cAMP accelerates the decay of stretch-activated inward currents in guinea-pig urinary bladder myocytes.
1. Myocytes from the urinary bladder were stretched longitudinally by 5-20%. At -50 mV, stretch induced whole-cell inward currents (Iin) between -100 and -600 pA. Iin decayed slowly with time to 93 +/- 20% (mean +/- S.E.M., n = 6) of the initial value in 1 min. The mechanisms of this 'adaptation' and its modulation by dibutyryl cAMP (dBcAMP) were analysed with whole-cell and single channel currents. 2. When the cells were internally perfused with 100 microM 8-bromo-cAMP (8BrcAMP), stretch induced an Iin of the usual amplitude that decayed completely within 40 +/- 13 s. When 200 microM dBcAMP was bath applied 10 s after the start of the stretch, Iin decayed to zero within 85 +/- 18 s. 3. dBcAMP increased the K+ current through Ca(2+)-activated BK channels (IK(Ca)) at 0 mV with a time course that correlated well with the decay of Iin, and block of IK(Ca) by TEA suppressed the dBcAMP-induced decay of Iin. In the presence of intracellular BAPTA, dBcAMP increased the stretch-induced Iin. The results suggest that adaptation is caused by superimposition of IK(Ca) which is increased through elevation of near-membrane [Ca2+] and by cAMP-dependent phosphorylation. 4. Single channel analysis was carried out with 140 mM KCl electrode solution and at -50 mV. Stretch-activated channels (SACs) were recorded during pulses of negative pressures between -2 and -5 kPa. Activity (NPo) of SACs was constant for at least 4 min, e.g. evidence for adaptation was missing. dBcAMP (200 microM) increased NPo of SACs by 142 +/- 35% (n = 16). 5. dBcAMP increased NPo via frequency of openings and channel open time. In five of sixteen patches, dBcAMP induced openings without suction. Similar effects were induced by the catalytic subunit of cAMP-dependent protein kinase (PKAc), applied to inside-out patches. 6. NPo, normalized by its maximum, increased with more negative pressure along an S-shaped curve. dBcAMP increased the sensitivity of SACs to stretch by shifting the point of half-maximal activity from -3.2 to -2.6 kPa. 7. The augmentation of NPo by dBcAMP is attributed to the phosphorylation of SACs promoting their opening. Adaptation of Iin is discussed as a 'secondary' effect of stretch-activated channels: Ca2+ influx through SACs increases the Ca2+ concentration that activates BK channels whose Ca2+ sensitivity is increased by cAMP.
Wellner
M C
MC
Department of Physiology, University of Cologne, Germany.
Isenberg
G
G
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D000242:Cyclic AMP / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D006168:Guinea Pigs; D015640:Ion Channel Gating / Q000187:drug effects; D008564:Membrane Potentials / Q000187:drug effects; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D015221:Potassium Channels / Q000187:drug effects / Q000378:metabolism; D019789:Tetraethylammonium; D013757:Tetraethylammonium Compounds / Q000494:pharmacology; D001743:Urinary Bladder / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
D015220:Calcium Channels; D015221:Potassium Channels; D013757:Tetraethylammonium Compounds; D015124:8-Bromo Cyclic Adenosine Monophosphate; D004533:Egtazic Acid; D003994:Bucladesine; D019789:Tetraethylammonium; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid