Activation of the cGMP/PKG pathway inhibits electrical activity in rabbit urethral interstitial cells of Cajal by reducing the spatial spread of Ca2+ waves.
In the present study we used a combination of patch clamping and fast confocal Ca2+ imaging to examine the effects of activators of the nitric oxide (NO)/cGMP pathway on pacemaker activity in freshly dispersed ICC from the rabbit urethra, using the amphotericin B perforated patch configuration of the patch-clamp technique. The nitric oxide donor, DEA-NO, the soluble guanylyl cyclase activator YC-1 and the membrane-permeant analogue of cGMP, 8-Br-cGMP inhibited spontaneous transient depolarizations (STDs) and spontaneous transient inward currents (STICs) recorded under current-clamp and voltage-clamp conditions, respectively. Caffeine-evoked Cl- currents were unaltered in the presence of SP-8-Br-PET-cGMPs, suggesting that activation of the cGMP/PKG pathway does not block Cl- channels directly or interfere with Ca2+ release via ryanodine receptors (RyR). However, noradrenaline-evoked Cl- currents were attenuated by SP-8-Br-PET-cGMPs, suggesting that activation of cGMP-dependent protein kinase (PKG) may modulate release of Ca2+ via IP3 receptors (IP3R). When urethral interstitial cells (ICC) were loaded with Fluo4-AM (2 microm), and viewed with a confocal microscope, they fired regular propagating Ca2+ waves, which originated in one or more regions of the cell. Application of DEA-NO or other activators of the cGMP/PKG pathway did not significantly affect the oscillation frequency of these cells, but did significantly reduce their spatial spread. These effects were mimicked by the IP3R blocker, 2-APB (100 microm). These data suggest that NO donors and activators of the cGMP pathway inhibit electrical activity of urethral ICC by reducing the spatial spread of Ca2+ waves, rather than decreasing wave frequency.
Sergeant
G P
GP
Smooth Muscle Research Centre, Dundalk Institute of Technology, Dublin Road, Dundalk, Co. Louth, Ireland.
Johnston
Louise
L
McHale
N G
NG
Thornbury
K D
KD
Hollywood
M A
MA
D000200:Action Potentials / Q000502:physiology*; D000818:Animals; D001683:Biological Clocks / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology*; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000378:metabolism*; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism*; D005260:Female; D008297:Male; D009433:Neural Inhibition / Q000502:physiology; D009569:Nitric Oxide / Q000378:metabolism*; D011817:Rabbits; D015398:Signal Transduction / Q000502:physiology; D014521:Urethra / Q000502:physiology*
D009569:Nitric Oxide; D017869:Cyclic GMP-Dependent Protein Kinases; D006152:Cyclic GMP; D002118:Calcium