Why do Potassium Currents "run-up" in whole cell patch clamp?

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Morgado
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Why do Potassium Currents "run-up" in whole cell patch clamp?

I am studying potassium currents in A7r5 cells (vascular smooth muscle cells from rat aorta) using whole-cell patch clamp.
After whole-cell configuration was acquired, a “run-up” of potassium currents ( from 200 pA to 1500 pA or often much more, 3000 pA or even 5000 pA) often occurred during 5-10 minutes. Anyone knows why this happens? Any papers about this run-up?
Thanks for any help.
Morgado

Fraser Moss
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Are you including ATP in your

Are you including ATP in your patch pipette?
There are examples of ATP inhibited potassium currents (IK) in coronary smooth muscle cells.
In the case below, patching with ATP free solutions results in run-up of the potassium current.
Pflugers Arch. 1994 May;427(1-2):110-20.
Characterization of the ATP-inhibited K+ current in canine coronary smooth muscle cells.
Xu X, Lee KS.
Cardiovascular Diseases Research, Upjohn Laboratories, Kalamazoo, Michigan 49007.
Intracellular adenosine triphosphate (ATP)-inhibited K+ currents (IK, ATP) in canine coronary artery smooth muscle cells were characterized in the whole-cell configuration using the suction pipette method. Cells dialysed internally with solutions containing 5 mM ATP (ATPi) showed little detectable whole-cell current at potentials more negative than -30 mV. However, cells dialysed with ATPi-free solutions developed a time- and voltage-independent current which reached a maximum of 132 +/- 25 pA at -40 mV about 10 min following patch rupture. After "run-up", the current showed little "run-down". Concentration-dependent inhibition by ATPi yielded an inhibition constant (Ki) of 350 microM and a Hill coefficient of 2.3. In ATPi-free solutions, the large current at -40 mV was reduced by glibenclamide with a Ki of 20 nM and a Hill coefficient of 0.95. Conversely, in 1 mM ATPi solutions, the small current at -40 mV was increased by P-1075 from 8 +/- 2 pA to 143 +/- 33 pA, with a dissociation constant (Kd) of 0.16 microM and a Hill coefficient of 1.7. The effect of P-1075 was antagonized by glibenclamide. Maximal current density elicited by either ATPi depletion or external application of the channel opener P-1075 was similar with slope conductances of 81 +/- 10 pS/pF and 76 +/- 13 pS/pF respectively in the potential range of -90 to -40 mV. External Ca2+ had no effect on this current. Finally, in 1 mM ATPi, 20 and 50 microM adenosine increased the current slope conductance by 36 +/- 15% and 73 +/- 10% respectively between -90 to -40 mV. The IK, ATP, although very small in these cells, was extremely effective in causing membrane potential hyperpolarization.
PMID: 8058459
It is comonly thought that run-up and run-down phenomena are caused by changes in the basal levels of phosphorylation of potassium channels (which phenomemon happens depends on the kinase(s) involved and the contents of your patch pipette).
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Fraser Moss
Fraser Moss's picture
here are 7 papers that

here are 7 papers that example potassium channel run-up
Search: postassium channel "run-up" at http://www.ncbi.nlm.nih.gov/pubmed/
1: Collins A, Larson MK.
K(ir)2.2 inward rectifier potassium channels are inhibited by an endogenous factor in Xenopus oocytes independently from the action of a mitochondrial uncoupler.
J Cell Physiol. 2008 Nov 18.
PMID: 19016473

2: Hong SJ, Chang CC.
Inhibition of quantal release from motor nerve by wortmannin.
Br J Pharmacol. 1999 Sep;128(1):142-8.
PMID: 10498845

3: Inoue M, Sakamoto Y, Yano A, Imanaga I.
Cyanide suppression of inwardly rectifying K+ channels in guinea pig chromaffin cells involves dephosphorylation.
Am J Physiol. 1997 Jul;273(1 Pt 1):C137-47.
PMID: 9252451

4: Donnelly DF.
Modulation of glomus cell membrane currents of intact rat carotid body.
J Physiol. 1995 Dec 15;489 ( Pt 3):677-88.
PMID: 8788933

5: Xu X, Lee KS.
Characterization of the ATP-inhibited K+ current in canine coronary smooth muscle cells.
Pflugers Arch. 1994 May;427(1-2):110-20.
PMID: 8058459

6: Tiaho F, Nargeot J, Richard S.
Repriming of L-type calcium currents revealed during early whole-cell patch-clamp recordings in rat ventricular cells.
J Physiol. 1993 Apr;463:367-89.
PMID: 8246188

7: Duchatelle-Gourdon I, Lagrutta AA, Hartzell HC.
Effects of Mg2+ on basal and beta-adrenergic-stimulated delayed rectifier potassium current in frog atrial myocytes.
J Physiol. 1991 Apr;435:333-47.
PMID: 1685188

Morgado
Morgado's picture
Yes, I am including 5 mM ATP

Yes, I am including 5 mM ATP in the patch pipette.
I think the run up potassium currents are not due to KATP channels, because they are not inhibited by glibenclamide. And I am sure they are not leak currents.
> It is comonly thought that run-up and run-down phenomena are caused by changes in the basal levels of phosphorylation of potassium channels (which phenomemon happens depends on the kinase(s) involved and the contents of your patch pipette)
I think I should try to add a kinase inhibitor (PKA or PKG inhibitors) to the pipette solution to see if the run up disappears.
Thanks for your help