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Synergistic activation of G protein-gated inwardly rectifying potassium channels by the betagamma subunits of G proteins and Na(+) and Mg(2+) ions.

Petit-Jacques J, Sui JL, Logothetis DE - J. Gen. Physiol. (1999)

Bottom Line: Native and recombinant G protein-gated inwardly rectifying potassium (GIRK) channels are directly activated by the betagamma subunits of GTP-binding (G) proteins.The presence of phosphatidylinositol-bis-phosphate (PIP(2)) is required for G protein activation.At high levels of PIP(2), synergistic interactions among Na(+), Mg(2+), and G(betagamma) subunits resulted in severalfold stimulated levels of channel activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Mount Sinai School of Medicine of the New York University, New York, New York 10029, USA.

ABSTRACT
Native and recombinant G protein-gated inwardly rectifying potassium (GIRK) channels are directly activated by the betagamma subunits of GTP-binding (G) proteins. The presence of phosphatidylinositol-bis-phosphate (PIP(2)) is required for G protein activation. Formation (via hydrolysis of ATP) of endogenous PIP(2) or application of exogenous PIP(2) increases the mean open time of GIRK channels and sensitizes them to gating by internal Na(+) ions. In the present study, we show that the activity of ATP- or PIP(2)-modified channels could also be stimulated by intracellular Mg(2+) ions. In addition, Mg(2+) ions reduced the single-channel conductance of GIRK channels, independently of their gating ability. Both Na(+) and Mg(2+) ions exert their gating effects independently of each other or of the activation by the G(betagamma) subunits. At high levels of PIP(2), synergistic interactions among Na(+), Mg(2+), and G(betagamma) subunits resulted in severalfold stimulated levels of channel activity. Changes in ionic concentrations and/or G protein subunits in the local environment of these K(+) channels could provide a rapid amplification mechanism for generation of graded activity, thereby adjusting the level of excitability of the cells.

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Related in: MedlinePlus

Mg2+ ions gate the ATP-modified GIRK channels. NPo, NFo, and MTo plots (bin = 5 s) of GIRK channel activity in an inside-out patch excised from an oocyte expressing GIRK1/GIRK4. 10 mM Mg2+, 2.5 mM MgATP, and 20 mM Na+ were applied via the bath as indicated by the bars. The membrane was clamped at −80 mV and 5 μM acetylcholine was present in the pipette. NPo, NFo, and MTo in 10 mM Mg2+ were calculated such that the amplitude levels were set to match the reduced amplitude of the channel openings in 10 mM Mg2+.
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Figure 3: Mg2+ ions gate the ATP-modified GIRK channels. NPo, NFo, and MTo plots (bin = 5 s) of GIRK channel activity in an inside-out patch excised from an oocyte expressing GIRK1/GIRK4. 10 mM Mg2+, 2.5 mM MgATP, and 20 mM Na+ were applied via the bath as indicated by the bars. The membrane was clamped at −80 mV and 5 μM acetylcholine was present in the pipette. NPo, NFo, and MTo in 10 mM Mg2+ were calculated such that the amplitude levels were set to match the reduced amplitude of the channel openings in 10 mM Mg2+.

Mentions: In certain experiments, 5 mM MgATP increased the activity of the GIRK channels in the absence of Na+ ions (e.g., Sui et al. 1996). 5 mM MgATP in the solution corresponds to a free Mg2+ ion concentration of ∼2.1 mM (Vivaudou et al. 1991). This observation prompted us to test whether Mg2+ ions alone were able to gate the channel that had been modified by ATP. In Fig. 3, in an inside-out patch from an oocyte coexpressing the channel subunits GIRK1/GIRK4, Mg2+ ions (10 mM) had no significant effects on channel activity in the absence of ATP. After washout of Mg2+, the channel was activated by the combination of MgATP (2.5 mM; corresponding to ∼1.1 mM free Mg2+) and Na+ ions (20 mM). MgATP application was maintained and, upon withdrawal of Na+ ions, channel activity became comparable to basal levels. Application of Mg2+ ions (10 mM), in the continuous presence of MgATP (2.5 mM), increased channel activity to levels similar to those obtained with Na+ ions (as confirmed by sequential application of 10 mM of each of the ions at the end of the experiment). Withdrawal of Mg2+ ions caused channel activity to return to basal levels (n = 3). The MTo of the channel activity was increased from ∼1 to ∼2 ms by the application of MgATP, but was not further modified during the gating by Mg2+ or Na+ ions.


Synergistic activation of G protein-gated inwardly rectifying potassium channels by the betagamma subunits of G proteins and Na(+) and Mg(2+) ions.

Petit-Jacques J, Sui JL, Logothetis DE - J. Gen. Physiol. (1999)

Mg2+ ions gate the ATP-modified GIRK channels. NPo, NFo, and MTo plots (bin = 5 s) of GIRK channel activity in an inside-out patch excised from an oocyte expressing GIRK1/GIRK4. 10 mM Mg2+, 2.5 mM MgATP, and 20 mM Na+ were applied via the bath as indicated by the bars. The membrane was clamped at −80 mV and 5 μM acetylcholine was present in the pipette. NPo, NFo, and MTo in 10 mM Mg2+ were calculated such that the amplitude levels were set to match the reduced amplitude of the channel openings in 10 mM Mg2+.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2230539&req=5

Figure 3: Mg2+ ions gate the ATP-modified GIRK channels. NPo, NFo, and MTo plots (bin = 5 s) of GIRK channel activity in an inside-out patch excised from an oocyte expressing GIRK1/GIRK4. 10 mM Mg2+, 2.5 mM MgATP, and 20 mM Na+ were applied via the bath as indicated by the bars. The membrane was clamped at −80 mV and 5 μM acetylcholine was present in the pipette. NPo, NFo, and MTo in 10 mM Mg2+ were calculated such that the amplitude levels were set to match the reduced amplitude of the channel openings in 10 mM Mg2+.
Mentions: In certain experiments, 5 mM MgATP increased the activity of the GIRK channels in the absence of Na+ ions (e.g., Sui et al. 1996). 5 mM MgATP in the solution corresponds to a free Mg2+ ion concentration of ∼2.1 mM (Vivaudou et al. 1991). This observation prompted us to test whether Mg2+ ions alone were able to gate the channel that had been modified by ATP. In Fig. 3, in an inside-out patch from an oocyte coexpressing the channel subunits GIRK1/GIRK4, Mg2+ ions (10 mM) had no significant effects on channel activity in the absence of ATP. After washout of Mg2+, the channel was activated by the combination of MgATP (2.5 mM; corresponding to ∼1.1 mM free Mg2+) and Na+ ions (20 mM). MgATP application was maintained and, upon withdrawal of Na+ ions, channel activity became comparable to basal levels. Application of Mg2+ ions (10 mM), in the continuous presence of MgATP (2.5 mM), increased channel activity to levels similar to those obtained with Na+ ions (as confirmed by sequential application of 10 mM of each of the ions at the end of the experiment). Withdrawal of Mg2+ ions caused channel activity to return to basal levels (n = 3). The MTo of the channel activity was increased from ∼1 to ∼2 ms by the application of MgATP, but was not further modified during the gating by Mg2+ or Na+ ions.

Bottom Line: Native and recombinant G protein-gated inwardly rectifying potassium (GIRK) channels are directly activated by the betagamma subunits of GTP-binding (G) proteins.The presence of phosphatidylinositol-bis-phosphate (PIP(2)) is required for G protein activation.At high levels of PIP(2), synergistic interactions among Na(+), Mg(2+), and G(betagamma) subunits resulted in severalfold stimulated levels of channel activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Mount Sinai School of Medicine of the New York University, New York, New York 10029, USA.

ABSTRACT
Native and recombinant G protein-gated inwardly rectifying potassium (GIRK) channels are directly activated by the betagamma subunits of GTP-binding (G) proteins. The presence of phosphatidylinositol-bis-phosphate (PIP(2)) is required for G protein activation. Formation (via hydrolysis of ATP) of endogenous PIP(2) or application of exogenous PIP(2) increases the mean open time of GIRK channels and sensitizes them to gating by internal Na(+) ions. In the present study, we show that the activity of ATP- or PIP(2)-modified channels could also be stimulated by intracellular Mg(2+) ions. In addition, Mg(2+) ions reduced the single-channel conductance of GIRK channels, independently of their gating ability. Both Na(+) and Mg(2+) ions exert their gating effects independently of each other or of the activation by the G(betagamma) subunits. At high levels of PIP(2), synergistic interactions among Na(+), Mg(2+), and G(betagamma) subunits resulted in severalfold stimulated levels of channel activity. Changes in ionic concentrations and/or G protein subunits in the local environment of these K(+) channels could provide a rapid amplification mechanism for generation of graded activity, thereby adjusting the level of excitability of the cells.

Show MeSH
Related in: MedlinePlus