Sarcoplasmic Reticulum Function in Smooth Muscle - Physiological ...

132 SUSAN WRAY AND THEODOR BURDYGA
suppression of RyR3 did not alter Ca spark activity. In
urinary bladder myocytes, RyR2s are required for generation
of Ca sparks (314). Native RyR3s are either not
involved (133, 314) or may even have an inhibitory effect
on Ca sparks (407). However, when both RyR1 and RyR2
are inhibited with antisense oligonucleotides, and under
conditions of increased SR Ca loading, RyR3 can be activated
by caffeine or agonists (475, 476). RyR3 gene knockout
significantly inhibits the contractile response to hypoxia
but not the norepinephrine-induced Ca and contractile
responses, in pulmonary artery smooth muscle cells
(805).
The function of RyR3 in Ca signaling is complicated
by alternative splicing of RyR3. The expression of a short
isoform of RyR3 (RyR3s) in HEK293 cells can inhibit both
the full-length RyR3 (RyR3L) and RyR2 subtypes (315). In
native smooth muscle, the dominant negative effect of the
spliced isoform of RyR3 inhibits RyR2 activation and Ca
signals in duodenum (138) and the activity of RyR3 in
mouse pregnant myometrium (139). These observations
reveal a novel mechanism by which a splice variant of one
RyR3 isoform may heteromerize with the other RyR variants
and suppress the activity of these RyR isoforms via a
dominant negative effect. Thus tissue-specific expression
of RyR3 splice variants is likely to account for some of the
pharmacological and functional heterogeneities of RyR3,
for example, the lack of Ca sparks and caffeine sensitivity
in rat myometrium, despite expression of all three forms
of RyRs (88).
On the basis of the available information, RyR2 appears
to be the most important component of Ca sparks in
smooth muscle, and recent findings on RyR3 splice variants
have helped unravel some of the perplexity in Ca
signaling in smooth muscle. Calcium sparks are discussed
in detail in section IX.
C. IP 3-Sensitive Ca release channels
Binding of G protein-coupled receptor agonists to their
receptors in smooth muscles leads to activation of phosphatidylinositol-specific
phospholipase C (PLC), which catalyzes
the hydrolysis of phosphatidylinositol 4,5-bisphosphate
(PIP 2) to yield IP 3 and diacylglycerol (DAG) (48). The
IP 3 releases Ca from intracellular stores by binding to IP 3R,
which are IP 3-gated Ca release channels, while DAG activates
protein kinase C (PKC). Activation of IP 3Rs via activation
of L-type Ca channels independent of Ca entry, socalled
metabotropic Ca channel-induced Ca release, has
been suggested in arterial myocytes but requires confirmation
by other studies (149), and is not discussed further.
1. Expression
Three distinct IP3R gene products (types 1–3) have
been identified in different mammalian species (283, 546).
All three isoforms of IP 3R are structurally and functionally
related (193). The IP 3R isoforms differ in their sensitivities
to breakdown by cellular proteases (IP 3R2 relatively
more resistant that IP 3R1 and IP 3R3) (765). These
receptors exist as tetrameric structures, with a monomeric
molecular mass of 300 kDa. Unlike RyRs, IP 3Rs
are poorly expressed in skeletal and cardiac muscle but
extensively expressed in brain and other tissues including
smooth muscles (317, 471, 498, 546, 695, 760, 794). In
smooth muscles, the density of IP 3Ris100 times less
than in brain (437, 802). In visceral smooth muscles (356,
379–381), IP 3Rs are expressed in greater quantity than
RyRs with an overall stoichiometric ratio of IP 3Rs to RyRs
of 10–12:1, while in vascular smooth muscles the ratio
is 3-4:1 (62).
Smooth muscle cells express multiple IP 3R isoforms,
although as with RyRs, the level of expression of different
subtypes is tissue and species dependent. Using reversetranscriptase
PCR analysis, Morel et al. (493) showed that
rat portal vein expressed IP 3R1 and IP 3R2, whereas rat
ureter expressed predominantly IP 3R1 and IP 3R3 (493),
although in an earlier study it was reported that rat ureteric
myocytes expressed all three isoforms of IP 3Rs (60).
In the thoracic aorta and mesenteric arteries, IP 3R1 was
the only isoform found, while IP 3R1 and weak expression
of IP 3R2 were reported for smooth muscle cells of basilar
artery (324). The type 1 isoform predominates in guinea
pig intestinal smooth muscle cells (222) as well as in adult
porcine (297) and rat aorta (693), whereas type 3 predominates
in neonatal aorta (693). Subtype-specific IP 3R antibodies
revealed that the expression of IP 3R1 was similar
in cultured aortic cells and aorta homogenate, but expression
of IP 3R2 and IP 3R3 types was increased threefold in
cultured cells (693). Immunostaining and functional studies
performed on isolated cells and cells in situ in rat
portal vein showed two subpopulations of cells, one
which predominantly expressed IP 3R1 and a second predominantly
expressing IP 3R2. Distinct patterns of the Ca
responses were generated by the two subpopulations of
myocytes in response to agonist (493).
2. Localization
Physiol Rev VOL 90 JANUARY 2010 www.prv.org
In some smooth muscles, IP 3R1 has been localized
throughout the cell, i.e., central and peripheral SR, as
visualized using EM (191, 515, 693, 729), and in the others
it had a mainly subplasmalemmal location (222). IP 3R2
has a diffuse cytoplasmic distribution, similar to IP 3R1
(and IP 3R3), but also occurs as dense patches in the
peripheral cytoplasm, a pattern that IP 3R1 did not exhibit
(668). IP 3Rs were also arranged in clusters in rat ureteric
myocytes, as well as throughout the cell (60). In vascular
myocytes, IP 3R2 is distributed peripherally and associated
with the nucleus in proliferating cells (668, 694). In
portal vein, IP 3R2 was also found closely associated with