Fig. 2. (A) Activation of the channel by IP₃, but not by the biologically inactive enantiomer L-IP₃. After the formation of a lipid bilayer membrane across the aperture of the bilayer chambers, vesicles are added to the cis chamber. Vesicle fusion events with the bilayer are monitored by observing the appearance of channel activity (a), in this case at an applied voltage of +60 mV. The cis solution (1 M KCl, 10 mM CaCl₂, 200 mM MOPS/BTP, pH 7.2) is replaced with 1 mM EGTA, 200 mM MOPS/BTP, pH 7.2 to remove all permeant ions except Ca²⁺. Trans-solution remained the same (50 mM Ca(OH)₂, 200 mM MOPS/BTP, pH 7.2). No spontaneous Ca²⁺ channel activity is present after wash out (+60 mV clamp) (b). To demonstrate the presence of IP₃-activated Ca²⁺ channel activity, the receptor agonist D-IP₃ is added to the cis chamber. Addition of IP₃ induces Ca²⁺ channel activity, with a conductance of about 10 pS (+60 mV clamp) (c). Channel activity is not present after removal of D-IP₃ by wash-out (+40 mV clamp) (d). Ca²⁺ channels are not activated by addition of L-IP₃ (+40 mV clamp) (e). The level marked closed on current traces represents zero current. The bars represent the current in pA (vertical) and time in seconds (horizontal). (B) The small conductance Ca²⁺ channel is inhibited by 2-APB but not by TMB-8. IP₃ activates a small conductance Ca²⁺ channel (+50 mV clamp) (a). TMB-8 (200 µM) does not inhibit the channel (+70 mV clamp) (b), but 2-APB (25 µM) does (+50 mV clamp) (c).