Sarcoplasmic Reticulum Function in Smooth Muscle - Physiological ...
Sarcoplasmic Reticulum Function in Smooth Muscle - Physiological ...
Sarcoplasmic Reticulum Function in Smooth Muscle - Physiological ...
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136 SUSAN WRAY AND THEODOR BURDYGA<br />
that persisted after reduc<strong>in</strong>g SR Ca content to 40% of its<br />
<strong>in</strong>itial value. At extremely low [IP 3], Ca efflux via the leak<br />
exceeded IP 3-<strong>in</strong>duced Ca efflux. These authors also speculated<br />
that there is heterogeneity of Ca stores with respect<br />
to this leak, perhaps between peripheral and central<br />
SR. Additional support for a nonrelease channel leak has<br />
come from experiments <strong>in</strong> permeabilized pancreatic ac<strong>in</strong>ar<br />
cells, where <strong>in</strong>hibitors of RyR, IP 3R, and Ca release by<br />
NAADP were used, and still an unchanged basal leak<br />
occurred from the ER (408). This group also found a<br />
consistent leak rate over a broad range of SR Ca loads<br />
(486). Although it may be SERCA isoform specific, there is<br />
little reason to th<strong>in</strong>k that the leak is due to reverse mode<br />
of SERCA <strong>in</strong> smooth muscle cells, as has been suggested<br />
for cardiac muscle (627, 646).<br />
To date, there appear to have been but a handful of<br />
papers exam<strong>in</strong><strong>in</strong>g the nature and extent of the SR Ca leak<br />
<strong>in</strong> smooth muscles. The amount of Ca reported upon<br />
block<strong>in</strong>g SERCA has varied, with substantial depletion of<br />
the SR seen <strong>in</strong> cells from uterus (633), stomach (753), and<br />
(cultured) aorta (478, 706) and very little depletion <strong>in</strong><br />
bladder (218), (toad) gastric (661), and (cultured) uterus<br />
(792). These differences are consistent with estimates<br />
made <strong>in</strong> other cell types of leak rates from 10 to 200<br />
mol/m<strong>in</strong> (see Camello et al., Ref. 101).<br />
For A7r5 cells the rate of loss was, expressed as a<br />
maximal fractional Ca content loss, 22%/m<strong>in</strong> (478). Thus it<br />
is clear that <strong>in</strong> several smooth muscles, the SR leak could<br />
rapidly (few m<strong>in</strong>utes) deplete the SR of Ca. However, it<br />
should be cautioned that none of these measurements has<br />
been made under physiological conditions, as isolated,<br />
permeabilized cells, with a pharmacopeia of <strong>in</strong>hibitors<br />
are required for these studies. Thus, <strong>in</strong> smooth muscle<br />
tissues, the contribution of the leak to sett<strong>in</strong>g lum<strong>in</strong>al<br />
Ca levels and its modulation dur<strong>in</strong>g stimulation to affect<br />
functional responses rema<strong>in</strong>s to be better <strong>in</strong>vestigated.<br />
F. Summary<br />
In this section we have reviewed SR Ca release mechanisms,<br />
their modulation by pharmacological and endogenous<br />
agents, and the process of “leak.” Molecular and<br />
pharmacological tools along with knockout animals have<br />
greatly <strong>in</strong>creased our understand<strong>in</strong>g of how Ca is released<br />
from the SR of smooth muscle cells. With the existence of<br />
these different Ca-releas<strong>in</strong>g mechanisms, along with the<br />
different mechanisms of modulation, isoform expression,<br />
and cellular distribution, smooth muscle myocytes appear<br />
to be the most complex of all cells. Certa<strong>in</strong>ly striated<br />
myocytes as well as neuronal and secretory cell types are<br />
not as rich <strong>in</strong> their Ca release processes. As we have<br />
noted elsewhere, we consider this to be a reflection of the<br />
wide range of functions and phenotypic lability <strong>in</strong> smooth<br />
muscle cells.<br />
VI. LUMINAL CALCIUM<br />
A. Introduction<br />
Physiol Rev VOL 90 JANUARY 2010 www.prv.org<br />
Some of the first evidence that the SR <strong>in</strong> smooth<br />
muscle is a Ca storage site was obta<strong>in</strong>ed histochemically<br />
after calcium had been precipitated (147, 259, 567). Better<br />
spatial resolution and the ability to look simultaneously at<br />
several ions came with the technique of electron-probe<br />
microanalysis, used to determ<strong>in</strong>e elemental concentrations<br />
<strong>in</strong> different regions of the myocyte. This approach<br />
led to measurements of [Ca] <strong>in</strong> the SR of 30–50 mmol/kg<br />
dry wt or 2–5 mM wet wt (69, 371), i.e., considerably<br />
higher than <strong>in</strong> the cytoplasm. These studies also detected<br />
a decrease <strong>in</strong> SR [Ca] with stimulation (655). There are,<br />
however, numerous limitations to this approach, such as<br />
the need for fixation and specialized apparatus and the<br />
small dynamic range. Determ<strong>in</strong>ation of 45 Ca efflux can<br />
provide some <strong>in</strong>dication of SR function (393, 477), but<br />
large background counts from 45 Ca bound to Ca-b<strong>in</strong>d<strong>in</strong>g<br />
prote<strong>in</strong>s, and nonspecific leaks, severely limit its usefulness.<br />
In cardiac muscle, 19 F-NMR has been used with a<br />
difluor<strong>in</strong>ated version of BAPTA. The advantage of this<br />
technique is that by alternat<strong>in</strong>g the obta<strong>in</strong><strong>in</strong>g of 19 F data<br />
with 31 P spectra, near-simultaneous <strong>in</strong>formation on ATP<br />
and phosphocreat<strong>in</strong>e and pH can also be obta<strong>in</strong>ed.<br />
B. Fluorescent Indicators for SR Lum<strong>in</strong>al<br />
Ca Measurement<br />
Other methods have been sought for measur<strong>in</strong>g SR<br />
[Ca], of which fluorescent <strong>in</strong>dicators appear the most<br />
promis<strong>in</strong>g. As Zou et al. (813) put it, “there is a strong<br />
need to develop Ca sensors capable of real-time quantitative<br />
Ca concentration measurements <strong>in</strong> specific subcellular<br />
environments without us<strong>in</strong>g natural Ca b<strong>in</strong>d<strong>in</strong>g prote<strong>in</strong>s<br />
such as calmodul<strong>in</strong>, which themselves participate as<br />
signal<strong>in</strong>g molecules <strong>in</strong> cells.” In this paper they describe<br />
the development of one such set of sensors with K d values<br />
rang<strong>in</strong>g from 0.4 to 2 mM. The [Ca] <strong>in</strong> the SR is such that<br />
K d values <strong>in</strong> this range are required. This along with<br />
selectivity (for Ca and the SR) and considerations of what<br />
is bound and what is free Ca, make measur<strong>in</strong>g SR/ER Ca<br />
nontrivial. For a commentary on these issues around SR<br />
[Ca] determ<strong>in</strong>ation, see Bygrave and Benedetti (96), and<br />
for a general review of [Ca] measurement, see Takahashi<br />
et al. (684).<br />
Lum<strong>in</strong>al [Ca] measurements to date <strong>in</strong> smooth muscles<br />
have only been reported for a limited number of<br />
tissues. Most of the studies have used the approach of<br />
fluorescent <strong>in</strong>dicators such as Mag-fura 2 (K d 49 M;<br />
Ref. 667), which was orig<strong>in</strong>ally developed as a probe for<br />
magnesium, and fluo 5-N, Furaptra (e.g., Ref. 667). Some<br />
<strong>in</strong>dicators orig<strong>in</strong>ally considered for cytoplasmic studies