Hypothesis / aims of study
Enhanced spontaneous bladder contractions (SCs) are a key mechanism underlying detrusor overactivity (DO). The bladder mucosa enhances SCs in detrusor muscle via diffusible signals or direct cell contact (1). Nitric oxide (NO), an inhibitory modulator of the lower urinary tract, is released from neuronal and non-neuronal sources and influences smooth muscle contractility, including suppression of SCs (2,3). This study investigated how NO influences contractile activity in pig bladder strips with and without mucosa.
Study design, materials and methods
Female pig bladders were obtained from a local abattoir and placed in cold Krebs bicarbonate (KB) solution. From each bladder dome, longitudinally oriented tissue strips were prepared: intact (I; detrusor + mucosa), denuded (D; detrusor with mucosa removed), mucosa alone (M), and reconstructed (R; denuded with previously isolated mucosa reattached). Strips were mounted in organ baths under a fixed (1.5 g) load in KB solution at 37°C. The effects of L-NG-nitro arginine methyl ester (L-NAME, 100 μM, nitric oxide synthase inhibitor), sodium nitroprusside (SNP, 10 μM; nitric oxide donor), or vehicle control (KB solution) on SCs were measured after incubating the four preparations for up to 30 mins. Tissues were then stimulated with 10 µM carbachol or 120 mM potassium chloride (KCl) in the continued presence of SNP, L-NAME or KB solution, and maximum tonic tensions were recorded. Mean amplitude (g tension/mg tissue) and frequency (events in 5 mins) of SCs, as well as tonic responses to carbachol and KCl with and without L-NAME and SNP, were recorded and compared across preparations. All data are expressed as mean ± SD. n = number of preparations. Statistical analyses used non-parametric t-test/ANOVA and appropriate post hoc tests.
Results
SNP significantly reduced the frequencies of SCs in intact, mucosa, and reconstructed strips within 5 mins, with recovery by 30 mins post incubation (Figure 1A). The same response to SNP was not detected in denuded strips where the mucosa was removed (Figure 1A). SNP had no significant effect on the amplitude of SCs in all preparations (Figure 1B).
L-NAME significantly enhanced the frequencies of SCs in intact, mucosa and reconstructed preparations, but not in denuded strips, after 30 min of incubation (Figure 2A). L-NAME had a significant effect on the amplitude of SCs in intact strips only (Figure 2B)
The tonic responses to CCh and KCl were not altered in the presence of SNP or L-NAME in all bladder strips.
Interpretation of results
These findings support a role for mucosa-derived NO signalling in modulating SCs in the bladder wall. The lack of response in denuded strips suggests that the mucosa is the primary site of NO target/ production influencing such phasic activity.
The reversible inhibition of SC frequency by SNP may reflect NO’s short half-life in aqueous environments and a dynamic NO-mediated signalling pathway.
That tonic responses to carbachol and KCl were unaffected by NO modulation, suggesting NO selectively influences spontaneous, not evoked, contractions—indicating distinct regulatory mechanisms for SCs and tonic responses within the bladder wall.
Reattaching mucosa to denuded detrusor did not alter responses to NO modulators, suggesting a physical connection between layers is not required for NO-driven SC modulation.
Concluding message
This study demonstrates that NO selectively modulates spontaneous, but not evoked, pig bladder contractions, and that this effect is dependent on the presence of the mucosa. The absence of NO-mediated effects in denuded strips and their restoration in reconstructed tissues indicate that mucosa-derived NO plays a critical role in regulating detrusor phasic activity via a diffusible, rather than contact-dependent, mechanism. These findings enhance our understanding of mucosal regulation of bladder function and support the concept that altered NO signalling within the mucosa may contribute to the pathophysiology of detrusor overactivity.