Figure 1.

Reconstitution of α₂-AR coupling to N-type Ca²⁺ channels by expression of PTX-insensitive Gα_oA mutants. Time course of current inhibition and superimposed current traces in the absence (1 and 3) or presence (2 and 4) of 10 μM NE recorded from control neurons (no PTX) (A), control (500 ng/ml PTX, overnight) (B), PTX-pretreated neurons coexpressing Gα_oA(C351G) and Gβ₁γ₂ (C). The cDNAs encoding PTX-insensitive Gα mutants and Gβ₁γ₂ were injected directly into nuclei of SCG neurons. The Ca²⁺ current was evoked every 10 s by a double-pulse voltage protocol (see Inset in A) consisting of two identical test pulses (+10 mV from a holding potential of −80 mV) separated by a large, depolarizing conditioning pulse to +80 mV. The amplitudes of currents generated by pre- (●) and post- (○) pulses were plotted. (D) Summary of prepulse facilitation in the absence or presence of NE. Prepulse facilitation was calculated by the ratio of the postpulse to prepulse current amplitude measured isochronally at 10 ms after the start of the test pulses. (E) Summary of NE-induced Ca²⁺ current inhibition. The C-terminal cysteine residue of Gα_oA also was mutated to serine or isoleucine. Inhibition (%) was calculated by using the amplitudes of currents determined isochronally 10 ms after the start of the prepulse. In both D and E, data are presented as mean ± SEM and numbers in parentheses indicate the number of neurons tested.