Fig. 5.

The Ca²⁺-dependence of Doc2b determines that of spontaneous release in rescued DKO neurons. (A) Typical mEPSC recordings in DKO cells rescued with GFP (control), Doc2b^WT or Doc2b^D218,220N. Spontaneous mEPSCs were recorded in network cultures of hippocampal neurons in the presence of TTX and gabazine. Increasing concentrations of extracellular Ca²⁺ were administered to the same cell. (B) Mean mEPSC frequencies ± sem in DKO cells expressing Doc2b^WT (n=10), Doc2b^D218,220N (n=26) or GFP as a control (n=21) (C) Mean mEPSC frequencies of the same constructs expressed in wildtype cells. The average mEPSC frequency varied between experiments, but our experimental design prevents confounding effects thereof (see Fig S2E) (D) Mean mEPSC frequencies in DKO cells expressing Doc2b^WT (n=35) or mutant Doc2b^K237,319E (n=31) (E) Ca²⁺-dependent liposome-binding by the isolated C2A domain of Doc2b^WT and Doc2b^D218,220N (n=6). *** (p<0.005); ** (p<0.01), * (p<0.05).