Fig. 3.

C-terminal truncation on hPanx1 indeed allows Ca²⁺ influx through hemichannels. (A) Cx45/Panx1 KO HeLa cells were cotransfected with fl-hPanx1 and Lck-GCaMP3 and Ca²⁺ transients were evaluated by fluorescence live cell time-lapse in response to pH 8.5. ATP was used for activating purinergic ionotropic receptors, n = 3. (B) Similarly, Lck-GCamP3 and Δ371hPanx1 were transfected, showing a rapid increase in GCamP3 signal when hemichannels were activated with alkaline pH, which was blocked with CBX, n = 3. (C and D) By using Fura-2, previous results were confirmed, showing that alkaline pH only allows an increase in Fura ratio in truncated hPanx1 transfectants, but not with fl-hPanx1 isoform. CBX partially inhibited the calcium signal evaluated with Fura-2. All recordings were made in the presence of 1 mM Na₃VO₄, a Ca²⁺ pump blocker, n = 4. (E) The area under the curve (AUC in arbitrary units: A.U.) showed a significant increase in response to alkaline pH only in Δ371hPanx1 transfectants. This response was not affected in the presence of Xestospongin C (XetC), an IP₃R antagonist but was almost completely suppressed by DCFS, n = 4. (F) There is a high correlation (R² = 0.86) between the expression level of Δ371hPanx1 and the response of hemichannels to alkaline pH recorded with Fura-2. *P ≤ 0.05, **P ≤ 0.01.