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Calcium channels in smooth muscle cells
drugs: the dihydropyridines (Worley et al., 1986; Yatani el
al., 1987).
Receptor-operated channels may also admit calcium,
although the amounts probably vary depending on the
smooth muscle and the receptor involved. The main function
The importance of calcium ions for contraction of muscle of receptor-operated channels is probably to shift the membegan to be appreciated when attempts were made to con- brane potential into (or out of, in the case of relaxant substruct physiological salt solutions suitable for maintaining stances) the potential range where potential-sensitive calcium
contraction of muscle tissues in vitro (e.g. Ringer, 1896). channels operate. Smooth muscle depolarized by high-potasMuch later, experiments on smooth muscles were carried out sium solution can admit calcium when carbachol, a muscarwhich created some confusion concerning the mechanism of inic-receptor stimulant, is applied (Durbin & Jenkinson,
tension generation in this muscle type: recordings with 196 1).Single-cell studies show that ATP receptor-operated
microelectrodes showed a good correlation between the level channels can admit divalent cations including calcium
of membrane potential, frequency of action potential dis- (Benham et al., 1987) and single-channel studies have shown
charge, and tension in phasic visceral smooth muscle (Bul- that appreciable amounts of calcium may enter through ATPbring, 1955), but it was soon demonstrated in phasic receptor-operated channels (Benham & Tsien, 1987). We
uterine smooth muscle (Evans et al., 1958) that it could find that muscarinic-receptor-operated channels behave in a
contract well to acetylcholine in depolarizing high-potassium similar way.
solutions. This paradox was resolved by the suggestion that
In single smooth muscle cells the importance of release of
calcium may enter the smooth muscle cell both via channels calcium-stores can be amply demonstrated. In vascular
gated by potential (‘potential-sensitive channels’) and muscle noradrenaline can release stored calcium, and in
through channels gated by receptor (‘receptor-operated intestinal smooth muscle, acetylcholine can have a very simichannels’) (Bolton, 1979; van Breeman et al., 1979). A fur- lar effect (Benham et al., 1985; Benham & Bolton, 1986).
ther problem was the variable degree to which contractions However, the importance of this release for the contractile
to receptor activation (e.g. by acetylcholine via muscarinic response of the whole muscle to low concentrations of stimureceptors in visceral muscle or by noradrenaline via a- lants needs to be established; it is likely that the release of
receptors in vascular muscle) were resistant to calcium-free calcium in response to receptor activation involves inositol
conditions (Edman & Schild, 1962). Some smooth muscle 1,4,5-trisphosphate production from phosphatidylinositol
responses are very resistant (e.g. Tomita et al., 1985). We 4,5-biphosphate by the action of phospholipase C (Itoh et
now believe this is caused by the sequestration of calcium in al., 1988).
storage sites within the smooth muscle cell from where it can
be efficiently and rapidly released following receptor activaSupported by the Medical Research Council.
tion.
Potential-sensitive calcium channels seem to be ubiquiAaronson, P. I., Benham, C. D., Bolton, T. B., Hess, P., Lang, R. J. &
tously present on smooth muscle cells, although their density
Tsien, R. W. ( 1 9 8 6 )J . Physiol. (London) 377,3601’.
is lower if the smooth muscle type is electrically inexcitable Benham, C. D. & Bolton, T. B. ( 1 9 8 6 ) J. I’hysiol. (London) 3 8 1 ,
(e.g. many vascular muscles d o not generate action potentials
3 8 5 -406
very readily either spontaneously or in response to electrical Benham, C. D. & Tsien, R. W. ( 1 9 8 7 ) Nuture (London) 3 2 8 ,
depolarization) (Bolton et al., 1988).These calcium channels
275-278
open in response to depolarization of the membrane and are Benhain, C. D.. Bolton, T. B. & Lang, R. J. ( 1 9 8 5 )Nuture (London)
316,345-347
responsible for the upstroke of the action potential where
Benham,
C. D., Bolton, T. B., Byrne, N. G. & Large, W. A. ( 1 9 8 7 )
this is seen in excitable smooth muscles. In non- or less excitJ . Physiol. (London) 3 8 7 , 4 7 3 - 4 8 8
able smooth muscle, depolarization causes the opening of Bolton,
T. B. ( 1 9 7 9 )Physiol. Rev. 59,606-7 18
these channels also, but action potentials d o not occur. Prob- Bolton, T. B., Aaronson, P. 1. & MacKenzie, I. ( 1 9 8 8 ) Ann. N. Y.
ably at least two types of potential-sensitive calcium channel
Acud. Sci. in the press
exist (Aaronson et al., 1986); the exact contributions of these Bulbring, E. ( 1955) J. I’hysiol. (London) 128,200-22 1
to the total inward calcium current is in doubt as is the sensi- Durbin, R. P. & Jenkinson, D. H. ( 1 9 6 1 )J. Physiol. (London) 157,
74-89
tivity of individual channel types to calcium-entry blocking
T. B. BOLTON, 1. MAcKENZIE, P. 1. AARONSON and
S. P. LIM
St George S Hospital Medical School, London S WI 7 ORE,
U.K.
1088
625th MEETING. LONDON
Edman. K. A. P. & Schild, H. 0. (1962) J. I'hysiol. (London) 161,
424-44 1
Evans, D. H. L., Schild, H. 0.& Thesleff, S. (1958) J. I'hysiol. 143,
474-485
Itoh. T., Hirata, M., Kitamura, K. & Kuriyama, H. (1088) Biochem.
Soc. Trans. 16,488-489
Ringer, S. ( 1896)J. I'hysiol. (London) 18,425-429
Tomita, T., Ashoori, E & Takai, A. ( I 985) in Calmodulin Anrugonisrs und C'elliilur Physiology, pp. 363-378, Academic Press, Inc.,
London
493
van Breeman, C., Aaronson, P. & Loutzenhiser, R. (1979)
Pharrnacol. Rev. 30,167-208
Worley, J. R., Deitmer, J. W. & Nelson, M. T. (1986) /'roc. Nutl.
Acad. Sci. U.S.A. 83,5746-5750
Yatani, A., Seidel, C. L., Allen, J. & Brown, A. M. (1987)C'irc. Res.
60,523-533
Received 27 January 1988
Measurements of intracellular calcium concentration in mammalian vascular smooth muscle cells
during agonist-induced contractions
KATHLEEN G. MORGAN, FRANK V. BROZOVICH
and MEEI JY H JlANG
Departmerit of Medicirie, Harvurd Medical School, Beth Israel
Hospital, Boston, MA 02215, U.S.A.
We have previously reported that qualitatively different
[Ca?'1, profiles can be obtained during qualitatively identical
smooth muscle contractions induced by different agonists.
The agonist-specificity of the [Ca*+],profile has been confirmed with both the luminescent calcium indicator aequorin
and the fluorescent calcium indicators quin-2 and fura-2 in
either intact strips or enzymically isolated or cultured cells
(Morgan, 1987; Bradley & Morgan, 1987; Griendling et al.,
1986). We have recently correlated the agonist-specific
[Caz+],profiles with the temporal profiles of isometric force,
myosin light chain phosphorylation, and maximum unloaded
shortening velocity (Brozovich & Morgan, 1987; Jiang &
Morgan, 1988). Phosphorylation was determined by twodimensional polyacrylamide-gel electrophoresis and maximum shortening velocity was determined by the slack test.
All measurements were made in ferret aorta at body temperature. In one set of experiments, [Ca?+],(as determined
with aequorin) and force were determined; in a second set of
experiments, both force and the maximum unloaded
shortening velocity were measured; while in a third set, force
and myosin phosphorylation were determined. Data from
these experiments were pooled so that they could be compared at various times after stimulation.
On addition of 1 W 5M-phenylephrine, there was a rapid
rise in [Ca? 1,. myosin phosphorylation and shortening
velocity, so that all three parameters peaked during the
development of force. However, during the maintenance of
force, all three parameters fell to a lower plateau level. These
findings are consistent with the muscle entering the 'latch
state' (Dillon et a/., 108 1), i.e. muscle force is maintained at a
steady-state level concomitant with a fall in [Ca'+],, myosin
phosphorylation and muscle shortening velocity.
In marked contrast, during potassium depolarization with
moderate concentrations of potassium, there were no initial
transients, but rather a monophasic sustained parallel rise in
+
Abbreviation used: [Ca'+],, intracellular calcium ion concentration.
Vol. 16
[Ca2+I,,myosin phosphorylation, and shortening velocity was
observed. The potassium response is of interest since it
represents a unique description of a condition in which the
intact muscle does not appear to enter the 'latch state'.
Thus, from a qualitative perspective, changes in myosin
phosphorylation and shortening velocity roughly parallel the
observed changes in [Ca2+],.However, on a quantitative
level, when a concentration of potassium was chosen to produce an increase in [Ca2+Ii,which approximated the steadystate change in [Ca"], produced by 1 W SM-phenylephrine, it
was found that the steady-state force and shortening velocity
were much greater in the presence of phenylephrine than
could be explained by the steady-state elevation in [Ca2+],or
myosin phosphorylation. Thus, these results point to a cornplex relationship not only between [Ca2'Ii and steady-state
force, but also between [Ca'+], and shortening velocity. It is
possible that during the potassium contracture, a simple relationship exists between [Ca2 ],-mediated phosphorylation
and force as well as between phosphorylation and velocity,
but that phenylephrine recruits additional [Ca2+ji-dependent
regulatory systems.
+
We thank the National Institutes of Health for financial support
(HL31704) and the American Heart Association for an Established
lnvestigatorship to K.G.M.
Bradley, A. B. & Morgan, K. G. (1987) A m . J. /'hy.sio/. 385,
437-448
Brozovich, F. V. & Morgan, K. G. ( I 987) Biophys. J. 51,339A
DeFeo, T. T., Brigns, G. M. & Morgan, K. G. (1987) A m . J. I'hvsiol.
253,1456-14%-1
Dillon, P. F., Aksoy, M. O., Driska, S. P. & Murphy, R. A. ( 1 983)
Science 211,495-497
Griendling, K. K., Rttenhouse, S. E., Brock, T. A,. Ekstein, L. S.,
Ginbrone, M. A. & Alexander, R. W. (1986)J. Hiol. C'hem. 261,
590 1-5906
Jiang, M. J. & Morgan, K. G. (1987) A m . J. I'hysiol. 253,
1365-1 371
Jiang, M. J. & Morgan, K. G. (1988) Riophys. J. 53,597
Morgan, K. G. (1987)A m . J. Cardiol. 59,28A-29A
I
Received 27 January 1988