Mitochondrial Ca2+ uptake increases Ca2+ release from inositol 1,4,5-trisphosphate receptor clusters in smooth muscle cells

Abstract

Smooth muscle activities are regulated by inositol 1,4,5-trisphosphate (InsP3)-mediated increases in cytosolic Ca2+ concentration ([Ca2+]c). Local Ca2+ release from an InsP3 receptor (InsP3R) cluster present on the sarcoplasmic reticulum is termed a Ca2+ puff. Ca2+ released via InsP3R may diffuse to adjacent clusters to trigger further release and generate a cellwide (global) Ca2+ rise. In smooth muscle, mitochondrial Ca2+ uptake maintains global InsP 3-mediated Ca2+ release by preventing a negative feedback effect of high [Ca2+] on InsP3R. Mitochondria may regulate InsP3-mediated Ca2+ signals by operating between or within InsP3R clusters. In the former mitochondria could regulate only global Ca2+ signals, whereas in the latter both local and global signals would be affected. Here whether mitochondria maintain InsP 3-mediated Ca2+ release by operating within (local) or between (global) InsP3R clusters has been addressed. Ca2+ puffs evoked by localized photolysis of InsP3 in single voltage-clamped colonic smooth muscle cells had amplitudes of 0.5-4.0 F/F 0, durations of -112 ms at half-maximum amplitude, and were abolished by the InsP3R inhibitor 2-aminoethoxydiphenyl borate. The protonophore carbonyl cyanide 3-chloropheylhydrazone and complex I inhibitor rotenone each depolarized ΔψM to prevent mitochondrial Ca2+ uptake and attenuated Ca2+ puffs by -66 or -60%, respectively. The mitochondrial uniporter inhibitor, RU360, attenuated Ca2+ puffs by -62%. The "fast" Ca2+ chelator 1,2-bis(o-aminophenoxy) ethane-N,N,N′,N′-tetraacetic acid acted like mitochondria to prolong InsP3-mediated Ca2+ release suggesting that mitochondrial influence is via their Ca2+ uptake facility. These results indicate Ca2+ uptake occurs quickly enough to influence InsP3R communication at the intra-cluster level and that mitochondria regulate both local and global InsP3-mediated Ca2+ signals. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.