Abstract
Small conductance Ca2+-activated K+ (SK) and intermediate conductance Ca2+-activated K+ (IK) channels are thought to be involved in detrusor smooth muscle (DSM) excitability and contractility. Using naphtho[1,2-d]thiazol-2-ylamine (SKA-31), a novel and highly specific SK/IK channel activator, we investigated whether pharmacological activation of SK/IK channels reduced guinea pig DSM excitability and contractility. We detected the expression of all known isoforms of SK (SK1-SK3) and IK channels at mRNA and protein levels in DSM by single-cell reverse transcription-polymerase chain reaction and Western blot. Using the perforated patch-clamp technique on freshly isolated DSM cells, we observed that SKA-31 (10 μM) increased SK currents, which were blocked by apamin (1 μM), a selective SK channel inhibitor. In current-clamp mode, SKA-31 (10 μM) hyperpolarized the cell resting membrane potential, which was blocked by apamin (1 μM) but not by 1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole (TRAM-34) (1 μM), a selective IK channel inhibitor. SKA-31 (10 nM-10 μM) significantly inhibited the spontaneous phasic contraction amplitude, frequency, duration, and muscle force in DSM isolated strips. The SKA-31 inhibitory effects on DSM contractility were blocked by apamin (1 μM) but not by TRAM-34 (1 μM), which did not per se significantly affect DSM spontaneous contractility. SK channel activation with SKA-31 reduced contractions evoked by electrical field stimulation. SKA-31 effects were reversible upon washout. In conclusion, SK channels, but not IK channels, mediate SKA-31 effects in guinea pig DSM. Pharmacological activation of SK channels reduces DSM excitability and contractility and therefore may provide a novel therapeutic approach for controlling bladder dysfunction.
Footnotes
This study was supported by the National Institutes of Health National Institute of Diabetes and Digestive and Kidney Diseases [Grants DK084284, DK083687] (to G.V.P.).
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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ABBREVIATIONS:
- OAB
- overactive bladder
- DSM
- detrusor smooth muscle
- DS
- dissection solution
- PSS
- physiological salt solution
- VDCC
- L-type voltage-gated Ca2+ channel
- SK
- small conductance Ca2+-activated K+
- IK
- intermediate conductance Ca2+-activated K+
- BK
- large conductance voltage- and Ca2+-activated K+
- RT-PCR
- reverse transcription-polymerase chain reaction
- TTX
- tetrodotoxin
- BSA
- bovine serum albumin
- DMSO
- dimethyl sulfoxide
- EFS
- electrical field stimulation
- RMP
- resting membrane potential
- NS
- nonsignificant
- SKA-31
- naphtho[1,2-d]thiazol-2-ylamine
- TRAM-34
- 1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole
- bp
- base pairs
- NS309
- 6,7-dichloro-1H-indole-2,3-dione-3-oxime
- NS4591
- 4,5-dichloro-1,3-diethyl-1,3-dihydro-benzoimidazol-2-one
- n
- number of DSM strips or cells
- N
- number of guinea pigs
- CP
- competing peptide.
- Received July 20, 2011.
- Accepted October 13, 2011.
- Copyright © 2012 by The American Society for Pharmacology and Experimental Therapeutics
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