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Fig. 5. Functional properties of terminal differentiated hmNSC. (A-B) Dopamine production and release was measured in hmNSCs differentiated using the neuronal induction protocol for 14 days. (A) Representative chromatograms of HPLC-ECD determination of dopamine in medium conditioned for 3 days (left) and extracellular buffer with 56 mM KCl conditioned for 45 minutes (right). (B) Quantification of dopamine in medium conditioned for 3 days (left), in extracellular buffer conditioned for 45 minutes (center), and in extracellular buffer with 56 mM KCl conditioned for 45 minutes (right). #, P<0.05 when compared to extracellular buffer dopamine levels (paired t-test). (C,D) Electrophysiological recordings on hmNSCs differentiated using the glial induction protocol. For voltage-clamp measurements, cells were held at –80 mV and hyperpolarized or depolarized in 10 mV steps between –160 and +70 mV. (C) Example of a sustained outward current shown without and with (inset) leak subtraction using a –P/4 protocol (left). Current-voltage relationship of the normalized outward currents recorded with leak subtraction (n=7, right panel). (D) Example of an inward current without and with leak subtraction (inset: only currents for depolarizing steps to –40, –20, 0, 20 and 60 mV are shown) (left). Peak current-voltage relationship for the same cell with leak subtraction (right panel). The line represents a fit to the following equation: I(V)/I_max= g(V–V_rev)/1+exp[(V–V_0.5)/k_V], where I (I_max) is the (maximum) membrane current; g, the maximum conductance; V, the applied voltage; V_rev, the reversal potential for Na⁺; V_0.5, the potential of half-maximal activation and k_V, a slope factor.

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