Mechanism underlying bupivacaine inhibition of G protein-gated inwardly rectifying K ⁺ channels

Abstract

Local anesthetics, commonly used for treating cardiac arrhythmias, pain, and seizures, are best known for their inhibitory effects on voltage-gated Na ؉ channels. Cardiovascular and central nervous system toxicity are unwanted side-effects from local anesthetics that cannot be attributed to the inhibition of only Na ؉ channels. Here, we report that extracellular application of the membranepermeant local anesthetic bupivacaine selectively inhibited G protein-gated inwardly rectifying K ؉ channels (GIRK:Kir3) but not other families of inwardly rectifying K ؉ channels (ROMK:Kir1 and IRK:Kir2). Bupivacaine inhibited GIRK channels within seconds of application, regardless of whether channels were activated through the muscarinic receptor or directly via coexpressed G protein G␤␥ subunits. Bupivacaine also inhibited alcohol-induced GIRK currents in the absence of functional pertussis toxin-sensitive G proteins. The mutated GIRK1 and GIRK2 (GIRK1͞2) channels containing the high-affinity phosphatidylinositol 4,5-bisphosphate (PIP2) domain from IRK1, on the other hand, showed dramatically less inhibition with bupivacaine. Surprisingly, GIRK1͞2 channels with high affinity for PIP2 were inhibited by ethanol, like IRK1 channels. We propose that membrane-permeant local anesthetics inhibit GIRK channels by antagonizing the interaction of PIP2 with the channel, which is essential for G␤␥ and ethanol activation of GIRK channels. L ocal anesthetics are commonly used for treating cardiac arrhythmias, alleviating pain, and controlling seizures (1). Accidental overdose of local anesthetics, however, produces cardiovascular and central nervous system toxicity. The initial phase of bupivacaine overdose leads to tachycardia in the heart and convulsions in the brain (2). Local anesthetics are well known for their inhibitory effects on voltage-gated Na ϩ channels (3). The suppression of voltage-gated Na ϩ channels would be expected to reduce membrane excitability. The inhibition of K ϩ channels, however, increases membrane excitability and could therefore contribute to bupivacaine-induced tachycardia and convulsions. Indeed, some local anesthetics inhibit voltage-gated K ϩ channels (4-6). The effect of local anesthetics on native inwardly rectifying K ϩ channels has been equivocal (5-8). Interestingly, several investigators have observed that a permanently charged derivative of lidocaine, QX-314, suppresses the activity of G protein-gated inwardly K ϩ currents when introduced directly into the cytoplasm of neurons (9-11). QX-314 is not clinically useful, however, because it does not cross the cell membrane. We report here that the family of G protein-gated inwardly rectifying K ϩ channels (GIRK), but not other families of inwardly rectifying K ϩ channels, are inhibited by the clinically relevant membrane-permeant local anesthetics.GIRK channels (also referred to as Kir3) are members of a family of inwardly rectifying K ϩ channels that contain seven different groups (Kir1-7) (12). Like all inwardly rectifying K ϩ channels, GIRK channels ...

Inhibition of G Protein-Activated Inwardly Rectifying K+ Channels by Various Antidepressant Drugs

“…In addition, the adenosine-induced GIRK currents were not significantly affected by intracellularly applied desipramine (90.8710.6% of pretreated control current, paired t-test, p40.1, n ¼ 5, Figure 6c), whereas the GIRK currents were significantly inhibited by intracellularly applied QX-314 as reported previously (Zhou et al, 2001;Kobayashi et al, 2003). The results, therefore, suggest that extracellular desipramine can directly inhibit GIRK channels activated by the XA1 receptors.…”

Inhibition of G Protein-Activated Inwardly Rectifying K+ Channels by Ifenprodil

“…The percentage inhibition by ifenprodil was similar to that of basally active GIRK1/2 channels (p40.05 at each concentration, Student's t-test), suggesting interaction of ifenprodil with GIRK channels but not the kOR. In addition, the U50488H-induced GIRK currents were not significantly affected by intracellularly applied ifenprodil (90.4712.7% of untreated control current, paired t-test, p40.1, n ¼ 4, Figure 5c), whereas such GIRK currents were significantly inhibited by intracellularly applied QX-314 as reported previously (Zhou et al, 2001;Kobayashi et al, 2003). The results indicate that intracellular ifenprodil could not inhibit GIRK channels and G-proteins mediated by kOR activation.…”

“…In oocytes expressing GIRK1/2 channels, the GIRK currents induced by ethanol were attenuated in the presence of ifenprodil, with an IC 50 value of 6.171.8 mM and an n H value of 0.6970.09, in a reversible manner (88.972.7% inhibition at 100 mM, n ¼ 6; Figure 6). In addition, since the carboxyl terminal domains of GIRK channels are crucial for the ethanol sensitivity of the channel (Lewohl et al, 1999;Zhou et al, 2001), we examined whether intracellular ifenprodil affects ethanol activation of GIRK channels. However, the ethanol-induced GIRK currents were not significantly affected by intracellularly applied ifenprodil (91.778.0% of untreated control current, paired t-test, p40.1, n ¼ 4, Figure 6c).…”

Molecular mechanism underlying ethanol activation of G-protein–gated inwardly rectifying potassium channels

“…Thus, alcoholinduced changes in affinity of GIRK for PIP 2 may recapitulate some of the same events induced by Gβγ binding. The cross-talk between alcohol, Gβγ, and the low affinity for PIP 2 suggest that GIRK channels have evolved to be sensitive to small molecules that either inhibit (48,49) or activate (3-5) the channel.…”