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NCE. mM [Ca]. NMDA, AMPA rec. nAChR. CNG. VR/temp rec. VDAC. Stretch receptor. TRP. mM [Ca]. SOC. IP3R. SERCA. 100 nM [Ca]. ROC. VGCC. mM [Ca]. PMCA. RyR. SERCA. Sarcoplasmic Reticulum (SR) / T Tubule System. Twitch. Summation. Tetanus. Role of Ca ++ in contraction. - PowerPoint PPT Presentation

TRANSCRIPT

Page 1: VGCC
Page 2: VGCC

Why Calcium?

Double positive charge provides increased affinity for negatively charged proteinsbut lower affinity than larger divalent cations such as Cu, Zn, or Mn. Thecoordination chemistry of Ca is higher and more flexible than for Mg.

The fact that Ca complexes with inorganic compounds and to proteins suggeststhat the maintenance of low [Ca] intracellularly would require less energy than forother cations. The maintenance of a large transmembrane gradient is critical fora second messanger ion.

This large transmembrane gradient provides the signal-to-noise ratio required forefficient signal transduction.

Resting intracellular [Ca] is ~100 nM versus mM extracellular [Ca] or aconcentration gradient of ~10,000.

Page 3: VGCC

VGCC

ROC

SOC

TRP

Stretch receptor

CNGnAChR

NMDA, AMPA rec

VR/temp rec

IP3R

RyR

VDAC

SERCA

SERCA

NCE

PMCA

mM [Ca]

100 nM [Ca]

mM [Ca]

mM [Ca]mM [Ca]

Page 4: VGCC
Page 5: VGCC
Page 6: VGCC

Sarcoplasmic Reticulum (SR) / T Tubule System

Page 7: VGCC
Page 8: VGCC
Page 9: VGCC
Page 10: VGCC

Twitch

Page 11: VGCC

Summation

Page 12: VGCC

Tetanus

Page 13: VGCC

Role of Ca++ in contraction

Page 14: VGCC
Page 15: VGCC
Page 16: VGCC
Page 17: VGCC

SR SR

T-tubule T-tubuleA B

SR

Muscle contraction

T-tubuleVDCC

RyRFKBP12/12.6

Ca release

CRyR Accessory proteinsFKBP12/12.6KinasesPhosphatasesAdaptor/anchoring/targeting proteinsSorcinS100sTriadinJunctinAnnexin VICalmodulin

VDCC

RyR1

RyR2

Page 18: VGCC

vertebrate skeletal muscle contraction

Page 19: VGCC
Page 20: VGCC

Excitation-contraction coupling

Ryanodine receptors

Page 21: VGCC

Properties of Calcium Release Channels

Ryanodine Receptors

RyR1 RyR2 RyR3Size (amino acids)

of monomer 5,037 4,970 4,870Size (daltons) of

monomer ~565,000 ~560,000 ~560,000Sedimentationcoefficient of

tetramer30 S 30 S 30 S

Stoichiometry ofFKBP/RyR 4 4 4

Single channelconductancein Ca 50 mM

~120 pS ~120 pS ~100 pS

Single channelconductancein Cs 250 mM

~540 pS ~540 pS ?

Endogenousmodulators

M Ca activates yes yes yesmM Ca inhibits yes yes yesmM Mg inhibits yes yes yes

kinases yes yes ?phosphatases yes yes ?

DHPR interaction yes ? ?calmodulin yes yes yes

adeninenucleotides yes yes yes

MgATP yes yes yesNO yes yes ?

Page 22: VGCC

Pharmacology of Ryanodine Receptors

RyR1 RyR2 RyR3 Site of actionXanthines(caffeine) activates activates activates Ca activation sites

Ryanodine/ryanoids

subconductancestate

subconductancestate

subconductance state

carboxyterminus

ruthenium red inhibits inhibits inhibitsCa binding

site(s)/channelpore

Antraquinones(doxorubicin) activates activates activates ?

FK506 &rapamycin activates activates activates FKBP12/12.6Purinergicagonists/

antagonists(adenosine)

activates activates activates ATP binding sites

Calmodulinantagonist

inhibits inhibits inhibits calmodulin bindingsite

Local anesthetics(tetracaine) inhibits inhibits inhibits ?Dantrolene inhibits inhibits ? ?

Phenol derivatives(4-chloro-m-

cresol)activates activates ? ?

NO generatingcompounds Activates/inhibits Activates/inhibits ? ?

Page 23: VGCC

RyR

Page 24: VGCC

SRMembrane

Cytosol

Lumen

NH2

CO2H5000

4000

3000

2000

1000 500

4500

3500

1500

2500FKBPPP2A

PP1

mAKAPRII-PKA

PKA/CamKIIPO4 siteCalmodulin

Rya

no

din

e

P

Ca-Inactivation

Ca-Activation

GIG

DHPR/RyR1

MH/CCD

MH/CCD

MH/CCD

M1 M2 M3 M4

RyR2

Page 25: VGCC
Page 26: VGCC

Voltage-gated ion channels

Page 27: VGCC
Page 28: VGCC
Page 29: VGCC
Page 30: VGCC
Page 31: VGCC
Page 32: VGCC
Page 33: VGCC
Page 34: VGCC

RyR2

SR

Lumen

Ca2+

Ca2+

P

P

P

1AR 2AR

AC

cAMP

Regulation of EC coupling by adrenergicsignaling --“fight or flight”

Page 35: VGCC

RyR2 macromolecular complex

Page 36: VGCC

RyR macromolecular complexes are RyR macromolecular complexes are held together byheld together by

leucine/isoleucine zippersleucine/isoleucine zippers

Page 37: VGCC

RyR2/calcium release channel phosphorylated RyR2/calcium release channel phosphorylated by addition of cAMP aloneby addition of cAMP alone

Page 38: VGCC
Page 39: VGCC

PKA PKA phosphorylation phosphorylation activates RyR2 activates RyR2

and induces and induces subconductance subconductance

statesstates

Page 40: VGCC
Page 41: VGCC

Cytosolic [Ca]

RyR

2 o

pen

pro

bab

ilit

yR

yR2

op

en p

rob

abil

ity

ECC gainCa transientcontractility

DADs - SCD[Ca]

Time

Iso CHF

Page 42: VGCC
Page 43: VGCC
Page 44: VGCC

IP3R RyR

Skel muscle + +++

Smooth m. +++ +

Neurons +++ +++

IP3 Activates None

Ryanodine None Locks open/closes

Caffeine (5 mM) Inhibits Activates

Ca2+ IP3R1 - biphasic

IP3R2/3 - opens

biphasic

RR None inhibits

Heparin inhibits activates

Page 45: VGCC

Excitation-secretion coupling

Inositol 1,4,5-trisphosphatereceptors

Page 46: VGCC

IP3 Receptors

IP3R1 IP3R2 IP3R3Size (amino acids)

of monomer 2,749 2,691 2,685Size (daltons) of

monomer 313,000 ~300,000 ~300,000Stoichiometry of

FKBP/RyR 4 4 ?

Endogenousmodulators

IP3 activates activates activatesnM Ca activates yes yes yesM Ca inhibits yes yes noATP <2 mM activates activates activatesATP >2 mM inhibits inhibits inhibits

tyrosine kinases activates ? ?phosphatases ? ? ?

Page 47: VGCC

Pharmacology of IP3 Receptors

IP3R1 IP3R2 IP3R3 Site of action(caffeine) inhibits inhibits inhibits Ca activation sitesheparin inhibits inhibits inhibits ?2-APB inhibits inhibits inhibits ?

xestospongin inhibits inhibits inhibits ?

Page 48: VGCC

IP3 signaling

Page 49: VGCC
Page 50: VGCC

Stable transfectant Stable transfectant

Jurkat T cells do notJurkat T cells do not

express IP3R1express IP3R1

IP3R1IP3R1

Page 51: VGCC
Page 52: VGCC
Page 53: VGCC
Page 54: VGCC

Activation of IP3-gated CaActivation of IP3-gated Ca2+2+ channel by channel by fynfyn

Page 55: VGCC
Page 56: VGCC

Bcl-2

P

Bcl-2Bcl-2

Bcl-2

Rec

Growth Factor

Plasma Membrane

Cytoplasm

PIP2PLC-

IP3ER Bad

P

Degradation

14-3-3

Bcl-xBad

Ca2+

IP3R

CalcineurinSERCA

Ca2+

Cytosolic TF

NuclearTF

Survival

Nucleus

Cell Death

DNADigestion

M M

Akt

Cytoskeletal Breakdown

Cytochrome CCaspases

CAD

Bad

Survival

+ X ?

Calcium signaling during apoptosis Calcium signaling during apoptosis

Page 57: VGCC

VGCC

Stretch receptor

CNGnAChR

IP3R

RyR

VDAC

SERCA

SERCA

NCE

PMCA

mM [Ca]

100 nM [Ca]

mM [Ca]

mM [Ca]mM [Ca]

T-tubule

Sarcoplasmicreticulum

Endoplasmicreticulum

mitochondria