Radiation therapy (RT) is necessary for stage IIB and greater cervical cancers. Within 6 months of diagnosis, 53% of women will receive pelvic RT. Although external beam and brachytherapy radiation techniques can deliver adequate radiation to the targeted neoplasm, healthy neighboring tissues in the pelvis may become ionized leading to urinary incontinence. Tissue injury from RT is both an acute and chronic process. In male rats, prostatic radiation induced an early decrease and late increase in bladder contractility. RT is shown to induce cystitis in several pre-clinical models, yet the pathological mechanisms leading to urinary incontinence are largely unknown.  The aim of this study is to determine how a single dose of pelvic radiation administered to a rat model impacts bladder and urethra contractions at 4 and 9 weeks post RT. We hypothesize RT will decrease bladder and urethra contractions by 9 weeks post RT.

Adult female Sprague Dawley rats were anesthetized, given a single dose of 20 Gy radiation selectively targeted to the pelvis and allowed to recover for 4 or 9 wks (4RT n=8; 9RT n=8). Age matched controls (4CON n=3; 9CON n=4) underwent the same procedure without receiving any radiation. Bladders, internal urethral sphincters (IUS), and external urethral sphincters (EUS) were excised. The bladder was cut into strips and the urethral sphincters were cut into 2 mm rings, and mounted in tissue baths. Contractility was measured using a high potassium solution (KCl, 120 mM), carbachol (10-9 to 10-4 M; bladder only), caffeine (40 mM), and electric field stimulation (EFS, 1-48 Hz at fixed parameters) in the absence or presence of a ryanodine receptor antagonist, dantrolene (10-5 M) to determine striated muscle contribution, or sodium channel inhibitor, tetrodotoxin (TTX, 10-6 M) to determine if contractions are of neuronal origin and not a result of direct smooth muscle stimulation.

Overall, there were no differences between the bladder and sphincter data for 4CON and 9CON groups; therefore, both groups were combined and referred to collectively as CON. Within the bladder, RT decreased EFS contractions at both 4 and 9 weeks (Figure 1, CON: 58.8±7.84, 4RT: 41.5±4.13, 9RT: 38.0±2.58mN; p<0.05). In contrast, RT did not change KCl or carbachol-induced bladder contractions. Conversely in the IUS, RT increased EFS contractions after 4 weeks, but contractions returned to CON values by 9 weeks (CON: 2.0±0.34, 4RT: 3.1±0.55, 9RT: 2.1±0.27mN; p<0.05). EFS IUS contractions were unaffected by dantrolene incubation, confirming the absence of striated muscle contribution. IUS contractions to KCl, and caffeine did not change following RT. Multiple RT induced changes were noted in the EUS. At 4 weeks post RT, no changes were noted in EUS KCl contractions; however, by 9 weeks KCl contractions doubled (CON: 3.9±1.04, 4RT: 4.1±0.71, 9RT: 8.9±0.50mN; p<0.05). Caffeine induced contractions increased only 9 weeks after RT (CON: 5.3±1.08, 4RT: 9.0±0.69, 9RT: 11.4±1.82mN; p<0.05). RT caused a threefold increase in 4RT and 9RT EFS contractions (Figure 2, CON: 3.2±1.06, 4RT: 7.6±1.67, 9RT: 10.34±2.22mN; p<0.05). When incubated with dantrolene to eliminate striated muscle EFS contribution, no change in EFS contractions were noted at 4 or 9 weeks post RT (CON: 1.6±0.35, 4RT: 3.0±0.75, 9RT: 3.9± 0.99mN). To confirm neuronal origin, all EFS contractions within the bladder, IUS, and EUS were inhibited following incubation with TTX.

Pelvic RT decreases nerve mediated bladder contractions yet increases nerve mediated urethral sphincter contractions. Interestingly, most changes occur within the EUS – the urethral sphincter under voluntary striated muscle control.

The adaptive physiological changes in smooth muscle signaling of the bladder and urethral sphincters following radiation has been poorly described in previous literature but is crucial to understand the development of urinary incontinence. Further studies may provide mechanistic detail necessary to prevent or treat RT induced incontinence and improve the quality of life for cervical cancer survivors