Bioeng 6460
Electrophysiology and Bioelectricity
Modeling
of
EC-Coupling and Contraction
Frank B. Sachse
fs@cvrti.utah.edu
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Overview
• Quiz
• Excitation-Contraction Coupling
• Anatomy
• Cross Bridge Binding
• Coupling
• Experimental Studies
• Outline of Experiment
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 2
Phases of Sarcomere Calcium Handling
Begin of contraction
Ca release
Triggering
Relaxation
Ca uptake
Filling
A:
Actin
M:
Myosin
Z:
Z-Disc
SL:
Sarcolemma
TTS: Transversal
tubules
SPR: Sarcoplasmic
Reticulum
Ca2+ Release
Ca2+ Uptake
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 3
Calcium Handling and EC-Coupling
Transversal
Tubule
Cell Membrane
S a rc
op
R e ti c l a s m i c
u lu m
Troponin
Mi
NCX:
ATP:
RyR:
PLB:
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h
toc
on
io
dr
n
Sodium-calcium exchanger
Ionic pump driven by ATP-hydrolysis
Calcium channel (ryanodine receptor)
Phospholamban
(Bers, Nature Insight Review Articles, 2002, modified)
Bioeng 6460: Electrophysiology and Bioelectricity - Page 4
Quiz: Role of Mitochondria
Work in pairs of 2
Time 5 min
Mitochondria take part of the cellular calcium handling.
Why were they not considered in electrophysiological models?
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 5
Sarcomeres in Cardiac Muscle (Fawcett & McNutt 69)
Sarcoplasmic reticulum
Terminal cisternae
Z-Disc
Mitochondrion Transversal tubuli
Sarcoplasmic reticulum
Dyad
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 6
Sarcomeres in Skeletal Muscle (Fawcett & McNutt 69)
Sarcolemma Transversal
tubules
I-Band
Sarcoplasmic
reticulum
A-Band
Terminal cisternae
Z-Disc
Triad
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 7
Electron Micrograph of Sarcomere in Striated Muscle
A - anisotrop
I - isotrop
(Modified from Lodish et al.,
Molecular Cell Biology,
2004)
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 8
Myofilaments
(Modified from Lodish et al.,
Molecular Cell Biology,
2004)
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 9
Proteins of Sarcomere
2 !m
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 10
EC-Coupling: Involved Proteins
2 µm
Actin
Myosin
Actin
TnC
TnI
Actin
TnT
Tm
Head-to-tail
overlap
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Tn: Troponin
Tm: Tropomyosin
A: Actin
Z: Z-Disk
(adapted from Gordon et al. 2001)
Bioeng 6460: Electrophysiology and Bioelectricity - Page 11
Force Development: Sliding Filament Theory
Cellular force development by sliding myofilaments (Huxley 1957), i.e. actin and
myosin, located in sarcomere
Attachment of myosin heads to actin
Detachment
Filament sliding
Spanning of myosin heads
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 12
Force Development: Sliding Filament Theory
http://www.sci.sdsu.edu/movies/actin_myosin.html
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 13
Actin-Myosin Interaction
A
ADP
Pi
Actin
Adenosine diphosphate
Phosphate
M
ATP
Myosin
Adenosine triphosphate
(Bers, Excitation-Contraction Coupling and Cardiac Contractile Force, 1991, modified)
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 14
EC-Coupling
Rest
small Ca2+
Stimulus
Contraction
Ca2+ release from SR
High concentration of
intracellular Ca2+
high Ca2+
Force development
Contraction of
sarcomere/cell
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 15
Contraction of Myocyte by Electrical Stimulation
Microscopic imaging of isolated ventricular cell from guinea pig
http://www-ang.kfunigraz.ac.at/˜schaffer
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 16
Measurement of Force Development in Single Cell
Myocyte glued between
glass plates
Force transmission to
measurement system
Stimulator
Fluid at body
temperature
Mechanical Fixation
(variable strain)
Glass plates
Measured force
per myocyte:
0.15-6.0 µN
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 17
Measurement Techniques
Permeabilization of sarcolemma/skinning of myocytes by saponin or Triton X-100
Direct control of intracellular concentrations of ions, drugs etc.
[
] [
Ca2+ = Ca2+
i
]
o
!
Transillumination of myocyte or muscle strands with laser light
Diffraction pattern ~ sarcomere length
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 18
Sarcomere Length Measurement via Laser Diffraction
(Figures from Lecarpentier et al., Real-Time Kinetics
of Sarcomere Relaxation by Laser Diffraction, AJP,
1985)
More information:
http://muscle.ucsd.edu/musintro/diffraction.shtml
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 19
Mathematical Modeling of Myofilament Sliding (Huxley)
f
A+ M " A#M
g
Number
of bzgl.
myosins
notungekoppeltem
bound to actin Myosin
A + M: Anzahl
von
Actin
Number
of myosins bound
actin gekoppelt)
A # M: Anzahl
Querbrücken
(Actin /to
Myosin
f,g: Ratenkonstanten
fürbinding
Bindung
Ablösung
Rate constants for
andbzw.
unbinding
d [A # M]
= f(Max[A # M] # [A # M]) # g[A # M]
dt
Max[A # M]: Maximale
Anzahl of
anmyosin
Querbrücken
Maximal number
proteins
bound to actin filament
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Bioeng 6460: Electrophysiology and Bioelectricity - Page 20