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Inhibition of acetylcholine-activated K+ currents by U73122 is mediated by the inhibition of PIP2 -channel interaction
- Source :
- British Journal of Pharmacology. 134:1066-1072
- Publication Year :
- 2001
- Publisher :
- Wiley, 2001.
-
Abstract
- We have investigated the effect of {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122, a specific inhibitor of phospholipase C (PLC), on acetylcholine-activated K+ currents (IKACh) in mouse atrial myocytes. In perforated patch clamp mode, IKACh was activated by 10 μM acetylcholine. When atrial myocytes were pretreated with {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 or {"type":"entrez-nucleotide","attrs":{"text":"U73343","term_id":"1688125","term_text":"U73343"}}U73343, IKACh was inhibited dose-dependently (half-maximal inhibition at 0.12±0.0085 and 0.16±0.0176 μM, respectively). The current-voltage relationships for IKACh in the absence and in the presence of {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 showed that the inhibition occurred uniformly from −120 to +40 mV, indicating a voltage-independent inhibition. When {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 was applied after IKACh reached steady-state, a gradual decrease in IKACh was observed. The time course of the current decrease was well fitted to a single exponential, and the rate constant was proportional to the concentration of {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122. When KACh channels were directly activated by adding 1 mM GTPγS to the bath solution in inside-out patches, {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 (1 μM) decreased the open probability significantly without change in mean open time. When KACh channels were activated independently of G-protein activation by 20 mM Na+, open probability was also inhibited by {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122. Voltage-activated K+ currents and inward rectifying K+ currents were not affected by {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122. These findings show that inhibition by {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 and {"type":"entrez-nucleotide","attrs":{"text":"U73343","term_id":"1688125","term_text":"U73343"}}U73343 of KACh channels occurs at a level downstream of the action of Gβγ or Na+ on channel activation. The interference with phosphatidylinositol 4,5-bisphosphate (PIP2)-channel interaction can be suggested as a most plausible mechanism. Keywords: {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122, {"type":"entrez-nucleotide","attrs":{"text":"U73343","term_id":"1688125","term_text":"U73343"}}U73343, acetylcholine-activated K+ current, phospholipase C, phospholipase C inhibitor, atrial myocytes, phosphatidylinositol 4,5-bisphosphate, patch-clamp Introduction {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 has been shown to inhibit phospholipase C (PLC) at low micromolar concentrations (Smith et al., 1990; Bleasdale et al., 1990), and it is one of the most widely used PLC inhibitors. Very often the structurally related {"type":"entrez-nucleotide","attrs":{"text":"U73343","term_id":"1688125","term_text":"U73343"}}U73343 is used as negative control, since it does not inhibit PLC (Smith et al., 1990; Bleasdale et al., 1990). Recent studies, however, indicate that the action of {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 and {"type":"entrez-nucleotide","attrs":{"text":"U73343","term_id":"1688125","term_text":"U73343"}}U73343 may not be specific as originally thought, since both agents also inhibit phospholipase D activity (Bosch et al., 1998), and since {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 elicits intracellular calcium release (Muto et al., 1997). Furthermore, the non-selective actions of {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 were also shown for a variety of receptor-mediated signal transductions, such as adenosine A1-receptor (Walker et al., 1998) and histamine H1-receptor (Hughes et al., 2000). ACh-activated K+ currents (IKACh) are responsible for the inhibitory effect of cardiac function by ACh (Jan & Jan, 1997; Yamada et al., 1998). Activation of IKACh by ACh receptors (mAChR) is mediated via the pertussis toxin-sensitive G-protein. G-protein-ion channel coupling mechanisms have been widely investigated for IKACh and its molecular equivalent G-protein-gated inwardly rectifying K+ channels (GIRK), and it is now believed that the direct binding of G protein Gβγ subunits to the channel protein opens GIRK channels (Huang et al., 1995; Krapivinsky et al., 1995; Kunkel & Peralta, 1995; Inanobe et al., 1995). In addition to Gβγ subunits, GIRK channels can also be activated by Na+ ions through G-protein-independent pathway (Sui et al., 1996; 1998). Both Gβγ subunits and Na+ ion cause a stabilization of phosphatidylinositol 4,5-bisphosphate (PIP2)-channel interaction, which is absolutely required for channel opening (Huang et al., 1998). In the present study, we investigated the effects of {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 and {"type":"entrez-nucleotide","attrs":{"text":"U73343","term_id":"1688125","term_text":"U73343"}}U73343 on IKACh in mouse atrial myocytes and found that both inhibit IKACh independently of PLC inhibition. The inhibition was almost complete at concentrations similar to or below those used for PLC inhibition. The target for {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 action was investigated using inside-out patch recording, showing that the inhibition by {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 occurs even when the channels are activated by Na+ independently of G-protein activation. These results suggest that {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 inhibits KACh channels possibly by the interference with PIP2-channel interaction.
Details
- ISSN :
- 00071188
- Volume :
- 134
- Database :
- OpenAIRE
- Journal :
- British Journal of Pharmacology
- Accession number :
- edsair.doi...........1efd7620ae5622832274343f6848afeb
- Full Text :
- https://doi.org/10.1038/sj.bjp.0704347