Peripheral Neuromodulation For Overactive Bladder

IntroductionOveractive bladder (OAB) is defined as the presence of urinary urgency, typically accompanied by frequency and nocturia, with or without urgency incontinence. The etiology and pathophysiology of OAB are still unknown. Conservative treatment for OAB includes behavioral modification and medical therapy such as anti-muscarinics. Though clinically effective, anti-muscarinics still have common side effects including dry mouth, constipation, blurred vision, and cognitive impairment leading to poor compliance and unmet expectations. Neuromodulation is an attractive alternative when conservative treatment fails.Neuromodulation works through afferent nerve stimulation to the sacral nerve root (S2-4) to modulate the function of bladder and/or urethral sphincters to improve the symptoms of OAB. Sacral neuromodulation (SNM) was the first pathway for clinical neuromodulation for OAB. Common complications of SNM include pain and/or infection at the implantation site, lead migration and battery failure. Therefore, less invasive pathways including tibial nerve and pudendal nerve emerged for neuromodulation. However, the mechanisms underlying neuromodulation of OAB are still uncertain. Most studies focus on two aspects:the anatomical site of action and nerotransmitters involved in neuromodulation.The lower urinary tract is innervated by an integrated afferent and efferent neuronal complex of peripheral neural circuits involving sympathetic, parasympathetic, and somatic neurons. It is known that reflex bladder activity is controlled by two distinct neural pathways-a spinobulbospinal micturition reflex pathway activated by non-nociceptive Aδ afferents and a spinal micturition reflex pathway activated by nociceptive C-fiber afferents. The potential sites of neuromodulation action consist of spinal cord and peripheral nerve system, and supraspinal system e.g. brain and pontine. A clinical study found that early SNM implantation in patients with spinal cord injury (SCI) prevented detrusor overactivity and urinary incontinence suggesting that SNM works at spinal cord. Acetic acid (AA) induced nociceptive bladder reflex in cats. Bladder overactivity was inhibited by tibial nerve stimulation (TNS) and pudendal nerve stimulation (PNS) mediated by inhibitory effect of interneurons in spinal cord on parasympathetic efferent pathway.In cats with chronic spinal cord injury, electrical stimulation of the pudendal nerve on one side at different frequencies induced either inhibitory or excitatory effects on bladder activity. Low frequency (3-10Hz) stimulation elicited an inhibitory effect, while higher frequency (20-40Hz) stimulation produced a facilitatory effect on bladder activity. Excitatory and inhibitory pathways can be activated differentially at the level of the pudendal nerve according to stimulus frequency. The ability to control selectively both continence and micturition with a single electrode on a peripheral nerve is an exciting prospect for neurorehabilitation.Animal studies have found that various neurotransmitters are involved in neuromodulation for OAB, e.g. inhibitory amino acids y-aminobutyric acid (GABA),5-hydroxytryptamine (5-HT), opioid peptide, and glutamate. It has been reported that AA was used to irritate the bladder and induce nociceptive bladder overactivity, while saline was used to distend the bladder and induce non-nociceptive bladder activity in cats. TNS has an inhibitory effect on both nociceptive and non-nociceptive bladder activities. Furthermore, opioid receptors are involved in nociceptive rather than non-nociceptive bladder reflex. However, opioid receptors are not involved in pudendal inhibition of bladder overactivity but metabotropic glutamate5receptors are partially involved. Therefore, it is speculated that multiple neurotransmitters are involved in pudendal inhibition of nociceptive and/or non-nociceptive bladder reflexes.GABA which is a major inhibitory neurotransmitter at both spinal and supraspinal synapses has been implicated in the control of micturition in animals. In cats, application of GABA or GABAA receptor agonists by the intrathecal route, iontophoretically to bladder parasympathetic preganglionic neurons in the sacral spinal cord inhibits preganglionic neuron firing and/or reflex bladder activity. Because PNS also elicits inhibitory postsynaptic potentials in bladder parasympathetic preganglionic neurons in the cat, we hypothesize that pudendal neuromodulation might inhibit bladder overactivity by stimulating spinal GABAergic inhibitory mechanisms.This study consists of two parts:(1) Picrotoxin (a GABAA receptor antagonist) was administered intravenously (i.v.) or intrathecally (i.t.) to examine the role of GABAA receptors in PNS inhibition by using anesthetized cats.(2) The effects of tibial and pudendal nerve stimulation on reflex bladder activity were investigated in cats with acute cord injury at T9-10under a-chloralose anesthesia. Under isovolumetric contraction, electrical stimulation of tibial and pudendal nerve on one side at different frequencies were performed to study bladder activity. Hexamethonium bromide was intravenously administered followed by intrathecal injection lidocaine at L6-7to examine the amplitude changes of bladder contraction. This novel animal model was established to study bladder activity mediated by nociceptive spinal reflex and reveal the mechanism of somatic neuromodulation for OAB.Part1:Differential Role of Spinal GABAA Receptors in Pudendal Nerve Inhibition of Nociceptive and Nonnociceptive Bladder Reflexes in CatsObjectiveTo examine the role of spinal GABAA receptors in PNS inhibition of OABMethod1. Surgical procedureThirty-three cats were used in this study. After the animals were anesthetized, a tracheotomy was performed and a tube was inserted to keep the airway patent. A catheter was inserted into right carotid artery to monitor systemic blood pressure. Right and left cephalic veins were catheterized for i.v. administration of drugs and fluid. The ureters were isolated and cut for external drainage. A double lumen catheter was inserted through the urethra into the bladder. One lumen was connected to a pump to slowly infuse saline or0.25%AA. The other lumen was attached to a pressure transducer to measure bladder pressure. The right pudendal nerve was isolated and a tripolar cuff electrode was implanted around the pudendal nerve and connected to a stimulator via a constant voltage stimulus isolator, In10cats, a small incision was made to remove the L3spinal process and expose the spinal cord. Then, the spinal dura was pierced and a fine catheter was inserted caudally underneath the dura to position catheter tip at SI sacral spinal cord for i.t. injection of picrotoxin.2. Stimulation protocolUniphasic rectangular pulses (5Hz frequency,0.2ms pulse width) were used to stimulate the pudendal nerve. The stimulation threshold (T) was defined as the minimal intensity for inducing anal sphincter twitching. PNS of multiple intensity thresholds (2T/4T) was used to inhibit bladder activity.3. Drug administrationIn the first group (N=13), repeated cystometrogram (CMGs) were performed with infusion of0.25%AA to irritate the bladder, activate nociceptive bladder afferent C-fibers, and induce bladder overactivity. Picrotoxin administered intravenously or intrathecally to examine the role of GABAA receptors in PNS inhibition. In the second group (N=20), repeated saline CMGs were performed to activate non-nociceptive Aδ afferent initiated micturition reflexes. Picrotoxin was administered intravenously or intrathecally to examine the role of GABAA receptors in PNS inhibition. CMG data was collected for statistical analysis.4. Data analysisCMGs were performed by slowly infusing the bladder with saline to determine the bladder capacity that is defined as the bladder volume threshold required to induce a micturition contraction of large amplitude (>30cmH2O) and long duration (>20seconds). For each CMG bladder capacity was normalized to the initial saline control capacity in the same animal, which allowed for comparisons between animals. The bladder capacities were averaged for each condition and reported with standard error of the mean. Student T-test or ANOVA followed by Dunnett or Bonferroni post-tests was used to determine statistical significance (p<0.05). Result1. AA irritation significantly (p<0.01) reduced bladder capacity to34.3±7.1%of the saline control capacity; while PNS at2T and4T significantly (p<0.01) increased AA bladder capacity to84.0±7.8%and93.2±15.0%, respectively, of the saline control.2. Picrotoxin (0.4mg, i.t.) did not change AA bladder capacity but completely removed PNS inhibition of AA-induced bladder overactivity.3. Picrotoxin (i.v.) only increased AA bladder capacity at a high dose (0.3mg/kg) but significantly (p<0.05) reduced2T PNS inhibition at low doses (0.01-0.1mg/kg).4. During saline cystometry, PNS significantly (p<0.01) increased bladder capacity to147.0±7.6%at2T and172.7±8.9%at4T of control capacity; and picrotoxin (0.4mg, i.t. or0.03-0.3mg/kg, i.v.) also significantly (p<0.05) increased bladder capacity.5. Picrotoxin treatment did not alter PNS inhibition during saline infusion.ConclusionAA-induce bladder overactivety can be inhibitited by PNS and spinal GABAA receptor is involved in PNS inhibition of nociceptive bladder reflex mediated by C-fiber. PNS also has an inhibitory effect on saline distance induced bladder reflex and spinal GABAA receptor is not involved in PNS inhibition of non-nociceptive bladder reflex mediated by Aδ fiber. Spinal GABAA receptors have a tonic facilitatory role of in the control of normal bladder reflex activity. These results indicate that spinal GABAA receptors have different roles in controlling nociceptive and non-nociceptive reflex bladder activities and in PNS inhibition of these activities.Part2:Somatic Neuromodulation of Bladder Activity Mediated by Nociceptive Spinal Reflex in CatsObjectiveTo establish an animal model to study bladder activity mediated by nociceptive spinal reflex and study the mechanism of somatic neuromodulation for OAB Method1. Surgical procedureTwelve cats were used in this study. After the animals were anesthetized, a tracheotomy was performed and a tube was inserted to keep the airway patent. A catheter was inserted into right carotid artery to monitor systemic blood pressure. Right cephalic vein was catheterized for i.v. administration of drugs and fluid. The ureters were isolated and cut for external drainage. A double lumen catheter was inserted through the urethra into the bladder. One lumen was connected to a pump to slowly infuse saline or0.25%AA. The other lumen was attached to a pressure transducer to measure bladder pressure. The right pudendal nerve and left tibial nerve were isolated and a tripolar and bipolar cuff electrodes were implanted around nerves and connected to a stimulator via a constant voltage stimulus isolator. A small incision was made to remove the T9-10spinal process and expose the spinal cord for complete SCI later. Laminectomy was performed at the level of L6-7spinal cord for i.t. administration of lidocaine.2. Stimulation protocolUniphasic rectangular pulses (5Hz frequency,0.2ms pulse width) were used to stimulate the pudendal and tibial nerve. The stimulation threshold (T) was defined as the minimal intensity for inducing anal sphincter twitching or left toe movement. PNS and TNS of multiple intensity thresholds (2T/4T) were used to inhibit bladder activity.3. CystometrogramRepeated CMGs were performed with infusion of saline to determine bladder control capacity. After SCI,0.25%AA was infused to irritate the bladder and induce bladder overactivity. When the bladder capacity stabilized, four CMGs were performed with AA infusion:(1) control CMG without PNS,(2) CMG during2T-PNS,(3) CMG during4T-PNS, and (4) control CMG without PNS to determine any post-stimulation effect. When the bladder capacity stabilized, four CMGs were performed with AA infusion:(1) control CMG without TNS,(2) CMG during2T-TNS,(3) CMG during4T-TNS, and (4) control CMG without PNS to determine any post-stimulation effect. 4. The effect of PNS and TNS with different frequencies on isovolumetric bladder activity2T-PNS and4T-TNS (0.2ms pulse width) with different frequencies (0.5,1,5,10,20,40Hz) were performed to deterime the effect of somatic neuromodulation on isovolumetric bladder activity.5. The effect of hexamethonium bromide (i.v.) and lidocaine (i.t.) on isovolumetric bladder activityHexamethonium bromide (10mg/kg) was administered intravenously to examine the changes of bladder pressure. Five minutes later, laminectomy was performed at the level of L6-7spinal cord for i.t. administration of lidocaine to examine the changes of bladder pressure.6. Data analysisFor each CMG bladder capacity was normalized to the initial saline control capacity in the same animal, which allowed for comparisons between animals. Bladder activity was defined as the ratio between area under curve (AUC) during stimulation and AUC before stimulation to determine the effect of PNS and TNS with different frequencies on isovolumetric bladder activity. The amplitude of bladder contraction was measured to study the effect of hexamethonium bromide (i.v.) and lidocaine (i.t.) on isovolumetric bladder activity. All data was expressed with standard error of the mean. Student T-test or ANOVA followed by Dunnett or Bonferroni post-tests was used to determine statistical significance (p<0.05).Result1. After acute SCI, AA irritation significantly (p<0.01) reduced bladder capacity to68.8±6.4%of the saline control capacity. PNS at2T and4T significantly (p<0.01) increased A A bladder capacity to92.4±12.0%and107.6±1.7%, respectively, of the saline control; while TNS at2T and4T did not alter bladder capacity.2. After intravenous injection of hexamethonium bromide, the isovolumetric bladder activity was significantly decreased from19.3±2.9to8.4±1.9cndH2O (p<0.01). After intrathecal injection of2%lidocaine, the isovolumetric bladder activity was significantly decreased from8.4±1.9to3.9±1.0cmH20(p<0.01).3. The isovolumetric bladder activity was significantly inhibited by2T-PNS at0.5,1,5and40Hz.2T-PNS at10or20Hz and4T-TNS at0.5,1,5,10,20or40Hz did not alter isovolumetric bladder activity. ConclusionOur results demonstrate that AA-induce nociceptive bladder activity after acute SCI can be inhibited by PNS; while TNS has no effect on bladder overactivity. The inhibitory effect of PNS is frequency dependent. The spinal nociceptive bladder activity can be partially blocked by intravenous hexamethonium bromide injection. This novel animal model of nociceptive bladder activity provides a new approach to study the mechanism of somatic neuromodulation for OAB.