FIG. 2.

Temporal dynamics of CNO-induced Ca²⁺ oscillations in hM3Dq-ATDC5 cells during long-term stimulatory culture. (A) Stimulation with 500 nM CNO at day 1 of culture elicited mean primary Ca²⁺ peak intensities of ∼33 a.u., which quickly dropped to ∼15 a.u. at the second peak and decayed linearly to ∼5 a.u. for later peaks. At day 3 of culture and beyond, significantly reduced primary peak intensities (∼13–18 a.u.; p < 0.05, one-way ANOVA with Tukey's post hoc test), which decayed to 2–3 a.u. at later peaks, were observed. (B) Peak-to-peak periods exhibited similar dynamics across all culture timepoints; the first peak interval (i.e., between the primary and initial secondary peak) was always longer than all subsequent intervals, which decayed to a plateau between 25 and 45 s. The initial inter-peak period decreased—non-significantly—from ∼160 s at day 1 to ∼130–135 s at days 3 and 7, and significantly to ∼105 s at day 14 (p < 0.01, one-way ANOVA with Tukey's post hoc test). (C) Mean Ca²⁺ oscillation height decreased significantly from ∼11 a.u. at day 1 to ∼5 a.u. at day 3 and beyond. (D) Mean oscillation peak-to-peak period gradually decreased from ∼116 s at day 1 to ∼108 s at day 3, then to ∼105 s at day 7 and ∼85 s at day 14. *p < 0.05, **p < 0.01, ***p < 0.001; one-way ANOVA with Tukey's post hoc test. The statistical comparisons indicated in red fell just short of significance, p < 0.10.