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MP94-19: A POTENTIAL NOVEL MECHANISM FOR DETRUSOR UNDER-ACTIVITY MEDIATED BY MYOSIN LIGHT CHAIN PHOSPHORYLATION AND AMP-DEPENDENT KINASE

Tuesday, May 16
7:00 AM - 9:00 AM
Location: BCEC: Room 253AB

Presentation Authors: Randy Vince*, Ramanan Remesh, John Speich, Adam Klausner, Amy Miner, Paul Ratz, Richmond, VA

Introduction: Muscarinic receptor stimuli such as carbachol (CCh) cause increases in detrusor smooth muscle (DSM) myosin light chain (MLC) phosphorylation and contraction. Notably, upon stimulation with CCh, DSM force rises rapidly to a peak value, then gradually declines (fades) despite the continued presence of CCh. The mechanism behind the fade in force s not understood. There is evidence that, in addition to causing contraction, muscarinic receptor stimulation can activate AMPK, and AMPK has been shown to negatively regulate smooth muscle contraction. The purpose of this study was to test the hypothesis that DSM contraction fade is due to the delayed activation of AMPK

Methods: Mouse bladder with mucosa were cut into rings ~3 mm wide, and strips of DSM free from underlying mucosa were removed from rabbit bladders. Each tissue was placed in an organ bath connected to a force transducer and length-adjuster and subjected to a length-tension protocol to identify the length (Lref) that produced the strongest active force induced by KCl. Each ring was subsequently set to 95% Lref. Three tissues were then contracted by exposure to 10 microM CCh and quick-frozen at 5, 30 and 180 seconds. A fourth tissue was not contracted and quick-frozen to assess the basal-state. Tissues were subsequently processed to quantify the level of phosphoproteins that are indices of activation of AMPK (ACC-pS79 and AMPK-p172) and of actomyosin crossbridges (MLC-pS19 and MYPT1-pT853).

Results: Compared to the basal-state, CCh induced a strong increase in force in mouse bladder and rabbit DSM that peaked at, respectively, 60 sec and 20 sec, before declining to ~50% of the peak values within 180 sec (n=3). MLC-p also displayed a biphasic response. AMPK-pand ACC-p displayed a delayed increase corresponding with the decrease in contractile force.

Conclusions: Muscarinic receptor stimulus CCh caused a rapid increase in force in mouse bladder and rabbit DSM that faded with time despite the presence of CCh. This fade in force correlated with a delayed increase in AMPK activity as assessed by increases in the phosphoproteins, ACC-p and AMPK-p. Because AMPK has been shown to inhibit smooth muscle contraction, the correlation of an increase in AMPK activity with the decrease in force supports the hypothesis that AMPK may be responsible for fade in DSM force. Because AMPK activity is also elevated during ischemia/hypoxia, a condition known to cause bladder underactivity, these data warrant further studies to investigate the potential role of AMPK in bladder underactivity

Source Of Funding: none

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MP94-19: A POTENTIAL NOVEL MECHANISM FOR DETRUSOR UNDER-ACTIVITY MEDIATED BY MYOSIN LIGHT CHAIN PHOSPHORYLATION AND AMP-DEPENDENT KINASE



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