FIG. 1.

Primary and oscillatory calcium signaling responses of hM3Dq-ATDC5 cells subjected to once-daily 500 nM CNO administration during long-term stimulatory culture. (A) CNO-evoked primary Ca²⁺ responses remained largely unchanged across 14 days of daily stimulatory culture; however, oscillatory Ca²⁺ signaling behaviors changed over time. The number of cells exhibiting 2+ peaks significantly increased through day 7 of culture (compared with days 1 and 14), whereas extensive oscillatory behaviors significantly diminished at day 14; the number of cells exhibiting 5+ peaks remained stable through day 7 (at ∼28–30%), then dropped to ∼11% at day 14; and the number of cells exhibiting 9+ peaks was suppressed from day 3 onward, dropping from ∼14% to <5%. (B) The average number of oscillations per cell gradually diminished through day 7 of culture, dropping from 4.2 peaks/cell to ∼3.5 peaks, then to ∼2.2 peaks/cell at day 14. *p < 0.05, **p < 0.01, ***p < 0.001; one-way ANOVA with Tukey's post hoc test. CNO, clozapine N-oxide.