The role of cAMP dependent protein kinase in modulating spontaneous intracellular Ca waves in interstitial cells of Cajal from the rabbit urethra |
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Authors: | Bernard T Drumm Gerard P Sergeant Mark A Hollywood Keith D Thornbury Noel G McHale Brian J Harvey |
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Institution: | 1. Smooth Muscle Research Centre, Dundalk Institute of Technology, Dublin Road, Co. Louth, Ireland;2. Molecular Medicine Laboratories, Royal College of Surgeons in Ireland, Beaumont Hospital, Dublin 9, Co. Dublin, Ireland |
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Abstract: | Interstitial cells of Cajal (ICC) serve as electrical pacemakers in the rabbit urethra. Pacemaking activity in ICC results from spontaneous intracellular Ca2+ waves that rely on Ca2+ release from endoplasmic reticulum (ER) stores. The purpose of this study was to investigate if the action of protein kinase A (PKA) affected the generation of Ca2+ waves in ICC. Intracellular Ca2+] was measured in fluo-4 loaded ICC, freshly isolated from the rabbit urethra using a Nipkow spinning disc confocal microscope. Application of the PKA inhibitor H-89 (10 μM) significantly inhibited the generation of spontaneous Ca2+ waves in ICC and this was associated with a significant decrease in the ER Ca2+ load, measured with 10 mM caffeine responses. Ca2+ waves could be rescued in the presence of H-89 by stimulating ryanodine receptors (RyRs) with 1 mM caffeine but not by activation of inositol 1,4,5 tri-phosphate receptors (IP3Rs) with 10 μM phenylephrine. Increasing intracellular PKA with the cAMP agonists forskolin and 8-bromo-cAMP failed to yield an increase in Ca2+ wave activity. We conclude that PKA may be maximally active under basal conditions in ICC and that inhibition of PKA with H-89 leads to a decreased ER Ca2+ load sufficient to inactivate IP3Rs but not RyRs. |
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Keywords: | Urethra ICC Ca2+ waves PKA |
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