title: Ca(2+) clearance by plasmalemmal NCLX, Li(+)-permeable Na(+)/Ca(2+) exchanger, is required for the sustained exocytosis in rat insulinoma INS-1 cells.
abstract: Na(+)/Ca(2+) exchangers are key players for Ca(2+) clearance in pancreatic β-cells, but their molecular determinants and roles in insulin secretion are not fully understood. In the present study, we newly discovered that the Li(+)-permeable Na(+)/Ca(2+) exchangers (NCLX), which were known as mitochondrial Na(+)/Ca(2+) exchangers, contributed to the Na(+)-dependent Ca(2+) movement across the plasma membrane in rat INS-1 insulinoma cells. Na(+)/Ca(2+) exchange activity by NCLX was comparable to that by the Na(+)/Ca(2+) exchanger, NCX. We also confirmed the presence of NCLX proteins on the plasma membrane using immunocytochemistry and cell surface biotinylation experiments. We further investigated the role of NCLX on exocytosis function by measuring the capacitance increase in response to repetitive depolarization. Small interfering (si)RNA-mediated downregulation of NCLX did not affect the initial exocytosis, but significantly suppressed sustained exocytosis and recovery of exocytosis. XIP (NCX inhibitory peptide) or Na(+) replacement for inhibiting Na(+)-dependent Ca(2+) clearance also selectively suppressed sustained exocytosis. Consistent with the idea that sustained exocytosis requires ATP-dependent vesicle recruitment, mitochondrial function, assessed by mitochondrial membrane potential (ΔΨ), was impaired by siNCLX or XIP. However, depolarization-induced exocytosis was hardly affected by changes in intracellular Na(+) concentration, suggesting a negligible contribution of mitochondrial Na(+)/Ca(2+) exchanger. Taken together, our data indicate that Na(+)/Ca(2+) exchanger-mediated Ca(2+) clearance mediated by NCLX and NCX is crucial for optimizing mitochondrial function, which in turn contributes to vesicle recruitment for sustained exocytosis in pancreatic β-cells.
authors.affiliation: Department of Physiology and Biomembrane Plasticity Research Center, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul, 110-799, Republic of Korea.
authors.forename: Young-Eun
authors.initials: YE
authors.lastname: Han
authors.affiliation: Department of Physiology and Biomembrane Plasticity Research Center, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul, 110-799, Republic of Korea.
authors.forename: Shin-Young
authors.initials: SY
authors.lastname: Ryu
authors.affiliation: Department of Physiology and Biomembrane Plasticity Research Center, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul, 110-799, Republic of Korea.
authors.forename: Sun-Hyun
authors.initials: SH
authors.lastname: Park
authors.affiliation: Department of Physiology and Biomembrane Plasticity Research Center, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul, 110-799, Republic of Korea.
authors.forename: Kyu-Hee
authors.initials: KH
authors.lastname: Lee
authors.affiliation: Department of Physiology and Biomembrane Plasticity Research Center, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul, 110-799, Republic of Korea.
authors.forename: Suk-Ho
authors.initials: SH
authors.lastname: Lee
authors.affiliation: Department of Physiology and Biomembrane Plasticity Research Center, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul, 110-799, Republic of Korea. wonkyung@snu.ac.kr.
authors.forename: Won-Kyung
authors.initials: WK
authors.lastname: Ho
mesh_terms: D000200:Action Potentials; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D005089:Exocytosis; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000378:metabolism*; D008094:Lithium / Q000378:metabolism*; D053078:Membrane Potential, Mitochondrial; D051379:Mice; D008810:Mice, Inbred C57BL; D010455:Peptides / Q000494:pharmacology; D051381:Rats; D019831:Sodium-Calcium Exchanger / Q000235:genetics / Q000378:metabolism*
chemical_list: D010455:Peptides; D019831:Sodium-Calcium Exchanger; C110374:sodium-calcium exchanger 1; C066745:exchanger inhibitory peptide; D008094:Lithium; D002118:Calcium
keywords: Ca2+ transport; Capacitance; Exocytosis; Na+/Ca2+ exchanger; Pancreatic β-cell