The Duration and Amplitude of the Plateau Phase of ATP- and ADP-Evoked Ca²⁺ Signals Are Modulated by Ectonucleotidases in in situ Endothelial Cells of Rat Aorta

Abstract

ATP has a long-lasting vasodilatory effect, possibly due to its capability to induce a prolonged increase in the intracellular Ca²⁺ concentration ([Ca²⁺]_i) in endothelial cells (EC) and activate constitutive nitric oxide synthase. However, contradictory data have been reported regarding the time course of ATP-evoked Ca²⁺ signals in in situ EC. In particular, short-duration Ca²⁺ signals have been reported, which might be thought to be unable to sustain a prolonged, NO-induced vasodilation. The current experiments were therefore performed in in situ EC of rat aorta in order to more fully define the time course of ATP-evoked Ca²⁺ signals. 20 µM ATP evoked a short-lasting Ca²⁺ signal. However, medium stirring, high agonist concentrations, inhibition of ectonucleotidases and application of a poorly hydrolyzable agonist evoked long-lasting Ca²⁺ signals (up to 20 min at 37°C). These studies suggest that ATP is able to sustain a prolonged [Ca²⁺]_i increase, unless ectonucleotidase activity reduces the agonist concentration near the EC surface to subthreshold values, quickly cutting the Ca²⁺ signal. Furthermore, the amplitude of the long-lasting phase of the Ca²⁺ signal depended on the balance between agonist degradation by ectonucleotidases and agonist transport, by diffusion and convection, from bulk solution to the EC surface.

Connexins and M3 Muscarinic Receptors Contribute to Heterogeneous Ca2+Signaling in Mouse Aortic Endothelium

“…Such spreading of ACh-induced Ca 2+ signals is in accordance with dye transfer experiments showing extensive coupling between endothelial cells from the mouse aorta [10,11]. In line with the work of Berra-Romani et al [29], our results show homogenous endothelial Ca 2+ signaling in response to ATP stimulation. In contrast to these results, Kameritsch et al, recently reported heterogeneous Ca 2+ responses to ATP and demonstrated a role for endothelial gap junctions in the spreading of Ca 2+ signaling in response to ATP stimulation [30].…”

The Mechanism of Injury-Induced Intracellular Calcium Concentration Oscillations in the Endothelium of Excised Rat Aorta

“…B Addition of a low dose of the InsP 3 -mobilizing agonist, ATP (20 µ M ), resumed Ca 2+ oscillations in nonspiking cells (upper and lower traces), whereas it did not modify the interspike interval of ongoing transients (middle trace). Note that, at the concentration employed here, ATP did not stimulate Ca 2+ entry in the endothelium of excised rat aorta [23]. C Ryanodine (20 µ M ), a selective ryanodine receptor inhibitor at this dose, did not cause any detectable change in the spiking response to injury. …”

“…6A), but it did not hamper the long-lasting decay phase [7]. Moreover, Ca 2+ oscillations resumed in 27 out of 27 cells exposed to a low dose (20 µ M ) of the InsP 3 -releasing agonist ATP [23] (fig. 6B, upper and lower traces).…”

Ca2+ Signalling in Damaged Endothelium: Do Connexin Hemichannels Aid in Filling the Gap?

“…Whereas the former exhibit a transient Ca 2+ wave [48,49], which is mainly due to Ca 2+ mobilization from inositol-1,4,5-trisphosphate (InsP 3 )-sensitive receptors (InsP 3 Rs), the latter display a biphasic signal [33,50,51], comprising a rapid Ca 2+ spike followed by an intermediate plateau level due to Ca 2+ influx from the extracellular space. In addition, depending on either the physical nature of the stimulus or on the agonist dose, the Ca 2+ signal may adopt an oscillatory pattern driven by the interplay between Ca 2+ release from InsP 3 Rs and Ca 2+ entry through plasma membrane (PM) cationic channels [52][53][54][55].…”