Localized Ca2+-release signals (puffs) and propagated Ca2+ waves were chara
cterized in rat ureteric myocytes by confocal microscopy. Ca2+ puffs were e
voked by photorelease of low concentrations of Ins(1,4,5)P-3 from a caged p
recursor and by low concentrations of acetylcholine; they were also observe
d spontaneously in Ca2+-overloaded myocytes. Ca2+ puffs showed some variabi
lity in amplitude, time course and spatial spread, suggesting that Ins(1,4,
5)P-3-gated channels exist in clusters containing variable numbers of chann
els and that within these clusters a variable number of channels can be rec
ruited. Immunodetection of Ins(1,4,5)P-3 receptors revealed the existence o
f several spots of fluorescence in the confocal cell sections, supporting t
he existence of clusters of Ins(1,4,5)P-3 receptors. Strong Ins(1,4,5)P-3 p
hotorelease and high concentrations of acetylcholine induced Ca2+ waves tha
t originated from an initiation site and propagated in the whole cell by sp
atial recruitment of neighbouring Ca2+-release sites. Both Ca2+ puffs and C
a2+ waves were blocked selectively by intracellular applications of heparin
and an anti-Ins(1,4,5)P-3-receptor antibody, but were unaffected by ryanod
ine and intracellular application of an anti-ryanodine receptor antibody, m
RNAs encoding for the three subtypes of Ins(1,4,5)P-3 receptor and subtype
3 of ryanodine receptor were detected in these myocytes, and the maximal bi
nding capacity of [H-3]Ins(1,4,5)P-3 was 10- to 12-fold higher than that of
[H-3]-ryanodine. These results suggest that Ins(1,4,5)P-3-gated channels m
ediate a continuum of Ca2+ signalling in smooth-muscle cells expressing a h
igh level of Ins(1,4,5)P-3 receptors and no subtypes 1 and 2 of ryanodine r
eceptors.