Cascade Bioassay Evidence for the Existence of Urothelium-Derived Inhibitory Factor in Guinea Pig Urinary Bladder

Abstract

Our aim was to investigate whether guinea pig urothelium-derived bioactivities compatible with the existence of urothelium-derived inhibitory factor could be demonstrated by in vitro serial bioassay and whether purinergic P1 receptor agonists, nitric oxide, nitrite or prostaglandins might explain observed activities. In a cascade superfusion system, urothelium-denuded guinea pig ureters were used as bioassay tissues, recording their spontaneous rhythmic contractions in presence of scopolamine. Urothelium-intact or -denuded guinea pig urinary bladders were used as donor tissues, stimulated by intermittent application of carbachol before or during the nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME), the adenosine/P1 nucleoside receptor antagonist 8-(p-sulfophenyl)theophylline (8-PST) or the cyclo-oxygenase inhibitor diclofenac infused to bath donor and bioassay tissues. The spontaneous contractions of bioassay ureters were unaltered by application of carbachol 1–5 µM in the presence of scopolamine 5–30 µM. When carbachol was applied over the urothelium-denuded bladder, the assay ureter contraction rate was unaltered. Introducing carbachol over the everted urothelium-intact bladder significantly inhibited the contraction frequency of the assay ureter, suggesting the transfer of an inhibitory activity from the bladder to the assay ureter. The transmissible inhibitory activity was not markedly antagonized by L-NAME, 8-PST or diclofenac, while L-NAME nearly abolished nitrite release from the urothelium-intact bladder preparations. We suggest that urothelium-derived inhibitory factor is a transmissible entity over a significant distance as demonstrated in this novel cascade superfusion assay and seems less likely to be nitric oxide, nitrite, an adenosine receptor agonist or subject to inhibition by administration of a cyclo-oxygenase inhibitor.

Mucosal modulation of contractility in bladder strips from normal and overactive rat models and the effect of botulinum toxin A on overactive bladder strips

“…Hawthorn et al reported smaller carbachol‐evoked contractions in intact strips of pig bladder compared with mucosa‐free strips; moreover, mucosa‐free strip contractions were inhibited by the presence of a second intact strip indicating release of a modulatory signal. The exact identity of this diffusible UDIF is not yet known although several candidates have been proposed and excluded …”

“…The exact identity of this diffusible UDIF is not yet known although several candidates have been proposed and excluded. 8 Botulinum toxin type A (BoNT-A) is used to treat antimuscarinic refractory detrusor overactivity (DO) where involuntary detrusor contractions can be observed and overactive bladder syndrome (OAB) where DO may or may not be demonstrable. 9 The established mechanism of action of BoNT-A on neurones involves cleavage of synaptosome-associated protein-25 (SNAP-25), a component of the neuronal SNARE complex, preventing neuronal transmitter release.…”

Pharmacological Modulation of Mucosa-Related Impairment of β-Adrenoceptor-Mediated Relaxation in Human Detrusor

“…There was also a tendency to higher force development in response to carbachol, but this difference did not reach statistical significance. These results suggest that endogenous NO may attenuate the level of stimulated tension development [3] and confirm that NO is probably not the urothelium-derived relaxing factor [4,25,26]. Moreover, NO is unlikely to be involved in noradrenaline-induced relaxation because L-NAME had no effect on the concentration-relaxation curves with or without urothelium.…”

Inhibitory Effects of Urothelium‐related Factors

“…The study using human bladder strips showed that the factor was distinct from nitric oxide and it persisted in the presence of beta‐adrenoceptor blockade or cyclooxygenase inhibition. The existence of the urothelium‐derived inhibitory factor in the guinea pig urinary bladder was also demonstrated by assay on the spontaneous contracted guinea pig ureters in a cascade superfusion bioassay system, which offers further possibilities to identify the factor . The results indicated a transmissible inhibitory factor over a significant distance and seemed less likely to be nitric oxide, nitrite, an adenosine receptor agonist or cyclooxygenase products.…”