Wide long lasting perinuclear Ca 2+ release events generated by an interaction between ryanodine and IP 3 receptors in canine Purkinje cells

Abstract

The purpose of this study was to determine whether IP 3Rs contribute to the generation of wide long lasting perinuclear Ca 2+ release events in canine Purkinje cells. Spontaneous Ca 2+ release events (elevations of basal [Ca 2+] equivalent to F/F 0 3.4SD over F 0) were imaged using Fluo-4AM and 2D confocal microscope. Only cells free of Ca 2+ waves were analyzed. Subsarcolemmal region (SSL) was defined as 5 μm from cell edges. Core was the remaining cell. The majority of events (94%, 0.0035 ± 0.0007 events (ev)/μm 2/s, N = 34 cells) were detected within a single frame (typical events, TE). However, a subpopulation (6.0%, 0.00022 ± 0.00005 ev/μm 2/s, N = 41 cells: wide long lasting events, WLE) lasted for several frames, showed a greater spatial extent (51.0 ± 3.9 vs. TE 9.0 ± 0.3 μm 2, P < 0.01) and higher amplitude (F/F 0 1.38 ± 0.02 vs. TE 1.20 ± 0.003, P < 0.01). WLE event rate was increased by phenylephrine (10 μM, P < 0.01), inhibited by 2APB and U73122 ( P < 0.05), and abolished by tetracaine (1 mM) and ryanodine (100 μM). While SSL WLEs were scattered randomly, Core WLEs ( n = 69 events) were predominantly distributed longitudinally 18.2 ± 1.6 μm from the center of nuclei. Immunocytochemistry showed that IP 3R1s were located not only at SSL region but also near both ends of nucleus overlapping with RyRs. In Purkinje cells, wide long lasting Ca 2+ release events occur in SSL and in specific perinuclear regions. They are likely due to RyRs and IP 3R1s evoked Ca 2+ release and may play a role in Ca 2+ dependent nuclear processes.