Skip to main content
. 2007 Mar 15;581(Pt 2):479–493. doi: 10.1113/jphysiol.2006.123414

Figure 2. hERG current modulation by methcholine and ionomycin can be attenuated by inhibition or down-regulation of PKC.

Figure 2

A, mean hERG current in response to methacholine (Mch) in HEK-M3 cells following inhibition of PKC with 300 nm bis-1. The protocol and tail current analysis are the same as described in Fig. 1B. Bis-1 was applied for 3 min before co-application with methacholine. B, hERG current modulation is attenuated by 300 nm bis-1. Mean percentage values for tail current remaining in control conditions after 5 min (untreated), and upon application of the following: 300 nm bis-1 for 3 min; 1 μm bis-1 for 3 min; methacholine (Mch) for 2 min; 3 min perfusion with bis-1 followed by co-application of bis-1 with Mch for 2 min; 5 μm ionomycin (IM) for 2 min; 3 min perfusion with bis-1 followed by co-application of IM for 2 min. C, representative Western blots showing PKC isotype expression in hERG-HEK cells treated with 0.1% DMSO (control), 1 μm PMA, or 1 μm 4αPMA for 24 h. Control and PMA blots were performed in pairs to enable direct comparisons to be made. 4αPMA blots were performed separately. Similar results were observed in 4 other experiments. The lower panels show the blots after being stripped and re-probed for β-actin to show protein loading across all lanes. D, down-regulation of PKC isotypes abolishes hERG current modulation by Mch and IM. hERG current modulation in untreated cells and cells incubated in 1 μm PMA for 24 h (PMA). Tail current amplitudes were measured after > 2 min application of Mch, IM (5 μm) or forskolin (40 μm) and expressed as a percentage of current before test compound application. Experiments with Mch activation of muscarinic receptors were performed in HEK-M3 cells transiently expressing hERG and all other experiments were performed on hERG-HEK cells. **Significantly different from untreated (P < 0.01); ‡significantly different from Mch-only response (P < 0.01); †significantly different from IM-only response (P < 0.05). Note, for clarity not all significant differences are presented.