Figure 7. Normal Growth Medium Supports Constitutive InsP₃R Ca²⁺ Release.

(A and B) DT40-KO cells and XeB-treated HEK-293 cells have higher levels of phospho-PDH (n = 3). (C – E) DCA treatment normalizes PDH hyper-phosphorylation (C; 2 mM for 1h), enhances OCR (D; added acutely at “a”) and suppress autophagy (E; 2 mM for 1h) in KO cells to WT levels. (n = 3). (F) Effect of transient transfection (48 h) with specific or irrelevant (NT) siRNA on InsP₃R-1 expression in SH-SY5Y cells. InsP₃R-1/tubulin (n = 3) expressed relative to basal levels. (G – J) Effects of InsP₃R-1 knockdown (G and H) or XeB (10 μM, 1 h) (I and J) on autophagy and P-AMPK in SH-SY5Y cells (n=3). (K) Effects of InsP₃R-1 knockdown on basal and maximum OCR in SH-SY5Y cells (n=3). (L) Representative fluorescence recordings of Ca²⁺ release events in unstimulated SH-SY5Y cells in either Hank’s solution or normal culture media. (M) Quantification of Ca²⁺ release events in SH-SY5Y cells exposed either to Hank’s or normal media (n=3 experiments). (N) Effects of XeB (10 μM for 30 min) on spontaneous Ca²⁺ release events recorded in SH-SY5Y cells in normal media (n=3 experiments). **p < 0.01 ***p < 0.001. See also Figure S5 and Video S1.