Discrete localized fluorescence transients due to openings of a single
plasma membrane Ca$^2+$ permeable cation channel were recorded
using wide-field digital imaging microscopy with fluo-3 as the Ca$^2+$
indicator. These transients were obtained while simultaneously recording
the unitary channel currents using the whole-cell current-recording
configuration of the patch-clamp technique. This cation channel in
smooth muscle cells is opened by caffeine (Guerrero, A., F.S. Fay,
and J.J. Singer. 1994. J. Gen. Physiol. 104:375-394). The localized
fluorescence transients appeared to occur at random locations on
the cell membrane, with the duration of the rising phase matching
the duration of the channel opening. Moreover, these transients were
only observed in the presence of sufficient extracellular Ca$^2+$,
suggesting that they are due to Ca$^2+$ influx from the bathing
solution. The fluorescence transient is characterized by an initial
fast rising phase when the channel opens, followed by a slower rising
phase during prolonged openings. When the channel closes there is
an immediate fast falling phase followed by a slower falling phase.
Computer simulations of the underlying events were used to interpret
the time course of the transients. The rapid phases are mainly due
to the establishment or removal of Ca$^2+$ and Ca$^2+$-bound
fluo-3 gradients near the channel when the channel opens or closes,
while the slow phases are due to the diffusion of Ca$^2+$ and
Ca$^2+$-bound fluo-3 into the cytoplasm. Transients due to short
channel openings have a "Ca$^2+$ spark-like" appearance, suggesting
that the rising and early falling components of sparks (due to openings
of ryanodine receptors) reflect the fast phases of the fluorescence
change. The results presented here suggest methods to determine the
relationship between the fluorescence transient and the underlying
Ca$^2+$ current, to study intracellular localized Ca$^2+$
handling as might occur from single Ca$^2+$ channel openings,
and to localize Ca$^2+$ permeable ion channels on the plasma
membrane.
%0 Journal Article
%1 Zou_1999_575
%A Zou, H.
%A Lifshitz, L. M.
%A Tuft, R. A.
%A Fogarty, K. E.
%A Singer, J. J.
%D 1999
%J J. Gen. Physiol.
%K 10498675 Agonists, Aniline Animals, Bufo Caffeine, Calcium Calcium, Cations, Cell Central Channel Channels, Compounds, Computer Computer-Assisted, Conductivity, Cytoplasm, Cytosol, Dyes, Electric Electrophysiology, Factors, Fluorescence, Fluorescent Gating, Gov't, Image In Ion Membr, Membrane, Microscopy, Muscle, Myocytes, Nervous Non-P.H.S., P.H.S., Patch-Clamp Processing, Resear, Research Simulation, Smooth Smooth, Spectrometry, Stimulants, Support, System Techniques, Time U.S. Vitro, Xanthenes, ane, ch marinus,
%N 4
%P 575--588
%T Imaging Ca$^2+$ entering the cytoplasm through a single opening
of a plasma membrane cation channel.
%U http://www.jgp.org/cgi/content/full/114/4/575
%V 114
%X Discrete localized fluorescence transients due to openings of a single
plasma membrane Ca$^2+$ permeable cation channel were recorded
using wide-field digital imaging microscopy with fluo-3 as the Ca$^2+$
indicator. These transients were obtained while simultaneously recording
the unitary channel currents using the whole-cell current-recording
configuration of the patch-clamp technique. This cation channel in
smooth muscle cells is opened by caffeine (Guerrero, A., F.S. Fay,
and J.J. Singer. 1994. J. Gen. Physiol. 104:375-394). The localized
fluorescence transients appeared to occur at random locations on
the cell membrane, with the duration of the rising phase matching
the duration of the channel opening. Moreover, these transients were
only observed in the presence of sufficient extracellular Ca$^2+$,
suggesting that they are due to Ca$^2+$ influx from the bathing
solution. The fluorescence transient is characterized by an initial
fast rising phase when the channel opens, followed by a slower rising
phase during prolonged openings. When the channel closes there is
an immediate fast falling phase followed by a slower falling phase.
Computer simulations of the underlying events were used to interpret
the time course of the transients. The rapid phases are mainly due
to the establishment or removal of Ca$^2+$ and Ca$^2+$-bound
fluo-3 gradients near the channel when the channel opens or closes,
while the slow phases are due to the diffusion of Ca$^2+$ and
Ca$^2+$-bound fluo-3 into the cytoplasm. Transients due to short
channel openings have a "Ca$^2+$ spark-like" appearance, suggesting
that the rising and early falling components of sparks (due to openings
of ryanodine receptors) reflect the fast phases of the fluorescence
change. The results presented here suggest methods to determine the
relationship between the fluorescence transient and the underlying
Ca$^2+$ current, to study intracellular localized Ca$^2+$
handling as might occur from single Ca$^2+$ channel openings,
and to localize Ca$^2+$ permeable ion channels on the plasma
membrane.
@article{Zou_1999_575,
abstract = {Discrete localized fluorescence transients due to openings of a single
plasma membrane {C}a$^{2+}$ permeable cation channel were recorded
using wide-field digital imaging microscopy with fluo-3 as the {C}a$^{2+}$
indicator. These transients were obtained while simultaneously recording
the unitary channel currents using the whole-cell current-recording
configuration of the patch-clamp technique. This cation channel in
smooth muscle cells is opened by caffeine (Guerrero, A., F.S. Fay,
and J.J. Singer. 1994. J. Gen. Physiol. 104:375-394). The localized
fluorescence transients appeared to occur at random locations on
the cell membrane, with the duration of the rising phase matching
the duration of the channel opening. Moreover, these transients were
only observed in the presence of sufficient extracellular {C}a$^{2+}$,
suggesting that they are due to {C}a$^{2+}$ influx from the bathing
solution. The fluorescence transient is characterized by an initial
fast rising phase when the channel opens, followed by a slower rising
phase during prolonged openings. When the channel closes there is
an immediate fast falling phase followed by a slower falling phase.
Computer simulations of the underlying events were used to interpret
the time course of the transients. The rapid phases are mainly due
to the establishment or removal of {C}a$^{2+}$ and {C}a$^{2+}$-bound
fluo-3 gradients near the channel when the channel opens or closes,
while the slow phases are due to the diffusion of {C}a$^{2+}$ and
{C}a$^{2+}$-bound fluo-3 into the cytoplasm. Transients due to short
channel openings have a "{C}a$^{2+}$ spark-like" appearance, suggesting
that the rising and early falling components of sparks (due to openings
of ryanodine receptors) reflect the fast phases of the fluorescence
change. The results presented here suggest methods to determine the
relationship between the fluorescence transient and the underlying
{C}a$^{2+}$ current, to study intracellular localized {C}a$^{2+}$
handling as might occur from single {C}a$^{2+}$ channel openings,
and to localize {C}a$^{2+}$ permeable ion channels on the plasma
membrane.},
added-at = {2009-06-03T11:20:58.000+0200},
author = {Zou, H. and Lifshitz, L. M. and Tuft, R. A. and Fogarty, K. E. and Singer, J. J.},
biburl = {https://www.bibsonomy.org/bibtex/26451aba4de3c566760495d9307329599/hake},
description = {The whole bibliography file I use.},
file = {Zou_1999_575.pdf:Zou_1999_575.pdf:PDF},
interhash = {b8e745d3da01e810e124f0751122335f},
intrahash = {6451aba4de3c566760495d9307329599},
journal = {J. Gen. Physiol.},
key = 263,
keywords = {10498675 Agonists, Aniline Animals, Bufo Caffeine, Calcium Calcium, Cations, Cell Central Channel Channels, Compounds, Computer Computer-Assisted, Conductivity, Cytoplasm, Cytosol, Dyes, Electric Electrophysiology, Factors, Fluorescence, Fluorescent Gating, Gov't, Image In Ion Membr, Membrane, Microscopy, Muscle, Myocytes, Nervous Non-P.H.S., P.H.S., Patch-Clamp Processing, Resear, Research Simulation, Smooth Smooth, Spectrometry, Stimulants, Support, System Techniques, Time U.S. Vitro, Xanthenes, ane, ch marinus,},
month = Oct,
number = 4,
pages = {575--588},
pmid = {10498675},
timestamp = {2009-06-03T11:21:39.000+0200},
title = {Imaging {C}a$^{2+}$ entering the cytoplasm through a single opening
of a plasma membrane cation channel.},
url = {http://www.jgp.org/cgi/content/full/114/4/575},
volume = 114,
year = 1999
}