A corticotropin-releasing factor receptor antagonist improves urodynamic dysfunction produced by social stress or partial bladder outlet obstruction in male rats

Abstract

Barrington's nucleus, in the pons, regulates micturition through spinal projections to preganglionic parasympathetic neurons. The stress neuropeptide CRF is prominent in these projections and has an inhibitory influence. Social stress in rats causes urinary retention and abnormal urodynamics resembling those produced by partial bladder outlet obstruction (pBOO), and this is associated with CRF upregulation in Barrington's nucleus. Here, we examined the role of CRF in social stress- and pBOO-induced urodynamic dysfunction by assessing the ability of a CRF₁ receptor antagonist to alter these effects. Male rats exposed to repeated resident-intruder stress were administered vehicle or a CRF₁ antagonist (NBI-30775) daily prior to the stress. Urodynamic function was recorded in the unanesthetized state 72 h after the final stress. NBI-30775 prevented the increased intermicturition interval, micturition volume, and bladder capacity produced by social stress, but not the increase in CRF expression in Barrington's nucleus neurons. The urinary dysfunction was also partly prevented by shRNA targeting of CRF in Barrington's nucleus, suggesting that stress-induced urinary dysfunction results, in part, from CRF upregulation in Barrington's nucleus and enhanced postsynaptic effects in the spinal cord. Finally, NBI-30775 improved urodynamic function of rats that had pBOO of 2-wk duration when administered daily during the second week but did not block the increase in CRF expression in Barrington's nucleus neurons. These findings implicate a role for Barrington's nucleus CRF in stress- and pBOO-induced urodynamic changes and suggest that CRF₁ antagonists may be useful therapeutic agents for the treatment of urinary dysfunction.

Keywords: Barrington's nucleus; cystometry; resident-intruder; urinary.

Fig. 1.

Sample cystometry indicating how endpoints were determined. Bladder pressure (BP; mmHg) (top), bladder capacity (BC; μl) (middle), and micturition volume (MV; μl) (bottom). Top: arrowheads show the points at which resting pressure (RP), micturition threshold (MT), and micturition pressure (MP) were determined. Middle: vertical line labeled “BC (μl)” indicates how bladder capacity was determined. Bottom: horizontal line labeled “IMI (s)” indicates how intermicturition was derived. See materials and methods for details.

Fig. 2.

Representative cystometry traces from different experimental groups. A–D: in all traces, the abscissae indicate time from 0 to 500 s. The top trace in each part shows BP (mmHg); the middle trace shows BC (μl), as defined by the volume of saline infused into the bladder between micturition cycles, and the bottom trace shows MV (μl). For comparison, the ordinates were scaled to be equivalent for all groups.

Fig. 3.

Quantification of the effects of different treatments on urodynamics of control and stressed rats. A–C: micturition interval, bladder capacity, and micturition volume, respectively. D: MT, MP, and RP for all groups in one graph. Results of a two-way ANOVA comparing stress and drug treatment are presented in the text. A one-way ANOVA was also performed comparing all groups, including the AAV-shRNA groups, and symbols in the figure refer to results of post hoc tests from this analysis. *P < 0.05, **P < 0.01, †P < 0.06. Control+vehicle (n = 10), control/NBI (n = 5), stress+vehicle (n = 9), stress+NBI (n = 9), stress+shRNA (n = 8), and stress+control RNA (n = 7).

Fig. 4.

NBI-30775 had no effect on the stress-induced increase in the number of CRF-immunoreactive neurons in Barrington's nucleus. A: fluorescent photomicrographs of CRF-immunoreactive Barrington's nucleus neurons of representative control (top) and stressed (bottom) rats treated with vehicle. Top shows dorsal and medial appears to the left in all photomicrographs. B: bars represent the mean number of CRF-immunoreactive Barrington's nucleus neurons for controls treated with vehicle and stressed rats treated with vehicle or NBI-30775. Newman-Keuls post hoc tests: *P < 0.05 vs. control/vehicle.

Fig. 5.

CRF mRNA expression in Barrington's nucleus is reduced in stressed rats with infusion of AAV-shRNA-CRF into Barrington's nucleus. A: darkfield photomicrographs showing the CRF mRNA hybridization signal in Barrington's nucleus of stressed rats that were infused with AAV-shRNA-CRFscrambled (top) and AAV-shRNA-CRF (bottom). The two photomicrographs were taken using identical exposure times, and brightness and contrast of the composite photograph were adjusted after the composite was flattened, so adjustments were identical for both examples. B: bar graph indicates the mean number of CRF mRNA-expressing Barrington's nucleus neurons for rats infused with shRNA-CRF scrambled (n = 4) and shRNA-CRF (n = 6). *P < 0.05.

Fig. 6.

Representative cystometrograms from sham and partial bladder outlet obstruction (pBOO) rats. Shown are traces of a vehicle-treated sham rat (A), a vehicle-treated pBOO rat (B), and a pBOO rat treated with NBI-30775 (C). A–C: all traces of the abscissae indicate time from 0 to 500 s. The top trace of each part shows BP (mmHg), the middle trace shows BC (μl), as defined by the volume of saline infused into the bladder between micturition cycles, and the bottom trace shows MV (μl). For comparison, the ordinates were scaled to be equivalent for all groups.

Fig. 7.

NBI-30775 attenuates the urodynamic effects of pBOO. Bars represent the mean values for sham rats treated with vehicle (n = 6), and pBOO rats treated with vehicle (n = 9) or NBI-30775 (n = 10) for intermicturition interval (IMI; s), BC (μl), MV (μl), MT (mmHg), MP (mmHg), and RP (mmHg). Student-Newman-Keuls post hoc test: *P < 0.05 vs. pBOO/NBI. †P < 0.05 vs. sham/vehicle. ‡P = 0.06 vs. sham/vehicle.

Fig. 8.

CRF mRNA expression in Barrington's nucleus is increased in pBOO rats. A: dark-field photomicrographs showing the CRF mRNA hybridization signal in Barrington's nucleus of a sham (top) and a pBOO rat (bottom). The two photomicrographs were taken using identical exposure times and brightness, and contrast of the composite photograph was done after the composite was flattened, so adjustments were identical for both examples. B: bar graph indicates the mean number of CRF mRNA expressing Barrington's nucleus neurons for sham rats administered NBI-30775 (n = 7), pBOO rats administered vehicle (n = 9), and pBOO rats administered NBI-30775 (n = 6). *P < 0.05.