Dissociation of P₂ purinoceptor‐mediated increase in intracellular Ca²⁺ level from myosin light chain phosphorylation and contraction in rat aorta

1

The effects of P₂ agonists, adenosine‐5′‐triphosphate (ATP), α, β‐methylene‐adenosine‐5′‐triphosphate (α, β‐me‐ATP) and adenosine‐5‐O‐(3‐thiotriphosphate) (ATPγS), on the intracellular free Ca²⁺ level ([Ca²⁺]_i), myosin light chain (MLC) phosphorylation and force of contraction were examined in vascular smooth muscle of rat aorta.

The effects of P₂ agonists, adenosine‐5′‐triphosphate (ATP), α, β‐methylene‐adenosine‐5′‐triphosphate (α, β‐me‐ATP) and adenosine‐5‐O‐(3‐thiotriphosphate) (ATPγS), on the intracellular free Ca²⁺ level ([Ca²⁺]_i), myosin light chain (MLC) phosphorylation and force of contraction were examined in vascular smooth muscle of rat aorta.

ATP (0.1 μm‐1 mM), α, β‐me‐ATP (0.1–100 μm) and ATPγS (1–100 μm) induced transient increases followed by sustained increase in [Ca²⁺]_i. The effects of these agonists were concentration‐dependent. Compared with the effects of a high concentration of KCl (17.5–72.4 mM), the contractions induced by these P₂ purinoceptor agonists were smaller at a given [Ca²⁺]_i.

In the absence of extracellular Ca²⁺ (with 0.5 mM EGTA), ATPγS (10 μm) induced large transient increase in [Ca²⁺]_i with only small contraction in Ca²⁺‐free solution. In contrast, α, β‐me‐ATP (10 μm) induced only a very small increase in [Ca²⁺]_i and contraction.

ATP (1 mM), α, β‐me‐ATP (10 μm) and ATPγS (10 μm), added during stimulation with 0.1 μm noradrenaline, induced additional and transient increases in [Ca²⁺]_i which were also not associated with contraction.

High K⁺ (72.4 mM) increased MLC phosphorylation with a similar time course to that of the increase in [Ca²⁺]_i (peak phosphorylation was 56% when [Ca²⁺]_i increased to 100%). In contrast, the time course of the increase in MLC phosphorylation due to ATP (1 mM) did not coincide with that of the large increases in [Ca²⁺]_i; MLC phosphorylation increased to only 31% when [Ca²⁺]_i increased to 163%. The MLC phosphorylation due to α, β‐me‐ATP (10 μm) and ATPγS (10 μm), measured at peak [Ca²⁺]_i, were only 19% and 14%, respectively, irrespective of a large increase in [Ca²⁺]_i (138% and 188%, respectively).

The absence of a clear relationship between P₂‐purinoceptor‐mediated increase in [Ca²⁺]_i (either by Ca²⁺ influx or Ca²⁺ release) and MLC phosphorylation or force generation appears to imply that elevation in [Ca²⁺]_i does not contribute to these responses.

DOI: 10.1111/j.1476-5381.1996.tb15436.x

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