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| This page contain abstracts from more than 30 published research papers that have used the AE801 Sensor to obtain the data needed for the research. UI - 99388113
AU - Campbell SP
AU - Williams DA
AU - Frueh BR
AU - Lynch GS
TI - Contractile activation characteristics of single permeabilized fibres
from levator palpebrae superioris, orbicularis oculi and vastus
lateralis muscles from humans [In Process Citation]
LA - Eng
DA - 19990925
DP - 1999 Sep 1
IS - 0022-3751
TA - J Physiol (Lond)
PG - 615-22
SB - M
CY - ENGLAND
VI - 519 Pt 2
JC - JQV
AA - AUTHOR
AB - 1. We investigated the contractile activation characteristics of single
membrane-permeabilized fibres from the following muscles from humans:
the levator palpebrae superioris (LPS), an extraocular muscle; the
orbicularis oculi (OO), a facial muscle; and the vastus lateralis (VL),
a major muscle of the thigh. 2. Single permeabilized muscle fibres were
isolated from each of the different muscles, attached to a sensitive
force transducer and activated by rapid immersion in buffered solutions
of varying [Ca2+] and [Sr2+]. Fibres were allocated into discrete
populations based on their contractile characteristics, including their
differential force responses during Ca2+ and Sr2+ activation. 3. With
the exception of one fibre from the LPS, all 152 fibres sampled from
the three different human muscles could be classified into either
population I (slow, type I) or population II (fast, type II) based on
their force-pCa(pSr) relations. The LPS muscle fibre which was unable
to be classified into the two major fibre populations displayed a
combination of the typical force-pCa(pSr) relations for mammalian fast
and slow muscle fibres. 4. Although fibres from the LPS, OO and VL
muscles had similar differential sensitivities to Ca2+and Sr2+, the
steepness of the force-pCa(pSr) curves for fibres from the LPS and OO
muscles were highly variable compared with those for fibres from the VL
muscle. Specific forces (N cm-2) of the smaller diameter fibres from
the LPS and OO muscles were significantly lower than those of fibres
from the VL muscle. 5. The differences in the contractile activation
characteristics between fibres from the VL muscle and those of fibres
from facial (OO) muscles and extraocular (LPS) muscles, reflect the
differences in their fibre composition that are responsible for their
functional specificity.
AD - Muscle and Cell Physiology Laboratory, Department of Physiology, The
University of Melbourne, Parkville, Victoria 3052, Australia.
RO - O:099
PMID- 0010457076
PID - PHY_9327
4100- http://www.journals.cup.org/owa_dba/owa/approval?sjid=PHY&said=9327&spii
=S0022375199093278
SO - J Physiol (Lond) 1999 Sep 1;519 Pt 2:615-22
UI - 99388104
AU - Edman KA
TI - The force bearing capacity of frog muscle fibres during stretch: its
relation to sarcomere length and fibre width [In Process Citation]
LA - Eng
DA - 19990925
DP - 1999 Sep 1
IS - 0022-3751
TA - J Physiol (Lond)
PG - 515-26
SB - M
CY - ENGLAND
VI - 519 Pt 2
JC - JQV
AA - AUTHOR
AB - 1. Single fibres isolated from the anterior tibialis muscle of Rana
temporaria were tetanized (0.9-1.8 C) while a marked ( approximately 1
mm) segment was held at constant length by feedback control. Force
enhancement was produced by applying a controlled stretch ramp to the
fibre segment during the tetanus plateau, the steady force reached
during stretch being used as a measure of the maximum force that the
myosin cross-bridges can hold before they detach. 2. The amplitude of
force enhancement during stretch did not vary in proportion to the
isometric force as the sarcomere length was changed, maximum force
enhancement being attained near 2.4 &mgr;m sarcomere length compared
with 2.0 &mgr;m for the isometric force. 3. The influence of fibre
width on the force enhancement-sarcomere length relationship was
evaluated by normalizing force enhancement to the tetanic (pre-stretch)
force in this way allowing for the differences in myofilament overlap
at the various lengths. The amplitude of force enhancement (normalized
to the tetanic force) increased by approximately 70 % as the relative
width of the myofilament lattice was reduced from a nominal value of
1.05 at a sarcomere length of 1.8 &mgr;m to 0.85 at a sarcomere length
of 2.8 &mgr;m. 4. Changes in fibre width equivalent to those produced
by altering the sarcomere length were produced by varying the tonicity
of the extracellular medium. Force enhancement, normalized to the
control isometric force at each tonicity, exhibited a width dependence
that agreed well with that described in the previous point. Stretch
ramps applied to frog skinned muscle fibres during calcium-induced
contracture likewise resulted in a greater force enhancement during
stretch after reducing the fibre width by osmotic compression. 5. The
results suggest that the strength of binding of the myosin cross-
bridges, unlike the isometric force, varies with the lateral distance
between the myofilaments.
AD - Department of Pharmacology, University of Lund, Solvegatan 10, S-223 62
Lund, Sweden.
RO - O:099
PMID- 0010457067
PID - PHY_8921
4100- http://www.journals.cup.org/owa_dba/owa/approval?sjid=PHY&said=8921&spii
=S0022375199089218
SO - J Physiol (Lond) 1999 Sep 1;519 Pt 2:515-26
UI - 99310898
AU - Parris JR
AU - Cobban HJ
AU - Littlejohn AF
AU - MacEwan DJ
AU - Nixon GF
TI - Tumour necrosis factor-alpha activates a calcium sensitization pathway
in guinea-pig bronchial smooth muscle.
LA - Eng
MH - p42(Mapk) Kinase/metabolism
MH - Animal
MH - Bronchi/*drug effects
MH - Calcium/metabolism/physiology
MH - Calcium Signaling/*drug effects
MH - Guinea Pigs
MH - In Vitro
MH - Male
MH - Muscle Contraction/drug effects
MH - Muscle, Smooth/*drug effects
MH - Myosin Light Chains/metabolism
MH - Recombinant Proteins/pharmacology
MH - Sphingomyelin Phosphodiesterase/metabolism
MH - Support, Non-U.S. Gov't
MH - Tumor Necrosis Factor/*pharmacology
RN - EC 2.7.10.- (p42(Mapk) Kinase)
RN - EC 3.1.4.12 (Sphingomyelin Phosphodiesterase)
RN - 0 (Myosin Light Chains)
RN - 0 (Recombinant Proteins)
RN - 0 (Tumor Necrosis Factor)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
DA - 19991020
DP - 1999 Jul 15
IS - 0022-3751
TA - J Physiol (Lond)
PG - 561-9
SB - M
CY - ENGLAND
VI - 518 ( Pt 2)
JC - JQV
AA - Author
EM - 199912
AB - 1. The effects of tumour necrosis factor-alpha (TNF) on guinea-pig
bronchial smooth muscle contractility were investigated. 2. The Ca2+-
activated contractile response of permeabilized bronchial smooth muscle
strips was significantly increased after incubation with 1 microgram ml-
1 TNF for 45 min. This TNF-induced effect was not due to a further
increase in intracellular Ca2+. 3. The TNF-induced Ca2+ sensitization
was, at least partly, the result of an increase in myosin light chain20
phosphorylation. 4. The intracellular signalling pathway involved in
this effect of TNF was further investigated. Sphingomyelinase, a
potential mediator of TNF, had no effect on Ca2+ sensitivity of
permeabilized bronchial smooth muscle. Also, p42/p44 mitogen-activated
protein kinase (p42/p44mapk), activated by TNF in some cell types, did
not show an increased activation in bronchial smooth muscle after TNF
treatment. 5. In conclusion, TNF may activate a novel signalling
pathway in guinea-pig bronchial smooth muscle leading to an increase in
myosin light chain20 phosphorylation and a subsequent increase in Ca2+
sensitivity of the myofilaments. This pathway does not appear to
involve sphingomyelinase-liberated ceramides or activation of
p42/p44mapk. Given the importance of TNF in asthma, this TNF-induced
Ca2+ sensitization of the myofilaments may represent a mechanism
responsible for airway hyper-responsiveness.
AD - Department of Biomedical Sciences, Institute of Medical Sciences,
University of Aberdeen, Aberdeen, UK.
PMID- 0010381600
PID - PHY_8883
4100- http://www.journals.cup.org/owa_dba/owa/approval?sjid=PHY&said=8883&spii
=S0022375199088833
SO - J Physiol (Lond) 1999 Jul 15;518 ( Pt 2):561-9
UI - 99288149
AU - He ZH
AU - Chillingworth RK
AU - Brune M
AU - Corrie JE
AU - Webb MR
AU - Ferenczi MA
TI - The efficiency of contraction in rabbit skeletal muscle fibres,
determined from the rate of release of inorganic phosphate.
LA - Eng
MH - Adenosine Triphosphate/analogs & derivatives/metabolism
MH - Adenosinetriphosphatase/*metabolism
MH - Animal
MH - In Vitro
MH - Kinetics
MH - Muscle Contraction/*physiology
MH - Muscle Fibers/*physiology
MH - Muscle, Skeletal/*physiology
MH - Phosphates/*metabolism
MH - Photolysis
MH - Rabbits
MH - Sarcomeres/*physiology
MH - Temperature
RN - EC 3.6.1.3 (Adenosinetriphosphatase)
RN - 0 (Phosphates)
RN - 56-65-5 (Adenosine Triphosphate)
RN - 67030-27-7 (P(3)-1-(2-nitro)phenylethyladenosine 5'-triphosphate)
PT - JOURNAL ARTICLE
DA - 19990802
DP - 1999 Jun 15
IS - 0022-3751
TA - J Physiol (Lond)
PG - 839-54
SB - M
CY - ENGLAND
VI - 517 ( Pt 3)
JC - JQV
AA - Author
EM - 199910
AB - 1. The relationship between mechanical power output and the rate of ATP
hydrolysis was investigated in segments of permeabilized fibres
isolated from rabbit psoas muscle. 2. Contractions were elicited at 12
degrees C by photolytic release of ATP from the P3 -1-(2-nitrophenyl)
ester of ATP (NPE-caged ATP). Inorganic phosphate (Pi) release was
measured by a fluorescence method using a coumarin-labelled phosphate
binding protein. Force and sarcomere length were also monitored. 3.
ATPase activity was determined from the rate of appearance of Pi during
each phase of contraction. The ATPase rate was 10.3 s-1 immediately
following release of ATP and 5. 1 s-1 during the isometric phase prior
to the applied shortening. It rose hyperbolically with shortening
velocity, reaching 18.5 s-1 at a maximal shortening velocity > 1 ML s-1
(muscle lengths s-1). 4. Sarcomeres shortened at 0.09 ML s-1
immediately following the photolytic release of ATP and at 0.04 ML s-1
prior to the period of applied shortening. The high initial ATPase rate
may be largely attributed to initial sarcomere shortening. 5. During
shortening, maximal power output was 28 W l-1. Assuming the free energy
of hydrolysis is 50 kJ mol-1, the efficiency of contraction was
calculated from the power output at each shortening velocity. The
maximum efficiency was 0.36 at a shortening velocity of 0.27 ML s-1,
corresponding to a force level 51 % of that in the isometric state. 6.
At the maximal shortening velocity, only 10 % of the myosin heads are
attached to the thin filaments at any one time.
AD - National Institute for Medical Research, The Ridgeway, Mill Hill,
London NW7 1AA, UK.
PMID- 0010358123
PID - PHY_8978
4100- http://www.journals.cup.org/owa_dba/owa/approval?sjid=PHY&said=8978&spii
=S0022375199089784
SO - J Physiol (Lond) 1999 Jun 15;517 ( Pt 3):839-54
UI - 99288148
AU - Hanley PJ
AU - Young AA
AU - LeGrice IJ
AU - Edgar SG
AU - Loiselle DS
TI - 3-Dimensional configuration of perimysial collagen fibres in rat
cardiac muscle at resting and extended sarcomere lengths.
LA - Eng
MH - Analysis of Variance
MH - Animal
MH - Collagen/*ultrastructure
MH - Heart/physiology
MH - Heart Ventricle
MH - Image Processing, Computer-Assisted
MH - Microscopy, Confocal
MH - Microscopy, Electron
MH - Models, Structural
MH - Myocardium/cytology/*ultrastructure
MH - Rats
MH - Rats, Wistar
MH - Sarcomeres/*physiology/*ultrastructure
MH - Support, Non-U.S. Gov't
RN - 9007-34-5 (Collagen)
PT - JOURNAL ARTICLE
DA - 19990802
DP - 1999 Jun 15
IS - 0022-3751
TA - J Physiol (Lond)
PG - 831-7
SB - M
CY - ENGLAND
VI - 517 ( Pt 3)
JC - JQV
AA - Author
EM - 199910
AB - 1. We have used fluorescence confocal laser scanning microscopy to
attain the three-dimensional (3-D) microstructure of perimysial
collagen fibres over the range of sarcomere lengths (1.9-2.3
micrometers) in which passive force of cardiac muscle increases
steeply. 2. A uniaxial muscle preparation (right ventricular trabecula
of rat) was used so that the 3-D collagen configuration could be
readily related to sarcomere length. Transmission electron microscopy
showed that these preparations were structurally homologous to
ventricular wall muscle. 3. Trabeculae were mounted on the stage of an
inverted microscope and fixed at various sarcomere lengths. After a
trabecula was stained with the fluorophore Sirius Red F3BA and embedded
in resin, sequential optical sectioning enabled 3-D reconstruction of
its perimysial collagen fibres. The area fraction of these fibres,
determined from the cross-sections of seven trabeculae, was 10.5 +/-
3.9 % (means +/- s.d.). 4. The reconstructed 3-D images show that
perimysial collagen fibres are wavy (as distinct from coiled) cords
which straighten considerably as the sarcomere length is increased from
1.85 +/- 0.06 micrometer (near-resting length) to 2.3 +/- 0.04
micrometer (means +/- s.d., n = 4). These observations are consistent
with the notion that the straightening of these fibres is responsible
for limiting extension of the cardiac sarcomere to a length of
approximately 2.3 micrometers.
AD - Department of Physiology, School of Medicine and Health Science,
University of Auckland, Auckland, New Zealand. p_hanley98@hotmail.com
PMID- 0010358122
PID - PHY_9009
4100- http://www.journals.cup.org/owa_dba/owa/approval?sjid=PHY&said=9009&spii
=S0022375199090092
SO - J Physiol (Lond) 1999 Jun 15;517 ( Pt 3):831-7
UI - 99272460
AU - Yoshii A
AU - Iizuka K
AU - Dobashi K
AU - Horie T
AU - Harada T
AU - Nakazawa T
AU - Mori M
TI - Relaxation of contracted rabbit tracheal and human bronchial smooth
muscle by Y-27632 through inhibition of Ca2+ sensitization.
LA - Eng
MH - Amides/*pharmacology
MH - Animal
MH - Bronchi/*metabolism
MH - Calcium/*antagonists & inhibitors
MH - Carbachol/pharmacology
MH - Cholinergic Agents/pharmacology
MH - Detergents/pharmacology
MH - Dose-Response Relationship, Drug
MH - Endothelin-1/pharmacology
MH - G-Proteins/metabolism
MH - Guanosine 5'-O-(3-Thiotriphosphate)/pharmacology
MH - Human
MH - Muscle Contraction
MH - Muscle, Smooth/*metabolism
MH - Myosin-Light-Chain Kinase/metabolism
MH - Octoxynol/pharmacology
MH - Oxazoles/pharmacology
MH - Phosphodiesterase Inhibitors/pharmacology
MH - Phospholipase C/pharmacology
MH - Protein-Serine-Threonine Kinases/metabolism
MH - Pyridines/*pharmacology
MH - Rabbits
MH - Signal Transduction
MH - Support, Non-U.S. Gov't
MH - Trachea/*metabolism
MH - 1-Methyl-3-isobutylxanthine/pharmacology
RN - EC 2.7.1.117 (Myosin-Light-Chain Kinase)
RN - EC 2.7.10 (Protein-Serine-Threonine Kinases)
RN - EC 2.7.10.- (Rho-associated kinase)
RN - EC 3.1.4.3 (Phospholipase C)
RN - 0 (Amides)
RN - 0 (Cholinergic Agents)
RN - 0 (Detergents)
RN - 0 (Endothelin-1)
RN - 0 (G-Proteins)
RN - 0 (Oxazoles)
RN - 0 (Phosphodiesterase Inhibitors)
RN - 0 (Pyridines)
RN - 101932-71-2 (calyculin A)
RN - 138381-45-0 (Y 27632)
RN - 28822-58-4 (1-Methyl-3-isobutylxanthine)
RN - 37589-80-3 (Guanosine 5'-O-(3-Thiotriphosphate))
RN - 51-83-2 (Carbachol)
RN - 7440-70-2 (Calcium)
RN - 9002-93-1 (Octoxynol)
PT - JOURNAL ARTICLE
DA - 19990707
DP - 1999 Jun
IS - 1044-1549
TA - Am J Respir Cell Mol Biol
PG - 1190-200
SB - M
CY - UNITED STATES
IP - 6
VI - 20
JC - AOB
AA - Author
EM - 199909
AB - The mechanism of Ca2+ sensitization of contraction has not been
elucidated in airway smooth muscle (SM). To determine the role of a
small G protein, rhoA p21, and its target protein, rho-associated
coiled coil-forming protein kinase (ROCK), in receptor-coupled Ca2+
sensitization of airway SM, we studied the effect of (+)-(R)-trans-4-(1-
aminoethyl)-N-(4-pyridyl)cyclohexane carboxamide dihydrochloride,
monohydrate (Y-27632), a ROCK inhibitor, on isometric contractions in
rabbit tracheal and human bronchial SM. Y-27632 completely reversed 1
microM carbachol (CCh)-induced contraction of intact trachea with a
concentration producing half-maximum inhibition of effect (IC50) of
1.29 +/- 0.2 microM (n = 5). Although 4beta-phorbol 12,13-dibutyrate (1
microM)-induced Ca2+ sensitization was relatively resistant to Y-27632
in alpha-toxin-permeabilized trachea, CCh (100 microM) plus guanosine
triphosphate (GTP) (3 microM)- and guanosine 5'-O-(3'-thiotriphosphate)
(10 microM)-induced contractions were relaxed completely by Y-27632
with IC50 of 1.44 +/- 0.3 (n = 6) and 1.15 +/- 0.3 microM (n = 6).
Endothelin-1 (1 microM) plus GTP (3 microM)- developed force was also
reversed by Y-27632 with IC50 of 4. 10 +/- 1.1 microM (n = 6) in the
alpha-toxin-permeabilized bronchus. Both the rabbit and human SM
expressed rhoA p21, ROCK I, and its isoform ROCK II. Collectively,
rho/ROCK-mediated Ca2+ sensitization plays a central role in the
sustained phase of airway SM contraction, and selective inhibition of
this pathway may become a new strategy to resolve airflow limitation in
asthma.
AD - First Department of Internal Medicine, Faculty of Medicine, School of
Medicine, Gunma University, Maebashi, Gunma, Japan.
PMID- 0010340938
SO - Am J Respir Cell Mol Biol 1999 Jun;20(6):1190-200
UI - 99045480
AU - He ZH
AU - Ferenczi MA
AU - Brune M
AU - Trentham DR
AU - Webb MR
AU - Somlyo AP
AU - Somlyo AV
TI - Time-resolved measurements of phosphate release by cycling cross-
bridges in portal vein smooth muscle.
LA - Eng
MH - Adenosine Triphosphate/analogs & derivatives/metabolism
MH - Adenosinetriphosphatase/metabolism
MH - Animal
MH - Biophysics
MH - In Vitro
MH - Kinetics
MH - Muscle Contraction/physiology
MH - Muscle, Smooth, Vascular/*metabolism/physiology
MH - Myosin/metabolism
MH - Permeability
MH - Phosphates/*metabolism
MH - Phosphorylation
MH - Photolysis
MH - Portal Vein/metabolism/physiology
MH - Rabbits
MH - Support, Non-U.S. Gov't
MH - Support, U.S. Gov't, P.H.S.
RN - EC 3.6.1.3 (Adenosinetriphosphatase)
RN - 0 (Myosin)
RN - 0 (Phosphates)
RN - 56-65-5 (Adenosine Triphosphate)
RN - 67030-27-7 (P(3)-1-(2-nitro)phenylethyladenosine 5'-triphosphate)
PT - JOURNAL ARTICLE
ID - HL19242/HL/NHLBI
DA - 19990125
DP - 1998 Dec
IS - 0006-3495
TA - Biophys J
PG - 3031-40
SB - M
CY - UNITED STATES
IP - 6
VI - 75
JC - A5S
AA - Author
EM - 199903
AB - The rate of release of inorganic phosphate (Pi) from cycling cross-
bridges in rabbit portal-anterior mesenteric vein smooth muscle was
determined by following the fluorescence of the Pi-reporter, MDCC-PBP
(Brune, M., J. L. Hunter, S. A. Howell, S. R. Martin, T. L. Hazlett, J.
E. T. Corrie, and M. R. Webb. 1998. Biochemistry. 37:10370-10380).
Cross-bridge cycling was initiated by photolytic release of ATP from
caged-ATP in Triton-permeabilized smooth muscles in rigor. When the
regulatory myosin light chains (MLC20) had been thiophosphorylated, the
rate of Pi release was biphasic with an initial rate of 80 microM s-1
and amplitude 108 microM, decreasing to 13.7 microM s-1. These rates
correspond to fast and slow turnovers of 1.8 s-1 and 0.3 s-1, assuming
84% thiophosphorylation of 52 microM myosin heads. Activation by Ca2+-
dependent phosphorylation subsequent to ATP release resulted in slower
Pi release, paralleling the rate of contraction that was also slower
than after thiophosphorylation, and was also biphasic: 51 microM s-1
and 13.2 microM s-1. These rates suggest that the activity of myosin
light chain kinase and phosphatase ("pseudo-ATPase") contributes <20%
of the ATP usage during cross-bridge cycling. The extracellular "ecto-
nucleotidase" activity was reduced eightfold by permeabilization,
conditions in which the ecto-ADPase was 17% of the ecto-ATPase.
Nevertheless, the remaining ecto-ATPase activity reduced the precision
of the estimate of cross-bridge ATPase. We conclude that the transition
from fast to slow ATPase rates reflects the properties and forces
directly acting on cross-bridges, rather than the result of a time-
dependent decrease in activation (MLC20 phosphorylation) occurring in
intact smooth muscle. The mechanisms of slowing may include the effect
of positive strain on cross-bridges, inhibition of the cycling rate by
high affinity Mg-ADP binding, and associated state hydrolysis.
AD - National Institute for Medical Research, The Ridgeway, Mill Hill,
London NW7 1AA, United Kingdom.
PMID- 0009826623
SO - Biophys J 1998 Dec;75(6):3031-40
UI - 99007358
AU - He Z
AU - Stienen GJ
AU - Barends JP
AU - Ferenczi MA
TI - Rate of phosphate release after photoliberation of adenosine 5'-
triphosphate in slow and fast skeletal muscle fibers.
LA - Eng
MH - Adenosine Diphosphate/pharmacology
MH - Adenosine Triphosphate/*analogs & derivatives/metabolism
MH - Animal
MH - Calcium/pharmacology
MH - Carrier Proteins/metabolism
MH - Coumarins/metabolism
MH - Fluorescent Dyes
MH - Kinetics
MH - Muscle Contraction/physiology
MH - Muscle Fibers, Fast-Twitch/*metabolism
MH - Muscle Fibers, Slow-Twitch/*metabolism
MH - Muscle, Skeletal/*physiology
MH - Phosphates/*metabolism
MH - Photolysis
MH - Psoas Muscles/physiology
MH - Rabbits
MH - Sarcomeres/metabolism
MH - Support, Non-U.S. Gov't
RN - 0 (phosphate-binding proteins)
RN - 0 (Carrier Proteins)
RN - 0 (Coumarins)
RN - 0 (Fluorescent Dyes)
RN - 0 (Phosphates)
RN - 156571-46-9 (N-(2-(1-maleimidyl)ethyl)-7-(diethylamino)coumarin-3-
carboxamide)
RN - 56-65-5 (Adenosine Triphosphate)
RN - 58-64-0 (Adenosine Diphosphate)
RN - 67030-27-7 (P(3)-1-(2-nitro)phenylethyladenosine 5'-triphosphate)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
DA - 19981204
DP - 1998 Nov
IS - 0006-3495
TA - Biophys J
PG - 2389-401
SB - M
CY - UNITED STATES
IP - 5
VI - 75
JC - A5S
AA - Author
EM - 199902
AB - Inorganic phosphate (Pi) release was determined by means of a
fluorescent Pi-probe in single permeabilized rabbit soleus and psoas
muscle fibers. Measurements of Pi release followed photoliberation of
approximately 1.5 mM ATP by flash photolysis of NPE-caged ATP in the
absence and presence of Ca2+ at 15 degrees C. In the absence of Ca2+,
Pi release occurred with a slow rate of 11 +/- 3 microM . s-1 (n = 3)
in soleus fibers and 23 +/- 1 microM . s-1 (n = 10) in psoas fibers. At
saturating Ca2+ concentrations (pCa 4.5), photoliberation of ATP was
followed by rapid force development. The initial rate of Pi release was
0.57 +/- 0.05 mM . s-1 in soleus (n = 13) and 4.7 +/- 0.2 mM . s-1 in
psoas (n = 23), corresponding to a rate of Pi release per myosin head
of 3.8 s-1 in soleus and 31.5 s-1 in psoas. Pi release declined at a
rate of 0.48 s-1 in soleus and of 5.2 s-1 in psoas. Pi release in
soleus was slightly faster in the presence of an ATP regenerating
system but slower when 0.5 mM ADP was added. The reduction in the rate
of Pi release results from an initial redistribution of cross-bridges
over different states and a subsequent ADP-sensitive slowing of cross-
bridge detachment.
AD - National Institute for Medical Research, The Ridgeway, Mill Hill,
London NW7 1AA, United Kingdom.
PMID- 0009788934
SO - Biophys J 1998 Nov;75(5):2389-401
UI - 98399940
AU - Slawnych MP
AU - Morishita L
AU - Bressler BH
TI - Image-analysis-based assessment of the effects of the "Ca2+-jump"
technique on sarcomere uniformity.
LA - Eng
MH - Animal
MH - Calcium/*diagnostic use
MH - Image Processing, Computer-Assisted
MH - In Vitro
MH - Muscle Contraction/physiology
MH - Muscle Fibers/*physiology/*ultrastructure
MH - Muscle Relaxation/physiology
MH - Muscle, Skeletal/*physiology/*ultrastructure
MH - Rabbits
MH - Sarcomeres/*physiology/*ultrastructure
MH - Solutions
MH - Spectroscopy, Fourier Transform Infrared
MH - Support, Non-U.S. Gov't
RN - 0 (Solutions)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
DA - 19981105
DP - 1998 Sep
IS - 8750-7587
TA - J Appl Physiol
PG - 955-61
SB - M
CY - UNITED STATES
IP - 3
VI - 85
JC - HEG
AA - Author
EM - 199901
AB - A new image analysis-based technique was used to quantitatively examine
the effects of the "Ca2+-jump" activation protocol on the maintenance
of fiber quality in skinned rabbit psoas muscle fiber segments.
Specifically, contractions in pCa 4.6 were preceded by short-duration
"preactivation" soaks in a solution in which EGTA was replaced with the
low-Ca2+ buffering capacity analog hexamethylenediamine-N, N, N', N'-
tetraacetate, which facilitated rapid Ca2+ equilibration within the
fiber segments. Fiber quality was assessed by examining the Fourier
spectra of the muscle fiber images before, during, and after
activation. Segment lengths were typically below 500 micrometer, thus
allowing the majority of the sarcomeres to be visualized in the field
of view (x200 and x400 magnification). The preactivation protocol
resulted in less deterioration of fiber quality with repetitive
activation. In addition, there was also a significant reduction in the
time required to reach the 50% level of maximum tension, with no
significant change in the maximum tension level.
AD - Department of Anatomy, University of British Columbia, Vancouver,
British Columbia, Canada V6T 1Z3. slawnych@cortex.biomed.mcgill.ca
PMID- 0009729569
SO - J Appl Physiol 1998 Sep;85(3):955-61
UI - 98365280
AU - Sun Y
AU - Caputo C
AU - Edman KA
TI - Effects of BAPTA on force and Ca2+ transient during isometric
contraction of frog muscle fibers.
LA - Eng
MH - Animal
MH - Calcium/*metabolism
MH - Chelating Agents/*pharmacology
MH - Egtazic Acid/*analogs & derivatives/pharmacology
MH - In Vitro
MH - Isometric Contraction/*drug effects
MH - Kinetics
MH - Muscle Fibers/drug effects/*physiology
MH - Muscle, Skeletal/drug effects/*physiology
MH - Rana temporaria
MH - Sarcomeres/drug effects/physiology
MH - Support, Non-U.S. Gov't
MH - Time Factors
RN - 0 (Chelating Agents)
RN - 67-42-5 (Egtazic Acid)
RN - 7440-70-2 (Calcium)
RN - 85233-19-8 (1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid)
PT - JOURNAL ARTICLE
DA - 19980916
DP - 1998 Aug
IS - 0002-9513
TA - Am J Physiol
PG - C375-81
SB - M
CY - UNITED STATES
IP - 2 Pt 1
VI - 275
JC - 3U8
AA - Author
EM - 199811
AB - The effects of 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid
(BAPTA) on force and intracellular Ca2+ transient were studied during
isometric twitches and tetanuses in single frog muscle fibers. BAPTA
was added to the bathing solution in its permeant AM form (50 and 100
microM). There was no clear correlation between the changes in force
and the changes in Ca2+ transient. Thus during twitch stimulation BAPTA
did not suppress the Ca2+ transient until the force had been reduced to
<50% of its control value. At the same time, the peak myoplasmic free
Ca2+ concentration reached during tetanic stimulation was markedly
increased, whereas the force was slightly reduced by BAPTA. The effects
of BAPTA were not duplicated by using another Ca2+ chelator, EGTA,
indicating that BAPTA may act differently as a Ca2+ chelator. Stiffness
measurements suggest that the decrease in mechanical performance in the
presence of BAPTA is attributable to a reduced number of active cross
bridges. The results could mean that BAPTA, under the conditions used,
inhibits the binding of Ca2+ to troponin C resulting in a reduced state
of activation of the contractile system.
AD - Department of Pharmacology, University of Lund, S-223 62 Lund, Sweden.
PMID- 0009688591
SO - Am J Physiol 1998 Aug;275(2 Pt 1):C375-81
UI - 98349546
AU - Palmer S
AU - Kentish JC
TI - Roles of Ca2+ and crossbridge kinetics in determining the maximum rates
of Ca2+ activation and relaxation in rat and guinea pig skinned
trabeculae [see comments]
LA - Eng
MH - Actomyosin/*metabolism
MH - Animal
MH - Calcium/*pharmacology/physiology
MH - Comparative Study
MH - Diastole/drug effects/physiology
MH - Egtazic Acid/analogs & derivatives/metabolism/radiation effects
MH - Guinea Pigs
MH - Heart/*drug effects
MH - Ion Transport
MH - Kinetics
MH - Male
MH - Myocardial Contraction/*drug effects
MH - Myocardium/*metabolism
MH - Myofibrils/*drug effects
MH - Photolysis
MH - Rats
MH - Rats, Wistar
MH - Species Specificity
MH - Support, Non-U.S. Gov't
MH - Troponin C/*metabolism
RN - 0 (Troponin C)
RN - 0 (2-nitrophenyl-EGTA)
RN - 67-42-5 (Egtazic Acid)
RN - 7440-70-2 (Calcium)
RN - 9013-26-7 (Actomyosin)
PT - JOURNAL ARTICLE
DA - 19980811
DP - 1998 Jul 27
IS - 0009-7330
TA - Circ Res
PG - 179-86
SB - M
CY - UNITED STATES
IP - 2
VI - 83
JC - DAJ
AA - Author
EM - 199810
AB - We examined the influences of Ca2+ and crossbridge kinetics on the
maximum rate of force development during Ca2+ activation of cardiac
myofibrils and on the maximum rate of relaxation. Flash photolysis of
diazo-2 or nitrophenyl-EGTA was used to produce a sudden decrease or
increase, respectively, in [Ca2+] within Triton-skinned trabeculae from
rat and guinea pig hearts (22 degrees C). Trabeculae from both species
had similar Ca2+ sensitivities, suggesting that the rate of
dissociation of Ca2+ from troponin C (k(off)) is similar in the 2
species. However, the rate of relaxation after diazo-2 photolysis was 5
times faster in the rat (16.1 +/- 0.9 s(-1), mean +/- SEM, n = 11) than
in the guinea pig (2.99 +/- 0.26 s(-1), n = 7). This indicates that the
maximum relaxation rate is limited by crossbridge kinetics rather than
by k(off). The maximum rates of rapid activation by Ca2+ after
nitrophenyl-EGTA photolysis (k(act)) and of force redevelopment after
forcible crossbridge dissociation (k(act)) were similar and were
approximately 5-fold faster in rat (k(act)= 14.4 +/- 0.9 s(-1), k(tr)=
13.0 +/- 0.6 s(-1)) than in guinea pig (k(act)= 2.57 +/- 0.14 s(-1),
k(tr)= 2.69 +/- 0.30 s(-1)) trabeculae. This too may be mainly due to
species differences in crossbridge kinetics. Both k(act) and k(tr)
increased as [Ca2+] increased. This Ca2+ dependence of the rates of
force development is consistent with current models for the Ca2+
activation of the crossbridge cycle, but these models do not explain
the similarity in the maximal rates of activation and relaxation within
a given species.
AD - Department of Pharmacology, United Medical and Dental Schools, St.
Thomas's Hospital, London, UK.
CM - Comment in: Circ Res 1998 Jul 27;83(2):230-2
PMID- 0009686757
SO - Circ Res 1998 Jul 27;83(2):179-86
UI - 98319626
AU - Walker LA
AU - Gailly P
AU - Jensen PE
AU - Somlyo AV
AU - Somlyo AP
TI - The unimportance of being (protein kinase C) epsilon.
LA - Eng
MH - Animal
MH - Calcium/metabolism
MH - Down-Regulation (Physiology)
MH - Enzyme Inhibitors/pharmacology
MH - Ferrets
MH - Isoenzymes/metabolism/*physiology
MH - Male
MH - Muscle Contraction/drug effects/*physiology
MH - Peptides, Cyclic/pharmacology
MH - Phosphoprotein Phosphatase/antagonists & inhibitors
MH - Portal Vein/enzymology/metabolism
MH - Protein Kinase C/metabolism/*physiology
MH - Support, Non-U.S. Gov't
MH - Support, U.S. Gov't, P.H.S.
RN - EC 2.7.1.- (protein kinase C epsilon)
RN - EC 2.7.1.- (Protein Kinase C)
RN - EC 3.1.3.16 (Phosphoprotein Phosphatase)
RN - EC 3.1.3.53 (myosin-light-chain phosphatase)
RN - 0 (Enzyme Inhibitors)
RN - 0 (Isoenzymes)
RN - 0 (Peptides, Cyclic)
RN - 7440-70-2 (Calcium)
RN - 77238-39-2 (microcystin)
PT - JOURNAL ARTICLE
ID - PO1 HL48807/HL/NHLBI
DA - 19980730
DP - 1998 Jul
IS - 0892-6638
TA - FASEB J
PG - 813-21
SB - M
SB - X
CY - UNITED STATES
IP - 10
VI - 12
JC - FAS
AA - Author
EM - 199810
AB - The purpose of our study was to determine the mechanism through which
phorbol esters and smooth muscle myosin phosphatase inhibitors can
induce contraction of smooth muscle in the absence of Ca2+. Protein
kinase C-epsilon (PKC-epsilon) was previously implicated in this
process based largely on its supposed absence in the ferret portal
vein, and a correlation was drawn between the presence of this isoform
and the ability of smooth muscle to contract independently of Ca2+ and
phosphorylation of the 20 kDa regulatory light chains of myosin
(MLC20). We demonstrate here, with two antibodies, one to the NH2
terminus and the other to the COOH terminus of PKC-epsilon, that
epsilon is present in both ferret portal vein and rabbit portal vein
smooth muscle, neither of which exhibits phorbol ester-induced
contraction in the absence of Ca2+. However, in the presence of clamped
submaximal Ca2+, phorbol es ter increased MLC20 phosphorylation from
17.7+/-1.7% to 46.4+/-3.6% in ferret portal vein smooth muscle and
evoked an increase in force. Prolonged (48 h) incubation of ferret
portal vein with phorbol esters completely down-regulated PKC-epsilon,
as shown by Western blots, and abolished the phorbol ester-evoked
contraction at submaximal Ca2+, but not Ca2+-independent, contractions
induced by the phosphatase inhibitor microcystin. Contractions induced
by microcystin in Ca2+-free solution were associated with increased
phosphorylation of myosin light chain kinase (MLCK). Activation of MLCK
by autophosphorylation in the absence of Ca2+ occurs in vitro (1). We
conclude that PKC-epsilon is neither necessary nor sufficient for Ca2+-
independent regulation of myosin II in smooth muscle, but contractions
induced by agents that inhibit smooth muscle myosin phosphatase in the
absence of Ca2+ may be mediated by MLCK autophosphorylated or activated
by another Ca2+-independent kinase.
AD - Department of Molecular Physiology and Biological Physics, University
of Virginia, Health Sciences Center, Charlottesville 22906-0011, USA.
PMID- 0009657521
SO - FASEB J 1998 Jul;12(10):813-21
UI - 98297685
AU - Sabido-David C
AU - Brandmeier B
AU - Craik JS
AU - Corrie JE
AU - Trentham DR
AU - Irving M
TI - Steady-state fluorescence polarization studies of the orientation of
myosin regulatory light chains in single skeletal muscle fibers using
pure isomers of iodoacetamidotetramethylrhodamine.
LA - Eng
MH - Animal
MH - Chickens
MH - Chromatography, High Pressure Liquid
MH - Fluorescence Polarization/methods
MH - Fluorescent Dyes
MH - Gizzard
MH - In Vitro
MH - Muscle Contraction/*physiology
MH - Muscle Fibers/cytology/*physiology
MH - Muscle Relaxation
MH - Muscle, Skeletal/cytology/*physiology
MH - Muscle, Smooth/metabolism
MH - Myosin Light Chains/*analysis
MH - Rabbits
MH - *Rhodamines
MH - Sarcomeres/physiology/ultrastructure
MH - Sensitivity and Specificity
MH - Solutions
MH - Support, Non-U.S. Gov't
RN - 0 (Fluorescent Dyes)
RN - 0 (Myosin Light Chains)
RN - 0 (Rhodamines)
RN - 0 (Solutions)
RN - 81235-33-8 (tetramethylrhodamine iodoacetamide)
PT - JOURNAL ARTICLE
DA - 19980824
DP - 1998 Jun
IS - 0006-3495
TA - Biophys J
PG - 3083-92
SB - M
CY - UNITED STATES
IP - 6
VI - 74
JC - A5S
AA - Author
EM - 199810
AB - The regulatory light chain (RLC) from chicken gizzard myosin was
covalently modified on cysteine 108 with either the 5- or 6-isomer of
iodoacetamidotetramethylrhodamine (IATR). Labeled RLCs were purified by
fast protein liquid chromatography and characterized by reverse-phase
high-performance liquid chromatography (HPLC), tryptic digestion, and
electrospray mass spectrometry. Labeled RLCs were exchanged into the
native myosin heads of single skinned fibers from rabbit psoas muscle,
and the ATR dipole orientations were determined by fluorescence
polarization. The 5- and 6-ATR dipoles had distinct orientations, and
model orientational distributions suggest that they are more than 20
degrees apart in rigor. In the rigor-to-relaxed transition (sarcomere
length 2.4 microm, 10 degrees C), the 5-ATR dipole became more
perpendicular to the fiber axis, but the 6-ATR dipole became more
parallel. This orientation change was absent at sarcomere length 4.0
microm, where overlap between myosin and actin filaments is abolished.
When the temperature of relaxed fibers was raised to 30 degrees C, the
6-ATR dipoles became more parallel to the fiber axis and less ordered;
when ionic strength was lowered from 160 mM to 20 mM (5 degrees C), the
6-ATR dipoles became more perpendicular to the fiber axis and more
ordered. In active contraction (10 degrees C), the orientational
distribution of the probe dipoles was similar but not identical to that
in relaxation, and was not a linear combination of the orientational
distributions in relaxation and rigor.
AD - The Randall Institute, King's College London, England.
PMID- 0009635762
SO - Biophys J 1998 Jun;74(6):3083-92
UI - 98252441
AU - Dekker LR
AU - Rademaker H
AU - Vermeulen JT
AU - Opthof T
AU - Coronel R
AU - Spaan JA
AU - Janse MJ
TI - Cellular uncoupling during ischemia in hypertrophied and failing rabbit
ventricular myocardium: effects of preconditioning.
LA - Eng
MH - Animal
MH - Arrhythmia/etiology
MH - Calcium/metabolism
MH - Heart Failure, Congestive/*physiopathology
MH - Heart Hypertrophy/*physiopathology
MH - *Ischemic Preconditioning, Myocardial
MH - Myocardial Ischemia/*physiopathology
MH - Rabbits
MH - Time Factors
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
DA - 19980528
DP - 1998 May 5
IS - 0009-7322
TA - Circulation
PG - 1724-30
SB - A
SB - M
CY - UNITED STATES
IP - 17
VI - 97
JC - DAW
AA - Author
EM - 199807
AB - BACKGROUND: Patients with heart failure show a very high incidence of
arrhythmias and sudden death that is often preceded by ischemia;
however, data on electrophysiological changes during ischemia in
failing myocardium are sparse. We studied electrical uncoupling during
ischemia in normal and failing myocardium. METHODS AND RESULTS: Tissue
resistance, intracellular Ca2+ concentration (Indo-1 fluorescence
ratio), and mechanical activity were simultaneously determined in
arterially perfused right ventricular papillary muscles from 11 normal
and 15 failing rabbits. Heart failure was induced by combined volume
and pressure overload. Before sustained ischemia, muscles were
subjected to control perfusion (non-PC) or ischemic preconditioning
(PC). The onset of uncoupling during ischemia was equal in non-PC
normal (13.6+/-0.9 minutes of ischemia) and non-PC failing hearts
(13.3+/-0.7 minutes of ischemia). PC postponed uncoupling in normal
hearts by 10 minutes. In failing hearts, however, PC caused a large
variability in the onset of uncoupling during ischemia (mean, 12.2+/-
2.1; range, 5 to 22 minutes of ischemia). The duration of uncoupling
process was prolonged in failing hearts (12.9+/-0.9 minutes) compared
with normal hearts (7.8+/-0.4 minutes). The degree of heart failure and
relative heart weight of the failing hearts significantly correlated
with the earlier uncoupling after PC and the duration of uncoupling. In
every experiment, the start of Ca2+ rise and contracture preceded
uncoupling during ischemia. CONCLUSIONS: The duration of the process of
ischemia-induced electrical uncoupling in failing hearts is prolonged
compared with that in normal hearts. Ischemic PC has detrimental
effects in severely failing papillary muscles because it advances the
moment of irreversible ischemic damage.
AD - Department of Clinical and Experimental Cardiology, Academic Medical
Center, Amsterdam, The Netherlands. LRDekker@AMC.UVA.NL
PMID- 0009591767
SO - Circulation 1998 May 5;97(17):1724-30
UI - 98225225
AU - Wu X
AU - Haystead TA
AU - Nakamoto RK
AU - Somlyo AV
AU - Somlyo AP
TI - Acceleration of myosin light chain dephosphorylation and relaxation of
smooth muscle by telokin. Synergism with cyclic nucleotide-activated
kinase.
LA - Eng
MH - Animal
MH - Cyclic AMP-Dependent Protein Kinases/*metabolism
MH - Cyclic GMP/analogs & derivatives/pharmacology
MH - Cyclic GMP-Dependent Protein Kinases/*metabolism
MH - Forskolin/pharmacology
MH - In Vitro
MH - Kinetics
MH - Muscle Proteins/genetics/isolation & purification/*pharmacology
MH - Muscle Relaxation
MH - Muscle, Smooth/*drug effects/metabolism/physiology
MH - Myosin-Light-Chain Kinase/*metabolism
MH - Phosphorylation
MH - Rabbits
MH - Recombinant Proteins/genetics/isolation & purification/pharmacology
MH - Sulfhydryl Compounds/metabolism
MH - Support, U.S. Gov't, P.H.S.
RN - EC 2.7.1.117 (Myosin-Light-Chain Kinase)
RN - EC 2.7.10.- (Cyclic AMP-Dependent Protein Kinases)
RN - EC 2.7.10.- (Cyclic GMP-Dependent Protein Kinases)
RN - 0 (telokin)
RN - 0 (Muscle Proteins)
RN - 0 (Recombinant Proteins)
RN - 0 (Sulfhydryl Compounds)
RN - 31356-94-2 (8-bromocyclic GMP)
RN - 66428-89-5 (Forskolin)
RN - 7665-99-8 (Cyclic GMP)
PT - JOURNAL ARTICLE
ID - PO1-HL48807/HL/NHLBI
ID - PO1-HL19242/HL/NHLBI
DA - 19980602
DP - 1998 May 1
IS - 0021-9258
TA - J Biol Chem
PG - 11362-9
SB - M
SB - X
CY - UNITED STATES
IP - 18
VI - 273
JC - HIV
AA - Author
EM - 199808
AB - Incorporation of 32P into telokin, a smooth muscle-specific, 17-18-kDa,
acidic (pI 4.2-4.4) protein, was increased by forskolin (20 microM) in
intact rabbit ileum smooth muscle (ileum) and by 8-bromo-cyclic GMP
(100 microM) in alpha-toxin-permeabilized ileum. Native telokin (5-20
microM), purified from turkey gizzard, and recombinant rabbit telokin,
expressed in Escherichia coli and purified to >90% purity, induced dose-
dependent relaxation, associated with a significant decrease in
regulatory myosin light chain phosphorylation, without affecting the
rate of thiophosphorylation of regulatory myosin light chain of ileum
permeabilized with 0.1% Triton X-100. Endogenous telokin was lost from
ileum during prolonged permeabilization (>20 min) with 0.1% Triton X-
100, and the time course of loss was correlated with the loss of 8-
bromo-cyclic GMP-induced calcium desensitization. Recombinant and
native gizzard telokins were phosphorylated, in vitro, by the catalytic
subunit of cAMP-dependent protein kinase, cGMP-dependent protein
kinase, and p42/44 mitogen-activated protein kinase; the recombinant
protein was also phosphorylated by calmodulin-dependent protein kinase
II. Exogenous cGMP-dependent protein kinase (0.5 microM) activated by 8-
bromo-cyclic GMP (50 microM) phosphorylated recombinant telokin (10
microM) when added concurrently to ileum depleted of its endogenous
telokin, and their relaxant effects were mutually potentiated.
Forskolin (20 microM) also increased phosphorylation of telokin in
intact ileum. We conclude that telokin induces calcium desensitization
in smooth muscle by enhancing myosin light chain phosphatase activity,
and cGMP- and/or cAMP-dependent phosphorylation of telokin up-regulates
its relaxant effect.
AD - Department of Molecular Physiology and Biological Physics, University
of Virginia Health Sciences Center, Charlottesville, Virginia 22906-
0011, USA.
PMID- 0009556631
SO - J Biol Chem 1998 May 1;273(18):11362-9
UI - 98260334
AU - Ohizumi Y
AU - Matsunaga K
AU - Nakatani K
AU - Kobayashi J
TI - Potent stimulation of myofilament force and ATPase activity of skeletal
muscle by eudistomin M, a novel Ca(++)-sensitizing agent from a
Caribbean tunicate.
LA - Eng
MH - Adenosinetriphosphatase/*metabolism
MH - Animal
MH - Calcium/*metabolism
MH - Carbolines/pharmacology
MH - Guinea Pigs
MH - Male
MH - Microfilaments/*drug effects
MH - Muscle, Skeletal/*drug effects
MH - Myosin/analysis
MH - Rabbits
MH - Support, Non-U.S. Gov't
RN - EC 3.6.1.3 (Adenosinetriphosphatase)
RN - 0 (Carbolines)
RN - 0 (Myosin)
RN - 7440-70-2 (Calcium)
RN - 88704-39-6 (eudistomin M)
PT - JOURNAL ARTICLE
DA - 19980608
DP - 1998 May
IS - 0022-3565
TA - J Pharmacol Exp Ther
PG - 695-9
SB - M
CY - UNITED STATES
IP - 2
VI - 285
JC - JP3
AA - Author
EM - 199808
AB - In the course of our survey of biologically active compounds from
natural sources, eudistomins were isolated from a Caribbean tunicate
Eudistoma olivaceum. In the present experiments, eudistomin M (Eud-M, >
10(-5) M) caused a concentration-dependent increase in the contractile
response of skinned fibers from guinea pig skeletal psoas muscles to
Ca++. The superprecipitation and ATPase activity of myosin B from fast
skeletal muscles of rabbit back and leg were potentiated by this
compound (> 10(-5) M) in a concentration-dependent manner. In skinned
fibers, superprecipitation and the ATPase activity of myosin B, Eud-M
shifted the concentration-response curve for Ca++ to the upper
direction. Ca(++)-, K(+)-EDTA- or Mg(++)-ATPase was not affected by Eud-
M. This compound had no effect on the ATPase activity of actomyosin
reconstituted from actin and myosin in the presence or absence of
troponin. However, the ATPase activity of actin-myosin-troponin-
tropomyosin reconstituted system was increased significantly by Eud-M.
These results suggest that Eud-M increases the Ca++ sensitivity of the
contractile apparatus in skeletal muscles at least partially mediated
through troponin-tropomyosin system and thus enhances the ATPase
activity of myosin B and the contractile force of myofilament.
AD - Department of Pharmaceutical Molecular Biology, Faculty of
Pharmaceutical Sciences, Tohoku University, Sendai, Japan.
PMID- 0009580615
SO - J Pharmacol Exp Ther 1998 May;285(2):695-9
UI - 98204620
AU - Regnier M
AU - Martyn DA
AU - Chase PB
TI - Calcium regulation of tension redevelopment kinetics with 2-deoxy-ATP
or low [ATP] in rabbit skeletal muscle.
LA - Eng
MH - Adenosine Triphosphate/*pharmacology
MH - Animal
MH - Biophysics
MH - Calcium/*pharmacology
MH - Deoxyadenine Nucleotides/*pharmacology
MH - In Vitro
MH - Kinetics
MH - Models, Biological
MH - Muscle Contraction/*drug effects/physiology
MH - Myosin ATPase/metabolism
MH - Psoas Muscles/*drug effects/*physiology
MH - Rabbits
MH - Support, U.S. Gov't, P.H.S.
RN - EC 3.6.1.32 (Myosin ATPase)
RN - 0 (Deoxyadenine Nucleotides)
RN - 1927-31-7 (2'-deoxyadenosine triphosphate)
RN - 56-65-5 (Adenosine Triphosphate)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
ID - HL52558/HL/NHLBI
ID - HL51277/HL/NHLBI
ID - NS08384/NS/NINDS
DA - 19980608
DP - 1998 Apr
IS - 0006-3495
TA - Biophys J
PG - 2005-15
SB - M
CY - UNITED STATES
IP - 4
VI - 74
JC - A5S
AA - Author
EM - 199808
AB - The correlation of acto-myosin ATPase rate with tension redevelopment
kinetics (k(tr)) was determined during Ca(+2)-activated contractions of
demembranated rabbit psoas muscle fibers; the ATPase rate was either
increased or decreased relative to control by substitution of ATP (5.0
mM) with 2-deoxy-ATP (dATP) (5.0 mM) or by lowering [ATP] to 0.5 mM,
respectively. The activation dependence of k(tr) and unloaded
shortening velocity (Vu) was measured with each substrate. With 5.0 mM
ATP, Vu depended linearly on tension (P), whereas k(tr) exhibited a
nonlinear dependence on P, being relatively independent of P at
submaximum levels and rising steeply at P > 0.6-0.7 of maximum tension
(Po). With dATP, Vu was 25% greater than control at Po and was elevated
at all P > 0.15Po, whereas Po was unchanged. Furthermore, the Ca(+2)
sensitivity of both k(tr) and P increased, such that the dependence of
k(tr) on P was not significantly different from control, despite an
elevation of Vu and maximal k(tr). In contrast, lowering [ATP] caused a
slight (8%) elevation of Po, no change in the Ca(+2) sensitivity of P,
and a decrease in Vu at all P. Moreover, k(tr) was decreased relative
to control at P > 0.75Po, but was elevated at P < 0.75Po. These data
demonstrate that the cross-bridge cycling rate dominates k(tr) at
maximum but not submaximum levels of Ca(2+) activation.
AD - Department of Bioengineering, University of Washington, Seattle 98195,
USA. mregnier@u.washington.edu
PMID- 0009545059
SO - Biophys J 1998 Apr;74(4):2005-15
UI - 98204590
AU - Uttenweiler D
AU - Weber C
AU - Fink RH
TI - Mathematical modeling and fluorescence imaging to study the Ca2+
turnover in skinned muscle fibers.
LA - Eng
MH - Animal
MH - Biophysics
MH - Caffeine/pharmacology
MH - Calcium/*metabolism
MH - Comparative Study
MH - Fluorescent Dyes
MH - Fura-2
MH - In Vitro
MH - Ion Transport/drug effects
MH - Kinetics
MH - Male
MH - Mathematics
MH - Mice
MH - Mice, Inbred BALB C
MH - *Models, Biological
MH - Muscle Fibers, Fast-Twitch/drug effects/*metabolism
MH - Sarcoplasmic Reticulum/metabolism
MH - Support, Non-U.S. Gov't
RN - 0 (Fluorescent Dyes)
RN - 58-08-2 (Caffeine)
RN - 7440-70-2 (Calcium)
RN - 96314-98-6 (Fura-2)
PT - JOURNAL ARTICLE
DA - 19980608
DP - 1998 Apr
IS - 0006-3495
TA - Biophys J
PG - 1640-53
SB - M
CY - UNITED STATES
IP - 4
VI - 74
JC - A5S
AA - Author
EM - 199808
AB - A mathematical model was developed for the simulation of the spatial
and temporal time course of Ca2+ ion movement in caffeine-induced
calcium transients of chemically skinned muscle fiber preparations. Our
model assumes cylindrical symmetry and quantifies the radial profile of
Ca2+ ion concentration by solving the diffusion equations for Ca2+ ions
and various mobile buffers, and the rate equations for Ca2+ buffering
(mobile and immobile buffers) and for the release and reuptake of Ca2+
ions by the sarcoplasmic reticulum (SR), with a finite-difference
algorithm. The results of the model are compared with caffeine-induced
spatial Ca2+ transients obtained from saponin skinned murine fast-
twitch fibers by fluorescence photometry and imaging measurements using
the ratiometric dye Fura-2. The combination of mathematical modeling
and digital image analysis provides a tool for the quantitative
description of the total Ca2+ turnover and the different contributions
of all interacting processes to the overall Ca2+ transient in skinned
muscle fibers. It should thereby strongly improve the usage of skinned
fibers as quantitative assay systems for many parameters of the SR and
the contractile apparatus helping also to bridge the gap to the intact
muscle fiber.
AD - Ruprecht-Karls-Universitat Heidelberg, II Institute of Physiology,
Germany.
PMID- 0009545029
SO - Biophys J 1998 Apr;74(4):1640-53
UI - 98151502
AU - Sugi H
AU - Iwamoto H
AU - Akimoto T
AU - Ushitani H
TI - Evidence for the load-dependent mechanical efficiency of individual
myosin heads in skeletal muscle fibers activated by laser flash
photolysis of caged calcium in the presence of a limited amount of ATP.
LA - Eng
MH - Acetic Acids/*metabolism
MH - Adenosine Diphosphate/metabolism
MH - Adenosine Triphosphate/*metabolism
MH - Animal
MH - Calcium/*metabolism
MH - Chelating Agents
MH - Ethylenediamines/*metabolism
MH - Kinetics
MH - Lasers
MH - Muscle Contraction/*physiology
MH - Muscle Fibers/*physiology/ultrastructure
MH - Muscle, Skeletal/*physiology
MH - Phosphates/metabolism
MH - Photolysis
MH - Rabbits
MH - Solutions
MH - Time Factors
RN - 0 (Acetic Acids)
RN - 0 (Chelating Agents)
RN - 0 (Ethylenediamines)
RN - 0 (Phosphates)
RN - 0 (Solutions)
RN - 117367-86-9 (1-(2-nitro-4,5-dimethoxyphenyl)-N,N,N',N'-
tetrakis((oxycarbonyl)methyl)-1,2-ethanediamine)
RN - 56-65-5 (Adenosine Triphosphate)
RN - 58-64-0 (Adenosine Diphosphate)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
DA - 19980409
DP - 1998 Mar 3
IS - 0027-8424
TA - Proc Natl Acad Sci U S A
PG - 2273-8
SB - M
SB - X
CY - UNITED STATES
IP - 5
VI - 95
JC - PV3
AA - Author
EM - 199806
AB - Although a contracting muscle regulates its energy output depending on
the load imposed on it ("Fenn effect"), the mechanism underlying the
load-dependent energy output remains obscure. To explore the
possibility that the mechanical efficiency, with which chemical energy
derived from ATP hydrolysis is converted into mechanical work, of
individual myosin heads changes in a load-dependent manner, we examined
the auxotonic shortening of glycerinated rabbit psoas muscle fibers,
containing ATP molecules almost equal in number to the myosin heads,
after laser-flash photolysis of caged calcium. Immediately before laser-
flash activation, almost all of the myosin heads in the fiber are in
the state M.ADP.Pi, and can undergo only one ATP hydrolysis cycle after
activation. When the fibers were activated to shorten under various
auxotonic loads, the length, force, and power output changes were found
to be scaled according to the auxotonic load. Both the power and energy
outputs were maximal under a moderate auxotonic load. The amount of
M.ADP.Pi utilized at a time after activation was estimated from the
amount of isometric force developed after interruption of fiber
shortening. This amount was minimal in the isometric condition and
increased nearly in proportion to the distance of fiber shortening.
These results are taken as evidence that the efficiency of
chemomechanical energy conversion in individual myosin heads changes in
a load-dependent manner.
AD - Department of Physiology, School of Medicine, Teikyo University,
Itabashi-ku, Tokyo 173, Japan. sugi@med.teikyo-u.ac.jp
PMID- 0009482875
SO - Proc Natl Acad Sci U S A 1998 Mar 3;95(5):2273-8
UI - 98171004
AU - Horikawa Y
AU - Goel A
AU - Somlyo AP
AU - Somlyo AV
TI - Mitochondrial calcium in relaxed and tetanized myocardium.
LA - Eng
MH - Animal
MH - Calcium/*metabolism/pharmacology
MH - Electric Stimulation
MH - Electron Probe Microanalysis/methods
MH - Freezing
MH - Hamsters
MH - Heart Ventricle
MH - In Vitro
MH - Kinetics
MH - Male
MH - Mitochondria, Heart/*metabolism/ultrastructure
MH - Myocardial Contraction/drug effects/*physiology
MH - Papillary Muscles/drug effects/*physiology/ultrastructure
MH - Rats
MH - Rats, Sprague-Dawley
MH - Ryanodine/pharmacology
MH - Support, U.S. Gov't, P.H.S.
RN - 15662-33-6 (Ryanodine)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
ID - HL-48807/HL/NHLBI
DA - 19980512
DP - 1998 Mar
IS - 0006-3495
TA - Biophys J
PG - 1579-90
SB - M
CY - UNITED STATES
IP - 3
VI - 74
JC - A5S
AA - Author
EM - 199807
AB - The elemental composition of rat cardiac muscle was determined with
electron probe x-ray microanalysis (EPMA) of rapidly frozen papillary
muscles and trabeculae incubated with ryanodine (1 microM) in either
1.2 or 10 mM [Ca2+]o-containing solutions, paced at 0.6 Hz or tetanized
at 10 Hz. Total mitochondrial calcium increased significantly, by 4.2
mmol/kg dry weight during a 7 s tetanus, only in muscles tetanized in
the presence of 10 mM [Ca2+]o when cytoplasmic Ca2+ is 1-4 microM
(Backx, P. H., W.-D. Gao, M. D. Azan-Backx, and E. Marban. 1995. The
relationship between contractile force and intracellular [Ca2+] in
intact rat trabeculae. J. Gen. Physiol. 105:1-19). Comparison of total
mitochondrial with free mitochondrial Ca2+ reported in the literature
indicates that the total/free ratio is approximately 6000 at
physiological or near-physiological levels of total mitochondrial
calcium. Increases in free mitochondrial [Ca2+] consistent with
regulation of mitochondrial enzymes should be associated with increases
in total mitochondrial calcium detectable with EPMA. However, such
increases in mitochondrial calcium occur only as the result of
prolonged, unphysiological elevations of cytosolic [Ca2+].
AD - Department of Molecular Physiology and Biological Physics, University
of Virginia Health Sciences Center, Charlottesville 22906-0011, USA.
PMID- 0009512053
SO - Biophys J 1998 Mar;74(3):1579-90
UI - 98170992
AU - Iwamoto H
TI - Thin filament cooperativity as a major determinant of shortening
velocity in skeletal muscle fibers.
LA - Eng
MH - Animal
MH - Calcium/metabolism
MH - In Vitro
MH - Isometric Contraction/drug effects/*physiology
MH - Muscle Fibers/drug effects/*physiology/ultrastructure
MH - Muscle, Skeletal/*physiology/ultrastructure
MH - Phosphates/pharmacology
MH - Rabbits
MH - Sarcomeres/drug effects/*physiology/ultrastructure
MH - Support, Non-U.S. Gov't
MH - Time Factors
RN - 0 (Phosphates)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
DA - 19980512
DP - 1998 Mar
IS - 0006-3495
TA - Biophys J
PG - 1452-64
SB - M
CY - UNITED STATES
IP - 3
VI - 74
JC - A5S
AA - Author
EM - 199807
AB - The mechanism underlying the calcium sensitivity of the velocity of
shortening of skeletal muscle fibers was investigated using a multiple
shortening protocol: within a single contraction, skinned rabbit psoas
fibers were made to shorten repetitively under a light load by briefly
stretching back to their initial length at regular intervals. At
saturating [Ca2+], the initial fast shortening pattern was repeated
reproducibly. At submaximal [Ca2+], the first shortening consisted of
fast and slow phases, but only the slow phase was observed in later
shortenings. When the fibers were held isometric after the first
shortening, the velocity of the second shortening recovered with time.
The recovery paralleled tension redevelopment, implying a close
relationship between the velocity and the number of the preexisting
force-producing cross-bridges. However, this parallelism was lost as
[Ca2+] was increased. Thus, the velocity was modified in a manner
consistent with the cooperative thin filament activation by strong
binding cross-bridges and its modulation by calcium. The present
results therefore provide evidence that the thin filament cooperativity
is primarily responsible for the calcium sensitivity of velocity. The
effect of inorganic phosphate to accelerate the slow phase of
shortening is also explained in terms of the cooperative activation.
AD - Department of Physiology, School of Medicine, Teikyo University, Tokyo,
Japan.
PMID- 0009512041
SO - Biophys J 1998 Mar;74(3):1452-64
UI - 98147195
AU - Dijkman MA
AU - Heslinga JW
AU - Sipkema P
AU - Westerhof N
TI - Perfusion-induced changes in cardiac contractility depend on capillary
perfusion.
LA - Eng
MH - Animal
MH - Blood Pressure
MH - Capillaries/*physiology
MH - Coronary Vessels/physiology
MH - Microspheres
MH - *Myocardial Contraction
MH - Oxygen Consumption
MH - Papillary Muscles/anatomy & histology/*physiology
MH - Perfusion
MH - Rats
MH - Rats, Wistar
PT - JOURNAL ARTICLE
DA - 19980317
DP - 1998 Feb
IS - 0002-9513
TA - Am J Physiol
PG - H405-10
SB - M
CY - UNITED STATES
IP - 2 Pt 2
VI - 274
JC - 3U8
AA - Author
EM - 199805
AB - The perfusion-induced increase in cardiac contractility (Gregg
phenomenon) is especially found in heart preparations that lack
adequate coronary autoregulation and thus protection of changes in
capillary pressure. We determined in the isolated perfused papillary
muscle of the rat whether cardiac muscle contractility is related to
capillary perfusion. Oxygen availability of this muscle is independent
of internal perfusion, and perfusion may be varied or even stopped
without loss of function. Muscles contracted isometrically at 27
degrees C (n = 7). During the control state stepwise increases in
perfusion pressure resulted in all muscles in a significant increase in
active tension. Muscle diameter always increased with increased
perfusion pressure, but muscle segment length was unaffected. Capillary
perfusion was then obstructed by plastic microspheres (15 microns).
Flow, at a perfusion pressure of 66.6 +/- 26.2 cmH2O, reduced from 17.6
+/- 5.4 microliters/min in the control state to 3.2 +/- 1.3
microliters/min after microspheres. Active tension developed by the
muscle in the unperfused condition before microspheres and after
microspheres did not differ significantly (-12.8 +/- 29.4% change).
After microspheres similar perfusion pressure steps as in control never
resulted in an increase in active tension. Even at the two highest
perfusion pressures (89.1 +/- 28.4 and 106.5 +/- 31.7 cmH2O) that were
applied a significant decrease in active tension was found. We conclude
that the Gregg phenomenon is related to capillary perfusion.
AD - Laboratory for Physiology, Vrije Universiteit Amsterdam, The
Netherlands.
PMID- 0009486241
SO - Am J Physiol 1998 Feb;274(2 Pt 2):H405-10
UI - 98062941
AU - Blanchard EM
AU - Iizuka K
AU - Christe M
AU - Conner DA
AU - Geisterfer-Lowrance A
AU - Schoen FJ
AU - Maughan DW
AU - Seidman CE
AU - Seidman JG
TI - Targeted ablation of the murine alpha-tropomyosin gene.
LA - Eng
MH - Animal
MH - Gene Targeting
MH - Male
MH - Mice
MH - Mice, Knockout
MH - Myocardium/pathology
MH - Support, Non-U.S. Gov't
MH - Tropomyosin/*genetics/physiology
RN - 0 (Tropomyosin)
PT - JOURNAL ARTICLE
DA - 19971231
DP - 1997 Dec
IS - 0009-7330
TA - Circ Res
PG - 1005-10
SB - M
CY - UNITED STATES
IP - 6
VI - 81
JC - DAJ
AA - Author
EM - 199803
AB - We created a mouse that lacks a functional alpha-tropomyosin gene using
gene targeting in embryonic stem cells and blastocyst-mediated
transgenesis. Homozygous alpha-tropomyosin "knockout" mice die between
embryonic day 9.5 and 13.5 and lack alpha-tropomyosin mRNA.
Heterozygous alpha-tropomyosin knockout mice have approximately 50% as
much cardiac alpha-tropomyosin mRNA as wild-type littermates but
similar alpha-tropomyosin protein levels. Cardiac gross morphology,
histology, and function (assessed by working heart preparations) of
heterozygous alpha-tropomyosin knockout and wild-type mice were
indistinguishable. Mechanical performance of skinned papillary muscle
strips derived from mutant and wild-type hearts also revealed no
differences. We conclude that haploinsufficiency of the alpha-
tropomyosin gene produces little or no change in cardiac function or
structure, whereas total alpha-tropomyosin deficiency is incompatible
with life. These findings imply that in heterozygotes there is a
regulatory mechanism that maintains the level of myofibrillar
tropomyosin despite the reduction in alpha-tropomyosin mRNA.
AD - Howard Hughes Medical Institute, Boston, Mass., USA.
PMID- 0009400381
SO - Circ Res 1997 Dec;81(6):1005-10
UI - 98042194
AU - Wannenburg T
AU - Janssen PM
AU - Fan D
AU - de Tombe PP
TI - The Frank-Starling mechanism is not mediated by changes in rate of
cross-bridge detachment.
LA - Eng
MH - Adenosine Triphosphate/*metabolism
MH - Animal
MH - Calcium/pharmacology
MH - Heart/*physiology
MH - In Vitro
MH - Kinetics
MH - *Models, Cardiovascular
MH - Myocardial Contraction/drug effects/*physiology
MH - Myocardium/metabolism
MH - Myosin ATPase/*metabolism
MH - NAD/metabolism
MH - Rats
MH - Regression Analysis
MH - Sarcomeres/physiology
MH - Support, Non-U.S. Gov't
MH - Support, U.S. Gov't, P.H.S.
RN - EC 3.6.1.32 (Myosin ATPase)
RN - 53-84-9 (NAD)
RN - 56-65-5 (Adenosine Triphosphate)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
ID - HL-52322/HL/NHLBI
ID - HL-03255/HL/NHLBI
DA - 19971216
DP - 1997 Nov
IS - 0002-9513
TA - Am J Physiol
PG - H2428-35
SB - M
CY - UNITED STATES
IP - 5 Pt 2
VI - 273
JC - 3U8
AA - Author
EM - 199802
AB - We tested the hypothesis that the Frank-Starling relationship is
mediated by changes in the rate of cross-bridge detachment in cardiac
muscle. We simultaneously measured isometric force development and the
rate of ATP consumption at various levels of Ca2+ activation in skinned
rat cardiac trabecular muscles at three sarcomere lengths (2.0, 2.1,
and 2.2 microns). The maximum rate of ATP consumption was 1.5 nmol.s-
1.microliter fiber vol-1, which represents an estimated
adenosinetriphosphatase (ATPase) rate of approximately 10 s-1 per
myosin head at 24 degrees C. The rate of ATP consumption was tightly
and linearly coupled to the level of isometric force development, and
changes in sarcomere length had no effect on the slope of the force-
ATPase relationships. The average slope of the force-ATPase
relationships was 15.5 pmol.mN-1.mm-1. These results suggest that the
mechanisms that underlie the Frank-Starling relationship in cardiac
muscle do not involve changes in the kinetics of the apparent
detachment step in the cross-bridge cycle.
AD - Section on Cardiology, Bowman Gray School of Medicine, Wake Forest
University, Winston-Salem, North Carolina 27157-1045, USA.
PMID- 0009374781
SO - Am J Physiol 1997 Nov;273(5 Pt 2):H2428-35
UI - 98042192
AU - Janssen PM
AU - de Tombe PP
TI - Protein kinase A does not alter unloaded velocity of sarcomere
shortening in skinned rat cardiac trabeculae.
LA - Eng
MH - Animal
MH - Calcium/metabolism
MH - Connective Tissue/*physiology
MH - Cyclic AMP-Dependent Protein Kinases/*pharmacology
MH - Enzyme Inhibitors/pharmacology
MH - Heart/*physiology
MH - In Vitro
MH - Kinetics
MH - Myocardial Contraction
MH - Rats
MH - Rats, Inbred Strains
MH - Receptors, Adrenergic, beta/physiology
MH - Regression Analysis
MH - Sarcomeres/drug effects/*physiology
MH - Support, Non-U.S. Gov't
MH - Support, U.S. Gov't, P.H.S.
MH - Time Factors
RN - EC 2.7.10.- (Cyclic AMP-Dependent Protein Kinases)
RN - 0 (Enzyme Inhibitors)
RN - 0 (Receptors, Adrenergic, beta)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
ID - HL-52322/HL/NHLBI
DA - 19971216
DP - 1997 Nov
IS - 0002-9513
TA - Am J Physiol
PG - H2415-22
SB - M
CY - UNITED STATES
IP - 5 Pt 2
VI - 273
JC - 3U8
AA - Author
EM - 199802
AB - Whether beta-adrenergic stimulation affects the cross-bridge cycling
rate independently of its effect on Ca2+ handling by the cardiac
myocyte is still unknown. An increase in cross-bridge cycling rate may
result in increased unloaded velocity of sarcomere shortening (V0). To
test this hypothesis directly, skinned rat cardiac trabeculae were
attached between a silicon strain gauge (approximately 3.5 kHz resonant
frequency) and a fast displacement motor. V0 was measured by a modified
"Edman slack test" during a single maximal activation using seven to
eight sarcomere-length step releases (measured by laser diffraction)
ranging between 0.12 and 0.20 micron (15.0 +/- 0.1 degrees C). beta-
Adrenergic stimulation was mimicked by exposing the trabeculae to the
catalytic subunit of protein kinase A (PKA). Treatment with PKA (3
micrograms/ml; 45 min) caused a significant (P < 0.01) increase (41 +/-
13%) in the Ca2+ concentration required for half-maximal steady-state
tension development. Neither maximum tension nor V0 was affected by
treatment with PKA, suggesting that beta-adrenergic stimulation does
not affect the rate-limiting step of cross-bridge cycling during
unloaded shortening in myocardium.
AD - Department of Physiology and Biophysics, University of Illinois at
Chicago 60607-7171, USA.
PMID- 0009374779
SO - Am J Physiol 1997 Nov;273(5 Pt 2):H2415-22
UI - 97460444
AU - Wier WG
AU - ter Keurs HE
AU - Marban E
AU - Gao WD
AU - Balke CW
TI - Ca2+ 'sparks' and waves in intact ventricular muscle resolved by
confocal imaging [see comments] [published erratum appears in Circ Res
1997 Nov;81(5):893]
LA - Eng
MH - Aniline Compounds
MH - Animal
MH - Calcium/*metabolism
MH - Fluorescent Dyes
MH - Heart Ventricle
MH - Microscopy, Confocal
MH - Myocardial Contraction
MH - Myocardium/*metabolism
MH - Rats
MH - Rats, Inbred BN
MH - Support, Non-U.S. Gov't
MH - Support, U.S. Gov't, P.H.S.
MH - Ventricular Function
MH - Xanthenes
RN - 0 (Aniline Compounds)
RN - 0 (Fluorescent Dyes)
RN - 0 (Xanthenes)
RN - 121714-13-4 (Fluo-3)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
ID - HL-02466/HL/NHLBI
ID - HL-29473/HL/NHLBI
ID - HL-50435/HL/NHLBI
ID - +
DA - 19971030
DP - 1997 Oct
IS - 0009-7330
TA - Circ Res
PG - 462-9
SB - M
CY - UNITED STATES
IP - 4
VI - 81
JC - DAJ
AA - Author
EM - 199801
AB - The [Ca2+]i transient in heart is now thought to involve the
recruitment and summation of discrete and independent "units" of Ca2+
release (Ca2+ "sparks") from the sarcoplasmic reticulum, each of which
is controlled locally by single coassociated L-type Ca2+ channels
("local control theory of excitation-contraction coupling"). All prior
studies on Ca2+ sparks, however, have been performed in single
enzymatically dissociated heart cells under nonphysiological
conditions. In order to understand the possible significance of Ca2+
sparks to normal working cardiac muscle, we used confocal microscopy to
record Ca2+ sparks, spatially averaged [Ca2+]i transients and Ca2+
waves in individual cells of intact rat right ventricular trabeculae
(composed of < 15 cells in cross section) microinjected with the Ca2+
indicator fluo 3 under physiological conditions ([Ca2+]o, 1 mmol/L;
temperature, 33 +/- 1 degree C). Twitch force was recorded
simultaneously. When stretched to optimal length (sarcomere length, 2.2
microns) and stimulated at 0.2 Hz, the trabeculae generated
approximately equal to 700 micrograms of force per cell. Spatially
averaged [Ca2+]i transients recorded from individual cells within a
trabecula were similar to those recorded previously from single cells.
The amplitude distribution of the peak ratio of Ca2+ sparks was
bimodal, with maxima at ratios of 1.8 +/- 0.3 and 2.7 +/- 0.2 (mean +/-
SD), respectively. The amplitude of the peak of Ca2+ sparks was
approximately equal to 170 nmol/L. Ca2+ sparks occurred at a frequency
of 12.0 +/- 0.8/s (mean +/- SEM) in line scans covering 94 sarcomeres.
Ca2+ waves occurred randomly at a frequency of 0.57 +/- 0.08/s and
propagated with a velocity of 29.5 +/- 1.7 microns/s. The extent of
Ca2+ wave propagation was 3.9 +/- 0.3 sarcomere lengths (sarcomere
length, 2.2 microns). Ca2+ sparks could be identified along the leading
edge of the waves at intervals of 1.30 +/- 0.11 sarcomere length. Our
observations suggest that (1) Ca2+ sparks, similar to those recorded in
single cells, occur in trabeculae under physiological conditions and
(2) coupling of Ca2+ spark generation between neighboring sites occurs
and may lead to (3) the development of Ca2+ waves, which propagate
under physiological conditions at a low velocity over limited
distances. The results suggest that concepts of excitation-contraction
coupling recently derived from isolated myocytes are applicable to
intact cardiac trabeculae [corrected].
AD - Department of Physiology, University of Maryland School of Medicine,
Baltimore 21201, USA. gwier001@umabnet.ab.umd.edu
CM - Comment in: Circ Res 1997 Oct;81(4):636-8
RO - M:MWS
PMID- 0009314826
LR - 19980402
SO - Circ Res 1997 Oct;81(4):462-9
UI - 97276757
AU - Shah AM
AU - Mebazaa A
AU - Yang ZK
AU - Cuda G
AU - Lankford EB
AU - Pepper CB
AU - Sollott SJ
AU - Sellers JR
AU - Robotham JL
AU - Lakatta EG
TI - Inhibition of myocardial crossbridge cycling by hypoxic endothelial
cells: a potential mechanism for matching oxygen supply and demand?
LA - Eng
MH - Adenosinetriphosphatase/metabolism
MH - Animal
MH - Calcium/metabolism
MH - *Cell Hypoxia
MH - Cells, Cultured
MH - Comparative Study
MH - Data Interpretation, Statistical
MH - Endothelium, Vascular/*cytology/*physiology
MH - Human
MH - In Vitro
MH - Microfilaments/metabolism/physiology
MH - *Myocardial Contraction
MH - Myocardial Stunning/metabolism/physiopathology
MH - Myocardium/cytology/*metabolism
MH - Myosin/metabolism
MH - Oxygen/*metabolism
MH - Sheep
MH - Signal Transduction/physiology
MH - Support, Non-U.S. Gov't
MH - Support, U.S. Gov't, P.H.S.
MH - Swine
RN - EC 3.6.1.3 (Adenosinetriphosphatase)
RN - 0 (Myosin)
RN - 7440-70-2 (Calcium)
RN - 7782-44-7 (Oxygen)
PT - JOURNAL ARTICLE
ID - RO-1 HL-39138-03/HL/NHLBI
DA - 19970516
DP - 1997 May
IS - 0009-7330
TA - Circ Res
PG - 688-98
SB - M
CY - UNITED STATES
IP - 5
VI - 80
JC - DAJ
AA - Author
EM - 199707
AB - Previous studies have shown that cardiac endothelial cells release
substances that influence myocardial contraction. Since PO2 is an
important stimulus that modulates endothelial function, we investigated
the effects of acute moderate hypoxia and reoxygenation on the release
of cardioactive factors by endothelial cells. Endothelial cells
cultured from several vascular beds were superfused with normoxic
(equilibrated with room air; PO2, approximately 160 mm Hg) or hypoxic
(PO2, 40 to 50 mm Hg) physiological buffer solution, and the
superfusates were reequilibrated to a PO2 of approximately 160 mm Hg
and then tested for their effects on various myocardial assays.
Endothelial cell viability and buffer ionic composition were unaltered
after the superfusion procedures. The superfusates of hypoxic
endothelial cells induced rapid, potent, reversible inhibition of
isolated cardiac myocyte contraction without reducing cytosolic Ca2+
transients. This activity was not lost after heating (95 degrees C) and
was present in low molecular weight (Mr, <500) superfusate fractions.
Hypoxic endothelial superfusate reduced unloaded shortening velocity of
human skinned soleus muscle fibers. It markedly depressed in vitro
actin motility over cardiac myosin and reduced the rate of actin-
activated cardiac myosin ATPase activity but had no effect on
corresponding smooth muscle myosin assays. Reoxygenation of hypoxic
endothelial cells resulted in loss of this inhibitory activity. These
data indicate that cultured endothelial cells respond to acute moderate
hypoxia by releasing an unidentified substance(s) that inhibits
myocardial crossbridge cycling, independent of Ca2+ or other second
messenger signaling pathways. Such a mechanism could have important
implications for the regulation of oxygen supply-demand balance in the
heart and be relevant to conditions such as myocardial hibernation.
AD - Department of Cardiology, University of Wales College of Medicine,
Cardiff, UK. shaham2@cf.ac.uk
PMID- 0009130450
SO - Circ Res 1997 May;80(5):688-98
UI - 97276756
AU - Palmer S
AU - Kentish JC
TI - Differential effects of the Ca2+ sensitizers caffeine and CGP 48506 on
the relaxation rate of rat skinned cardiac trabeculae.
LA - Eng
MH - Animal
MH - Azocines/*pharmacology
MH - Caffeine/*pharmacology
MH - Calcium/*metabolism
MH - Cardiotonic Agents/*pharmacology
MH - Chelating Agents/diagnostic use
MH - Comparative Study
MH - Heart/*drug effects
MH - In Vitro
MH - Male
MH - Myocardial Contraction/*drug effects
MH - Myofibrils/drug effects
MH - Photolysis
MH - Rats
MH - Rats, Wistar
MH - Software
MH - Statistics
MH - Support, Non-U.S. Gov't
RN - 0 (Azocines)
RN - 0 (Cardiotonic Agents)
RN - 0 (Chelating Agents)
RN - 124029-65-8 (diazo-2)
RN - 166020-57-1 (BA 41899)
RN - 58-08-2 (Caffeine)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
DA - 19970516
DP - 1997 May
IS - 0009-7330
TA - Circ Res
PG - 682-7
SB - M
CY - UNITED STATES
IP - 5
VI - 80
JC - DAJ
AA - Author
EM - 199707
AB - During heart failure, force production by the heart decreases. This may
be overcome by Ca2+-sensitizing drugs, which increase myofibril Ca2+
sensitivity without necessarily altering intracellular Ca2+
concentration. However, Ca2+ sensitizers slow the relaxation of intact
cardiac muscle. We used diazo-2, a caged chelator of Ca2+, to study the
effects of the Ca2+ sensitizers caffeine and CGP 48506 on the intrinsic
relaxation rate of cardiac myofibrils. Trabeculae from rat right
ventricles were skinned by 1% Triton X-100 and were activated in a 10-
microL bath. In steady state experiments, CGP 48506 (10 micromol/L)
shifted the force-pCa curve leftward by 0.41+/-0.03 pCa units (mean+/-
SEM, n=6). An identical shift was induced by caffeine (20 mmol/L).
Photolysis of diazo-2 by a flash of light (160 mJ, 310 to 400 nm)
caused an immediate decrease in Ca2+-activated force produced by the
trabeculae. Relaxation was fitted by a double-exponential decay, and
the rate constants were found to be independent of force and preflash
Ca2+ concentration. The initial fast rate, corresponding to
myofibrillar relaxation, was increased from 17.3+/-2.0 to 30.9+/-3.7 s(-
1) (n=4) by caffeine but was unaffected by CGP 48506 (16.6+/-1.7 and
14.4+/-2.3 s(-1) in the absence and presence of drug, respectively;
n=5). Thus, myofibril relaxation need not be slowed by Ca2+-sensitizing
agents but can even be accelerated. Despite similarities in their
effects on myofibril Ca2+ sensitivity, caffeine and CGP 48506 affect
the myofibrils at least partly via different mechanisms.
AD - Department of Pharmacology, United Medical and Dental Schools, St.
Thomas's Hospital, London, UK.
PMID- 0009130449
SO - Circ Res 1997 May;80(5):682-7
UI - 97171296
AU - Fredberg JJ
AU - Jones KA
AU - Nathan M
AU - Raboudi S
AU - Prakash YS
AU - Shore SA
AU - Butler JP
AU - Sieck GC
TI - Friction in airway smooth muscle: mechanism, latch, and implications in
asthma.
LA - Eng
MH - Airway Resistance/*physiology
MH - Animal
MH - Asthma/*physiopathology
MH - Dogs
MH - Friction
MH - Muscle, Smooth/*physiology
MH - Support, U.S. Gov't, P.H.S.
MH - Trachea/*physiology
PT - JOURNAL ARTICLE
ID - PO1 HL-33009/HL/NHLBI
ID - HL-45532/HL/NHLBI
DA - 19970409
DP - 1996 Dec
IS - 8750-7587
TA - J Appl Physiol
PG - 2703-12
SB - M
CY - UNITED STATES
IP - 6
VI - 81
JC - HEG
AA - Author
EM - 199706
AB - In muscle, active force and stiffness reflect numbers of actin-myosin
interactions and shortening velocity reflects their turnover rates, but
the molecular basis of mechanical friction is somewhat less clear. To
better characterize molecular mechanisms that govern mechanical
friction, we measured the rate of mechanical energy dissipation and the
rate of actomyosin ATP utilization simultaneously in activated canine
airway smooth muscle subjected to small periodic stretches as occur in
breathing. The amplitude of the frictional stress is proportional to
eta E, where E is the tissue stiffness defined by the slope of the
resulting force vs. displacement loop and eta is the hysteresivity
defined by the fatness of that loop. From contractile stimulus onset,
the time course of frictional stress amplitude followed a biphasic
pattern that tracked that of the rate of actomyosin ATP consumption.
The time course of hysteresivity, however, followed a different
biphasic pattern that tracked that of shortening velocity. Taken
together with an analysis of mechanical energy storage and dissipation
in the cross-bridge cycle, these results indicate, first, that like
shortening velocity and the rate of actomyosin ATP utilization,
mechanical friction in airway smooth muscle is also governed by the
rate of cross-bridge cycling; second, that changes in cycling rate
associated with conversion of rapidly cycling cross bridges to slowly
cycling latch bridges can be assessed from changes of hysteresivity of
the force vs. displacement loop; and third, that steady-state force
maintenance (latch) is a low-friction contractile state. This last
finding may account for the unique inability of asthmatic patients to
reverse spontaneous airways obstruction with a deep inspiration.
AD - Department of Environmental Health, Harvard School of Public Health,
Boston, Massachsetts 02115, USA. jfredber@hsph.harvard.edu
PMID- 0009018525
SO - J Appl Physiol 1996 Dec;81(6):2703-12
UI - 96355254
AU - Takeshima H
AU - Ikemoto T
AU - Nishi M
AU - Nishiyama N
AU - Shimuta M
AU - Sugitani Y
AU - Kuno J
AU - Saito I
AU - Saito H
AU - Endo M
AU - Iino M
AU - Noda T
TI - Generation and characterization of mutant mice lacking ryanodine
receptor type 3.
LA - Eng
MH - Animal
MH - Base Sequence
MH - Calcium/physiology
MH - Calcium Channels/*physiology
MH - Cell Division
MH - Genes, Structural
MH - Lymphocyte Transformation
MH - Mice
MH - Mice, Knockout
MH - Molecular Sequence Data
MH - Motor Activity
MH - Muscle Contraction
MH - Muscle Proteins/*physiology
MH - Muscle, Smooth, Vascular/physiology
MH - Restriction Mapping
MH - Spleen/cytology
MH - Support, Non-U.S. Gov't
RN - 0 (Calcium Channels)
RN - 0 (Muscle Proteins)
RN - 0 (Ryanodine Receptor Calcium Release Channel)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
DA - 19961003
DP - 1996 Aug 16
IS - 0021-9258
TA - J Biol Chem
PG - 19649-52
SB - M
SB - X
CY - UNITED STATES
IP - 33
VI - 271
JC - HIV
AA - Author
EM - 199612
AB - The ryanodine receptor type 3 (RyR-3) functions as a Ca2+-induced Ca2+
release (CICR) channel and is distributed in a wide variety of cell
types including skeletal muscle and smooth muscle cells, neurons, and
certain non-excitable cells. However, the physiological roles of RyR-3
are totally unclear. To gain an insight into the function of RyR-3 in
vivo, we have generated mice lacking RyR-3 by means of the gene
targeting technique. The mutant mice thus obtained showed apparently
normal growth and reproduction. Although Ca2+-induced Ca2+ release from
intracellular Ca2+ stores of the mutant skeletal muscle differed in
Ca2+ sensitivity from that of wild-type muscle, excitation-contraction
coupling of the mutant muscle seemed to be normal. Moreover, we could
not find any significant disturbance in the smooth muscle and
lymphocytes from the mutant mice. On the other hand, the mutant mice
showed increased locomotor activity, which was about 2-fold greater
than that of the control mice. These results indicate that the loss of
RyR-3 causes no gross abnormalities and suggest that the lack of RyR-3-
mediated Ca2+ signaling results in abnormalities of certain neurons in
the central nervous system.
AD - Department of Pharmacology, Faculty of Medicine, University of Tokyo,
Bunkyo-ku, Tokyo 113, Japan.
PMID- 0008702664
CU - 1997
SI - GENBANK/D84236
SI - GENBANK/D84237
SO - J Biol Chem 1996 Aug 16;271(33):19649-52
UI - 96305042
AU - Dekker LR
AU - Fiolet JW
AU - VanBavel E
AU - Coronel R
AU - Opthof T
AU - Spaan JA
AU - Janse MJ
TI - Intracellular Ca2+, intercellular electrical coupling, and mechanical
activity in ischemic rabbit papillary muscle. Effects of
preconditioning and metabolic blockade.
LA - Eng
MH - Animal
MH - Biomechanics
MH - Calcium/*metabolism
MH - Electric Conductivity
MH - Female
MH - Fluorescent Dyes
MH - Hydrogen-Ion Concentration
MH - Indoles
MH - Intracellular Membranes/metabolism/*physiology
MH - Iodoacetates/pharmacology
MH - Male
MH - Myocardial Contraction
MH - Myocardial Ischemia/metabolism/*physiopathology
MH - *Myocardial Reperfusion
MH - Papillary Muscles/drug effects/metabolism/*physiopathology
MH - Rabbits
MH - Time Factors
RN - 0 (Fluorescent Dyes)
RN - 0 (Indoles)
RN - 0 (Iodoacetates)
RN - 64-69-7 (Iodoacetic Acid)
RN - 7440-70-2 (Calcium)
RN - 96314-96-4 (indo-1)
PT - JOURNAL ARTICLE
DA - 19961212
DP - 1996 Aug
IS - 0009-7330
TA - Circ Res
PG - 237-46
SB - M
CY - UNITED STATES
IP - 2
VI - 79
JC - DAJ
AA - Author
EM - 199702
AB - During myocardial ischemia, electrical uncoupling and contracture
herald irreversible damage. In the present study, we tested the
hypothesis that an increase of intracellular Ca2+ is an important
factor initiating these events. Therefore, we simultaneously determined
tissue resistance, mechanical activity, pH(0), and intracellular Ca2+
(with the fluorescent indicator indo 1, Molecular Probes, Inc) in
arterially perfused rabbit papillary muscles. Sustained ischemia was
induced in three experimental groups: (1) control, (2) preparations
preconditioned with two 5-minute periods of ischemia followed by
reperfusion, and (3) preparations pretreated with 1 mmol/L iodoacetate
to block anaerobic metabolism and minimize acidification during
ischemia. In a fourth experimental group, intracellular Ca2+ was
increased under nonischemic conditions by perfusing with 0.1 mmol/L
ionomycin and 0.1 mumol/L gramicidin. Ca2+ transients and contractions
rapidly disappeared after the induction of ischemia. In the control
group, diastolic Ca2+ began to rise after 12.6 +/- 1.3 minutes of
ischemia; uncoupling, after 14.5 +/- 1.2 minutes of ischemia; and
contracture, after 12.6 +/- 1.5 minutes of ischemia (mean +/- SEM).
Preconditioning significantly postponed Ca2+ rise, uncoupling, and
contracture (21.5 +/- 4.0, 24.0 +/- 4.1, and 23.0 +/- 5.3 minutes of
ischemia, respectively). Pretreatment with iodoacetate significantly
advanced these events (1.9 +/- 0.7, 3.6 +/- 0.9, and 1.9 +/- 0.2
minutes of ischemia, respectively). In all groups, the onset of
uncoupling always followed the start of Ca2+ rise, whereas the start of
contracture was not different from the rise in Ca2+. Perfusion with
ionomycin and gramicidin permitted estimation of a threshold [Ca2+] for
electrical uncoupling of 685 +/- 85 nmol/L. In conclusion, the rise in
intracellular Ca2+ is the main trigger for cellular uncoupling during
ischemia. Contracture is closely associated with the increase of
intracellular Ca2+ during ischemia.
AD - Department of Experimental Cardiology, Academic Medical Center,
Amsterdam, The Netherlands.
PMID- 0008756000
CU - 1998
SO - Circ Res 1996 Aug;79(2):237-46
UI - 96025773
AU - Zimmermann B
AU - Somlyo AV
AU - Ellis-Davies GC
AU - Kaplan JH
AU - Somlyo AP
TI - Kinetics of prephosphorylation reactions and myosin light chain
phosphorylation in smooth muscle. Flash photolysis studies with caged
calcium and caged ATP.
LA - Eng
MH - Adenosine Triphosphate/*analogs & derivatives/metabolism
MH - Animal
MH - Calcium/*metabolism
MH - Calmodulin/metabolism
MH - Chelating Agents/*pharmacology
MH - Comparative Study
MH - Egtazic Acid/*analogs & derivatives/pharmacology
MH - In Vitro
MH - Kinetics
MH - Mesenteric Veins/metabolism
MH - Models, Biological
MH - Muscle, Smooth, Vascular/*metabolism
MH - Myosin Light Chains/*metabolism
MH - Phosphorylation
MH - Photolysis
MH - Rabbits
MH - Support, Non-U.S. Gov't
MH - Support, U.S. Gov't, P.H.S.
MH - Time Factors
RN - 0 (Calmodulin)
RN - 0 (Chelating Agents)
RN - 0 (Myosin Light Chains)
RN - 0 (2-nitrophenyl-EGTA)
RN - 56-65-5 (Adenosine Triphosphate)
RN - 67-42-5 (Egtazic Acid)
RN - 67030-27-7 (P(3)-1-(2-nitro)phenylethyladenosine 5'-triphosphate)
RN - 7440-70-2 (Calcium)
PT - JOURNAL ARTICLE
ID - HL 19242-19/HL/NHLBI
ID - HL 48807-03/HL/NHLBI
DA - 19951204
DP - 1995 Oct 13
IS - 0021-9258
TA - J Biol Chem
PG - 23966-74
SB - M
SB - X
CY - UNITED STATES
IP - 41
VI - 270
JC - HIV
AA - Author
EM - 199602
AB - The pre-myosin light chain (MLC20) phosphorylation components of the
lag phase (td) of contractile activation were determined in
permeabilized smooth muscles activated by photolytic release of ATP
from caged ATP and/or Ca2+ from 4-(2-nitrophenyl)-EGTA (NP-EGTA).
Calmodulin (CaM) shortened the td (470 ms at 0 added CaM) that followed
Ca2+ release, but its effect (td = approximately 200 ms) saturated at
40 microM. Photolysis of caged ATP following preequilibration with
identical [Ca4CaM] shortened td to 41 ms. The rate of phosphorylation
was very fast (3.5 s-1 at 22 degrees C in the presence of 5 microM
exogenous CaM) following photolysis of caged ATP, and, following Ca2+
release, phosphorylation was accelerated by CaM. Simultaneous
photolysis of caged ATP and NP-EGTA was followed by a td of 194 ms at 5
microM CaM and a rate of MLC20 phosphorylation intermediate between
these parameters following photolysis of, respectively, NP-EGTA and
caged ATP. In the presence of the normal, total endogenous CaM content
(37 +/- 4 microM) of protal vein smooth muscles td was 565 ms. Steady
state maximum force at pCa 5.5 was increased by much lower (100 nM)
exogenous [CaM] than was required (> 2.5 microM) to shorten the td. We
estimate the endogenous CaM available under steady state conditions in
vivo to be approximately 0.25 microM and probably less during a rapid
Ca2+ transient. We conclude that the [CaM] dependence of the kinetics
of MLC20 phosphorylation and force development (t1/2 and td) initiated
by Ca2+ reflects the recruitment of a slowly diffusible component of
total CaM. The relatively long duration of td (197 ms) at saturating
[CaM] suggests the contribution to td of an additional component,
possibly a prephosphorylation activation/isomerization of the Ca4CaM
myosin light chain kinase complex (Torok, K., and Trentham, D. R.
(1994) Biochemistry 33, 12807-12820). The relatively short delay (108
ms in the presence of 40 microM CaM) following simultaneous photolysis
of NP-EGTA and caged ATP suggests that preincubation with ATP (prior to
photolysis of NP-EGTA) may inhibit the formation of a preactive Ca2CaM
myosin light chain kinase complex.
AD - Department of Molecular Physiology and Biological Physics, University
of Virginia, Charlottesville 22908, USA.
PMID- 0007592592
SO - J Biol Chem 1995 Oct 13;270(41):23966-74
UI - 95339576
AU - Lesh RE
AU - Somlyo AP
AU - Owens GK
AU - Somlyo AV
TI - Reversible permeabilization. A novel technique for the intracellular
introduction of antisense oligodeoxynucleotides into intact smooth
muscle.
LA - Eng
MH - Animal
MH - Bacterial Toxins/metabolism
MH - Base Sequence
MH - Cell Nucleus/metabolism
MH - Comparative Study
MH - Cytoplasm/metabolism
MH - Electrophoresis, Polyacrylamide Gel
MH - Fluorescence
MH - Guinea Pigs
MH - Ileum
MH - In Vitro
MH - Male
MH - Microscopy, Confocal
MH - Microscopy, Electron
MH - Molecular Sequence Data
MH - Muscle Contraction
MH - Muscle, Smooth/*cytology/ultrastructure
MH - Oligonucleotides, Antisense/*administration & dosage/analysis/genetics
MH - *Organ Culture
MH - Permeability
MH - Rats
MH - Staphylococcus aureus
MH - Support, U.S. Gov't, P.H.S.
MH - Time Factors
RN - 0 (Bacterial Toxins)
RN - 0 (Oligonucleotides, Antisense)
PT - JOURNAL ARTICLE
ID - 1K11-AR-01871/AR/NIAMS
ID - 1PO1-HL-48807/HL/NHLBI
ID - PO1-HL-19242/HL/NHLBI
DA - 19950822
DP - 1995 Aug
IS - 0009-7330
TA - Circ Res
PG - 220-30
SB - M
CY - UNITED STATES
IP - 2
VI - 77
JC - DAJ
AA - Author
EM - 199510
AB - Antisense oligodeoxynucleotides (ODNs) have been used to modify gene
expression in vitro and are also promising therapeutic agents. Although
there are numerous reports of antisense ODN-mediated changes in protein
expression of cultured cells, use of these compounds to achieve
antisense regulation of specific proteins in intact tissue has been
limited. The aims of this study were (1) to define organ culture
conditions for ileum smooth muscle that would permit long-term
maintenance of force-generating capabilities and normal ultrastructure
and (2) to develop a method for efficient introduction of antisense
ODNs into intact tissue. Sheets of ODN-containing, reversibly
permeabilized rat outer longitudinal ileum were maintained in a serum-
free organ culture medium for 1 week without significant decreases in
tension response to membrane depolarization or carbachol stimulation;
the G protein-coupled calcium sensitization pathway was also intact
after 7 days. Reversible permeabilization, a method previously used to
load smooth and cardiac muscle with aequorin and heparin, was effective
for loading > 95% of ileum smooth muscle cells with a fluorescein-
conjugated antisense ODN (5'-AAGGGCCATTTTGTT-FITC-3'). Confocal
microscopy of reversibly permeabilized smooth muscle loaded with
fluorescent antisense ODNs revealed intense nuclear fluorescence and
less intense, homogeneous, cytoplasmic fluorescence. Internally
radiolabeled ODNs (homologous to the above sequence) showed complete
degradation between 4 and 16 hours after introduction into the cells.
In summary, we have demonstrated methods for long-term organ culture
and high-efficiency introduction of antisense ODNs into intact smooth
muscle sheets. Such methods have broad potential utility for
investigating many questions in smooth muscle biology. At present,
however, a major limitation of this approach is the short half-life of
phosphorothioated ODNs.
AD - Department of Anesthesiology, University of Virginia Health Sciences
Center, Charlottesville 22908, USA.
PMID- 0007614709
SO - Circ Res 1995 Aug;77(2):220-30
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