Abstract
In arterial myocytes the Ca2+ mobilizing messenger NAADP evokes spatially restricted Ca2+ bursts from a lysosome-related store that are subsequently amplified into global Ca2+ waves by Ca2+-induced Ca2+-release from the sarcoplasmic reticulum (SR) via ryanodine receptors (RyRs). Lysosomes facilitate this process by forming clusters that co-localize with a subpopulation of RyRs on the SR. We determine here whether RyR subtypes 1, 2 or 3 selectively co-localize with lysosomal clusters in pulmonary arterial myocytes using affinity purified specific antibodies. The density of: (1) alpha lgP120 labelling, a lysosome-specific protein, in the perinuclear region of the cell (within 1.5 mu m of the nucleus) was similar to 4-fold greater than in the sub-plasmalemmal (within 1.5 mu m of the plasma membrane) and similar to 2-fold greater than in the extra-perinuclear (remainder) regions; (2) RyR3 labelling within the perinuclear region was similar to 4- and similar to 14-fold greater than that in the extra-perinuclear and sub-plasmalemmal regions, and similar to 2-fold greater than that for either RyR1 or RyR2; (3) despite there being no difference in the overall densities of fluorescent labelling of lysosomes and RyR subtypes between cells, co-localization with alpha lgp120 labelling within the perinuclear region was similar to 2-fold greater for RyR3 than for RyR2 or RyR1; (4) co-localization between alpha lgp120 and each RyR subtype declined markedly outside the perinuclear region. Furthermore, selective block of RyR3 and RyR1 with dantrolene (30 mu M) abolished global Ca2+ waves but not Ca2+ bursts in response to intracellular dialysis of NAADP (10 nM). We conclude that a subpopulation of lysosomes cluster in the perinuclear region of the cell and form junctions with SR containing a high density of RyR3 to comprise a trigger zone for Ca2+ signalling by NAADP. (C) 2007 Elsevier Ltd. AR rights reserved.
Original language | English |
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Pages (from-to) | 190-201 |
Number of pages | 12 |
Journal | Cell Calcium |
Volume | 44 |
Issue number | 2 |
Early online date | 11 Jan 2008 |
DOIs | |
Publication status | Published - Aug 2008 |
Keywords
- lysosomes
- ryanodine receptors
- NAADP
- CA2+ release channel
- adenine-dinucleotide phosphate
- functional-characterization
- sarcoplasmic-reticulum
- 2-pool mechanism
- cells
- stores