Effects of a Physiological Insulin Concentration on the Endothelin-Sensitive Ca²⁺ Store in Porcine Coronary Artery Smooth Muscle

The effect of insulin to attenuate the Ca²⁺ and contractile response of vascular smooth muscle to a number of agonists has been described previously, but the Ca2+ regulatory mechanisms of insulin action remain unclear. We determined the effect of a physiological insulin concentration (300 pmol/L) on the Ca²⁺ response of vascular smooth muscle cells of the porcine right coronary artery to endothelin 1 (ET-1); furthermore, we examined the cellular Ca²⁺ stores affected by insulin (i.e., Ca²⁺ stores releasable by inositol 1,4,5-trisphosphate, caffeine, and ionomycin). We measured the Ca²⁺ responses of acutely isolated single smooth muscle cells with the fluorescent Ca²⁺ indicator Fura-2. Acute insulin exposure (20 min) significantly attenuated the Ca²⁺ response of single smooth muscle cells to 10 nmol/L ET-1. This inhibitory effect of insulin was observed both in the presence and absence of extracellular Ca²⁺. In contrast with the effects on ET-1-induced Ca²⁺ responses, insulin did not inhibit the Ca²⁺ response to 5 mmol/l caffeine, an agent that directly releases sarcoplasmic reticulum Ca²⁺ stores. Insulin was also without effect on the total cellular Ca²⁺ store released by 1 µmol/l ionomycin, a Ca²⁺-transporting ionophore. When ET-1 and caffeine were given in succession, a sizable caffeine-sensitive Ca²⁺ store could be released from insulin-treated cells but not control cells, indicating that the sarcoplasmic reticulum Ca²⁺ store of insulin-treated cells was not depleted by ET-1. Generalized depletion of the sarcoplasmic reticulum Ca²⁺ store is not one of the cellular mechanisms involved in the effect of insulin on coronary smooth muscle; instead, the effect may be due to an inhibitory influence on transmembrane signal transduction, such as diminished ET-1-induced inositol 1,4,5-trisphosphate production or reduced ability of this phosphoinositol to release stored Ca²⁺.