Lower urinary tract (LUT) symptoms secondary to benign prostatic hyperplasia (BPH) and obstruction (BPO) are highly prevalent in the aging male population. Currently available pharmacotherapies include α-adrenoceptor antagonists, aromatase inhibitors and phosphodiesterase type 5 inhibitors. However, refractoriness and limited efficacy are issues with these agents as demonstrated by the high number of BPO surgeries performed each year. Accordingly, there is a significant need for more effective therapeutic option. Our data indicate that daily treatment of aged mice with the soluble guanylate cyclase (sGC) activator, cinaciguat, reduces hyperplasia within the prostatic urethra and surrounding region encircled by the external urethral sphincter (EUS), relieving urethral compression. Therefore, our aim was to determine whether prostatic hyperplasia in mice correlates with functional outlet obstruction and the effects of cinaciguat treatment. To achieve this, we implanted telemetric pressure sensors into the bladders of adult and aged male mice. Pressure recordings from awake freely mobile mice were combined with customized metabolic cages equipped with high sensitivity load cells. These allowed recording of small voided volumes at high acquisition rates to facilitate extrapolation of urine flow rate. This novel methodology allowed us to demonstrate for the first time a naturally occurring BPO, defined by high micturition pressures and low flow rates, in aged male mice supporting their use as a preclinical model for the study of BPH/BPO.

Telemeter implantation and recordings. Adult (9 months) and aged (≥24 months) male C57Bl/6 mice (N=4 each) were anesthetized with isoflurane and an HDX telemeter (DSI Inc., Fig. 1A) implanted via a laparotomy with the pressure line inserted into the bladder lumen through the dome and secured with a purse string ligature (Fig. 1B). The telemeter with built-in battery pack was placed subcutaneously on the flank of the mouse and the surgical wounds sutured. Mice were allowed to recover with daily prophylactic antibiotic/analgesic treatments for seven-days before baseline recordings were obtained. For recordings, mice were placed in customized metabolic cages (Columbus Instruments) equipped with load cells capable of recording volumes as small as 5 µl at a sampling rate of 10 Hz. Load cells plates were lined with filter paper for urine spots to assess voiding behaviour. Telemeters were magnetically activated when the mice were placed in their cages and recordings taken over a full light/dark cycle. Data were acquired and analysed using LabChart8 software and expressed as mean ± standard deviation. Following acquisition of baseline recordings, a cohort of aged mice (N=4) received daily cinaciguat gavage (100-150 µl, 10 mg/kg/day in 0.5% methylcellulose + 10% DMSO in water) for a two-week period. Telemetric recordings were obtained weekly. Statistical differences in cystometric parameters were evaluated using Student’s t-test, where p<0.05 was considered significant.

In vivo telemetric bladder pressure recordings combined with urine spots tests from adult and aged male mice (Fig. 1C and 1D) demonstrated significant differences between the age groups. Specifically, aged mice showed higher number of voids, smaller voided volumes, slower urine flow rates and higher maximal voiding pressures compared to younger animals. Treatment of aged mice with cinaciguat showed a time dependent normalization of voided volume, flow rate, maximum voiding pressure and partially lowered voiding frequency (Fig. 2A-C). Summary of the cystometric parameters are shown in Table 1.

These data support a naturally occurring BPO pathology in aged male mice. It has been widely speculated that rodent models do not fully recapitulate obstructive pathology seen in humans due to the lack of a fibrous capsule. However, we have demonstrated cellular hyperplasia is present within the prostatic urethra of aged mice. The rodent prostatic urethra is encircled by the skeletal muscle composing the EUS which is responsible for outlet constriction; in humans, the EUS is below the prostate. Our novel methodology supports that aged rodents can exhibit naturally occurring BPO; the urodynamic hallmark being high bladder pressures and low urine flow rates, a phenomenon that has not been previously described in mice. Furthermore, cinaciguat ameliorates the BPO and LUT symptomology which may be due to effects on both the prostate and bladder.

We demonstrate for the first time that aged male mice exhibit cystometric parameters consistent with the diagnostic criteria of BPH-induced outlet obstruction in humans. Furthermore, cinaciguat treatment in aged mice improves LUT function which may be due to effects on relieving the obstruction and reducing bladder overactivity. This indicates sGC activators may have advantages over currently approved drugs with their multiple therapeutic actions and ability to circumvent refractoriness associated with impaired nitric oxide production.