Psychological stress induced bladder overactivity in female mice is associated with enhanced afferent nerve activity.

Psychological stress has been linked to the development and exacerbation of overactive bladder symptoms, as well as afferent sensitisation in other organ systems. Therefore, we aimed to investigate the effects of water avoidance stress on bladder afferent nerve activity in response to bladder filling and pharmaceutical stimulation with carbachol and ATP in mice. Adult female C57BL/6J mice were exposed to either water avoidance stress (WAS) for 1 h/day for 10 days or normal housing conditions. Voiding behaviour was measured before starting and 24-h after final stress exposure and then animals were euthanised to measure afferent nerve activity in association with bladder compliance, spontaneous phasic activity, contractile responses, as well as release of urothelial mediators. WAS caused increased urinary frequency without affecting urine production. The afferent nerve activity at low bladder pressures (4-7 mmHg), relevant to normal physiological filling, was significantly increased after stress. Both low and high threshold nerves demonstrated enhanced activity at physiological bladder pressures. Urothelial ATP and acetylcholine release and bladder compliance were unaffected by stress as was the detrusor response to ATP (1 mM) and carbachol (1 µM). WAS caused enhanced activity of individual afferent nerve fibres in response bladder distension. The enhanced activity was seen in both low and high threshold nerves suggesting that stressed animals may experience enhanced bladder filling sensations at lower bladder volumes as well as increased pain sensations, both potentially contributing to the increased urinary frequency seen after stress.

Chronic psychological stress and lower urinary tract symptoms.

It is well established that lower urinary tract symptoms (LUTS), particularly urinary urgency and incontinence, cause stress and anxiety for patients. However, there is mounting evidence that the relationship between these two factors is bidirectional and that chronic psychological stress itself can result in the development of symptoms such as urinary frequency, urgency, incontinence, and pelvic pain. This review considers the evidence that such a relationship exists and reviews the literature from clinical and animal studies to identify some of the mechanisms that might be involved. Inflammatory responses induced by chronic stress appear to offer the strongest link to bladder dysfunction. There is overwhelming evidence, both in patients and animal models, for a release of pro-inflammatory cytokines and chemokines during periods of chronic stress. Furthermore, cytokines have been shown to cause bladder dysfunction and pain via actions in the central nervous system and locally in the bladder. In the brain and spinal cord, pro-inflammatory cytokines influence the regulation of micturition pathways by corticotropin-releasing factor (CRF) and its receptors, while peripherally cytokines affect bladder function, directly causing detrusor hypertrophy and afferent nerve hypersensitivity. There is little information on which treatments may have most benefit for stressed/anxious patients with LUTS, but animal studies suggest traditional drugs for overactive bladder (solifenacin, mirabegron) are more effective on LUTS than anxiolytic drugs (fluoxetine, imipramine). The preliminary preclinical data for CRF receptor antagonists is not consistent. A clearer understanding of the mechanisms involved in stress-induced LUTS should provide a basis for improved treatment of this condition.

Hypersensitivity of bladder low threshold, wide dynamic range, afferent fibres following treatment with the chemotherapeutic drugs cyclophosphamide and ifosfamide.

The cytotoxic drugs cyclophosphamide (CPO) and ifosfamide (IFO) cause toxic urological effects due to the production of urinary metabolites that cause bladder inflammation. This study aimed to identify changes in the bladder afferent system following treatment with these drugs that might explain reported urological adverse effects. Intravesical pressure and afferent nerve activity were recorded during bladder distension and drug administration in isolated bladders from mice, 24 h after intraperitoneal treatment with cyclophosphamide (100 mg/kg), ifosphamide (200 mg/kg) or saline (control). In isolated bladders, total afferent nerve activity at maximum bladder distension was increased from 182 ± 13 imp/s in control animals, to 230 ± 14 imp/s in CPO-treated (p < 0.05) and 226 ± 17 imp/s in IFO-treated (p < 0.001) mice. Single fibre analysis revealed the increase resulted from an enhanced activity in low threshold, wide dynamic range fibres (23.3 ± 1.9 imp/s/fibre in controls to 31.5 ± 2.5 (p < 0.01) in CPO and 29.9 ± 2.0 imp/s/fibre (p < 0.05) in IFO treated). CPO treatment was accompanied by an increase in urinary frequency in vivo, but was not associated with increases in urothelial release of ATP or acetylcholine, bladder compliance or spontaneous muscle activity. Also, CPO-treatment did not affect afferent nerve responses or pressure responses to purinergic, muscarinic or nicotinic agonists. This is the first report of CPO and IFO-induced changes in specific populations of bladder afferents, namely an increase in low threshold, wide dynamic range fibres. These effects appear to be direct and not secondary to increases in smooth muscle activity or the release of urothelial mediators.

Novel insights into the mechanism of cyclophosphamide-induced bladder toxicity: chloroacetaldehyde's contribution to urothelial dysfunction in vitro.

The clinical use of cyclophosphamide and ifosfamide is limited by a resultant bladder toxicity which has been attributed to the metabolite acrolein. Another metabolite chloroacetaldehyde (CAA) associated with nephrotoxicity, has not been investigated for toxicity in the bladder and this study investigates the effects of acrolein and CAA on human urothelial cells in vitro. Human urothelial cells (RT4 and T24) were treated with acrolein or CAA and changes in cell viability, reactive oxygen species, caspase-3 activity and release of urothelial mediators ATP, acetylcholine, PGE2 were measured. The protective effects of N-acetyl cysteine (NAC) were also assessed. Both metabolites were toxic to human urothelial cells, however, CAA significantly decreased cell viability at a ten-fold lower concentration (10 µM) than acrolein (100 µM). This was associated with increased ROS production and caspase-3 activity. NAC protected cells from these changes. In RT4 cells 100 µM acrolein caused a significant increase in basal and stretch-induced ATP, Ach and PGE2 release. In T24 cells chloroacetaldehyde (10 µM) increased basal and stimulated ATP and PGE2 levels. Again, NAC protected against changes in urothelial mediator release following acrolein or CAA. This study is the first to report that CAA in addition to acrolein contributes to the urotoxicity of cyclophosphamide and ifosfamide. Both metabolites altered urothelial mediator levels which could contribute to the sensory and functional bladder changes experienced by patients after treatment with cyclophosphamide or ifosfamide. Alterations in urothelial cell viability and mediator release may be causally linked to oxidative stress, with NAC providing protection against these changes.

Three Gaseous Neurotransmitters, Nitric oxide, Carbon Monoxide, and Hydrogen Sulfide, Are Involved in the Neurogenic Relaxation Responses of the Porcine Internal Anal Sphincter.

BACKGROUND/AIMS: The internal anal sphincter (IAS) plays an important role in maintaining continence and a number of neurotransmitters are known to regulate IAS tone. The aim of this study was to determine the relative importance of the neurotransmitters involved in the relaxant and contractile responses of the porcine IAS. METHODS: Responses of isolated strips of IAS to electrical field stimulation (EFS) were obtained in the absence and presence of inhibitors of neurotransmitter systems. RESULTS: Contractile responses of the sphincter to EFS were unaffected by the muscarinic receptor antagonist, atropine (1 µM), but were almost completely abolished by the adrenergic neuron blocker guanethidine (10 µM). Contractile responses were also reduced (by 45% at 5 Hz, P < 0.01) following desensitisation of purinergic receptors with α,ß-methylene-ATP (10 µM). In the presence of guanethidine, atropine, and α,ß-methylene-ATP, the remaining relaxatory responses to EFS were examined. These responses were not altered by the cyclooxygenase inhibitor, indomethacin (5 µM), the vasoactive intestinal polypeptide receptor antagonist, [D-p-Cl-Phe(6),Leu(17)]-vasoactive intestinal peptide (PheLeu-VIP; 100 nM), or the purinoceptor antagonists, 8-phenyltheophyline (P1 receptors) or suramin (P2 receptors). However, relaxation responses were reduced by Nω-nitro-L-arginine (L-NNA; 100 µM), an inhibitor of nitric oxide synthesis (40-50% reduction), zinc protoprophyrin IX (10 µM), an inhibitor of carbon monoxide synthesis (20-40% reduction), and also propargylglycine (30 µM) and aminooxyacetic acid (30 µM), inhibitors of hydrogen sulphide synthesis (15-20% reduction). CONCLUSIONS: Stimulation of IAS efferent nerves releases excitatory and inhibitory neurotransmitters: noradrenaline is the predominant contractile transmitter with a smaller component from ATP, whilst 3 gases mediate relaxation responses to EFS, with the combined contributions being nitric oxide > carbon monoxide > hydrogen sulfide.