Inhibition of phosphodiesterase type 9 (PDE9) improves storage and voiding dysfunction in mice with spinal cord injury

Shimizu N1, Hashimoto M2, Suzuki T3, Takaoka E3, Kwon J3, Shimizu T3, Wada N3, Hirayama A4, Uemura H2, Kanai A J5, De Groat W C6, Yoshimura N3

Research Type

Basic Science / Translational

Abstract Category

Neurourology

Abstract 46
Basic Science: Pharmacology
Scientific Podium Short Oral Session 5
Wednesday 29th August 2018
10:52 - 11:00
Hall B
Spinal Cord Injury Voiding Dysfunction Physiology Detrusor Overactivity Pharmacology
1. Dept of Urology- University of Pittsburgh, Kindai University, Faculty of Medicine, 2. Dept of Urology-Kindai University, Faculty of Medicine, 3. Dept of Urology- University of Pittsburgh, 4. Dept of Urology- Kindai University Nara Hospital, Faculty of Medicine, 5. Dept of Medicine, University of Pittsburgh, 6. Dept of Pharmacology and Chemical Biology- University of Pittsburgh
Presenter
N

Nobutaka Shimizu

Links

Abstract

Hypothesis / aims of study
Chronic spinal cord injury (SCI) rostral to the lumbosacral level induces detrusor overactivity (DO) during the storage phase, which is mediated by spinal reflexes triggered by hyperexcitable C-fiber afferent pathways. During the voiding phase, inefficient voiding is commonly observed due to detrusor-sphincter dyssynergia (DSD) after SCI.  It has been shown that phosphodiesterase (PDE) inhibitors, which can modulate cGMP or cAMP levels in tissues, such as PDE type 5 inhibitors have been used for the treatment of lower urinary tract dysfunction (LUTD). PDE type 9 (PDE9) is one of a novel identified isozyme, which is expressed in brain [1], skeletal muscle [2], urinary tract [3]. However, it remains to be elucidated whether PDE9 is involved in LUTD induced by SCI. Therefore, we investigated the effects of a PDE9 inhibitor (PDE9i) to clarify the role of PDE9 in storage and voiding dysfunction using SCI mice.
Study design, materials and methods
Female C57BL/6N (8-9 weeks old) mice were used, and SCI was induced by complete transection of the Th8/9 spinal cord under isoflurane anesthesia. SCI mice were divided into 2 groups; (1) SCI mice treated with PDE9i (5 mg/kg/day) (n=7), (2) SCI mice with saline (n=5). Two weeks after SCI, PDE9i or saline (treatment or control group, respectively) was administered daily by i.p. injection for 14 days. After spinal cord transection, their bladders were manually squeezed to eliminate urine once daily for 4 weeks until cystometric evaluation. SCI mice were evaluated using single-filling cystometry (CMG) and external urethral sphincter (EUS)-electromyogram (EMG) under an awake condition. In single CMG recordings, the number of non-voiding contractions (NVCs), micturition pressure (MP), post-void residual volume (PVR) and voiding efficiency (VE) were evaluated in each SCI mouse (Figure 1).  In simultaneous CMG and EUS-EMG recordings, voiding contraction time, reduced EMG activity duration and the ratio of reduced EMG activity time to voiding contraction time were measured during the voiding phase to evaluate DSD in each SCI mouse. In real-time PCR analyses, L6 dorsal root ganglia (DRG), urethra, bladder mucosa and muscle were removed from saline treated SCI mice (n=8) and PDE9i treated SCI mice (n=8) as well as saline treated normal (spinal intact) mice (n=10), and the levels of PDE9, TRPV1, TNFα and VEGF transcripts were evaluated.
Results
Compared to saline treated SCI mice, NVCs during bladder filling were significantly reduced (Figure 1), and voiding efficiency was significantly improved with reduced residual urine in PDE9i treated SCI mice (Figure 2). In CMG and EUS-EMG recordings, the duration of reduced EMG activity or the ratio of reduced EMG activity time to voiding contraction time during the voiding phase was significantly increased between PDE9i treated and saline treated SCI mice. PDE9 transcripts were identified in L6 DRG, urethra and bladder mucosa and muscle. The levels of TRPV1 mRNA in DRG and bladder muscle were increased in SCI mice vs. spinal intact mice, and significantly decreased after PDE9i treatment in SCI mice. The TNFα mRNA levels in urethra, bladder mucosa and muscle were increased in SCI mice vs. spinal intact mice, and significantly decreased after PDE9i treatment in SCI mice. VEGF mRNA levels in DRG, bladder muscle and mucosa were increased in SCI mice vs. spinal intact mice, but not significantly changed after PDE9i treatment in SCI mice (Figure 3).
Interpretation of results
The treatment with PDE9i improved DO evident as a decrease in NVCs in association with the reduction of the expression of TRPV1 in L6 DRG, which is predominantly expressed in C-fiber afferent pathways, suggesting that the PDE9 activation is involved in the inflammatory changes in bladder afferent pathways. Also, PDE9i treatment reduced the expression of TNFα in urethra and bladder tissues and also improved inefficient voiding and DSD as shown by decreases in residual urine and EMG activity time in SCI mice. Overall, the results of this study demonstrated that PDE9 significantly contributes to LUTD as well as inflammatory changes in bladder afferent pathways after SCI and that PDE9i can improve SCI-induced voiding and storage dysfunction.
Concluding message
PDE9 inhibition improved SCI-induced detrusor overactivity and inefficient voiding, along with significant reductions in C-fiber afferent receptors such as TRPV1 and proinflammatory cytokines such as TNF in mice. Thus, PDE9 could be a therapeutic target for storage and voiding LUTD after SCI.
Figure 1
Figure 2
References
  1. J. Neurocytol. 31: 729-741 2002.
  2. Can. J. Physiol. Pharmacol. 80: 1132-1135 2002.
  3. B.J.U. int. 109: 934-940 2012
Disclosures
Funding NIH P01DK093424, DOD W81XWH-17-1-0403 Clinical Trial No Subjects Animal Species Mouse Ethics Committee University of Pittsburgh Institutional Animal Care and Use Committee