We conducted this study to evaluate the effects of an imidazoline 2 receptor (I2R) agonist, 2BFI, on lower urinary tract dysfunction (LUTD) in mice with spinal cord injury (SCI). It is reported that clonidine, activator of α2-adrenoceptors and imidazoline receptors, improved detrusor sphincter dyssynergia (DSD) caused by SCI through the action in the spinal cord [1]. Thus, we hypothesized that I2Rs could be involved in neurogenic LUTD after SCI.

C57BL/6N female mice at 9-10-weeks old were used. SCI mice underwent complete transection of the Th8-9 spinal cord. Mice were divided into three groups (1) spinal intact (SI), (2) SCI with vehicle, and (3) SCI with 2BFI treatment (20mg/kg, twice daily, i.p.), which was initiated 2 weeks after SCI. Four weeks after SCI, cystometrograms (CMG) and external urethral sphincter (EUS)-electromyography (EMG) were recorded. Thereafter, L6-S1 dorsal root ganglia (DRG) were harvested to evaluate the transcripts of TRPV1, TNF-alpha and iNOS using qPCR. Furthermore, using another set of mice, we tested intrathecal (i.t.) application of 2BFI (20µg per administration) at the L6 spinal cord level during CMG and EMG. In organ bath study, we cut the bladder into two longitudinal strips. Bladder body muscle strips were mounted longitudinally in a vertical double-jacketed organ bath with 37℃oxygenated 15 mL Krebs solution. Precontraction of muscle strips was induced by adding KCL (80μM) or Carbachol (10μM) to each bath. We waited for 30-40 minutes for precontraction stabilization and then, 2BFI application (1, 10, 100µM) (I2R agonist) was applied to each bath without or with BU224 (100µM) (I2R antagonist), which was applied 15 minutes prior to 2BFI.

SCI mice showed the significantly higher number of non-voiding contractions (NVCs) and lower voiding efficiency than SI mice. However, 2BFI-treated SCI mice exhibited a lower number of NVCs and improved voiding efficiency along with increased EUS relaxation time during voiding than vehicle-treated SCI mice. mRNA levels of TRPV1, TNF-alpha and iNOS in L6-S1 DRG were significantly higher in vehicle-treated SCI mice than in SI mice; however, decreased after 2BFI treatment. In the treatment with intrathecal application of 2BFI, significant reductions in residual urine and NVCs, and an increase of EUS relaxation time were seen after a transient stimulation period in SCI mice following i.t. application of 2BFI (Figure 1 and Table 1). In the organ bath study, 2BFI induced relaxation in muscle strips precontracted with KCL or Carbachol. BU224 reduced the relaxation effects of 2BFI.

We demonstrated that 2BFI (i.p. injection daily or intrathecal application) can improve DO, evident as NVC reduction, and DSD, evident as increase EUS relaxation time, in SCI mice. As shown in a previous report with clonidine [1], DSD was improved by i.t. application of 2BFI, indicating that I2R activation at spinal sites contributes to the improvement of vesicourethral coordination during voiding after SCI. The improvement of DO by 2BFI were accompanied by the decrease of the expression of a C-fiber afferent marker (TRPV1) and inflammatory mediators (TNF-alpha and iNOS) in L6-S1 DRG, as similarly shown in a previous study [2]. Furthermore, in the organ bath study, 2BFI significantly reduced detrusor contractility through peripheral effects of 2BFI, as evidenced by its relaxation effects on bladder muscle strips of SCI mice.

I2R activation, at the lumbosacral spinal cord and the bladder, could be effective for the treatment of both storage and voiding LUTD induced by SCI.

J Urol. 1998 Dec;160(6 Pt 1):2137-8.
Neurourol Urodyn. 2020 Jan;39(1):108-115.

Continence 12S (2024) 101434
DOI: 10.1016/j.cont.2024.101434