1. ATP-mediated Ca2+ signalling was studied in freshly isolated rat portal
vein myocytes by means of a laser confocal microscope and the patch-clamp t
echnique.
2. In vascular myocytes held at -60 mV, ATP induced a large inward current
that was supported mainly by activation of P2X1 receptors, although other P
2X receptor subtypes (P2X3, P2X4 and P2X5) were revealed by reverse transcr
iption-polymerase chain reaction.
3. Confocal Ca2+ measurements revealed that ATP-mediated Ca2+ responses sta
rted at initiation sites where spontaneous or triggered Ca2+ sparks were no
t detected, whereas membrane depolarizations triggered Ca2+ waves by repeti
tive activation of Ca2+ sparks from a single initiation site. mediated Ca2 release requires, at least, RYR2, but not RYR3.
4. ATP-mediated Ca2+ responses depended on Ca2+ influx through non-selectiv
e cation channels that activated, in turn, Ca2+ release from the intracellu
lar store via ryanodine receptors (RYRs). Using specific antibodies directe
d against the RYR subtypes we show that ATP-mediated Ca2+ release requires,
at least RYR2, but not RYR3.
5. Our results suggest that, in vascular myocytes, Ca2+ influx through P2X1
receptors may trigger Ca2+-induced Ca2+ release at intracellular sites whe
re RYRs are not clustered.