Selective positive modulation of the SK3 and SK2 subtypes of small conductance Ca2+-activated K+ channels.
OBJECTIVE
Positive modulators of small conductance Ca(2+)-activated K(+) channels (SK1, SK2, and SK3) exert hyperpolarizing effects that influence the activity of excitable and non-excitable cells. The prototype compound 1-EBIO or the more potent compound NS309, do not distinguish between the SK subtypes and they also activate the related intermediate conductance Ca(2+)-activated K(+) channel (IK). This paper demonstrates, for the first time, subtype-selective positive modulation of SK channels.


METHODS
Using patch clamp and fluorescence techniques we studied the effect of the compound cyclohexyl-[2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-pyrimidin-4-yl]-amine (CyPPA) on recombinant hSK1-3 and hIK channels expressed in HEK293 cells. CyPPA was also tested on SK3 and IK channels endogenously expressed in TE671 and HeLa cells.


RESULTS
CyPPA was found to be a positive modulator of hSK3 (EC(50) = 5.6 +/- 1.6 microM, efficacy 90 +/- 1.8 %) and hSK2 (EC(50) = 14 +/- 4 microM, efficacy 71 +/- 1.8 %) when measured in inside-out patch clamp experiments. CyPPA was inactive on both hSK1 and hIK channels. At hSK3 channels, CyPPA induced a concentration-dependent increase in the apparent Ca(2+)-sensitivity of channel activation, changing the EC(50)(Ca(2+)) from 429 nM to 59 nM.


CONCLUSIONS
As a pharmacological tool, CyPPA may be used in parallel with the IK/SK openers 1-EBIO and NS309 to distinguish SK3/SK2- from SK1/IK-mediated pharmacological responses. This is important for the SK2 and SK1 subtypes, since they have overlapping expression patterns in the neocortical and hippocampal regions, and for SK3 and IK channels, since they co-express in certain peripheral tissues.
Hougaard
C
C
NeuroSearch A/S, Pederstrupvej 93, Ballerup, Denmark.
Eriksen
B L
BL
Jørgensen
S
S
Johansen
T H
TH
Dyhring
T
T
Madsen
L S
LS
Strøbaek
D
D
Christophersen
P
P
D000465:Algorithms; D001562:Benzimidazoles / Q000494:pharmacology; D002460:Cell Line; D004594:Electrophysiology; D005456:Fluorescent Dyes; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects; D010091:Oximes / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D051640:Small-Conductance Calcium-Activated Potassium Channels / Q000187:drug effects*; D013793:Thallium
C507351:5,6-dichloro-1-ethyl-1,3-dihydro-2H-benzimidazol-2-one; C493327:6,7-dichloro-1H-indole-2,3-dione 3-oxime; D001562:Benzimidazoles; D005456:Fluorescent Dyes; D007211:Indoles; C496897:KCNN2 protein, human; C406328:KCNN3 protein, human; D010091:Oximes; D051640:Small-Conductance Calcium-Activated Potassium Channels; D013793:Thallium; C102635:1-ethyl-2-benzimidazolinone