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Isolation of Human Atrial Myocytes for Simultaneous Measurements of Ca2+ Transients and Membrane Currents
Published on: July 3, 2013
Adrenoceptor-mediated regulation of myofibrillar Ca2+ sensitivity through the GTP-binding protein-related mechanisms:
S Satoh1, S Kinugawa, H Tsutsui
1Research Institute of Angiocardiology and Cardiovascular Clinic, Kyushu University School of Medicine, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.satoshin@cardiol.med.kyushu-u.ac.jp
Abstract:
To investigate the mechanisms of receptor-mediated regulation of heart muscle contraction, we developed a tension-recording system using beta-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 microM isoproterenol plus 100 microM 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 microM metoprolol or 1 mM guanosine 5'-O-(2-thiodiphosphate) (GDPbetaS). Phenylephrine (10 microM) plus 100 microM GTP increased the Ca2+ sensitivity (pCa50; from 6.12 to 6. 28), and this effect was blocked by 1 microM phentolamine or 1 mM GDPbetaS. After Ca2+ loading into the SR, 10 microM phenylephrine plus 100 microM GTP in a low-ethylene- glycol-bis(beta-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA, 0. 1 mM) relaxing solution induced oscillatory contractions that were attenuated by either 1 microM phentolamine or pre-treatment with 10 microM inositol 1,4,5-trisphosphate. Our results demonstrate that beta1-adrenergic stimulation decreases myofibrillar Ca2+ sensitivity and that alpha1-adrenergic stimulation both increases the Ca2+ sensitivity and activates Ca2+ release from the agonist-sensitive SR through GTP-binding protein-related mechanisms.
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