On the mechanism of 4-aminopyridine action on the cloned mouse brain potassium channel mKv1.1
| Autor: | Brian D. Robertson, Gary J. Stephens, J C Garratt, David G. Owen |
|---|---|
| Rok vydání: | 1994 |
| Předmět: |
Patch-Clamp Techniques
Potassium Channels Physiology Intracellular pH Molecular Sequence Data Aminopyridines CHO Cells Transfection Membrane Potentials Mice Cricetulus Cricetinae Potassium Channel Blockers medicine Extracellular Animals Channel blocker Amino Acid Sequence Patch clamp 4-Aminopyridine Cloning Molecular Brain Chemistry Chemistry Potassium channel blocker Hydrogen-Ion Concentration Molecular biology Peptide Fragments Potassium channel Mechanism of action medicine.symptom Ion Channel Gating Research Article medicine.drug |
| Zdroj: | The Journal of Physiology. 477:187-196 |
| ISSN: | 0022-3751 |
| DOI: | 10.1113/jphysiol.1994.sp020183 |
| Popis: | 1. This study used the whole-cell patch clamp technique to investigate the mechanism of action of the K+ channel blocker 4-aminopyridine (4-AP) on the cloned K+ channel mouse Kv1.1 (mKv1.1) expressed in Chinese hamster ovary cells. 2. Cells transfected with mKv1.1 expressed a non-inactivating, delayed rectifier-type K+ current. 4-AP induced a dose-, voltage- and use-dependent block of mKv1.1. 3. 4-AP blockade of mKv1.1 was similar whether 4-AP was administered extracellularly (IC50 = 147 microM) or intracellularly (IC50 = 117 microM). 4. Inclusion of the first twenty amino acids of the N-terminus sequence of the Shaker B K+ channel ('inactivation peptide') in the patch electrode transformed mKv1.1 into a rapidly inactivating current. The time constant of decay for the modified current was dependent on the concentration of inactivation peptide, and under these conditions extracellular 4-AP had a reduced potency (IC50 values of 471 and 537 microM for 0.5 and 2 mg ml-1 inactivation peptide, respectively). 5. A permanently charged analogue of 4-AP, 4-aminopyridine methiodide (4-APMI), was found to block mKv1.1 when applied inside the cell, but was without effect when administered externally. 6. Decreasing the intracellular pH (pHi) to 6.4 caused an increase in 4-AP potency (IC50 = 76 microM), whereas at pHi 9.0, the 4-AP potency fell (IC50 = 295 microM). Conversely, increasing extracellular pH (pHo) to 9.0 caused an increase in 4-AP potency (IC50 = 93 microM), whereas at pHo 6.4, 4-AP potency decreased (IC50 = 398 microM). 7. Taken together, these findings support the hypotheses that the uncharged form of 4-AP crosses the membrane, and that it is predominantly the cationic form which acts on mKv1.1 channels intracellularly, possibly at or near to the binding site for the inactivation peptide. |
| Databáze: | OpenAIRE |
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