This is the renewal of an international collaboration that addressed key issues regarding pathophysiological mechanisms and therapeutic potentials for neurogenic bladder dysfunction (NBD; i.e., afferent sensitization) and detrusor overactivity (DO; i.e., micromotion). We have identified clinically relevant effects of phosphodiesterase type 5 (PDE5) inhibitors, sildenafil and tadalafil, on lower urinary tract (LUT) dysfunction in mice with acrolein- induced cystitis or T8-T9 spinal cord transection (SCT). We now demonstrate improved physiological parameters including decreased non-voiding contractions, afferent sensitization, micromotion and neuronal and stretch- induced ATP release; there are also increased intercontractile intervals and compliance. Moreover, we achieved these results whilst overcoming reliance on nitric oxide (NO?), by using the soluble guanylate cyclase (sGC) activator, BAY 58-2667 (cinaciguat), which counteracts key limitations of PDE5 inhibitors which render them clinically ineffective when there is nitrergic nerve damage or oxidative stress causing decreased NO? production and inactivation of sGC, respectively, both events commonly occur with NBD/DO. Sildenafil/tadalafil inhibit PDE5 as they are structural analogs of cGMP but are ineffective if NO? is not produced or the sGC heme is oxidized (Fe3+) as NO? can only bind to reduced heme (Fe2+). Cinaciguat, which is not an analog of cGMP, directly activates sGC and the conversion of GTP to cGMP in the absence of NO? or with an oxidized heme due to inhibition of CyB5R3 which keeps heme in the reduced state. Thus, cinaciguat may be effective in patients refractory to PDE5 inhibitors due to decreased NO? production or sGC inactivation. Clinical responses to PDE5 inhibitors used to treat LUT symptoms and now cinaciguat, involve increased protein kinase G (PKG) signaling which alters the behavior of several cell types. In afferent nerves it reduces firing, in efferent nerves it decreases ATP release, in urothelial cells it reduces stretch-induced ATP release, and in detrusor there appears to be an indirect relaxing effect on smooth muscle via interstitial cells (ICs). We propose to evaluate LUT sensory, motor and reflex behavior in C57BL/6 mice of both sexes in response to cinaciguat. For NBD, we will again use intravesical instillation of acrolein, a transient receptor potential ankyrin- 1 (TRPA1) receptor agonist and for DO, SCT. The University of Pittsburgh team will use in vivo and in vitro approaches to study peripheral LUT functions and the University of Bristol group, the in situ decerebrate arterially perfused mouse (DAPM) to study centrally mediated LUT reflexes, as they relate to the NO?-sGC-PKG pathway. We will also employ NADPH cytochrome b5 reductase 3 (CyB5R3) knockout mice, with selective deletions in smooth muscle and nerves rendering these relevant tissues unresponsive to NO? (and PDE5 inhibitors) but not sGC activators. Cinaciguat has passed phase 1 safety trials for non-urological pathologies so the proposed studies have considerable clinical relevance and translational potential.
This is a renewal of an international collaboration to study the soluble guanylate cyclase (sGC) activator, BAY 58-2667 (cinaciguat), in treating bladder afferent sensitization and detrusor overactivity due to micromotion. Using C57BL/6 mice of both sexes with acrolein-induced cystitis or T8-T9 spinal cord transection (SCT) we have demonstrated improved bladder function whilst overcoming reliance on nitric oxide (NO?) due to nitrergic nerve damage and decreased NO? production or oxidative stress and inactivation of sGC. The University of Pittsburgh team will use in vivo and in vitro approaches to study peripheral LUT functions and the University of Bristol group, an in situ decerebrate arterially perfused mouse (DAPM) to study centrally mediated LUT reflexes. Since cinaciguat has passed phase 1 safety trials for non-urological pathologies, the proposed studies have considerable clinical relevance and translational potential.
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