Figure 3

Ca²⁺ transients evoked by K⁺ depolarization or P2X ₃ receptors in WT and R192Q KI neurons. A, Examples of Ca²⁺ transients of trigeminal neurons evoked by KCl (20 mM, 2-s application) before (black trace) and after (red trace) application of ω-agatoxin (200 nM, 30 min). B, KI neurons show significant increase in KCl (20 mM, 2-s application) mediated Ca²⁺ transients compared to WT (*p = 0.005, n = 28 and n = 45, in WT and KI, respectively). Histograms also represent inhibition by ω-agatoxin of Ca²⁺ transients for WT (n = 14) and KI (n = 35) neurons. After ω-agatoxin responses of WT and KI neurons differ from their own controls (**p ≤ 0.001). C, Representative traces of α,β-meATP (10 μ M, 2-s application)-evoked Ca²⁺ transients before (black trace) and after (red trace) application of ω-agatoxin (200 nM, 30 min). D, Histograms show larger Ca²⁺ transients evoked by α,β-meATP (10 μ M, 2-s application) from KI (n = 26) than WT (n = 16) and neurons (*p = 0.04). Histograms also show that ω-agatoxin reduced Ca²⁺ transients of KI (n = 22) and WT (n = 9) neurons. **p ≤ 0.001 for each case. E, Microphotographs of immunofluorescence experiments depicting WT and KI trigeminal neurons in culture expressing P2X₃ receptors or Ca_V2.1 channels. Bar = 50 μ m. Histograms (right) show% of P2X₃- (top) or Ca_V2.1- (bottom) immunoreactive neurons (taking as 100% the β-tubulin III immunoreactive) (n = 5, p > 0.05 for P2X₃ receptors; n = 3, p > 0.05 for Ca_V2.1-expressing neurons). F, Histograms show% of Ca_V2.1-immunoreactive neurons (top; taken as 100%) which are immunopositive for P2X₃ (n = 7, p > 0.05) or% of P2X₃-immunoreactive neurons (bottom) which are immunopositive for Ca_V2.1 (n = 4, p > 0.05).