Fig. 5.

Mutation A454T decreases secretion efficiency. (A) Currents from two MPC 9/3L-AH cells expressing either WT (Upper) or A454T channels (Lower) in response to a train of five successive 200-ms depolarizing voltage steps to +20 mV delivered at 20 Hz. Arrows indicate the 0 current level. (B) Capacitance traces, plotted as a function of time, from the same cells showed in A. WT (○) and A454T (•) P/Q-expressing cells were stimulated at 20 Hz as described. (C) Amperometric recording from a MPC 9/3L-AH cell loaded with dopamine during the application of a local puff of a depolarizing high-K⁺ solution. Note the synchronous release of dopamine after the onset of the stimulus. (Inset) Higher time-scale resolution of the amperometric spike indicated with an asterisk. (D) Averaged data for Ca²⁺ influx normalized by the whole-cell capacitance [Q_Ca density (pC/pF)] elicited by the first depolarizing pulse, the first two depolarizing pulses, and all five depolarizing pulses (as indicated) under three different intracellular Ca²⁺-buffering conditions in WT (EGTA 0.1 mM, n = 8; EGTA 1 mM, n = 21; EGTA 5 mM, n = 7) and A454T MPC 9/3L-AH transfected cells (EGTA 0.1 mM, n = 10; EGTA 1 mM, n = 21; EGTA 5 mM, n = 7). (E) Exocytosis [ΔCm (fF)] normalized as a function of Ca²⁺ entry [Q_Ca density (pC/pF)] corresponding to the conditions described in D. *P < 0.05. (F) Averaged data for Ca²⁺ influx normalized by cell size [Q_Ca density (pC/pF)] elicited by the first depolarizing pulse, the first two depolarizing pulses, and all five depolarizing pulses (as indicated) under intermediate intracellular Ca²⁺-buffering conditions (1 mM EGTA) in MPC 9/3L-AH cells transfected with either the human WT (hWT, n = 12) or human A454T (hA454T, n = 7) P/Q channel. (G) Exocytosis [ΔCm (fF)] normalized as a function of Ca²⁺ entry [Q_Ca density (pC/pF)] corresponding to the conditions described in F. *P < 0.05.