1. The Carmeliet-Coraboeuf-Weidmann Lecture
Calcium signalling in the heart:
From basic mechanisms to arrhythmias
David Eisner

2. Watch Video

3. Watch Video

4. •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

5. intracellular Ca
Watch
Video

6. Ca cycling in the heart
3Na+
L-Ca
channel
NCX
Ca2+
SR
RyR
SERCA
phospholamban
Ca2+
Ca2+
cytoplasmic Ca
SR Ca ↑ po
caffeine
po
in steady state
influx = efflux
po

7. Measurement of Ca efflux and influx
0.8
Sarcolemmal Ca entry = exit
2+
[Ca ]i
(µM)
0.0
Im
(600 pA)
L-type Ca current
(Ca entry)
Na-Ca exchange
(Ca efflux)
25
pA
4
100 ms
sarcolemmal
Ca flux
(µmol/L)
0
0.5 sec

8. What controls the size of the Ca transient?
3Na+
L-Ca
channel
NCX
Ca2+
SR
RyR
Ca2+
SERCA
phospholamban
Ca2+
Ca2+

9. •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

10. Ca transient
(SR Ca)3
0.8
Ca transient
amplitude
(µM)
0.0
0
25
50
SR Ca content (µmol/L)
Trafford et al (2000). J. Physiol. 522, 259-270

11. Consequences of steep relationship
small changes of SR content inotropically significant
Ca transient
amplitude
(µM)
SR Ca content (µmol/L)

12. Consequences of steep relationship
small changes of SR content inotropically significant
Must control SR Ca content precisely
Ca transient
amplitude
(µM)
SR Ca content (µmol/L)

13. •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

14. 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?

15. SR Ca content controls sarcolemmal fluxes
(a)
(a)
(b)
SR full
SR empty
1
(b)
1
2+
[Ca ]i
-1
(µmol.l )
0
2+
[Ca ]i
-1
(µmol.l )
Im
(1.5 nA)
0
10 sec
1 sec
systolic Ca
systolic Ca
inactivate L-type current &
Ca influx
Ca efflux on NCX
influx
efflux
(a)
ICa-L
(1.25nA)
(b)
tail
current
(0.15nA)
(a)
(b)
50 ms
200 ms
Trafford et al (1997) Circ Res 81, 477-484

16. control of Ca entry and efflux by the systolic Ca transient
controls SR Ca content: a simple feedback mechanism
3Na+
L-Ca
channel
NCX
+
-
Ca2+
RyR
Ca2+
SERCA
Ca2+
Eisner et al (1998) Cardiovasc Res 38, 589-604
Eisner et al (2000) Circ Res 87, 1087-1094

17. Relevance of feedback mechanism to HF
3Na+
L-Ca
channel
NCX
+
-
Ca2+
RyR
Ca2+
SERCA
Ca2+
↑NCX
↑ RyR po ↓ SR Ca
↓SERCA

18. •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

19. Does systolic modulation of the RyR affect systolic Ca?
phosphorylation ↑ RyR po
contributes to inotropic effects of
β-adrenergic stimulation?
P
RyR
Ca2+
Ca2+
But does ↑ RyR po →↑ systolic Ca?

20. 500 µM caffeine
[Ca2+]i
1.5
RyR opening
(uM)
0.0
12
sarcolemmal
Ca flux
efflux
(µmol/L) = influx
0
efflux > influx
[[[
SR Ca
Ca transient
efflux
influx
Increasing RyRSR Ca
↓
110
opening does not
SR Ca increase Ca
content
transient in the
(µmol/L)
70
steady state
4 sec
efflux = influx
Ca efflux

21. •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

22. Heart failure makes RyRs leaky
Control
Heart failure
Open
Closed
What are the effects of leak on Ca cycling?
Marx et al (2000) Cell 12, 365-376

23. •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

24. Ca and arrhythmias
Heart Failure
Arrhythmias
use positive
inotropes to ↑SR
& systolic Ca
Why?

25. Ca leak → diastolic Ca waves
NCX
3 Na+
SR
3 Na+
Ca2+
Ca2+
RyR
SERCA
Ca2+
Ca2+
Ca2+
Watch video

26. Ca waves occur above a threshold SR Ca content
SR Ca content (µmol/L)
Díaz et al (1997). J. Physiol. 501, 3-16

27. wave
Ca waves are arrhythmogenic
Ca efflux on NCX
depolarizing current
DAD
Ferrier et al (1973; Circ. Res. 38, 156-162)

28. How do DADs produce arrhythmias?
1. Direct depolarization
Ectopic beat
40
Em (mV)
DAD
-90
4.0
Ca
(340/380 nm)
Ca wave
2.4
500 ms

29. How do DADs produce arrhythmias?
2. Increased APD heterogeneity
50
Vm
(mV)
-90
20
Ca
(F/Fo)
470 ms
640 ms
Ca wave
1
1s
Ca wave
systolic Ca release
ICa
APD
Johnson……Volders (2013) Circ Res. 112:246-256.

30. Ca waves arise from ↑ diastolic Ca leak from SR
↑SR Ca → ↑ Ca leak
• Digitalis toxicity
• Reperfusion arrhythmias
• Inotropic agents
↑ RyR po
↑ Ca leak
• Heart Failure
• RyR mutations CPVT
(catecholaminergic
polymorphic ventricular tachycardia)

31. CPVT
Priori, S. G. et al. Circulation 2002;106:69-74
(catecholaminergic polymorphic ventricular tachycardia)
 Why do leaky RyRs → arrhythmias?
 How does β stimulation → arrhythmias?

32. Two ways to produce Ca waves
↓ threshold
0
1
Wave
probability
normal
SR Ca
↑SR Ca
0
0
50
100
SR Ca content (µmol/L)
150

33. Lower SR threshold in RyR R4496C mouse
160
Ca wave
threshold
(µmol/L)
+iso
+iso
WT
R4496C
0
WT
R4496C
Kashimura et al (2010) Circ Res 107, 1483-1489

34. SR Ca content is higher after β- stimulation
80
**
SR Ca threshold
SR Ca
content
mol/L
0
iso
β- stimulation acts by ↑ SR content (via ↑ SERCA)
Venetucci, Trafford & Eisner (2007) Circ Res 100, 105-111

35. Origin of Ca waves in CPVT
RyR mutation
↓ threshold
0
1
β- stimulation
Wave
probability
normal
SR Ca
↑SR Ca
0
0
50
100
SR Ca content (µmol/L)
150

36. Summary
•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

37. Acknowledgements
Gina Galli
David Greensmith
Takeshi Kashimura
Rajiv Sankaranarayanan
Andrew Trafford
Luigi Venetucci
Carlo Napolitano
Silvia Priori
Dan Johnson
Paul Volders

38. 24 June 1983
2013

39. Elisabetta Cerbai
Paul Volders
Godfrey Smith
Felix Bonke Alex de Hemptinne
Lennart Bouman
Hilary Brown
Barbara Casadei
Guy Vassort
Antonio Zaza
Sylvain Richard
Karin Sipido
Alessandro Mugelli
Ursula Ravens