The Carmeliet-Coraboeuf-Weidmann Lecture

Calcium signalling in the heart: From basic mechanisms to arrhythmias

David Eisner

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• Introduction

• SR Ca is a major regulator of the Ca transient

size

• How is SR Ca controlled?

• Does modifying the RyR affect systolic Ca?

• Leaky RyR and systolic function

• The link between leaky RyRs and arrhythmias

intracellular Ca

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Ca cycling in the heart

Ca2+

SRRyR

L-Cachannel

Ca2+

SERCACa2+

3Na+

NCX

phospholamban

in steady state

influx = efflux

cytoplasmic Ca po

SR Ca ↑ po

caffeine po

L-type Ca current(Ca entry)

Na-Ca exchange(Ca efflux)

Measurement of Ca efflux and influx

Im(600 pA)

0.5 sec

0.0

[Ca2+]i(µM)

0.8

0

4

sarcolemmalCa flux

(µmol/L)

25 pA

100 ms

Sarcolemmal Ca entry = exit

Ca2+

SRRyR

L-Cachannel

Ca2+

SERCA

Ca2+

3Na+

NCX

phospholamban

What controls the size of the Ca transient?

Ca2

+

• Introduction

• SR Ca is a major regulator of the Ca transient

size

• How is SR Ca controlled?

• Does modifying the RyR affect systolic Ca?

• Leaky RyR and systolic function

• The link between leaky RyRs and arrhythmias

SR Ca content (µmol/L)

0 25 500.0

0.8

0 25 50

Ca transientamplitude(µM)

0.0

0.8

Trafford et al (2000). J. Physiol. 522, 259-270

(SR Ca)3Ca transient

Ca transientamplitude(µM)

SR Ca content (µmol/L)

Consequences of steep relationship

small changes of SR content inotropically significant

Ca transientamplitude(µM)

SR Ca content (µmol/L)

Consequences of steep relationship

small changes of SR content inotropically significant

Must control SR Ca content precisely

• Introduction

• SR Ca is a major regulator of the Ca transient

size

• How is SR Ca controlled?

• Does modifying the RyR affect systolic Ca?

• Leaky RyR and systolic function

• The link between leaky RyRs and arrhythmias

How is SR content controlled?

regulation of SERCA by phosphorylation of phospholamban (PKA & CaMKII)

“autoregulation” – effects of SR release on sarcolemmal fluxes

empty SR with caffeine: how does it refill?

(a)

(a)

1

[Ca2+]i(µmol.l-1)

0

10 sec

(a)

[Ca2+]i(µmol.l-1)

0

1

Im(1.5 nA)

(b)

1 sec

(b)ICa-L

(1.25nA)

50 ms

influx

tailcurrent

(0.15nA)

(b)

200 ms

efflux

Trafford et al (1997) Circ Res 81, 477-484

SR Ca content controls sarcolemmal fluxes

systolic Ca inactivate L-type current & Ca influx

systolic Ca Ca efflux on NCX

(a) (b)

SR empty SR full

RyR

L-Cachannel

Ca2+

SERCA

Ca2+

3Na+

NCX

-

Ca2+

Eisner et al (1998) Cardiovasc Res 38, 589-604

Eisner et al (2000) Circ Res 87, 1087-1094

control of Ca entry and efflux by the systolic Ca transient

controls SR Ca content: a simple feedback mechanism

+

RyR

L-Cachannel

Ca2+

SERCA

Ca2+

3Na+

NCX

-

Ca2+

Relevance of feedback mechanism to HF

+

↑NCX

↑ RyR po

↓SERCA

↓ SR Ca

• Introduction

• SR Ca is a major regulator of the Ca transient

size

• How is SR Ca controlled?

• Does modifying the RyR affect systolic Ca?

• Leaky RyR and systolic function

• The link between leaky RyRs and arrhythmias

RyR

Ca2+Ca2+

Does systolic modulation of the RyR affect systolic Ca?

P

phosphorylation ↑ RyR po contributes to inotropic effects of

β-adrenergic stimulation?

But does ↑ RyR po →↑ systolic Ca?

[Ca2+]i

(uM)

SR Cacontent

(µmol/L)70

110

sarcolemmalCa flux

0

120.0

1.5

500 µM caffeine

efflux > influx

efflux = influx↓SR Ca

(µmol/L)

4 sec

efflux = influx

RyR opening

Ca transient

Ca efflux

SR Ca

efflux

influx

[[[

Increasing RyR opening does

not increase Ca transient in the

steady state

• Introduction

• SR Ca is a major regulator of the Ca transient

size

• How is SR Ca controlled?

• Does modifying the RyR affect systolic Ca?

• Leaky RyR and systolic function

• The link between leaky RyRs and arrhythmias

Closed

Open

Marx et al (2000) Cell 12, 365-376

Heart failure makes RyRs leaky

Heart failure Control

What are the effects of leak on Ca cycling?

• Introduction

• SR Ca is a major regulator of the Ca transient

size

• How is SR Ca controlled?

• Does modifying the RyR affect systolic Ca?

• Leaky RyR and systolic function

• The link between leaky RyRs and arrhythmias

Ca and arrhythmias

Why?

Heart Failure

use positive inotropes to

↑SR & systolic Ca

Arrhythmias

Ca2+

Ca2+

3 Na+

Ca2+ Ca2+

SR

RyR

SERCA

Ca2+

3 Na+NCX

Ca leak → diastolic Ca waves

Watch video

Díaz et al (1997). J. Physiol. 501, 3-16

SR Ca content (µmol/L)

Ca waves occur above a threshold SR Ca content

Ferrier et al (1973; Circ. Res. 38, 156-162)

Ca efflux on NCX

DAD

Ca waves are arrhythmogenicwave

depolarizing current

How do DADs produce arrhythmias?

1. Direct depolarization

Ca (340/380 nm)

2.4

4.0

Em (mV)

-90

40

500 ms

Ca wave

DAD

Ectopic beat

2. Increased APD heterogeneity

How do DADs produce arrhythmias?

Johnson……Volders (2013) Circ Res. 112:246-256.

systolic Ca release ICa APD Ca wave

20

-90

50

Vm (mV)

Ca (F/Fo)

470 ms 640 ms

1 s

Ca wave

1

• Digitalis toxicity• Reperfusion arrhythmias • Inotropic agents

↑SR Ca → ↑ Ca leak

↑ RyR po ↑ Ca leak

Ca waves arise from ↑ diastolic Ca leak from SR

• Heart Failure• RyR mutations CPVT (catecholaminergic

polymorphic ventricular tachycardia)

CPVT

(catecholaminergic polymorphic ventricular tachycardia)

Priori, S. G. et al. Circulation 2002;106:69-74

Why do leaky RyRs → arrhythmias?

How does β stimulation → arrhythmias?

Two ways to produce Ca waves

SR Ca content (µmol/L)0 50 150100

0

1

Waveprobability

0

normalSR Ca

↑SR Ca

↓ threshold

WT WTR4496C R4496C

0

160

Ca wave threshold(µmol/L) +iso +iso

Lower SR threshold in RyR R4496C mouse

Kashimura et al (2010) Circ Res 107, 1483-1489

SR Ca content is higher after β- stimulation

SR Cacontentmol/L

0

80

iso

**

SR Ca threshold

β- stimulation acts by ↑ SR content (via ↑ SERCA)

Venetucci, Trafford & Eisner (2007) Circ Res 100, 105-111

Origin of Ca waves in CPVT

SR Ca content (µmol/L)0 50 150100

0

1

Waveprobability

0

normalSR Ca

↑SR Ca

↓ threshold

β- stimulation

RyR mutation

• Steep dependence of Ca release on SR content

• Even a small decrease of SR Ca is important in HF

• SR Ca controlled by effects of systolic Ca transient on NCX and L-type current

• systolic Ca due to β-adrenergic stimulation not due to effect on RyR

• Leaky RyR diastolic Ca

• deranged RyR per se not arrhythmogenic

• CPVT decreases threshold

Summary

Gina Galli

David Greensmith

Takeshi Kashimura

Rajiv Sankaranarayanan

Andrew Trafford

Luigi Venetucci

Carlo Napolitano

Silvia Priori

Dan Johnson

Paul Volders

Acknowledgements

198324 June 2013

Felix BonkeLennart Bouman

Hilary Brown

Ursula RavensAlessandro MugelliKarin SipidoSylvain Richard

Antonio Zaza

Barbara Casadei

Elisabetta CerbaiPaul Volders Alex de Hemptinne

Guy Vassort

Godfrey Smith