Adrenoceptor-mediated regulation of myofibrillar Ca²⁺ sensitivity through the GTP-binding protein-related mechanisms: tension recording in β-escin-skinned single rat cardiac cells with preserved receptor functions

Abstract

 To investigate the mechanisms of receptor-mediated regulation of heart muscle contraction, we developed a tension-recording system using β-escin-skinned single cardiac cells of rats and studied the effects of agonists on myofibrillar Ca²⁺ sensitivity and Ca²⁺ release from the sarcoplasmic reticulum (SR). In pCa/tension relations, 1 µM isoproterenol plus 100 µM guanosine 5′-triphosphate (GTP) decreased the myofibrillar Ca²⁺ sensitivity (pCa₅₀, the [Ca²⁺] required for half-maximal tension, as an indicator of the sensitivity; from 6.07 to 5.92); this effect was blocked by 1 µM metoprolol or 1 mM guanosine 5′-O-(2-thiodiphosphate) (GDPβS). Phenylephrine (10 µM) plus 100 µM GTP increased the Ca²⁺ sensitivity (pCa₅₀; from 6.12 to 6.28), and this effect was blocked by 1 µM phentolamine or 1 mM GDPβS. After Ca²⁺ loading into the SR, 10 µM phenylephrine plus 100 µM GTP in a low-ethylene- glycol-bis(β-aminoethylether)-N,N,N′,N′-tetraacetic acid (EGTA, 0.1 mM) relaxing solution induced oscillatory contractions that were attenuated by either 1 µM phentolamine or pre-treatment with 10 µM inositol 1,4,5-trisphosphate. Our results demonstrate that β₁-adrenergic stimulation decreases myofibrillar Ca²⁺ sensitivity and that α₁-adrenergic stimulation both increases the Ca²⁺ sensitivity and activates Ca²⁺ release from the agonist-sensitive SR through GTP-binding protein-related mechanisms.