Nociceptin/orphanin FQ receptor expression in clinical pain disorders and functional effects in cultured neurons.

The nociceptin/orphanin FQ peptide receptor (NOP), activated by its endogenous peptide ligand nociceptin/orphanin FQ (N/OFQ), exerts several effects including modulation of pain signalling. We have examined, for the first time, the tissue distribution of the NOP receptor in clinical visceral and somatic pain disorders by immunohistochemistry and assessed functional effects of NOP and µ-opioid receptor activation in cultured human and rat dorsal root ganglion (DRG) neurons. Quantification of NOP-positive nerve fibres within the bladder suburothelium revealed a remarkable several-fold increase in detrusor overactivity (P < 0.0001) and painful bladder syndrome patient specimens (P = 0.0014) compared with controls. In postmortem control human DRG, 75% to 80% of small/medium neurons (≤50 µm diameter) in the lumbar (somatic) and sacral (visceral) DRG were positive for NOP, and fewer large neurons; avulsion-injured cervical human DRG neurons showed similar numbers. NOP immunoreactivity was significantly decreased in injured peripheral nerves (P = 0.0004), and also in painful neuromas (P = 0.025). Calcium-imaging studies in cultured rat DRG neurons demonstrated dose-dependent inhibition of capsaicin responses in the presence of N/OFQ, with an IC50 of 8.6 pM. In cultured human DRG neurons, 32% inhibition of capsaicin responses was observed in the presence of 1 pM N/OFQ (P < 0.001). The maximum inhibition of capsaicin responses was greater with N/OFQ than µ-opioid receptor agonist DAMGO. Our findings highlight the potential of NOP agonists, particularly in urinary bladder overactivity and pain syndromes. The regulation of NOP expression in visceral and somatic sensory neurons by target-derived neurotrophic factors deserves further study, and the efficacy of NOP selective agonists in clinical trials.

Increased levels of SV2A botulinum neurotoxin receptor in clinical sensory disorders and functional effects of botulinum toxins A and E in cultured human sensory neurons.

BACKGROUND: There is increasing evidence that botulinum neurotoxin A may affect sensory nociceptor fibers, but the expression of its receptors in clinical pain states, and its effects in human sensory neurons, are largely unknown. METHODS: We studied synaptic vesicle protein subtype SV2A, a receptor for botulinum neurotoxin A, by immunostaining in a range of clinical tissues, including human dorsal root ganglion sensory neurons, peripheral nerves, the urinary bladder, and the colon. We also determined the effects of botulinum neurotoxins A and E on localization of the capsaicin receptor, TRPV1, and functional sensitivity to capsaicin stimuli in cultured human dorsal root ganglion neurons. RESULTS: Image analysis showed that SV2A immunoreactive nerve fibers were increased in injured nerves proximal to the injury (P = 0.002), and in painful neuromas (P = 0.0027); the ratio of percentage area SV2A to neurofilaments (a structural marker) was increased proximal to injury (P = 0.0022) and in neuromas (P = 0.0001), indicating increased SV2A levels in injured nerve fibers. In the urinary bladder, SV2A nerve fibers were found in detrusor muscle and associated with blood vessels, with a significant increase in idiopathic detrusor over-activity (P = 0.002) and painful bladder syndrome (P = 0.0087). Colon biopsies showed numerous SV2A-positive nerve fibers, which were increased in quiescent inflammatory bowel disease with abdominal pain (P = 0.023), but not in inflammatory bowel disease without abdominal pain (P = 0.77) or in irritable bowel syndrome (P = 0.13). In vitro studies of botulinum neurotoxin A-treated and botulinum neurotoxin E-treated cultured human sensory neurons showed accumulation of cytoplasmic vesicles, neurite loss, and reduced immunofluorescence for the heat and capsaicin receptor, TRPV1. Functional effects included dose-related inhibition of capsaicin responses on calcium imaging after acute treatment with botulinum neurotoxins A and E. CONCLUSION: Differential levels of SV2A protein expression in clinical disorders may identify potential new targets for botulinum neurotoxin therapy. In vitro studies indicate that treatment with botulinum neurotoxins A and E may affect receptor expression and nociceptor function in sensory neurons.

Increased cannabinoid receptor 1-immunoreactive nerve fibers in overactive and painful bladder disorders and their correlation with symptoms.

OBJECTIVE: To study the expression of cannabinoid receptor 1 (CB1) in human urinary bladder hypersensitivity and overactivity disorders, and correlate changes with symptoms. Cannabinoid receptor agonists have been shown to modulate urinary bladder contractility and reduce pain after bladder inflammation; their clinical efficacy on lower urinary tract symptoms was demonstrated in the Cannabinoids in Multiple Sclerosis study. METHODS: Bladder tissue specimens were obtained from patients with painful bladder syndrome (PBS, n=13), idiopathic detrusor overactivity (IDO, n=14), and from controls with asymptomatic microscopic hematuria (n=16). The severity of symptoms was assessed using the Pelvic Pain and Urgency/Frequency Questionnaire. Pain score was also recorded on a visual analogue scale. Specimens were immunostained using specific antibodies to CB1 and to neurofilaments as a structural maker. Detrusor and suburothelial nerve fiber density was quantified with a visual grading scale. The immunohistochemistry results were correlated with "Pain, Frequency, and Urgency" scores. RESULTS: CB1-immunoreative nerve fibers were significantly increased in the suburothelium of PBS (P=.0123) and IDO (P=.0013) specimens, and in detrusor layer in IDO (P=.0003), as compared with controls. CB1-immunoreactive suburothelial nerve fiber density correlated significantly with pain scores (Visual Analogue Scale) in PBS (r=.6878, P=.0347) and urgency scores in IDO (r=.6623, P=.0027). Neurofilaments-immunoreactive suburothelial nerve fibers were significantly increased in PBS (P=.019) and IDO (P=.05). CONCLUSIONS: The results of this study suggest that increased nerve fibers, which express CB1, may be related to bladder pain in PBS and urgency in IDO. Our findings support clinical trials of CB1 agonists in bladder disorders.

Complex hydrocephalus (combination of communicating and obstructive type): an important cause of failed endoscopic third ventriculostomy.

BACKGROUND: Hydrocephalus can be classified as purely obstructive, purely communicating or due to combinations of pathologies (obstruction in addition to defective absorption). Endoscopic third ventriculostomy (ETV) as an alternative to shunt procedures is an established treatment for obstructive hydrocephalus. However, patients who have combination of pathologies (complex hydrocephalus) could result in failure of ETV in spite of a patent stoma. The aim of this study was to prospectively evaluate the incidence of complex hydrocephalus in patients with obstructive hydrocephalus who failed an endoscopic third ventriculostomy. FINDINGS: Seventy one patients of obstructive hydrocephalus who underwent ETV in our institution were included in this study. Aetiology of hydrocephalus included congenital aqueductal stenosis in 42 and tubercular meningitis (TBM) in 29 patients. Failure of ETV was seen in 15 (21%) patients. These 15 patients included 6 (14.3%) from the congenital group and 9 (31.0%) patients from the TBM group. Iohexol CT ventriculography confirmed a patent stoma (suggesting a complex hydrocephalus) in 10 (66.7%) out of the 15 failed ETV cases. The incidence of complex hydrocephalus was more common in TBM group (8/29 patients, 27.60%) compared to congenital group (2/42 patients, 4.8%). The complex hydrocephalus patients with a patent ETV stoma were successfully managed by a lumbar peritoneal (LP) shunt. CONCLUSION: Ten out of the 71 patients (14%) with obstructive hydrocephalus who underwent an ETV had a complex hydrocephalus, which was the major (66.7%) cause for failure of ETV. Improving methods to detect the exact type of hydrocephalus pre-operatively could increase success rate of ETV and avoid an unnecessary operative procedure (ETV).

A survey of the peri-operative management of urological patients on clopidogrel.

INTRODUCTION: Peri-operative management of patients receiving platelet inhibitors, such as clopidogrel presents a dilemma to surgeons in every surgical specialty including urology. The risk of procedure-related bleeding while continuing clopidogrel needs to be weighed against the risk of thrombo-embolism after discontinuing it. The objective of the survey was to determine current UK practice regarding clopidogrel use/cessation in patients undergoing elective urological procedures. SUBJECTS AND METHODS: A 10-part questionnaire relating to pre- and postoperative clopidogrel use was mailed to all UK urology consultants listed in the British Association of Urological Surgeons' directory. RESULTS: A total of 570 questionnaires were sent and 297 (52%) were returned. The majority of respondents stop clopidogrel prior to TUR surgery (96.6%), major urological surgery (91.7%), TRUS biopsy (90.6%), ESWL (81.8%) and cystoscopy and biopsy (70.1%). The time clopidogrel was stopped pre-operatively and restarted postoperatively was very variable and dependent on local guidelines or urologist preference. Almost half (49.5%) of the respondents would stop clopidogrel irrespective of its indication and 40.7% never consulted a cardiologist/haematologist before stopping clopidogrel. Less than half (43.4%) had a protocol/guideline in place concerning stopping clopidogrel before surgery. Of respondents, 43% do not routinely prescribe bridging therapy after discontinuing clopidogrel. Over half (55%) reported bleeding complications in patients who continued their clopidogrel during urological procedures and 22 (7.4%) of respondents reported an adverse thrombo-embolic event after stopping clopidogrel. The vast majority of respondents (92.8%) felt evidence-based guidelines on clopidogrel use during the peri-operative period would be useful. CONCLUSIONS: This survey has highlighted a significant variation in practice with regards to pre- and postoperative management of clopidogrel in patients undergoing urological procedures. The results of this survey highlight the need for evidence-based guidelines for the peri-operative management of patients on clopidogrel.

The impact of nutritional status on the outcome of Indian patients undergoing neurosurgical shunt surgery.

Undernutrition is common in surgical patients, is frequently unrecognised and is strongly associated with adverse outcomes such as high rates of complications and mortality, worsening functional status and prolonged hospitalisation. Owing to the associated infection and symptoms such as repeated vomiting, a high prevalence of undernutrition is expected in hydrocephalus patients, which may contribute to their poor surgical outcomes. The aim of this study was to evaluate the influence of preoperative nutritional status on the outcome of Indian patients with hydrocephalus undergoing neurosurgical shunt surgery. One hundred and twenty-four consecutive patients undergoing scheduled hydrocephalus shunt surgery were studied prospectively. All patients underwent nutritional screening according to different parameters prior to surgery. The patients were classified into normally nourished and undernourished groups. The undernourished group was further subdivided into moderately and severely undernourished. The surgical outcome was compared between these groups. A high prevalence (53%) of undernutrition was observed in these patients. Postoperative complications such as shunt infection (P = 0.0023), shunt revision (P = 0.0074) and mortality (P = 0.0003) were significantly more common in undernourished patients compared with normally nourished patients. Serum albumin emerged as the most significant independent predictor of postoperative mortality. The present study demonstrated a high prevalence of undernutrition in hydrocephalus patients in India and its adverse influence on the outcome of shunt surgery. Early preoperative nutritional status screening and its optimisation may decrease the morbidity and mortality of shunt surgery for hydrocephalus.

Pediatric intramedullary schwannoma without neurofibromatosis.

Pediatric intramedullary schwannoma without neurofibromatosis is extremely rare with only five cases reported so far. We present this rare finding in an 8-year-old boy who presented with a sudden onset of weakness in all limbs. An intraoperative diagnosis of schwannoma enabled us to carry out a total excision of the tumor, which resulted in near complete recovery at 18 months follow-up. Although rare, this diagnosis should be considered when a child presents with a solitary intramedullary tumor, since its total resection can be achieved improving surgical outcome.

Endoscopic management of hypertensive intraventricular haemorrhage with obstructive hydrocephalus.

BACKGROUND: Intracranial haemorrhage accounts for 30-60 % of all stroke admissions into a hospital, with hypertension being the main risk factor. Presence of intraventricular haematoma is considered a poor prognostic factor due to the resultant obstruction to CSF and the mass effect following the presence of blood resulting in raised intracranial pressure and hydrocephalus. We report the results following endoscopic decompression of obstructive hydrocephalus and evacuation of haematoma in patients with hypertensive intraventricular haemorrhage. METHODS: During a two year period, 25 patients diagnosed as having an intraventricular haemorrhage with obstructive hydrocephalus secondary to hypertension were included in this study. All patients underwent endoscopic evacuation of the haematoma under general anaesthesia. Post operative evaluation was done by CT scan and Glasgow outcome scale. RESULTS: Of the 25 patients, thalamic haemorrhage was observed in 12 (48%) patients, while, 11 (44%) had a putaminal haematoma. Nine (36%) patients had a GCS of 8 or less pre-operatively. Resolution of hydrocephalus following endoscopic evacuation was observed in 24 (96%) patients. No complications directly related to the surgical technique were encountered in our study. At six months follow-up, a mortality rate of 6.3% and 55.5% was observed in patients with a pre-operative GCS of > or = 9 and < or = 8 respectively. Thirteen of the 16 (81.3%) patients with a pre-operative GCS >/= 9 had good recovery. CONCLUSION: Endoscopic technique offers encouraging results in relieving hydrocephalus in hypertensive intraventricular haemorrhage. Final outcome is better in patient with a pre-operative GCS of >9. Future improvements in instrumentation and surgical techniques, with careful case selection may help improve outcome in these patients.

Pain during ice water test distinguishes clinical bladder hypersensitivity from overactivity disorders.

BACKGROUND: The Bladder cooling reflex (BCR) i.e. uninhibited detrusor contractions evoked by intravesical instillation of cold saline, is a segmental reflex believed to be triggered by menthol sensitive cold receptors in the bladder wall, with the afferent signals transmitted by C fibres. The BCR is a neonatal reflex that becomes suppressed by descending signals from higher centres at approximately the time when the child gains full voluntary control of voiding. It re-emerges in adults with neurogenic detrusor overactivity as a consequence of loss of central descending inhibition, resulting from conditions such as spinal cord injury or multiple sclerosis. We have recently shown an increase of nerve fibres expressing the cool and menthol receptor TRPM8 in both overactive (IDO) and painful bladder syndrome (PBS), but its functional significance is unknown. We have therefore studied the bladder cooling reflex and associated sensory symptoms in patients with PBS and overactivity disorders. METHODS: The BCR, elicited by ice water test (IWT) was performed in patients with painful bladder syndrome (PBS, n = 17), idiopathic detrusor overactivity (IDO, n = 22), neurogenic detrusor overactivity (NDO, n = 4) and stress urinary incontinence (as controls, n = 21). The IWT was performed by intravesical instillation of cold saline (0 - 4 degrees C). A positive IWT was defined as presence of uninhibited detrusor contraction evoked by cold saline, associated with urgency or with fluid expulsion. Patients were asked to report and rate any pain and cold sensation during the test. RESULTS: A positive IWT was observed in IDO (6/22, 27.3%) and NDO (4/4, 100%) patients, but was negative in all control and PBS patients. Thirteen (76.5%) PBS patients reported pain during the IWT, with significantly higher pain scores during ice water instillation compared to the baseline (P = 0.0002), or equivalent amount of bladder filling (100 mls) with saline at room temperature (P = 0.015). None of the control or overactive (NDO/IDO) patients reported any pain during the IWT. CONCLUSION: The BCR in DO may reflect loss of central inhibition, which appears necessary to elicit this reflex; the pain elicited in PBS suggests afferent sensitisation, hence sensory symptoms are evoked but not reflex detrusor contractions. The ice water test may be a useful and simple marker for clinical trials in PBS, particularly for novel selective TRPM8 antagonists.

Transient receptor potential vanilloid receptor subtype 1 in painful bladder syndrome and its correlation with pain.

PURPOSE: Painful bladder syndrome is a chronic, debilitating bladder hypersensitivity disorder characterized by urinary frequency, urgency and bladder pain without an identifiable cause. Recent advances in understanding the molecular basis of hypersensitivity provide an opportunity to advance the understanding of and treatment for painful bladder syndrome. We studied the heat and capsaicin receptor transient receptor potential vanilloid receptor subtype 1 in the bladder in patients with painful bladder syndrome and their relationship to pain symptoms. MATERIALS AND METHODS: Bladder biopsies were obtained from 20 characterized subjects with painful bladder syndrome and 25 with asymptomatic microscopic hematuria as controls. Specimens were immunostained using specific antibodies to transient receptor potential vanilloid receptor subtype 1 and neurofilaments as a structural maker. Nerve fiber and urothelial staining were quantified with computerized image analysis. The results of immunohistochemistry were correlated with the pain score. RESULTS: There was a marked increase in suburothelial nerve fibers expressing transient receptor potential vanilloid receptor subtype 1 in painful bladder syndrome in comparison with that in controls (p <0.0001). The ratio of transient receptor potential vanilloid receptor subtype 1 fibers to neurofilaments was also significantly increased in painful bladder syndrome, suggesting over expression of transient receptor potential vanilloid receptor subtype 1 (p <0.0001). When all specimens studied were included, the pain score correlated significantly with the relative nerve fiber density of transient receptor potential vanilloid receptor subtype 1 in the suburothelium (r = 0.6862, p = 0.0002) as well as the ratio of transient receptor potential vanilloid receptor subtype 1 fibers to neurofilaments (r = 0.5554, p = 0.004). Urothelial transient receptor potential vanilloid receptor subtype 1 showed a tendency toward an increase in the painful bladder syndrome group but it did not achieve statistical significance. No correlation was found between transient receptor potential vanilloid receptor subtype 1 immunoreactivity of urothelium or neurofilament fibers and the pain score. CONCLUSIONS: This study shows increased transient receptor potential vanilloid receptor subtype 1 in nerve fibers of the bladder in painful bladder syndrome and a correlation of the pain score with the relative density of transient receptor potential vanilloid receptor subtype 1 nerve fibers. Transient receptor potential vanilloid receptor subtype 1 may have a role in the pathophysiology of painful bladder syndrome and it is a potential target for novel therapeutic agents.

Localization of M2 and M3 muscarinic receptors in human bladder disorders and their clinical correlations.

PURPOSE: We studied the cellular localization of muscarinic receptor subtypes 2 and 3 in the human bladder and related any changes in overactive and painful bladder syndromes to measures of clinical dysfunction. MATERIALS AND METHODS: Bladder specimens obtained from patients with painful bladder syndrome (11), idiopathic detrusor overactivity (12) and from controls with asymptomatic microscopic hematuria (16) were immunostained using specific antibodies to muscarinic receptor subtypes 2 and 3, and to vimentin, which is a marker for myofibroblasts. Immunostaining results were quantified with computerized image analysis and correlated with clinical dysfunction using frequency and urgency scores. RESULTS: Muscarinic receptor subtype 2 and 3 immunoreactivity was observed in the urothelium, nerve fibers and detrusor layers. In addition, strong myofibroblast-like cell staining, similar to vimentin, was present in the suburothelial region and detrusor muscle. A significant increase in suburothelial myofibroblast-like muscarinic receptor subtype 2 immunoreactivity was seen in patients with painful bladder syndrome (p = 0.0062) and idiopathic detrusor overactivity (p = 0.0002), and in muscarinic receptor subtype 3 immunoreactivity in those with idiopathic detrusor overactivity (p = 0.0122) with a trend in painful bladder syndrome. Muscarinic receptor subtype 2 and 3 immunoreactivity significantly correlated with the urgency score (p = 0.0002 and 0.0206, respectively) and muscarinic receptor subtype 2 immunoreactivity correlated with the frequency score (p = 0.0029). No significant difference was seen in urothelial and detrusor muscarinic receptor subtypes 2 and 3 or vimentin immunostaining. CONCLUSIONS: To our knowledge this is the first study to show the cellular localization of muscarinic receptor subtypes 2 and 3 in the human bladder. The increase in muscarinic receptor subtypes 2 and 3 immunostaining in myofibroblast-like cells in clinical bladder syndromes and its correlation with clinical scores suggests a potential role in pathophysiological mechanisms and the therapeutic effect of anti-muscarinic agents.

Cool and menthol receptor TRPM8 in human urinary bladder disorders and clinical correlations.

BACKGROUND: The recent identification of the cold-menthol sensory receptor (TRPM8; CMR1), provides us with an opportunity to advance our understanding of its role in the pathophysiology of bladder dysfunction, and its potential mediation of the bladder cooling reflex. In this study, we report the distribution of the cool and menthol receptor TRPM8 in the urinary bladder in patients with overactive and painful bladder syndromes, and its relationship with clinical symptoms. METHODS: Bladder specimens obtained from patients with painful bladder syndrome (PBS, n = 16), idiopathic detrusor overactivity (IDO, n = 14), and asymptomatic microscopic hematuria (controls, n = 17), were immunostained using specific antibodies to TRPM8; nerve fibre and urothelial immunostaining were analysed using fibre counts and computerized image analysis respectively. The results of immunohistochemistry were compared between the groups and correlated with the Pain, Frequency and Urgency scores. RESULTS: TRPM8-immunoreactive staining was observed in the urothelium and nerve fibres scattered in the suburothelium. The nerve fibre staining was seen in fine-calibre axons and thick (myelinated) fibres. There was marked increase of TRPM8-immunoreactive nerve fibres in IDO (P = 0.0249) and PBS (P < 0.0001) specimens, compared with controls. A significantly higher number of TRPM8-immunoreactive axons were also seen in the IDO (P = 0.0246) and PBS (P < 0.0001) groups. Urothelial TRPM8 and TRPM8-immunoreactive thick myelinated fibres appeared unchanged in IDO and PBS. The relative density of TRPM8-immunoreactive nerve fibres significantly correlated with the Frequency (r = 0.5487, P = 0.0004) and Pain (r = 0.6582, P < 0.0001) scores, but not Urgency score. CONCLUSION: This study demonstrates increased TRPM8 in nerve fibres of overactive and painful bladders, and its relationship with clinical symptoms. TRPM8 may play a role in the symptomatology and pathophysiology of these disorders, and may provide an additional target for future overactive and painful bladder pharmacotherapy.