 2 60 2 -40 006 Hz

230 KCNQ1KCNE1 XenopusOocyte 20-22 -80 5 30 na -70

232 KCNQ1KCNE1 HEK293 36 -80 2 -80 to 110 (10) 2 -60 6s

233 KCNQ1KCNE1 CHO na -80 80 -90 3 20 1 -50 005Hz

237 KCNQ1 CHO na -80 na -80 to 60 1 -40

239 KCNQ1KCNE1 HEK293 RT -70 1 -70 to 60 na -40

241 KCNQ1 andor minK MouseCOS-7 22-25 -80 3 -120 to 60 (20) 2 -30

242 hKvLQT1hminK HEK293 19-36 -80 3 -50 to 60 (10) 5 -50

242 hKvLQT1hminK HEK293 19-36 -50 1 or 2 50 na -50 10s

244 KCNQ1 andor KCNE1 CHO na -90 3 -70 to 70 (10) na -60

245 IKs GPMyocyte 35 -50 1 50

249 KCNQ1KCNE1 HEK293 33-34 -40 1 -40 to 80 (10) na -40

Iks (Kv71minK kvLQT1 KCNQ1KCNE1)

Summary of Iks protocolsbull Distribution all cell types but less in midmyocardial myocytes

bull Pharmacology Atrial and ventricular repolarizationHeart rate dependentbull subunit KCNQ1 (channel forming) Kv71 KvLQT1bull subunit KCNE1 (accessary unit activation of KCNQ1) minK (minimum potassium

subunit)

bull Channel sources1 transfected cell lines CHO HEK 293 Cos-7 cells and Xenopus Oocyte (2136 = 58)2 Myocyte (42) species ndash Canine VM Guinea pig VM rabbit VM and mouse VM (mode

GP)

bull Temperature RT = 1629 55 physiological temperature = 45

bull Pulse sequencesndash Holding potential range = -90 to -30 (mode -80)ndash Depolarization potential 20-60 mV ndash Duration of depolarization 50 ms ndash 10s (mode 2s)ndash Repolarization potential -80 to -10 mV (mode -40 mV)ndash Duration of repolarization 05 ndash 5s (mode 2s)

bull IKs measurement steady state current at the end of depolarization and tail currents

Pulse sequences IKsID Channel

CurrentSpeciescells Temp Vh

(mV)Prepulse (ms mV)

Activation (s mV)

deactivation(s mV)

Interval (s)

47 hKvLQT1hminK CHO RT -80 2 40 01 Hz

49 hKv711hKCNE1 CHO RT -80 2 20 05 -40 15s

50 hKv711hKCNE1 HEk293 RT -80 2 20 05 -40 15s

184 IKs canineepiardial cells 37 -50 2 40 na -30 na

214 Kv711minK HEK293 na -80 5 -60 to 60 5s -50 na

215 IKs caninemyocyte na -40 5 50

224 hKCNQ1hKCNE1 HEK293 22-23 -80 3 -60 to 60 (20) na -50 na

229 hCNE1-hKCNQ1 Transgenic mousemyocyte RT -75 20 -40(Na block) 2 60 2 -40 006 Hz

230 KCNQ1KCNE1 XenopusOocyte 20-22 -80 5 30 na -70

232 KCNQ1KCNE1 HEK293 36 -80 2 -80 to 110 (10) 2 -60 6s

233 KCNQ1KCNE1 CHO na -80 80 -90 3 20 1 -50 005Hz

237 KCNQ1 CHO na -80 na -80 to 60 1 -40

239 KCNQ1KCNE1 HEK293 RT -70 1 -70 to 60 na -40

241 KCNQ1 andor minK MouseCOS-7 22-25 -80 3 -120 to 60 (20) 2 -30

242 hKvLQT1hminK HEK293 19-36 -80 3 -50 to 60 (10) 5 -50

242 hKvLQT1hminK HEK293 19-36 -50 1 or 2 50 na -50 10s

244 KCNQ1 andor KCNE1 CHO na -90 3 -70 to 70 (10) na -60

245 IKs GPMyocyte 35 -50 1 50

249 KCNQ1KCNE1 HEK293 33-34 -40 1 -40 to 80 (10) na -40

Iks (Kv71minK kvLQT1 KCNQ1KCNE1)

Pulse sequences IKsID Channel

CurrentSpeciescells Temp Vh

(mV)Prepulse (ms mV)

Activation (s mV)

deactivation(s mV)

Interval (s)

47 hKvLQT1hminK CHO RT -80 2 40 01 Hz

49 hKv711hKCNE1 CHO RT -80 2 20 05 -40 15s

50 hKv711hKCNE1 HEk293 RT -80 2 20 05 -40 15s

184 IKs canineepiardial cells 37 -50 2 40 na -30 na

214 Kv711minK HEK293 na -80 5 -60 to 60 5s -50 na

215 IKs caninemyocyte na -40 5 50

224 hKCNQ1hKCNE1 HEK293 22-23 -80 3 -60 to 60 (20) na -50 na

229 hCNE1-hKCNQ1 Transgenic mousemyocyte RT -75 20 -40(Na block) 2 60 2 -40 006 Hz

230 KCNQ1KCNE1 XenopusOocyte 20-22 -80 5 30 na -70

232 KCNQ1KCNE1 HEK293 36 -80 2 -80 to 110 (10) 2 -60 6s

233 KCNQ1KCNE1 CHO na -80 80 -90 3 20 1 -50 005Hz

237 KCNQ1 CHO na -80 na -80 to 60 1 -40

239 KCNQ1KCNE1 HEK293 RT -70 1 -70 to 60 na -40

241 KCNQ1 andor minK MouseCOS-7 22-25 -80 3 -120 to 60 (20) 2 -30

242 hKvLQT1hminK HEK293 19-36 -80 3 -50 to 60 (10) 5 -50

242 hKvLQT1hminK HEK293 19-36 -50 1 or 2 50 na -50 10s

244 KCNQ1 andor KCNE1 CHO na -90 3 -70 to 70 (10) na -60

245 IKs GPMyocyte 35 -50 1 50

249 KCNQ1KCNE1 HEK293 33-34 -40 1 -40 to 80 (10) na -40

Iks (Kv71minK kvLQT1 KCNQ1KCNE1)

Pulse sequences IKsID Channel

CurrentSpeciescells Temp Vh

(mV)Prepulse (ms mV)

Activation(s mV)

Voltage dependent(s mV)

Deactivation(s mV)

Interval (s)

251 IKs GPMyocyte 22-2535-37 -40 2 60 2 -40 to 80 (20) na -40

252 IKs GPMyocyte 35 -80 200 -50 3 60 2 -40 60s

252 hminK XenopusOocyte 35 -80 10 20 10 -40 to 60 2 -80

256 IKs Rabbitmyocyte 34 -50 3 -40 to 50 na

260 hKCNQ1hKCNE1 HEK293 RT -80 25 -70 to 100 (10) 25 -40 15s

262 IKs GPMyocyte 35 -80 1 40 5 -10

263 Kv71 and minK CHO RT -80 2 40 na -40

264 IKs GPmyocyte na -80 200 -40 05 50 na -40

267 IKs GPmyocyte na -90 na 0 na 40 01- 015 0-80 -90

273 IKs Caninemyocyte 37 -40 3 50 na -40

274 IKs GPmyocyte 36 -30 05 50 or 2 70 05 (2) -30 to 70 na -30

275 KCNQ1KCNE1 CHO 21 -80 1 40 01Hz

276 hKCNQ1hminK CHO 22 -80 6 40 na -80 005Hz

278 IKs Canine myocyte 35-37 -50 3 -50 to 50 2s -40

283 KCNQ1KCNE1 HEK293 21-22 -80 3 40 3 -80 to 40 na -40

286 IKs GPmyocyte 34-36 -50 005 30 005 ndash3 -50 to 30 na -50

287 Iks GPmyocyte 36 -80 na -40 2 60 2 -40 to 80 na -40

Iks (Kv71minK kvLQT1 KCNQ1KCNE1)

Extracellular buffer solution of IKsID NaCl Na-

Gluc KCl CaCl2 MgCl2 HEPES glucose Minor components Blocker pH pH buffer

47 49 50

137 4 18 1 10 10 74 NaOH

184 145 4 18 18 10 11 NaH2PO4 1 E4031 (5uM) Cadmium (300 uM) 74 NaOH

214 140 4 2 1 10 74 NaOH

215 144 5 56 25 5 Dextrose 11 Nifedipine (5 uM) E4031 (1 uM) 74 NaOH

224 140 54 18 1 10 10 NaH2PO4 033 73 NaOH

229 132 48 2 12 10 5 E4031 nisoldipine 74 NaOH

230 96 2 18 1 5 74 NaOH

232 140 54 18 1 5 55 74 NaOH

233 17 120 4 2 1 10 10 74 NaOH

237 130 5 18 1 10 Na-Acetate 28 74 NaOH

239 150 54 18 4 10 74 NaOH

241 135 5 25 12 5 10 74 NaOH

242 132 4 18 12 10 111 74 NaOH

244 140 4 18 12 55 11 74 NaOH

249 140 54 18 05 5 55 NaH2PO4 033 74 NaOH

250 140 54 18 1 5 10 74 NaOH

Extracellular buffer solution of IKsID NaCl Na-

Gluc KCl CaCl2 MgCl2 HEPES glucose Minor components Blocker pH pH buffer

251 1 1 2 10 N-methly-d-glucamine 132

lanthanum chloride 005 mMcadmium chloride 02 mM 74 HCl

252 140 47 13 1 10 10 74 NaOH

256 09 5 N-methly-d-glucamine 149

Nisoldipine 3 uME4031 10 uM 74 HCl

260 140 54 18 1 5 10 NaH2PO4 03 73 NaOH

262 137 4 18 2 10 115 Pyrubic acid 5 Thiamine 01 74 NaOH

263 130 4 1 1 10 10 TTX 003 mM CoCl2 2 mM 74 NaOH

267 137 54 2 1 10 10 74 NaOH

273 135 47 10 10 KH2PO4 12 MgSO4 12 NaHCO3 44

Nifetidine 5 uM 4-aminopyridine 3 mM 72 NaOH

274 140 1 5 10 Cd2+ 02 mM 74 NaOH

275 130 54 18 1 10 5 74 NaOH

278 136 54 1 1 5 Dextrose 10 NaH2PO4 033 E4031 5 uM 74 NaOH

283 140 5 1 1 5 10 NaH2PO4 033 74 NaOH

286 09 5 5 N-methly-d-glucamine 149

Nisoldipine 04 uME4031 5 uM 74 NaOH

Pipette solution of IKsID KCl K-asp

K-glucona

te

K-glutama

teMgCl2 EGTA ATP GTP HEPES Na2-

PhosCre NaCl CaCl2 KH2PO4 pH pH buffer

49 130 5 5 4 10 72 KOH

184 20 125 1 10 5 5 71 KOH

214 110 142 10 4 4 517 72 KOH

215 45 100 1 10 na 5 72

224 20 110 1 5 5 01 10 5 72 KOH

229 110 11 5 10 55 73

230 3000

232 20 125 10 10

233 140 1 1 20 73 KOH

237 10 125 1 10 5 10 10 72 KOH

239 30 130 01 4 10 74

241 117 2 11 11 10 1 72

242 15 119 32 5 5 5 74 KOH

244 130 1 5 5 10 74 KOH

249 130 5 5 01 5 5 74 HCl

250 80 40 10 5 01 5 40 065 72 HCl

251 50 60 1 11 5 10 1 74 KOH

252 140 11 1 10 10 72 KOH

amphotericin-B

Pipette solution of IKsID KCl K-asp

K-glucona

te

K-glutama

teMgCl2 EGTA ATP GTP HEPES Na2-

PhosCre NaCl CaCl2 KH2PO4 pH pH buffer

256 80 40 10 5 5 5 065 72

262 150 1 5 4 5 5 73 KOH

263^ 42 80 5 5 5 3 10 72 KOH

267 135 5 5 5-10 10 19 72 KOH

273 45 100 1 10 3 5 72 KOH

274 40 106 1 5 4 5 72 KOH

276 20 110 1 10 5 10 10 1 72 KOH

278 20 110 1 10 5 01 10 5 72 KOH

283 20 110 1 5 5 01 1 5 72 KOH

286 50 1 5 5 01 5 2 10 72 KOH

^phosphatidylinositol 45-diphosphate (5 mgml 1) was added to the internal solution to avoid the rundown of the current 70 L-aspartic acid 70 KOH

Summary of IK1 protocolsbull Distribution higher in ventricle than atrial

bull Pharmacology establish resting membrane potential selective antiarrhythmia strategy

bull Channel sources1 Transfected cell lines CHO HEK 293 and Xenopus Oocyte (831 = 26)2 Myocyte (74) species ndash Canine VM rat VM Frog VM Fish VM Cat VM Guinea pig VM

and rabbit VM (mode Rat and GP)

bull Temperaturendash RT = 1020 50 ndash physiological temperature = 50

bull Pulse sequencesndash Holding range = -120 to -30 (mode -40)ndash Activation range -140 to 100 mV (mostly voltage dependence protocols)ndash Duration of activation 01 to 4s (mode 05s)

bull IK1 measurement steady state current at the end of depolarization

Pulse sequences IK1

ID ChannelCurrent

Speciescells Temp Vh(mV)

Potency (s mV)

Voltage -Dependence(m mV)

Interval Frequency

47 hKir21 HamsterCHO RT -30 1 -120 01 Hz

184 IK1 CanineMyocyte 37 -40 1 -100 to 0

206 IK1 RatMyocyte 24 -40 05 -120 to 20

207 IK1 RatMyocyte 35 -40 05 -10 to 100 (10) 5s

208 IK1 RatMyocyte 23 -40 03 -120 to 50 3s

209 IK1 FrogMyocyte 20-21 -100 na -120 to 50 02 Hz

210 IK1 HumanAM 32-34 -75 05 -100

213 IK1 FishMyocyte 18 -80 1 -120 to 30 5s

213 IK1 RatMyocyte 375 -40 1 -120 to 30 5s

214 hKIr21 22 23 HumanHEK293 na -40 na -100 to 100 5s

215 IK1 CanineMyocyte na -40 04 -135 to -65

216 IK1 RatMyocyte 20-22 -80 na -120 to -80

217 Kir21 23 HumanHEK293 na -80 4 -120 to -20

217 IK1 CatMyocyte na -40 3 -120 to -30

220 IK1 RatMyocyte RT -120 na -135 to -75

222 IK1 RatMyocyte 22-24 -80 4 -120 to 50

224 hKIr21 HumanHEK293 22-23 -40 na -120 to 0

Extracellular buffer solution of IKsID NaCl Na-

Gluc KCl CaCl2 MgCl2 HEPES glucose Minor components Blocker pH pH buffer

251 1 1 2 10 N-methly-d-glucamine 132

lanthanum chloride 005 mMcadmium chloride 02 mM 74 HCl

252 140 47 13 1 10 10 74 NaOH

256 09 5 N-methly-d-glucamine 149

Nisoldipine 3 uME4031 10 uM 74 HCl

260 140 54 18 1 5 10 NaH2PO4 03 73 NaOH

262 137 4 18 2 10 115 Pyrubic acid 5 Thiamine 01 74 NaOH

263 130 4 1 1 10 10 TTX 003 mM CoCl2 2 mM 74 NaOH

267 137 54 2 1 10 10 74 NaOH

273 135 47 10 10 KH2PO4 12 MgSO4 12 NaHCO3 44

Nifetidine 5 uM 4-aminopyridine 3 mM 72 NaOH

274 140 1 5 10 Cd2+ 02 mM 74 NaOH

275 130 54 18 1 10 5 74 NaOH

278 136 54 1 1 5 Dextrose 10 NaH2PO4 033 E4031 5 uM 74 NaOH

283 140 5 1 1 5 10 NaH2PO4 033 74 NaOH

286 09 5 5 N-methly-d-glucamine 149

Nisoldipine 04 uME4031 5 uM 74 NaOH

Pipette solution of IKsID KCl K-asp

K-glucona

te

K-glutama

teMgCl2 EGTA ATP GTP HEPES Na2-

PhosCre NaCl CaCl2 KH2PO4 pH pH buffer

49 130 5 5 4 10 72 KOH

184 20 125 1 10 5 5 71 KOH

214 110 142 10 4 4 517 72 KOH

215 45 100 1 10 na 5 72

224 20 110 1 5 5 01 10 5 72 KOH

229 110 11 5 10 55 73

230 3000

232 20 125 10 10

233 140 1 1 20 73 KOH

237 10 125 1 10 5 10 10 72 KOH

239 30 130 01 4 10 74

241 117 2 11 11 10 1 72

242 15 119 32 5 5 5 74 KOH

244 130 1 5 5 10 74 KOH

249 130 5 5 01 5 5 74 HCl

250 80 40 10 5 01 5 40 065 72 HCl

251 50 60 1 11 5 10 1 74 KOH

252 140 11 1 10 10 72 KOH

amphotericin-B

Pipette solution of IKsID KCl K-asp

K-glucona

te

K-glutama

teMgCl2 EGTA ATP GTP HEPES Na2-

PhosCre NaCl CaCl2 KH2PO4 pH pH buffer

256 80 40 10 5 5 5 065 72

262 150 1 5 4 5 5 73 KOH

263^ 42 80 5 5 5 3 10 72 KOH

267 135 5 5 5-10 10 19 72 KOH

273 45 100 1 10 3 5 72 KOH

274 40 106 1 5 4 5 72 KOH

276 20 110 1 10 5 10 10 1 72 KOH

278 20 110 1 10 5 01 10 5 72 KOH

283 20 110 1 5 5 01 1 5 72 KOH

286 50 1 5 5 01 5 2 10 72 KOH

^phosphatidylinositol 45-diphosphate (5 mgml 1) was added to the internal solution to avoid the rundown of the current 70 L-aspartic acid 70 KOH

Summary of IK1 protocolsbull Distribution higher in ventricle than atrial

bull Pharmacology establish resting membrane potential selective antiarrhythmia strategy

bull Channel sources1 Transfected cell lines CHO HEK 293 and Xenopus Oocyte (831 = 26)2 Myocyte (74) species ndash Canine VM rat VM Frog VM Fish VM Cat VM Guinea pig VM

and rabbit VM (mode Rat and GP)

bull Temperaturendash RT = 1020 50 ndash physiological temperature = 50

bull Pulse sequencesndash Holding range = -120 to -30 (mode -40)ndash Activation range -140 to 100 mV (mostly voltage dependence protocols)ndash Duration of activation 01 to 4s (mode 05s)

bull IK1 measurement steady state current at the end of depolarization

Pulse sequences IK1

ID ChannelCurrent

Speciescells Temp Vh(mV)

Potency (s mV)

Voltage -Dependence(m mV)

Interval Frequency

47 hKir21 HamsterCHO RT -30 1 -120 01 Hz

184 IK1 CanineMyocyte 37 -40 1 -100 to 0

206 IK1 RatMyocyte 24 -40 05 -120 to 20

207 IK1 RatMyocyte 35 -40 05 -10 to 100 (10) 5s

208 IK1 RatMyocyte 23 -40 03 -120 to 50 3s

209 IK1 FrogMyocyte 20-21 -100 na -120 to 50 02 Hz

210 IK1 HumanAM 32-34 -75 05 -100

213 IK1 FishMyocyte 18 -80 1 -120 to 30 5s

213 IK1 RatMyocyte 375 -40 1 -120 to 30 5s

214 hKIr21 22 23 HumanHEK293 na -40 na -100 to 100 5s

215 IK1 CanineMyocyte na -40 04 -135 to -65

216 IK1 RatMyocyte 20-22 -80 na -120 to -80

217 Kir21 23 HumanHEK293 na -80 4 -120 to -20

217 IK1 CatMyocyte na -40 3 -120 to -30

220 IK1 RatMyocyte RT -120 na -135 to -75

222 IK1 RatMyocyte 22-24 -80 4 -120 to 50

224 hKIr21 HumanHEK293 22-23 -40 na -120 to 0

Pipette solution of IKsID KCl K-asp

K-glucona

te

K-glutama

teMgCl2 EGTA ATP GTP HEPES Na2-

PhosCre NaCl CaCl2 KH2PO4 pH pH buffer

49 130 5 5 4 10 72 KOH

184 20 125 1 10 5 5 71 KOH

214 110 142 10 4 4 517 72 KOH

215 45 100 1 10 na 5 72

224 20 110 1 5 5 01 10 5 72 KOH

229 110 11 5 10 55 73

230 3000

232 20 125 10 10

233 140 1 1 20 73 KOH

237 10 125 1 10 5 10 10 72 KOH

239 30 130 01 4 10 74

241 117 2 11 11 10 1 72

242 15 119 32 5 5 5 74 KOH

244 130 1 5 5 10 74 KOH

249 130 5 5 01 5 5 74 HCl

250 80 40 10 5 01 5 40 065 72 HCl

251 50 60 1 11 5 10 1 74 KOH

252 140 11 1 10 10 72 KOH

amphotericin-B

Pipette solution of IKsID KCl K-asp

K-glucona

te

K-glutama

teMgCl2 EGTA ATP GTP HEPES Na2-

PhosCre NaCl CaCl2 KH2PO4 pH pH buffer

256 80 40 10 5 5 5 065 72

262 150 1 5 4 5 5 73 KOH

263^ 42 80 5 5 5 3 10 72 KOH

267 135 5 5 5-10 10 19 72 KOH

273 45 100 1 10 3 5 72 KOH

274 40 106 1 5 4 5 72 KOH

276 20 110 1 10 5 10 10 1 72 KOH

278 20 110 1 10 5 01 10 5 72 KOH

283 20 110 1 5 5 01 1 5 72 KOH

286 50 1 5 5 01 5 2 10 72 KOH

^phosphatidylinositol 45-diphosphate (5 mgml 1) was added to the internal solution to avoid the rundown of the current 70 L-aspartic acid 70 KOH

Summary of IK1 protocolsbull Distribution higher in ventricle than atrial

bull Pharmacology establish resting membrane potential selective antiarrhythmia strategy

bull Channel sources1 Transfected cell lines CHO HEK 293 and Xenopus Oocyte (831 = 26)2 Myocyte (74) species ndash Canine VM rat VM Frog VM Fish VM Cat VM Guinea pig VM

and rabbit VM (mode Rat and GP)

bull Temperaturendash RT = 1020 50 ndash physiological temperature = 50

bull Pulse sequencesndash Holding range = -120 to -30 (mode -40)ndash Activation range -140 to 100 mV (mostly voltage dependence protocols)ndash Duration of activation 01 to 4s (mode 05s)

bull IK1 measurement steady state current at the end of depolarization

Pulse sequences IK1

ID ChannelCurrent

Speciescells Temp Vh(mV)

Potency (s mV)

Voltage -Dependence(m mV)

Interval Frequency

47 hKir21 HamsterCHO RT -30 1 -120 01 Hz

184 IK1 CanineMyocyte 37 -40 1 -100 to 0

206 IK1 RatMyocyte 24 -40 05 -120 to 20

207 IK1 RatMyocyte 35 -40 05 -10 to 100 (10) 5s

208 IK1 RatMyocyte 23 -40 03 -120 to 50 3s

209 IK1 FrogMyocyte 20-21 -100 na -120 to 50 02 Hz

210 IK1 HumanAM 32-34 -75 05 -100

213 IK1 FishMyocyte 18 -80 1 -120 to 30 5s

213 IK1 RatMyocyte 375 -40 1 -120 to 30 5s

214 hKIr21 22 23 HumanHEK293 na -40 na -100 to 100 5s

215 IK1 CanineMyocyte na -40 04 -135 to -65

216 IK1 RatMyocyte 20-22 -80 na -120 to -80

217 Kir21 23 HumanHEK293 na -80 4 -120 to -20

217 IK1 CatMyocyte na -40 3 -120 to -30

220 IK1 RatMyocyte RT -120 na -135 to -75

222 IK1 RatMyocyte 22-24 -80 4 -120 to 50

224 hKIr21 HumanHEK293 22-23 -40 na -120 to 0

Pipette solution of IKsID KCl K-asp

K-glucona

te

K-glutama

teMgCl2 EGTA ATP GTP HEPES Na2-

PhosCre NaCl CaCl2 KH2PO4 pH pH buffer

256 80 40 10 5 5 5 065 72

262 150 1 5 4 5 5 73 KOH

263^ 42 80 5 5 5 3 10 72 KOH

267 135 5 5 5-10 10 19 72 KOH

273 45 100 1 10 3 5 72 KOH

274 40 106 1 5 4 5 72 KOH

276 20 110 1 10 5 10 10 1 72 KOH

278 20 110 1 10 5 01 10 5 72 KOH

283 20 110 1 5 5 01 1 5 72 KOH

286 50 1 5 5 01 5 2 10 72 KOH

^phosphatidylinositol 45-diphosphate (5 mgml 1) was added to the internal solution to avoid the rundown of the current 70 L-aspartic acid 70 KOH

Summary of IK1 protocolsbull Distribution higher in ventricle than atrial

bull Pharmacology establish resting membrane potential selective antiarrhythmia strategy

bull Channel sources1 Transfected cell lines CHO HEK 293 and Xenopus Oocyte (831 = 26)2 Myocyte (74) species ndash Canine VM rat VM Frog VM Fish VM Cat VM Guinea pig VM

and rabbit VM (mode Rat and GP)

bull Temperaturendash RT = 1020 50 ndash physiological temperature = 50

bull Pulse sequencesndash Holding range = -120 to -30 (mode -40)ndash Activation range -140 to 100 mV (mostly voltage dependence protocols)ndash Duration of activation 01 to 4s (mode 05s)

bull IK1 measurement steady state current at the end of depolarization

Pulse sequences IK1

ID ChannelCurrent

Speciescells Temp Vh(mV)

Potency (s mV)

Voltage -Dependence(m mV)

Interval Frequency

47 hKir21 HamsterCHO RT -30 1 -120 01 Hz

184 IK1 CanineMyocyte 37 -40 1 -100 to 0

206 IK1 RatMyocyte 24 -40 05 -120 to 20

207 IK1 RatMyocyte 35 -40 05 -10 to 100 (10) 5s

208 IK1 RatMyocyte 23 -40 03 -120 to 50 3s

209 IK1 FrogMyocyte 20-21 -100 na -120 to 50 02 Hz

210 IK1 HumanAM 32-34 -75 05 -100

213 IK1 FishMyocyte 18 -80 1 -120 to 30 5s

213 IK1 RatMyocyte 375 -40 1 -120 to 30 5s

214 hKIr21 22 23 HumanHEK293 na -40 na -100 to 100 5s

215 IK1 CanineMyocyte na -40 04 -135 to -65

216 IK1 RatMyocyte 20-22 -80 na -120 to -80

217 Kir21 23 HumanHEK293 na -80 4 -120 to -20

217 IK1 CatMyocyte na -40 3 -120 to -30

220 IK1 RatMyocyte RT -120 na -135 to -75

222 IK1 RatMyocyte 22-24 -80 4 -120 to 50

224 hKIr21 HumanHEK293 22-23 -40 na -120 to 0

Summary of IK1 protocolsbull Distribution higher in ventricle than atrial

bull Pharmacology establish resting membrane potential selective antiarrhythmia strategy

bull Channel sources1 Transfected cell lines CHO HEK 293 and Xenopus Oocyte (831 = 26)2 Myocyte (74) species ndash Canine VM rat VM Frog VM Fish VM Cat VM Guinea pig VM

and rabbit VM (mode Rat and GP)

bull Temperaturendash RT = 1020 50 ndash physiological temperature = 50

bull Pulse sequencesndash Holding range = -120 to -30 (mode -40)ndash Activation range -140 to 100 mV (mostly voltage dependence protocols)ndash Duration of activation 01 to 4s (mode 05s)

bull IK1 measurement steady state current at the end of depolarization

Pulse sequences IK1

ID ChannelCurrent

Speciescells Temp Vh(mV)

Potency (s mV)

Voltage -Dependence(m mV)

Interval Frequency

47 hKir21 HamsterCHO RT -30 1 -120 01 Hz

184 IK1 CanineMyocyte 37 -40 1 -100 to 0

206 IK1 RatMyocyte 24 -40 05 -120 to 20

207 IK1 RatMyocyte 35 -40 05 -10 to 100 (10) 5s

208 IK1 RatMyocyte 23 -40 03 -120 to 50 3s

209 IK1 FrogMyocyte 20-21 -100 na -120 to 50 02 Hz

210 IK1 HumanAM 32-34 -75 05 -100

213 IK1 FishMyocyte 18 -80 1 -120 to 30 5s

213 IK1 RatMyocyte 375 -40 1 -120 to 30 5s

214 hKIr21 22 23 HumanHEK293 na -40 na -100 to 100 5s

215 IK1 CanineMyocyte na -40 04 -135 to -65

216 IK1 RatMyocyte 20-22 -80 na -120 to -80

217 Kir21 23 HumanHEK293 na -80 4 -120 to -20

217 IK1 CatMyocyte na -40 3 -120 to -30

220 IK1 RatMyocyte RT -120 na -135 to -75

222 IK1 RatMyocyte 22-24 -80 4 -120 to 50

224 hKIr21 HumanHEK293 22-23 -40 na -120 to 0

Pulse sequences IK1

ID ChannelCurrent

Speciescells Temp Vh(mV)

Potency (s mV)

Voltage -Dependence(m mV)

Interval Frequency

47 hKir21 HamsterCHO RT -30 1 -120 01 Hz

184 IK1 CanineMyocyte 37 -40 1 -100 to 0

206 IK1 RatMyocyte 24 -40 05 -120 to 20

207 IK1 RatMyocyte 35 -40 05 -10 to 100 (10) 5s

208 IK1 RatMyocyte 23 -40 03 -120 to 50 3s

209 IK1 FrogMyocyte 20-21 -100 na -120 to 50 02 Hz

210 IK1 HumanAM 32-34 -75 05 -100

213 IK1 FishMyocyte 18 -80 1 -120 to 30 5s

213 IK1 RatMyocyte 375 -40 1 -120 to 30 5s

214 hKIr21 22 23 HumanHEK293 na -40 na -100 to 100 5s

215 IK1 CanineMyocyte na -40 04 -135 to -65

216 IK1 RatMyocyte 20-22 -80 na -120 to -80

217 Kir21 23 HumanHEK293 na -80 4 -120 to -20

217 IK1 CatMyocyte na -40 3 -120 to -30

220 IK1 RatMyocyte RT -120 na -135 to -75

222 IK1 RatMyocyte 22-24 -80 4 -120 to 50

224 hKIr21 HumanHEK293 22-23 -40 na -120 to 0

Pulse sequences IK1ID Channel

CurrentSpeciescells Temp Vh

(mV)Prepulse(ms mV)

Potency (ms mV)

Voltage -Dependence(ms mV)

IntervalFrequency

225 Kir21 HEK293 na -40 na -117 na -120 to 0

225 Kv21 CHO -70 na -70 to na

226 Kir21 HEK293 na -80 4 -140 to 0 (10)

245 IK1 GPMyocyte 35 -50 05 -80 to- 50 (ramp)

247 IK1 GPMyocyte na -40 025 -110 to 60 (10)

252 Ik1 GPMyocyte 35 -80 02 -120

256 IK1 RabbitMyocyte 34 -40 03 -100 to 50 01 Hz

262 IK1 GPMyocyte 35 na 01 -100

264 IK1 GPMyocyte -40-80 05 -40 05 -120 to 0

264 IK1 GPMyocyte -80 05 -40

267 IK1 GPMyocyte -40 na -140 to 0

273 IK1 CanineM 37 -80 04 -125 to 65

274 IK1 GPMyocyte 36 -80 -90 -40 05 -40 05 -120 to -10

281 Kir21 XenopusOocyte 22 -40 05 -120

287 IK1 GPMyocyte 36 -80 05 -40 01 Hz

-80 mV

-40 mV

Extracellular buffer solution of IK1ID NaCl KCl CaCl2 MgCl2 HEPES glucose Sepcial

components Blocker (mM) pH pH buffer

47 137 4 18 1 10 10 74 NaOH

184 132 4 2 12 20 11 Ouabain (3uM) nifedipine (5uM) 74 NaOH

206 135 54 18 1 10 10 NaH2PO4 (033) 74 NaOH

207 140 54 18 15 10 55 NaH2PO4 (033) 10 uM Nifedipine BaCl2 (1mM) 74 NaOH

208 136 54 18 1 10 10 Cd2+ (03) 4-AP (2) 74 NaOH

209 1105 25 2 1 10 10 Na-Pyrubate (5)CdCl2 (1) TTA (00006) 4-AP (2) TEA (10) CoCl2 (1) NiCl2 (05) SITS (2)

74 NaOH

210 137 4 18 12 10 dextrose (11) CdCl2 (02) 74 NaOH

213 150 54 18 12 10 10 TTX (05 uM) nifedipine (001) E4031 (0001) 76 NaOH

214-1 125 4 EDTA (4) KH2PO4 (28) K2HPO4 (72) 72 NaOH

214-2 140 5 18 1 6 6

215 144 56 25 12 5 Dextrose (11) Nifedipine ( 0005) E4031 (0001) 74 na

216 140 5 1 1 10 10 CdCl2 (02) 74 na

217 130 4 18 1 10 10 74 NaOH

217 136 54 01 1 10 11 CoCl2 (05) 74 na

Extracellular buffer solution of IK1ID NaCl KCl CaCl2 MgCl2 HEPES glucose Special comp Blocker (mM) pH pH buffer

220 135 54 09 09 10 10 NaH2Po4 (033) CoCl2 (2) 4-AP (3) 74 NaOH

222 135 4 18 1 2 11 TTX (001) Co2+ (2) 74 NaOH

224 140 54 18 1 10 10 NaH2Po4 (033) 73 NaOH

225 140 54 2 08 10 10 74 NaOH

226 130 4 18 1 10 10 74 NaOH

247 140 54 05 5 55 NaH2Po4 (033) Nisoldipine (04) 74 NaOH

252 140 47 13 1 10 10 74 NaOH

262 137 4 18 2 10 115 Pyruvic acid (5) thiamine (01) 74 NaOH

264 140 54 1 5 10 74 NaOH

267 137 54 2 1 10 10 74 NaOH

273 135 47 10 10KH2PO4 (12) MgSO4 (12) NaHCO3 (44)

Nifedipine (5) 4-aminopyridine (3) 74 NaOH

281 96 2 18 1 5 73 NaOH

Pipette solution of IK1ID KCl K-aspartate K-glutamate MgCl2 EGTA ATP GTP HEPES Special component pH pH

buffer

47 130 5 5 4 10 72 KOH

184 20 125 1 10 5 5 71 KOH

206 150 1 5 3 5 74 KOH

207 10 130 1 10 CaCl2 (05) glucose (10) Phosphocreatinine (10) 72 KOH

208 20 100 1 10 5 5 KOH (110) 72 KOH

209 150 5 5 10 Phosphocreatinine (5) 723 KOH

210 130 1 5 5 5 NaCl (7) 72 KOH

213 140 1 5 4 003 10 72 KOH

214 145 5 CaCl2 (1) 72 KOH

215 45 100 1 10 Not indicated 5 72

216 20 130 1 5 10 NaCl (5) 74

217-1 110 1 5 10 K4BAPTA (5) 72

217-2 123 5 K2HPO4 (72) KH2PO4 (8) Na3VO4 (01) KF (5) K4P2O7 (10) 72

217-3 40 80 10 3 02 5 KH2PO4 (10) MgSo4 (1) phosphocreatinine (5) 735

220 25 130 1 1 5 01 5 Phosphocreatinine (3) 725 KOH

222 130 05 3 04 25 72 KOH

224 20 110 1 5 5 01 10 Phosphocreatinine (5) 72 KOH

225 140 2 1 1 10 72 KOH

226 100 1 5 10 K4BAPTA (5) 72 KOH

245 25 500 5 72 KOH

247 30 70 3 5 5 01 5 KH2PO4 (10) 72 KOH

252 140 11 1 10 NaCl (10) 72 KOH

262 150 1 5 4 5 Hydroxybutyric acid (2) Phosphocreatine (5) 73 KOH

264 274 287 40 106 1 5 4 5 72 KOH

267 135 5 5 5-10 NaCl (10) CaCl2 (19) 72 KOH

273 45 100 1 10 3 5 72 KOH

Summary of minor K protocols

summary 28

IKr Ito1 IKur IKs IK1

Sample literatures 18 18 8 36 31

cells cell line gt myocyte cell line gt myocyte cell line gt myocyte cell line myocyte cell line lt myocyte

Temp RT lt Phy RT gt Phy(80 vs 20)

RT(100)

RT Phy( 55 vs 45)

RT Phy (50 vs 50)

Pulses (mode) for IC50

Vh= -80 mV Vd= 0-70 mV Td=1-4s Vr= -80 -+20 Tr= 2-6s

Vh= -90 to -40 (-80)Vd= 0-70 mV (50) Td= 150- 500 ms (300)

Vh= -80 to -50 (-80)Vd=20-60 mV (50) Td= 150- 1000 ms (300)

Vh= -90 to -30 (-80)Vd=20-60 mV (40) Td= 05- 10s (2)Vr= -80 to -10 (-40)Tr= 05 ndash 5 s (2s )

Vh= -120 to -30 (-40)Vd=-140 to 100 mV (hyperpolarizing start from -120)Td=01 ndash 4s (05 s)

measure Tail current Peak current Steady state current at the end of depolarization and tail current

Steady state current at the end of depolarization and tail current

Steady state current at the end of depolarization

pulse trace

Extracellular buffer IKr Ito Ikur Iks Ik1

NaCl

KCl

CaCl2

MgCl2

HEPES

glucose

Na2HPO4 () () () ()

Na-acetate ()

NMDG ()

Na-acetate () () ()

NMDG ()

dextrose () () () ()

K-gluconate () () ()

Na-pyruvate

EDTA ()

KH2PO4K2HPO4 () ()

Blockers () included in less than half of protocols summary 29

Intracellular buffer IKr Ito Ikur Iks Ik1

Current carrier K-aspartateK-gluconateK-glutamate

KCl

MgCl2

EGTA

ATP Na2ATPK2ATPMgATP

HEPES

GTP () () () ()

ATPregeneration system phosphocreatinine () () () ()

CaCl2 () () () ()

NaCl () () () ()

KOH () ()

KH2PO4 () () ()

glucose ()

K4BAPTA ()

summary 30

() included in less than half of protocols

Representative current traces Ito (Kir 43)IKr

IK1 (Kir21)

Ikur (Kv15)

Iks (Kv71minK kvLQT1)

ICa

INa

ID 214 217

ReferencesID Journal Year volume first page47 J Cariovasc Pharmacol 2010 56 42049 J Cardiolvasc Electrophysiol 2010 21 30150 J Pharmacol Exp Ther 2008 324 427100 Am J Physiol Heart Circ Physiol 1998 275 H2016160 Front Pharmacol 2013 4 1173 J Clin Invest 1997 99 1714174 J Cariovasc Pharmacol 1998 31 800177 Mol Pharmacol 2005 57 101178 J Mol Cell Cardiol 2001 33 923179 British Journal of Pharmacology 2000 130 1753180 J Mol Cell Cardiol 2002 34 1029184 Circulation 2004 110 904186 Cardiovascular Research 2006 69 116195 J Pharmacol Exp Ther 2013 344 23199 J Mol Cell Cardiol 2012 53 593205 Heart Rhythm 2009 6 1625206 J Cariovasc Pharmacol 2012 59 37207 Mol Cell Biochem 2011 352 117208 Cell Physiol Biochem 2010 26 967209 J Nat Prod 2006 69 558210 European Journal of Pharmacology 2006 529 47213 Comparative Biochemistry and physiology 2013 158 181214 Cardiovascular Research 2013 99 203215 European Journal of Pharmacology 2013 710 10216 Med Sci Mont 2011 17 BR165217 J Cariovasc Pharmacol 2011 57 407220 Gen Physiol Biophys 2005 24 311221 J Cardiolvasc Electrophysiol 2005 16 1227222 Am J Physiol Heart Circ Physiol 2005 288 H778224 Plos One 2012 7 e51550

ID Journal Year volume first page225 J Biol Chem 2009 284 15432226 PNAS 2008 105 1364229 J Biol Chem 2009 284 9140230 Pflugers Arch 2013 epub232 Circ Arrhythm Electrophysiol 2013 epub233 Hum Exp Toxicol 2013 epub237 Can J Physiol Pharmacol 2010 88 1181239 J cardiolvasc Pharmacol 2010 55 145241 Cardiovascular Research 2009 82 430242 Am J Physiol Heart Circ Physiol 2008 295 H1867244 Circulation Research 2006 98 1055245 J Pharmacol Exp Ther 2006 317 1054247 J Cariovasc Pharmacol 2005 46 167249 J Cariovasc Pharmacol 2004 43 178251 Anesth Analog 2003 96 1308252 Naunyn Schmiedebergs Arch Pharmacol 2002 365 29256 European Journal of Pharmacology 1999 376 189260 British Journal of Pharmacology 2010 159 93262 J Cariovasc Pharmacol 2004 44 244263 British Journal of Pharmacology 2005 146 146264 British Journal of Pharmacology 2000 131 1809267 J Pharmacol Exp Ther 1997 281 233273 British Journal of Pharmacology 2002 136 330274 British Journal of Pharmacology 2003 140 863275 European Journal of Pharmacology 2006 532 44276 J Cardiolvasc Electrophysiol 2005 16 329278 British Journal of Pharmacology 2004 142 1300281 British Journal of Pharmacology 1997 120 968283 British Journal of Pharmacology 2006 147 289286 J Cardiolvasc Electrophysiol 2005 16 314287 European Journal of Pharmacology 2000 401 137

• Protocols of cardiac ion channels INaL Ito1 IKur IKs IK1
• Notes
• Late Sodium Channel Current
• Late sodium channel protocols
• PowerPoint Presentation
• Summary of Ito1 protocols
• Pulse sequences Ito
• Kinetics Pulse Sequences Ito
• Extracellular buffer solution of Ito
• Pipette solution of Ito
• Summary of IKur (Kv1531) protocols
• Pulse sequences IKur
• Extracellular buffer solution of IKur
• Pipette solution of IKur
• Summary of Iks protocols
• Pulse sequences IKs
• Slide 17
• Extracellular buffer solution of IKs
• Slide 19
• Pipette solution of IKs
• Slide 21
• Summary of IK1 protocols
• Pulse sequences IK1
• Slide 24
• Extracellular buffer solution of IK1
• Slide 26
• Pipette solution of IK1
• Summary of minor K protocols
• Extracellular buffer
• Intracellular buffer
• Representative current traces
• References
• Slide 33

Extracellular buffer solution of IK1ID NaCl KCl CaCl2 MgCl2 HEPES glucose Special comp Blocker (mM) pH pH buffer

220 135 54 09 09 10 10 NaH2Po4 (033) CoCl2 (2) 4-AP (3) 74 NaOH

222 135 4 18 1 2 11 TTX (001) Co2+ (2) 74 NaOH

224 140 54 18 1 10 10 NaH2Po4 (033) 73 NaOH

225 140 54 2 08 10 10 74 NaOH

226 130 4 18 1 10 10 74 NaOH

247 140 54 05 5 55 NaH2Po4 (033) Nisoldipine (04) 74 NaOH

252 140 47 13 1 10 10 74 NaOH

262 137 4 18 2 10 115 Pyruvic acid (5) thiamine (01) 74 NaOH

264 140 54 1 5 10 74 NaOH

267 137 54 2 1 10 10 74 NaOH

273 135 47 10 10KH2PO4 (12) MgSO4 (12) NaHCO3 (44)

Nifedipine (5) 4-aminopyridine (3) 74 NaOH

281 96 2 18 1 5 73 NaOH

Pipette solution of IK1ID KCl K-aspartate K-glutamate MgCl2 EGTA ATP GTP HEPES Special component pH pH

buffer

47 130 5 5 4 10 72 KOH

184 20 125 1 10 5 5 71 KOH

206 150 1 5 3 5 74 KOH

207 10 130 1 10 CaCl2 (05) glucose (10) Phosphocreatinine (10) 72 KOH

208 20 100 1 10 5 5 KOH (110) 72 KOH

209 150 5 5 10 Phosphocreatinine (5) 723 KOH

210 130 1 5 5 5 NaCl (7) 72 KOH

213 140 1 5 4 003 10 72 KOH

214 145 5 CaCl2 (1) 72 KOH

215 45 100 1 10 Not indicated 5 72

216 20 130 1 5 10 NaCl (5) 74

217-1 110 1 5 10 K4BAPTA (5) 72

217-2 123 5 K2HPO4 (72) KH2PO4 (8) Na3VO4 (01) KF (5) K4P2O7 (10) 72

217-3 40 80 10 3 02 5 KH2PO4 (10) MgSo4 (1) phosphocreatinine (5) 735

220 25 130 1 1 5 01 5 Phosphocreatinine (3) 725 KOH

222 130 05 3 04 25 72 KOH

224 20 110 1 5 5 01 10 Phosphocreatinine (5) 72 KOH

225 140 2 1 1 10 72 KOH

226 100 1 5 10 K4BAPTA (5) 72 KOH

245 25 500 5 72 KOH

247 30 70 3 5 5 01 5 KH2PO4 (10) 72 KOH

252 140 11 1 10 NaCl (10) 72 KOH

262 150 1 5 4 5 Hydroxybutyric acid (2) Phosphocreatine (5) 73 KOH

264 274 287 40 106 1 5 4 5 72 KOH

267 135 5 5 5-10 NaCl (10) CaCl2 (19) 72 KOH

273 45 100 1 10 3 5 72 KOH

Summary of minor K protocols

summary 28

IKr Ito1 IKur IKs IK1

Sample literatures 18 18 8 36 31

cells cell line gt myocyte cell line gt myocyte cell line gt myocyte cell line myocyte cell line lt myocyte

Temp RT lt Phy RT gt Phy(80 vs 20)

RT(100)

RT Phy( 55 vs 45)

RT Phy (50 vs 50)

Pulses (mode) for IC50

Vh= -80 mV Vd= 0-70 mV Td=1-4s Vr= -80 -+20 Tr= 2-6s

Vh= -90 to -40 (-80)Vd= 0-70 mV (50) Td= 150- 500 ms (300)

Vh= -80 to -50 (-80)Vd=20-60 mV (50) Td= 150- 1000 ms (300)

Vh= -90 to -30 (-80)Vd=20-60 mV (40) Td= 05- 10s (2)Vr= -80 to -10 (-40)Tr= 05 ndash 5 s (2s )

Vh= -120 to -30 (-40)Vd=-140 to 100 mV (hyperpolarizing start from -120)Td=01 ndash 4s (05 s)

measure Tail current Peak current Steady state current at the end of depolarization and tail current

Steady state current at the end of depolarization and tail current

Steady state current at the end of depolarization

pulse trace

Extracellular buffer IKr Ito Ikur Iks Ik1

NaCl

KCl

CaCl2

MgCl2

HEPES

glucose

Na2HPO4 () () () ()

Na-acetate ()

NMDG ()

Na-acetate () () ()

NMDG ()

dextrose () () () ()

K-gluconate () () ()

Na-pyruvate

EDTA ()

KH2PO4K2HPO4 () ()

Blockers () included in less than half of protocols summary 29

Intracellular buffer IKr Ito Ikur Iks Ik1

Current carrier K-aspartateK-gluconateK-glutamate

KCl

MgCl2

EGTA

ATP Na2ATPK2ATPMgATP

HEPES

GTP () () () ()

ATPregeneration system phosphocreatinine () () () ()

CaCl2 () () () ()

NaCl () () () ()

KOH () ()

KH2PO4 () () ()

glucose ()

K4BAPTA ()

summary 30

() included in less than half of protocols

Representative current traces Ito (Kir 43)IKr

IK1 (Kir21)

Ikur (Kv15)

Iks (Kv71minK kvLQT1)

ICa

INa

ID 214 217

ReferencesID Journal Year volume first page47 J Cariovasc Pharmacol 2010 56 42049 J Cardiolvasc Electrophysiol 2010 21 30150 J Pharmacol Exp Ther 2008 324 427100 Am J Physiol Heart Circ Physiol 1998 275 H2016160 Front Pharmacol 2013 4 1173 J Clin Invest 1997 99 1714174 J Cariovasc Pharmacol 1998 31 800177 Mol Pharmacol 2005 57 101178 J Mol Cell Cardiol 2001 33 923179 British Journal of Pharmacology 2000 130 1753180 J Mol Cell Cardiol 2002 34 1029184 Circulation 2004 110 904186 Cardiovascular Research 2006 69 116195 J Pharmacol Exp Ther 2013 344 23199 J Mol Cell Cardiol 2012 53 593205 Heart Rhythm 2009 6 1625206 J Cariovasc Pharmacol 2012 59 37207 Mol Cell Biochem 2011 352 117208 Cell Physiol Biochem 2010 26 967209 J Nat Prod 2006 69 558210 European Journal of Pharmacology 2006 529 47213 Comparative Biochemistry and physiology 2013 158 181214 Cardiovascular Research 2013 99 203215 European Journal of Pharmacology 2013 710 10216 Med Sci Mont 2011 17 BR165217 J Cariovasc Pharmacol 2011 57 407220 Gen Physiol Biophys 2005 24 311221 J Cardiolvasc Electrophysiol 2005 16 1227222 Am J Physiol Heart Circ Physiol 2005 288 H778224 Plos One 2012 7 e51550

ID Journal Year volume first page225 J Biol Chem 2009 284 15432226 PNAS 2008 105 1364229 J Biol Chem 2009 284 9140230 Pflugers Arch 2013 epub232 Circ Arrhythm Electrophysiol 2013 epub233 Hum Exp Toxicol 2013 epub237 Can J Physiol Pharmacol 2010 88 1181239 J cardiolvasc Pharmacol 2010 55 145241 Cardiovascular Research 2009 82 430242 Am J Physiol Heart Circ Physiol 2008 295 H1867244 Circulation Research 2006 98 1055245 J Pharmacol Exp Ther 2006 317 1054247 J Cariovasc Pharmacol 2005 46 167249 J Cariovasc Pharmacol 2004 43 178251 Anesth Analog 2003 96 1308252 Naunyn Schmiedebergs Arch Pharmacol 2002 365 29256 European Journal of Pharmacology 1999 376 189260 British Journal of Pharmacology 2010 159 93262 J Cariovasc Pharmacol 2004 44 244263 British Journal of Pharmacology 2005 146 146264 British Journal of Pharmacology 2000 131 1809267 J Pharmacol Exp Ther 1997 281 233273 British Journal of Pharmacology 2002 136 330274 British Journal of Pharmacology 2003 140 863275 European Journal of Pharmacology 2006 532 44276 J Cardiolvasc Electrophysiol 2005 16 329278 British Journal of Pharmacology 2004 142 1300281 British Journal of Pharmacology 1997 120 968283 British Journal of Pharmacology 2006 147 289286 J Cardiolvasc Electrophysiol 2005 16 314287 European Journal of Pharmacology 2000 401 137

• Protocols of cardiac ion channels INaL Ito1 IKur IKs IK1
• Notes
• Late Sodium Channel Current
• Late sodium channel protocols
• PowerPoint Presentation
• Summary of Ito1 protocols
• Pulse sequences Ito
• Kinetics Pulse Sequences Ito
• Extracellular buffer solution of Ito
• Pipette solution of Ito
• Summary of IKur (Kv1531) protocols
• Pulse sequences IKur
• Extracellular buffer solution of IKur
• Pipette solution of IKur
• Summary of Iks protocols
• Pulse sequences IKs
• Slide 17
• Extracellular buffer solution of IKs
• Slide 19
• Pipette solution of IKs
• Slide 21
• Summary of IK1 protocols
• Pulse sequences IK1
• Slide 24
• Extracellular buffer solution of IK1
• Slide 26
• Pipette solution of IK1
• Summary of minor K protocols
• Extracellular buffer
• Intracellular buffer
• Representative current traces
• References
• Slide 33

Pipette solution of IK1ID KCl K-aspartate K-glutamate MgCl2 EGTA ATP GTP HEPES Special component pH pH

buffer

47 130 5 5 4 10 72 KOH

184 20 125 1 10 5 5 71 KOH

206 150 1 5 3 5 74 KOH

207 10 130 1 10 CaCl2 (05) glucose (10) Phosphocreatinine (10) 72 KOH

208 20 100 1 10 5 5 KOH (110) 72 KOH

209 150 5 5 10 Phosphocreatinine (5) 723 KOH

210 130 1 5 5 5 NaCl (7) 72 KOH

213 140 1 5 4 003 10 72 KOH

214 145 5 CaCl2 (1) 72 KOH

215 45 100 1 10 Not indicated 5 72

216 20 130 1 5 10 NaCl (5) 74

217-1 110 1 5 10 K4BAPTA (5) 72

217-2 123 5 K2HPO4 (72) KH2PO4 (8) Na3VO4 (01) KF (5) K4P2O7 (10) 72

217-3 40 80 10 3 02 5 KH2PO4 (10) MgSo4 (1) phosphocreatinine (5) 735

220 25 130 1 1 5 01 5 Phosphocreatinine (3) 725 KOH

222 130 05 3 04 25 72 KOH

224 20 110 1 5 5 01 10 Phosphocreatinine (5) 72 KOH

225 140 2 1 1 10 72 KOH

226 100 1 5 10 K4BAPTA (5) 72 KOH

245 25 500 5 72 KOH

247 30 70 3 5 5 01 5 KH2PO4 (10) 72 KOH

252 140 11 1 10 NaCl (10) 72 KOH

262 150 1 5 4 5 Hydroxybutyric acid (2) Phosphocreatine (5) 73 KOH

264 274 287 40 106 1 5 4 5 72 KOH

267 135 5 5 5-10 NaCl (10) CaCl2 (19) 72 KOH

273 45 100 1 10 3 5 72 KOH

Summary of minor K protocols

summary 28

IKr Ito1 IKur IKs IK1

Sample literatures 18 18 8 36 31

cells cell line gt myocyte cell line gt myocyte cell line gt myocyte cell line myocyte cell line lt myocyte

Temp RT lt Phy RT gt Phy(80 vs 20)

RT(100)

RT Phy( 55 vs 45)

RT Phy (50 vs 50)

Pulses (mode) for IC50

Vh= -80 mV Vd= 0-70 mV Td=1-4s Vr= -80 -+20 Tr= 2-6s

Vh= -90 to -40 (-80)Vd= 0-70 mV (50) Td= 150- 500 ms (300)

Vh= -80 to -50 (-80)Vd=20-60 mV (50) Td= 150- 1000 ms (300)

Vh= -90 to -30 (-80)Vd=20-60 mV (40) Td= 05- 10s (2)Vr= -80 to -10 (-40)Tr= 05 ndash 5 s (2s )

Vh= -120 to -30 (-40)Vd=-140 to 100 mV (hyperpolarizing start from -120)Td=01 ndash 4s (05 s)

measure Tail current Peak current Steady state current at the end of depolarization and tail current

Steady state current at the end of depolarization and tail current

Steady state current at the end of depolarization

pulse trace

Extracellular buffer IKr Ito Ikur Iks Ik1

NaCl

KCl

CaCl2

MgCl2

HEPES

glucose

Na2HPO4 () () () ()

Na-acetate ()

NMDG ()

Na-acetate () () ()

NMDG ()

dextrose () () () ()

K-gluconate () () ()

Na-pyruvate

EDTA ()

KH2PO4K2HPO4 () ()

Blockers () included in less than half of protocols summary 29

Intracellular buffer IKr Ito Ikur Iks Ik1

Current carrier K-aspartateK-gluconateK-glutamate

KCl

MgCl2

EGTA

ATP Na2ATPK2ATPMgATP

HEPES

GTP () () () ()

ATPregeneration system phosphocreatinine () () () ()

CaCl2 () () () ()

NaCl () () () ()

KOH () ()

KH2PO4 () () ()

glucose ()

K4BAPTA ()

summary 30

() included in less than half of protocols

Representative current traces Ito (Kir 43)IKr

IK1 (Kir21)

Ikur (Kv15)

Iks (Kv71minK kvLQT1)

ICa

INa

ID 214 217

ReferencesID Journal Year volume first page47 J Cariovasc Pharmacol 2010 56 42049 J Cardiolvasc Electrophysiol 2010 21 30150 J Pharmacol Exp Ther 2008 324 427100 Am J Physiol Heart Circ Physiol 1998 275 H2016160 Front Pharmacol 2013 4 1173 J Clin Invest 1997 99 1714174 J Cariovasc Pharmacol 1998 31 800177 Mol Pharmacol 2005 57 101178 J Mol Cell Cardiol 2001 33 923179 British Journal of Pharmacology 2000 130 1753180 J Mol Cell Cardiol 2002 34 1029184 Circulation 2004 110 904186 Cardiovascular Research 2006 69 116195 J Pharmacol Exp Ther 2013 344 23199 J Mol Cell Cardiol 2012 53 593205 Heart Rhythm 2009 6 1625206 J Cariovasc Pharmacol 2012 59 37207 Mol Cell Biochem 2011 352 117208 Cell Physiol Biochem 2010 26 967209 J Nat Prod 2006 69 558210 European Journal of Pharmacology 2006 529 47213 Comparative Biochemistry and physiology 2013 158 181214 Cardiovascular Research 2013 99 203215 European Journal of Pharmacology 2013 710 10216 Med Sci Mont 2011 17 BR165217 J Cariovasc Pharmacol 2011 57 407220 Gen Physiol Biophys 2005 24 311221 J Cardiolvasc Electrophysiol 2005 16 1227222 Am J Physiol Heart Circ Physiol 2005 288 H778224 Plos One 2012 7 e51550

ID Journal Year volume first page225 J Biol Chem 2009 284 15432226 PNAS 2008 105 1364229 J Biol Chem 2009 284 9140230 Pflugers Arch 2013 epub232 Circ Arrhythm Electrophysiol 2013 epub233 Hum Exp Toxicol 2013 epub237 Can J Physiol Pharmacol 2010 88 1181239 J cardiolvasc Pharmacol 2010 55 145241 Cardiovascular Research 2009 82 430242 Am J Physiol Heart Circ Physiol 2008 295 H1867244 Circulation Research 2006 98 1055245 J Pharmacol Exp Ther 2006 317 1054247 J Cariovasc Pharmacol 2005 46 167249 J Cariovasc Pharmacol 2004 43 178251 Anesth Analog 2003 96 1308252 Naunyn Schmiedebergs Arch Pharmacol 2002 365 29256 European Journal of Pharmacology 1999 376 189260 British Journal of Pharmacology 2010 159 93262 J Cariovasc Pharmacol 2004 44 244263 British Journal of Pharmacology 2005 146 146264 British Journal of Pharmacology 2000 131 1809267 J Pharmacol Exp Ther 1997 281 233273 British Journal of Pharmacology 2002 136 330274 British Journal of Pharmacology 2003 140 863275 European Journal of Pharmacology 2006 532 44276 J Cardiolvasc Electrophysiol 2005 16 329278 British Journal of Pharmacology 2004 142 1300281 British Journal of Pharmacology 1997 120 968283 British Journal of Pharmacology 2006 147 289286 J Cardiolvasc Electrophysiol 2005 16 314287 European Journal of Pharmacology 2000 401 137

• Protocols of cardiac ion channels INaL Ito1 IKur IKs IK1
• Notes
• Late Sodium Channel Current
• Late sodium channel protocols
• PowerPoint Presentation
• Summary of Ito1 protocols
• Pulse sequences Ito
• Kinetics Pulse Sequences Ito
• Extracellular buffer solution of Ito
• Pipette solution of Ito
• Summary of IKur (Kv1531) protocols
• Pulse sequences IKur
• Extracellular buffer solution of IKur
• Pipette solution of IKur
• Summary of Iks protocols
• Pulse sequences IKs
• Slide 17
• Extracellular buffer solution of IKs
• Slide 19
• Pipette solution of IKs
• Slide 21
• Summary of IK1 protocols
• Pulse sequences IK1
• Slide 24
• Extracellular buffer solution of IK1
• Slide 26
• Pipette solution of IK1
• Summary of minor K protocols
• Extracellular buffer
• Intracellular buffer
• Representative current traces
• References
• Slide 33

Summary of minor K protocols

summary 28

IKr Ito1 IKur IKs IK1

Sample literatures 18 18 8 36 31

cells cell line gt myocyte cell line gt myocyte cell line gt myocyte cell line myocyte cell line lt myocyte

Temp RT lt Phy RT gt Phy(80 vs 20)

RT(100)

RT Phy( 55 vs 45)

RT Phy (50 vs 50)

Pulses (mode) for IC50

Vh= -80 mV Vd= 0-70 mV Td=1-4s Vr= -80 -+20 Tr= 2-6s

Vh= -90 to -40 (-80)Vd= 0-70 mV (50) Td= 150- 500 ms (300)

Vh= -80 to -50 (-80)Vd=20-60 mV (50) Td= 150- 1000 ms (300)

Vh= -90 to -30 (-80)Vd=20-60 mV (40) Td= 05- 10s (2)Vr= -80 to -10 (-40)Tr= 05 ndash 5 s (2s )

Vh= -120 to -30 (-40)Vd=-140 to 100 mV (hyperpolarizing start from -120)Td=01 ndash 4s (05 s)

measure Tail current Peak current Steady state current at the end of depolarization and tail current

Steady state current at the end of depolarization and tail current

Steady state current at the end of depolarization

pulse trace

Extracellular buffer IKr Ito Ikur Iks Ik1

NaCl

KCl

CaCl2

MgCl2

HEPES

glucose

Na2HPO4 () () () ()

Na-acetate ()

NMDG ()

Na-acetate () () ()

NMDG ()

dextrose () () () ()

K-gluconate () () ()

Na-pyruvate

EDTA ()

KH2PO4K2HPO4 () ()

Blockers () included in less than half of protocols summary 29

Intracellular buffer IKr Ito Ikur Iks Ik1

Current carrier K-aspartateK-gluconateK-glutamate

KCl

MgCl2

EGTA

ATP Na2ATPK2ATPMgATP

HEPES

GTP () () () ()

ATPregeneration system phosphocreatinine () () () ()

CaCl2 () () () ()

NaCl () () () ()

KOH () ()

KH2PO4 () () ()

glucose ()

K4BAPTA ()

summary 30

() included in less than half of protocols

Representative current traces Ito (Kir 43)IKr

IK1 (Kir21)

Ikur (Kv15)

Iks (Kv71minK kvLQT1)

ICa

INa

ID 214 217

ReferencesID Journal Year volume first page47 J Cariovasc Pharmacol 2010 56 42049 J Cardiolvasc Electrophysiol 2010 21 30150 J Pharmacol Exp Ther 2008 324 427100 Am J Physiol Heart Circ Physiol 1998 275 H2016160 Front Pharmacol 2013 4 1173 J Clin Invest 1997 99 1714174 J Cariovasc Pharmacol 1998 31 800177 Mol Pharmacol 2005 57 101178 J Mol Cell Cardiol 2001 33 923179 British Journal of Pharmacology 2000 130 1753180 J Mol Cell Cardiol 2002 34 1029184 Circulation 2004 110 904186 Cardiovascular Research 2006 69 116195 J Pharmacol Exp Ther 2013 344 23199 J Mol Cell Cardiol 2012 53 593205 Heart Rhythm 2009 6 1625206 J Cariovasc Pharmacol 2012 59 37207 Mol Cell Biochem 2011 352 117208 Cell Physiol Biochem 2010 26 967209 J Nat Prod 2006 69 558210 European Journal of Pharmacology 2006 529 47213 Comparative Biochemistry and physiology 2013 158 181214 Cardiovascular Research 2013 99 203215 European Journal of Pharmacology 2013 710 10216 Med Sci Mont 2011 17 BR165217 J Cariovasc Pharmacol 2011 57 407220 Gen Physiol Biophys 2005 24 311221 J Cardiolvasc Electrophysiol 2005 16 1227222 Am J Physiol Heart Circ Physiol 2005 288 H778224 Plos One 2012 7 e51550

ID Journal Year volume first page225 J Biol Chem 2009 284 15432226 PNAS 2008 105 1364229 J Biol Chem 2009 284 9140230 Pflugers Arch 2013 epub232 Circ Arrhythm Electrophysiol 2013 epub233 Hum Exp Toxicol 2013 epub237 Can J Physiol Pharmacol 2010 88 1181239 J cardiolvasc Pharmacol 2010 55 145241 Cardiovascular Research 2009 82 430242 Am J Physiol Heart Circ Physiol 2008 295 H1867244 Circulation Research 2006 98 1055245 J Pharmacol Exp Ther 2006 317 1054247 J Cariovasc Pharmacol 2005 46 167249 J Cariovasc Pharmacol 2004 43 178251 Anesth Analog 2003 96 1308252 Naunyn Schmiedebergs Arch Pharmacol 2002 365 29256 European Journal of Pharmacology 1999 376 189260 British Journal of Pharmacology 2010 159 93262 J Cariovasc Pharmacol 2004 44 244263 British Journal of Pharmacology 2005 146 146264 British Journal of Pharmacology 2000 131 1809267 J Pharmacol Exp Ther 1997 281 233273 British Journal of Pharmacology 2002 136 330274 British Journal of Pharmacology 2003 140 863275 European Journal of Pharmacology 2006 532 44276 J Cardiolvasc Electrophysiol 2005 16 329278 British Journal of Pharmacology 2004 142 1300281 British Journal of Pharmacology 1997 120 968283 British Journal of Pharmacology 2006 147 289286 J Cardiolvasc Electrophysiol 2005 16 314287 European Journal of Pharmacology 2000 401 137

• Protocols of cardiac ion channels INaL Ito1 IKur IKs IK1
• Notes
• Late Sodium Channel Current
• Late sodium channel protocols
• PowerPoint Presentation
• Summary of Ito1 protocols
• Pulse sequences Ito
• Kinetics Pulse Sequences Ito
• Extracellular buffer solution of Ito
• Pipette solution of Ito
• Summary of IKur (Kv1531) protocols
• Pulse sequences IKur
• Extracellular buffer solution of IKur
• Pipette solution of IKur
• Summary of Iks protocols
• Pulse sequences IKs
• Slide 17
• Extracellular buffer solution of IKs
• Slide 19
• Pipette solution of IKs
• Slide 21
• Summary of IK1 protocols
• Pulse sequences IK1
• Slide 24
• Extracellular buffer solution of IK1
• Slide 26
• Pipette solution of IK1
• Summary of minor K protocols
• Extracellular buffer
• Intracellular buffer
• Representative current traces
• References
• Slide 33

Extracellular buffer IKr Ito Ikur Iks Ik1

NaCl

KCl

CaCl2

MgCl2

HEPES

glucose

Na2HPO4 () () () ()

Na-acetate ()

NMDG ()

Na-acetate () () ()

NMDG ()

dextrose () () () ()

K-gluconate () () ()

Na-pyruvate

EDTA ()

KH2PO4K2HPO4 () ()

Blockers () included in less than half of protocols summary 29

Intracellular buffer IKr Ito Ikur Iks Ik1

Current carrier K-aspartateK-gluconateK-glutamate

KCl

MgCl2

EGTA

ATP Na2ATPK2ATPMgATP

HEPES

GTP () () () ()

ATPregeneration system phosphocreatinine () () () ()

CaCl2 () () () ()

NaCl () () () ()

KOH () ()

KH2PO4 () () ()

glucose ()

K4BAPTA ()

summary 30

() included in less than half of protocols

Representative current traces Ito (Kir 43)IKr

IK1 (Kir21)

Ikur (Kv15)

Iks (Kv71minK kvLQT1)

ICa

INa

ID 214 217

ReferencesID Journal Year volume first page47 J Cariovasc Pharmacol 2010 56 42049 J Cardiolvasc Electrophysiol 2010 21 30150 J Pharmacol Exp Ther 2008 324 427100 Am J Physiol Heart Circ Physiol 1998 275 H2016160 Front Pharmacol 2013 4 1173 J Clin Invest 1997 99 1714174 J Cariovasc Pharmacol 1998 31 800177 Mol Pharmacol 2005 57 101178 J Mol Cell Cardiol 2001 33 923179 British Journal of Pharmacology 2000 130 1753180 J Mol Cell Cardiol 2002 34 1029184 Circulation 2004 110 904186 Cardiovascular Research 2006 69 116195 J Pharmacol Exp Ther 2013 344 23199 J Mol Cell Cardiol 2012 53 593205 Heart Rhythm 2009 6 1625206 J Cariovasc Pharmacol 2012 59 37207 Mol Cell Biochem 2011 352 117208 Cell Physiol Biochem 2010 26 967209 J Nat Prod 2006 69 558210 European Journal of Pharmacology 2006 529 47213 Comparative Biochemistry and physiology 2013 158 181214 Cardiovascular Research 2013 99 203215 European Journal of Pharmacology 2013 710 10216 Med Sci Mont 2011 17 BR165217 J Cariovasc Pharmacol 2011 57 407220 Gen Physiol Biophys 2005 24 311221 J Cardiolvasc Electrophysiol 2005 16 1227222 Am J Physiol Heart Circ Physiol 2005 288 H778224 Plos One 2012 7 e51550

ID Journal Year volume first page225 J Biol Chem 2009 284 15432226 PNAS 2008 105 1364229 J Biol Chem 2009 284 9140230 Pflugers Arch 2013 epub232 Circ Arrhythm Electrophysiol 2013 epub233 Hum Exp Toxicol 2013 epub237 Can J Physiol Pharmacol 2010 88 1181239 J cardiolvasc Pharmacol 2010 55 145241 Cardiovascular Research 2009 82 430242 Am J Physiol Heart Circ Physiol 2008 295 H1867244 Circulation Research 2006 98 1055245 J Pharmacol Exp Ther 2006 317 1054247 J Cariovasc Pharmacol 2005 46 167249 J Cariovasc Pharmacol 2004 43 178251 Anesth Analog 2003 96 1308252 Naunyn Schmiedebergs Arch Pharmacol 2002 365 29256 European Journal of Pharmacology 1999 376 189260 British Journal of Pharmacology 2010 159 93262 J Cariovasc Pharmacol 2004 44 244263 British Journal of Pharmacology 2005 146 146264 British Journal of Pharmacology 2000 131 1809267 J Pharmacol Exp Ther 1997 281 233273 British Journal of Pharmacology 2002 136 330274 British Journal of Pharmacology 2003 140 863275 European Journal of Pharmacology 2006 532 44276 J Cardiolvasc Electrophysiol 2005 16 329278 British Journal of Pharmacology 2004 142 1300281 British Journal of Pharmacology 1997 120 968283 British Journal of Pharmacology 2006 147 289286 J Cardiolvasc Electrophysiol 2005 16 314287 European Journal of Pharmacology 2000 401 137

• Protocols of cardiac ion channels INaL Ito1 IKur IKs IK1
• Notes
• Late Sodium Channel Current
• Late sodium channel protocols
• PowerPoint Presentation
• Summary of Ito1 protocols
• Pulse sequences Ito
• Kinetics Pulse Sequences Ito
• Extracellular buffer solution of Ito
• Pipette solution of Ito
• Summary of IKur (Kv1531) protocols
• Pulse sequences IKur
• Extracellular buffer solution of IKur
• Pipette solution of IKur
• Summary of Iks protocols
• Pulse sequences IKs
• Slide 17
• Extracellular buffer solution of IKs
• Slide 19
• Pipette solution of IKs
• Slide 21
• Summary of IK1 protocols
• Pulse sequences IK1
• Slide 24
• Extracellular buffer solution of IK1
• Slide 26
• Pipette solution of IK1
• Summary of minor K protocols
• Extracellular buffer
• Intracellular buffer
• Representative current traces
• References
• Slide 33

Intracellular buffer IKr Ito Ikur Iks Ik1

Current carrier K-aspartateK-gluconateK-glutamate

KCl

MgCl2

EGTA

ATP Na2ATPK2ATPMgATP

HEPES

GTP () () () ()

ATPregeneration system phosphocreatinine () () () ()

CaCl2 () () () ()

NaCl () () () ()

KOH () ()

KH2PO4 () () ()

glucose ()

K4BAPTA ()

summary 30

() included in less than half of protocols

Representative current traces Ito (Kir 43)IKr

IK1 (Kir21)

Ikur (Kv15)

Iks (Kv71minK kvLQT1)

ICa

INa

ID 214 217

ReferencesID Journal Year volume first page47 J Cariovasc Pharmacol 2010 56 42049 J Cardiolvasc Electrophysiol 2010 21 30150 J Pharmacol Exp Ther 2008 324 427100 Am J Physiol Heart Circ Physiol 1998 275 H2016160 Front Pharmacol 2013 4 1173 J Clin Invest 1997 99 1714174 J Cariovasc Pharmacol 1998 31 800177 Mol Pharmacol 2005 57 101178 J Mol Cell Cardiol 2001 33 923179 British Journal of Pharmacology 2000 130 1753180 J Mol Cell Cardiol 2002 34 1029184 Circulation 2004 110 904186 Cardiovascular Research 2006 69 116195 J Pharmacol Exp Ther 2013 344 23199 J Mol Cell Cardiol 2012 53 593205 Heart Rhythm 2009 6 1625206 J Cariovasc Pharmacol 2012 59 37207 Mol Cell Biochem 2011 352 117208 Cell Physiol Biochem 2010 26 967209 J Nat Prod 2006 69 558210 European Journal of Pharmacology 2006 529 47213 Comparative Biochemistry and physiology 2013 158 181214 Cardiovascular Research 2013 99 203215 European Journal of Pharmacology 2013 710 10216 Med Sci Mont 2011 17 BR165217 J Cariovasc Pharmacol 2011 57 407220 Gen Physiol Biophys 2005 24 311221 J Cardiolvasc Electrophysiol 2005 16 1227222 Am J Physiol Heart Circ Physiol 2005 288 H778224 Plos One 2012 7 e51550

ID Journal Year volume first page225 J Biol Chem 2009 284 15432226 PNAS 2008 105 1364229 J Biol Chem 2009 284 9140230 Pflugers Arch 2013 epub232 Circ Arrhythm Electrophysiol 2013 epub233 Hum Exp Toxicol 2013 epub237 Can J Physiol Pharmacol 2010 88 1181239 J cardiolvasc Pharmacol 2010 55 145241 Cardiovascular Research 2009 82 430242 Am J Physiol Heart Circ Physiol 2008 295 H1867244 Circulation Research 2006 98 1055245 J Pharmacol Exp Ther 2006 317 1054247 J Cariovasc Pharmacol 2005 46 167249 J Cariovasc Pharmacol 2004 43 178251 Anesth Analog 2003 96 1308252 Naunyn Schmiedebergs Arch Pharmacol 2002 365 29256 European Journal of Pharmacology 1999 376 189260 British Journal of Pharmacology 2010 159 93262 J Cariovasc Pharmacol 2004 44 244263 British Journal of Pharmacology 2005 146 146264 British Journal of Pharmacology 2000 131 1809267 J Pharmacol Exp Ther 1997 281 233273 British Journal of Pharmacology 2002 136 330274 British Journal of Pharmacology 2003 140 863275 European Journal of Pharmacology 2006 532 44276 J Cardiolvasc Electrophysiol 2005 16 329278 British Journal of Pharmacology 2004 142 1300281 British Journal of Pharmacology 1997 120 968283 British Journal of Pharmacology 2006 147 289286 J Cardiolvasc Electrophysiol 2005 16 314287 European Journal of Pharmacology 2000 401 137

• Protocols of cardiac ion channels INaL Ito1 IKur IKs IK1
• Notes
• Late Sodium Channel Current
• Late sodium channel protocols
• PowerPoint Presentation
• Summary of Ito1 protocols
• Pulse sequences Ito
• Kinetics Pulse Sequences Ito
• Extracellular buffer solution of Ito
• Pipette solution of Ito
• Summary of IKur (Kv1531) protocols
• Pulse sequences IKur
• Extracellular buffer solution of IKur
• Pipette solution of IKur
• Summary of Iks protocols
• Pulse sequences IKs
• Slide 17
• Extracellular buffer solution of IKs
• Slide 19
• Pipette solution of IKs
• Slide 21
• Summary of IK1 protocols
• Pulse sequences IK1
• Slide 24
• Extracellular buffer solution of IK1
• Slide 26
• Pipette solution of IK1
• Summary of minor K protocols
• Extracellular buffer
• Intracellular buffer
• Representative current traces
• References
• Slide 33

Representative current traces Ito (Kir 43)IKr

IK1 (Kir21)

Ikur (Kv15)

Iks (Kv71minK kvLQT1)

ICa

INa

ID 214 217

ReferencesID Journal Year volume first page47 J Cariovasc Pharmacol 2010 56 42049 J Cardiolvasc Electrophysiol 2010 21 30150 J Pharmacol Exp Ther 2008 324 427100 Am J Physiol Heart Circ Physiol 1998 275 H2016160 Front Pharmacol 2013 4 1173 J Clin Invest 1997 99 1714174 J Cariovasc Pharmacol 1998 31 800177 Mol Pharmacol 2005 57 101178 J Mol Cell Cardiol 2001 33 923179 British Journal of Pharmacology 2000 130 1753180 J Mol Cell Cardiol 2002 34 1029184 Circulation 2004 110 904186 Cardiovascular Research 2006 69 116195 J Pharmacol Exp Ther 2013 344 23199 J Mol Cell Cardiol 2012 53 593205 Heart Rhythm 2009 6 1625206 J Cariovasc Pharmacol 2012 59 37207 Mol Cell Biochem 2011 352 117208 Cell Physiol Biochem 2010 26 967209 J Nat Prod 2006 69 558210 European Journal of Pharmacology 2006 529 47213 Comparative Biochemistry and physiology 2013 158 181214 Cardiovascular Research 2013 99 203215 European Journal of Pharmacology 2013 710 10216 Med Sci Mont 2011 17 BR165217 J Cariovasc Pharmacol 2011 57 407220 Gen Physiol Biophys 2005 24 311221 J Cardiolvasc Electrophysiol 2005 16 1227222 Am J Physiol Heart Circ Physiol 2005 288 H778224 Plos One 2012 7 e51550

ID Journal Year volume first page225 J Biol Chem 2009 284 15432226 PNAS 2008 105 1364229 J Biol Chem 2009 284 9140230 Pflugers Arch 2013 epub232 Circ Arrhythm Electrophysiol 2013 epub233 Hum Exp Toxicol 2013 epub237 Can J Physiol Pharmacol 2010 88 1181239 J cardiolvasc Pharmacol 2010 55 145241 Cardiovascular Research 2009 82 430242 Am J Physiol Heart Circ Physiol 2008 295 H1867244 Circulation Research 2006 98 1055245 J Pharmacol Exp Ther 2006 317 1054247 J Cariovasc Pharmacol 2005 46 167249 J Cariovasc Pharmacol 2004 43 178251 Anesth Analog 2003 96 1308252 Naunyn Schmiedebergs Arch Pharmacol 2002 365 29256 European Journal of Pharmacology 1999 376 189260 British Journal of Pharmacology 2010 159 93262 J Cariovasc Pharmacol 2004 44 244263 British Journal of Pharmacology 2005 146 146264 British Journal of Pharmacology 2000 131 1809267 J Pharmacol Exp Ther 1997 281 233273 British Journal of Pharmacology 2002 136 330274 British Journal of Pharmacology 2003 140 863275 European Journal of Pharmacology 2006 532 44276 J Cardiolvasc Electrophysiol 2005 16 329278 British Journal of Pharmacology 2004 142 1300281 British Journal of Pharmacology 1997 120 968283 British Journal of Pharmacology 2006 147 289286 J Cardiolvasc Electrophysiol 2005 16 314287 European Journal of Pharmacology 2000 401 137

• Protocols of cardiac ion channels INaL Ito1 IKur IKs IK1
• Notes
• Late Sodium Channel Current
• Late sodium channel protocols
• PowerPoint Presentation
• Summary of Ito1 protocols
• Pulse sequences Ito
• Kinetics Pulse Sequences Ito
• Extracellular buffer solution of Ito
• Pipette solution of Ito
• Summary of IKur (Kv1531) protocols
• Pulse sequences IKur
• Extracellular buffer solution of IKur
• Pipette solution of IKur
• Summary of Iks protocols
• Pulse sequences IKs
• Slide 17
• Extracellular buffer solution of IKs
• Slide 19
• Pipette solution of IKs
• Slide 21
• Summary of IK1 protocols
• Pulse sequences IK1
• Slide 24
• Extracellular buffer solution of IK1
• Slide 26
• Pipette solution of IK1
• Summary of minor K protocols
• Extracellular buffer
• Intracellular buffer
• Representative current traces
• References
• Slide 33

ReferencesID Journal Year volume first page47 J Cariovasc Pharmacol 2010 56 42049 J Cardiolvasc Electrophysiol 2010 21 30150 J Pharmacol Exp Ther 2008 324 427100 Am J Physiol Heart Circ Physiol 1998 275 H2016160 Front Pharmacol 2013 4 1173 J Clin Invest 1997 99 1714174 J Cariovasc Pharmacol 1998 31 800177 Mol Pharmacol 2005 57 101178 J Mol Cell Cardiol 2001 33 923179 British Journal of Pharmacology 2000 130 1753180 J Mol Cell Cardiol 2002 34 1029184 Circulation 2004 110 904186 Cardiovascular Research 2006 69 116195 J Pharmacol Exp Ther 2013 344 23199 J Mol Cell Cardiol 2012 53 593205 Heart Rhythm 2009 6 1625206 J Cariovasc Pharmacol 2012 59 37207 Mol Cell Biochem 2011 352 117208 Cell Physiol Biochem 2010 26 967209 J Nat Prod 2006 69 558210 European Journal of Pharmacology 2006 529 47213 Comparative Biochemistry and physiology 2013 158 181214 Cardiovascular Research 2013 99 203215 European Journal of Pharmacology 2013 710 10216 Med Sci Mont 2011 17 BR165217 J Cariovasc Pharmacol 2011 57 407220 Gen Physiol Biophys 2005 24 311221 J Cardiolvasc Electrophysiol 2005 16 1227222 Am J Physiol Heart Circ Physiol 2005 288 H778224 Plos One 2012 7 e51550

ID Journal Year volume first page225 J Biol Chem 2009 284 15432226 PNAS 2008 105 1364229 J Biol Chem 2009 284 9140230 Pflugers Arch 2013 epub232 Circ Arrhythm Electrophysiol 2013 epub233 Hum Exp Toxicol 2013 epub237 Can J Physiol Pharmacol 2010 88 1181239 J cardiolvasc Pharmacol 2010 55 145241 Cardiovascular Research 2009 82 430242 Am J Physiol Heart Circ Physiol 2008 295 H1867244 Circulation Research 2006 98 1055245 J Pharmacol Exp Ther 2006 317 1054247 J Cariovasc Pharmacol 2005 46 167249 J Cariovasc Pharmacol 2004 43 178251 Anesth Analog 2003 96 1308252 Naunyn Schmiedebergs Arch Pharmacol 2002 365 29256 European Journal of Pharmacology 1999 376 189260 British Journal of Pharmacology 2010 159 93262 J Cariovasc Pharmacol 2004 44 244263 British Journal of Pharmacology 2005 146 146264 British Journal of Pharmacology 2000 131 1809267 J Pharmacol Exp Ther 1997 281 233273 British Journal of Pharmacology 2002 136 330274 British Journal of Pharmacology 2003 140 863275 European Journal of Pharmacology 2006 532 44276 J Cardiolvasc Electrophysiol 2005 16 329278 British Journal of Pharmacology 2004 142 1300281 British Journal of Pharmacology 1997 120 968283 British Journal of Pharmacology 2006 147 289286 J Cardiolvasc Electrophysiol 2005 16 314287 European Journal of Pharmacology 2000 401 137

• Protocols of cardiac ion channels INaL Ito1 IKur IKs IK1
• Notes
• Late Sodium Channel Current
• Late sodium channel protocols
• PowerPoint Presentation
• Summary of Ito1 protocols
• Pulse sequences Ito
• Kinetics Pulse Sequences Ito
• Extracellular buffer solution of Ito
• Pipette solution of Ito
• Summary of IKur (Kv1531) protocols
• Pulse sequences IKur
• Extracellular buffer solution of IKur
• Pipette solution of IKur
• Summary of Iks protocols
• Pulse sequences IKs
• Slide 17
• Extracellular buffer solution of IKs
• Slide 19
• Pipette solution of IKs
• Slide 21
• Summary of IK1 protocols
• Pulse sequences IK1
• Slide 24
• Extracellular buffer solution of IK1
• Slide 26
• Pipette solution of IK1
• Summary of minor K protocols
• Extracellular buffer
• Intracellular buffer
• Representative current traces
• References
• Slide 33

• Protocols of cardiac ion channels INaL Ito1 IKur IKs IK1
• Notes
• Late Sodium Channel Current
• Late sodium channel protocols
• PowerPoint Presentation
• Summary of Ito1 protocols
• Pulse sequences Ito
• Kinetics Pulse Sequences Ito
• Extracellular buffer solution of Ito
• Pipette solution of Ito
• Summary of IKur (Kv1531) protocols
• Pulse sequences IKur
• Extracellular buffer solution of IKur
• Pipette solution of IKur
• Summary of Iks protocols
• Pulse sequences IKs
• Slide 17
• Extracellular buffer solution of IKs
• Slide 19
• Pipette solution of IKs
• Slide 21
• Summary of IK1 protocols
• Pulse sequences IK1
• Slide 24
• Extracellular buffer solution of IK1
• Slide 26
• Pipette solution of IK1
• Summary of minor K protocols
• Extracellular buffer
• Intracellular buffer
• Representative current traces
• References
• Slide 33