THE NEW TREATMENT OF THE URINARY BLADDER OVERACTIVITY DISORDER WITH BOTOX A-CYSTOSCOPIC INJECTIONS
Semir A. S. Al Samarrai
Botulinum Neurotoxin : A short historical review:
On 14 December 1895 in Ellezelles, Belgium, three persons died after consuming smoked ham sausages. A student of Roberto Koch isolated the responsible organism and named Bacillus botulinum (1), today known as Clostridium botulinum.
In 1946, Edward Schantz succeeded in extracting BoNT-A in crystal form (2, 3). In the 1960s, Alan Scott and Schantz tested the possibility of treating strabismus in monkeys.
Finally in 1978, after the American Federal Drug Agency (AFDA) permitted its use, Scott used BoNT-A injection successfully for treating strabismus in human (4). Carruthers in 1996 published the first paper about its use in cosmetic therapy (5).
In the mid-1990s, it was applied by Naumman for the treatment of hyperhydrosis (6). After its successful application in patients with detrusor sphincter dyssynergia, the research team of Schurch in Zurich (1997) started to show promising results after having done their first injection of BoNT-A in the urinary bladder for treating neurogenic incontinence in patients with spinal cord injuries (7), and for patients suffering from idiopathic overactive bladder, respectively (8).
Mechanism of action in the Bladder wall and muscle:
The pathophysiologic basis of Detrusor overactivity remains incompletely understood; indeed, it is probably a multifactorial problem, with a balance of predisposing and compensatory processes participating to elicit a fluctuating clinical picture.
Furthermore, functional changes at several levels in the regulatory operations of the lower urinary tract (LUT) give rise to seemingly similar clinical features, perhaps as a consequence of limited repertoire of behavior which can be discerned clinically (Detrusor overactivity, urgency and incontinence).
When recognizing the uncertainty as to precisely how botulinum-A-neurotoxin (BoNT-A) achieves its beneficial effect, it is important to consider that overactive urinary bladder is still enigmatic.
A) The Neurogenic Hypothesis:
Understanding of the reflex basis of the micturation cycle was elaborated in the later part of the 19th century by Barrington, who identified the pontine brainstem as the site of ultimate control. This insight indicated the importance of the “pontine micturation centre” (PMC) in regulating the state of the entire LUT, controlling the fundamental determining step of switching between the storing and voiding status.
Consequently, brainstem-mediated inhibition of bladder at spinal and peripheral level, with excitation of the bladder outlet, was perceived to underpin LUT reservoir function, also the LUT-Dysfunction is derived from CNS deficit, this is manifested by abnormal excitation or loss inhibition. Thus, the various strands by which the CNS could underpin overactivity with the bladder muscles were drawn together as “the neurogenic hypothesis”, this can be summarized as explaining overactive contractions by the emergence of abnormal reflex activity. Several processes within the CNS could lead to this. The physiological inhibition of lower centres by the brainstem is fundamental, and it is clear that any lesion affecting such inhibition would allow an abnormal level contractile activity to be expressed peripherally.
Depending on the precise location of any deficit in the CNS, loss of bladder inhibition could allow bladder muscle overactivity to emerge in the bladder. Lesions in other locations could simultaneously affect outlet function, the latter either being reduced (predisposing to Detrusor Overactivity incontinence) or increased (causing Detrusor Overacitvity (DO) with an elevated postvoid residue volume). Theoretically, either abnormal excitation of the pontine micturation centre (PMC) by higher centres, or loss inhibition, could facilitate premature switching of the storage phase over to voiding. At any level of the CNS, reduced inhibition or increased excitation could result from reorganization of reflexes, exemplified by the emergence of C-fiber-mediated activity in spinal injury (9).
B) The Myogenic Hypothesis:
A radical departure from the neurogenic hegemony came with the recognition that similar bladder behavior to bladder muscle Detrusor Overactivity (DO) can arise with no efferent input (10), and that nonneurogenic processes can elicit the same widespread uninhibited detrusor contraction arising without volitional control. The initial review in which the myogenic hypothesis was fully elaborated (11) still represents one of the clearest expositions of the fundamental considerations in overactive bladder disorder, which have to be explained to understand its mechanism basis. Of these, particular emphasis must be placed on increased peripheral excitability, exaggerated propagation and triggering.
Increased excitability refers to the observation of higher levels of spontaneous contraction in detrusor muscle strips isolated from people with DO. The increased excitability could result from alterations in the resting cellular membrane potential, or changes in the cell-surface receptors (‘supersensitivity’).
Certainly, various studies have shown a change in the sensitivity to standardized stimuli (12-16), although this is not invariably the case (17, 18). This supersensitivity could be a result of changes in intracellular pathways, or increased cell expression of surface proteins such as the muscarinic receptors.
The ability of smooth muscle of the bladder to increase cell surface receptor expression is an important consideration, as it signifies the ability of smooth muscle to adapt to change in the paracellular environment. For example, denervation of smooth muscle leads to reduced cholinergic stimulation and elicits up-regulation of cell-surface-cholinergic receptors. Consequently, exogenous application of equivalent doses of cholinergic agonist elicits a more powerful contractile response in denervated muscle than muscle removed from an otherwise normal specimen.
In principle, this might underlie loss of clinical efficacy in long-term use in people with DO who initially responded to anticholinergic drugs. In effect, sustained blockade of muscarinic receptors would produce loss of functional influence of the nerve endings on the muscle cells, leading to receptor up-regulation, which might eventually overcome the effect of the muscarinic antagonist. Subsequent discontinuation of anticholinergic drug would then lead to a ‘rebound’ exacerbation of symptoms. By itself, increased smooth muscle cell excitability of the bladder will not result in Detrusor Overactivity (DO), as spontaneous excitation is normally limited in extent by failure of excitation to spread more than a few millimeters (reflected in a short space constant) (19, 20). Thereby, only a small proportion of the bladder wall would participate in any spontaneous contraction, insufficient to affect intravesical pressure.
However, in people with DO, the propagation of spontaneous excitation appears to disperse over a more substantial proportion of the bladder wall, thereby carrying the potential to generate pressure change (11). This might be a result of the altered detrusor ultrastructures in DO (21), which includes increased expressional intracellular communication channels (22). Triggering refers to the fact that a pure increase in excitability at the cellular level will still not cause increased spontaneous activity unless something actually sets it off (the “trigger”).
Given the response of DO to anticholinergics, acetylcholine (Ach) must be involved at some point, and it is in triggering that Ach is most likely involved in the myogenic hypothesis, where altered properties of smooth muscle are said to underpin DO, the inhibitory effect of BoNT-A on vesicular Ach release (see below) can be observed to be beneficial.
C) Peripheral Cellular Physiology:
For some time the bladder has been regarded as a relatively “simple organ” with the bladder wall muscle (Detrusor) and parasympathetic nerve endings seen as the only relevant bladder structures for the urination process. However, the recent past has seen a burgeoning recognition of the cellular complexity of the bladder wall. The urethelium is a remarkable example of the fundamental reappraisal that is occurring. The presumption that the urethelium is simply a watertight barrier between the stored urine and the bladder wall is now recognized substantially to underestimate its functional potential (23, 24).
Particularly intriguing is the ability of urethelial cells to release mediator substances, including Ach and ATP, and substances that might suppress smooth muscle contraction. The extent of mediator release appears to be influenced by distention, supporting a role in priming or facilitating the voiding contraction.
Clearly, the profile of receptor agonists and inhibitors in the vicinity of the smooth muscle cell will affect its behaviour. Thus, urothelial transmitter release is interesting in the pathophysiological understanding of normal and overactive bladder activity. More subtly, simple anatomical considerations show that the detrusor muscle is separated from the urothelium by a region containing a considerable density of nerve endings. Thus, the site of action of urothelial mediator release might be mediated by peripheral nerves, presumed to be afferents. Interest in putative role of the urothelium in DO is whetted by possibility of manipulating urothelial transmitter release, e.g. through their expression of several surface proteins, including vanilloid, nicotinic and muscarinic receptors (25-27).
The presence of interstitial cells (ICs) is a further intriguing observation (28). Putatively the bladder ICs belong to a multipotent group of functional regulatory cells (29), some of which are capable of pacemaking (exemplified by the interstitial cells of Cajal in the gut) and propagation of excitation. The terminology of this cell class is often misapplied, but myofibroblasts are a specialized cell group which overlays with the ICs (30); some of the bladder ICs might be myofibroblasts, but not all are (31).
Bladder ICs occur in the detrusor layer and the suburothelial region. In the detrusor muscle, they constitute a widespread ramifying network; they are recognized to precipitate calcium waves in the muscle bundle (32). In the suburothelial region, the ICs again provide a widespread communicating network (22). They have been proposed to constitute part of a “sensory transduction triad”; along with the urothelium and afferent nerves (33). The hypothesis suggests that the three cell types act in concert to drive a graded response to bladder filling whose sensitivity (gain) can be modulated as a result of IC contraction (effectively off-loading tension in the afferents). Subtypes of ICs express M3 receptors (34), vanilloid and related receptors (35), which might represent a locus for potential pharmacological manipulation. However, as a possible drug target, it must be remembered that ICs are also in the urethra (36, 37). Thus, beneficial effects on urine storage, by influencing bladder ICs could be counteracted if urethral function is also affected by collateral effects on urethral ICs.
Yet further the reappraisal relates to the peripheral innervations. Historically, the peripheral ganglia nerve consigned to the status of mere “relay” (a relay is an engineering concept, counteracting loss of electrical potential in long pathways by inserting an amplifier which essentially passes on an input unaltered, bar the restoration of potential difference). This dismissive attitude is inappropriate as the complexity of intrinsic wiring, range of inputs and outputs, and presence of neuronal subpopulations clearly indicates a potential for neuronal modulation in the signal distribution at the level of the intramural and perivesical ganglia.
In addition, the recognition of nonandronergic, noncholinergic transmitter substances (38-40) allowed investigators to suggest more functionally significant modulation of neuromuscular transmission and possible relevance in the pathophysiology of DO (41). Furthermore, the transduction of stimuli into afferent information is more complex than previously recognized, both functionally (42) and anatomically (43).
D) Peripheral Integrative Physiology: the autonomous modules:
The sheer scope of peripheral cellular complexity is somewhat confusing. Nonetheless, the cellular complexity does not necessarily alter the fundamental tenets as alluded to above, i.e. that higher CNS centres normally exert tonic inhibition of lower centres, and that spontaneous excitation is relatively infrequent and does not spread far enough to involve a significant proportion of the bladderwall. However, what it does do is provide additional peripheral elements that could have the same contribution to DO as the smooth muscle changes proposed in the myogenic hypothesis.
Thus, exaggerated spontaneous activity could arise from changes in the ICs driving the detrusor (rather than from changes in detrusor cells as in the myogenic hypothesis). Propagation over abnormal distances could result from dissemination through the ramifying network of IC processes (rather than by direct muscle cell to muscle cell communication). Triggering could result from various sources, such as altered urethelial cell mediator release, changes in the transmitter profile of the peripheral innervations, or from the ICs. This is, in effect, the basis of the peripheral autonomy hypothesis as it relates to DO (44), which can be summarized as attributing to various cellular elements (myogenic, neurogenic, ICs and the urethelium) the potential to be responsible for changes in normal physiology that give rise to the processes of triggering, exaggerated excitability and excessive propagation that underpin overactive detrusor contractions. The integrative (peripheral autonomy) hypothesis is thus derived from and closely allied to (indeed, incorporates) the myogenic hypothesis. However, it extends further, as it hypothesis that the complexity of cellular elements must function integratively for normal physiological function, relevant in generation of sensation during urine storage and in reducing the energy expenditure required for efficient emptying during voiding (45). It proposes that the ICs and peripheral innervation interact to form a “myovesical plexus”, which determines crucial physiological aspects of urine storage and voiding (44, 45). Thereby, the theoretical scope for manipulating normal activity and overactive contractions is substantially broadened.
E) The Effect of Botulinum-Toxin (BoNT-A) of Nerve endings:
Having discussed the hypothetical origins of the Detrusor Overactivity of Bladderwall (DO), the cellular effects of BoNT-A must be understood before it is possible to gauge how it could influence DO.
Exposure to BoNT-A results in cascade of processes;
1. Binding to receptors on the membranes of specific nerve endings
2. Internalization via receptor-mediated endocytosis
3. pH-moderated translocation to the cystosol
4. Zinc-dependent endoprotease activity, cleaving, polypeptides essential for transmitter exocytosis.
The specifity of BoNT-A for cholinergic neurons in vivo is due to the presence of specific membrane acceptors/receptors (46). The synaptic vesicle protein SV2 might be the binding site (47), hence BoNT-A might enter more active neurons preferentially.
Mechanisms of action of Botulinum Toxin in treating Bladder Overactivity:
The cellular components in the physical sphere of diffusion after intravesical injection are now easy to ascertain. In fact, when describing intravesical injection, the term”intradetrusor” is exactly correct. All Endo-Urologists are truly able to stipulate the exact location of the injected agent at the time of the procedure. Accordingly, the specific term “intramural” signifies the certaintly about the precise degree to which the intradetrusor and the suburethelial compartments are involved.
The purified BoNT-A has a molecular mass of 150 kDa, the pharmaceutical products have a greater molecular mass. The size of the 900 kDa complex of one of the commercial BoNT-A products reduces diffusion of the toxin within the target muscle (48), along with systemic spread (49); this is relevant when considering the likely cellular targets for the treatment of Detrusor Overactivity (DO) of the urinary bladder.
Numerous factors have to be weighed up including:
1) Safety: a high dose may increase systemic dissemination.
2) Efficacy: insufficient toxin may not diffuse sufficiently to address the clinical indication.
3) Duration of response: administered dose affects duration in some context (50).
4) Health economic considerations.
There are various peripheral loci at which exocytotic transmitter release occurs in the LUT, which could potentially fall within the reach of peripherally injected toxin. These are:
1) Release of neuromuscular transmitters from efferent nerve endings (efferent).
2) Release of transmitters and modulatory substances from the presynaptic nerve terminals within the myovesical plexus (afferent).
3) Release of active mediators from urothelium into the realm of the subjacent myovesical plexus structures (afferent).
4) Interneuron activity in peripheral ganglia or the spinal cord (efferent and afferent).
It is debatable to classify the above points into either “afferent” or “efferent”, given the propensity for fibres attributed to one or other class to express characteristics of the other (e.g. release of efferent transmitters by afferent nerve) (51).
The term “efferent”, simply stated, is based on influencing muscle contractility; given that BoNT-A causes muscle paralysis, reduced muscle contractility in detrusor muscle should reduce overactive contractions.
“Afferent” suggests that reduced sensory information from the lower urinary tract will increase the reflex bladder volume associated with the switching of the pontine micturation center (PMC) from storage to voiding mode.
Release of neuromuscular transmitters from Efferent Nerves:
The effects of botulinum exotoxaemia are largely a result of motor paralysis of skeletal muscle. After localized injection, axotomy-like changes in the motor neurons occur in a few weeks. BoNT-A also affects release in autonomic organs, including several transmitter substances from the bladder (52-55). Accordingly, it is logical to presume that the clinical use of intravesical BoNT-A in the setting of Detrusor Overactivity induces a partial motor paralysis of the bladder. High enough doses would likely counteract voiding activity, assuming all neuromuscular transmitter release is inhibited, necessitating intermittent or indwelling catheterization.
Peripheral Afferent Mechanisms
For some people with idiopathic Bladder Overactivity to achieve the “ideal” of abolished symptoms and preservation of voiding, some other mechanism beyond partial detrusor paralysis is probably involved in the clinical response; because the quantity of sensory information reaching the CNS at any given bladder volume is reduced by the BoNT-A injections, such that a higher intravesical volume can be achieved before the PMC gauges the necessity to instigate switching from storage to voiding mode. It is plausible that fundamental alterations of lower urinary tract activity can arise without the detrusor itself being directly affected (56).
The specificity of BoNT-A for cholinergic neurons is due the presence of specific proteins, such that absence of the relevant surface protein will prevent cellular sensitivity to the toxin.
There is a background release of Ach in the bladder under conditions modeling the storage phase (57).
Overactive urinary bladder is the clinical manifestation of multifactorial pathophysiological processes, reflecting complex CNS and peripheral cellular physiology. BoNT-A alters the release of Ach from cholinergic nerves, and this efferent block might explain the treatment response.
Recently the Drugs regulator of the National Institute for Health and Clinical Excellence (NICE) has ruled that those with or without urge-incontinence should have access to the BoNT-A injections intravesically, if other anticholinergic treatment or other therapy methods of control prove ineffective. The Botox (BoNT-A) treatment might give dramatic improvements for many patients with DO and allows them to resume a normal lifestyle.
The International Continence Society (ICS) has defined painful bladder syndrome as the complaint of suprapubic pain related to bladder filling accompanied by other symptoms such as daytime and night time voiding frequency in the absence of proven urinary infection or other obvious pathology (58).
The European Society for the study of Interstitial Cystitis (ESSIC) in an attempt to use a consistent pain syndrome terminology has preferred the term bladder pain syndrome (BPS). BPS is defined as pelvic pain, pressure, or discomfort perceived to be related to the urinary bladder accompanied by at least one other urinary symptom such as persistent urge to void or increased daytime and night time voiding frequency (59).
The definition of interstitial cystitis (IC) should be restricted to cases that include typical cystoscopic and histological features (58, 59). The etiology of BPS/IC is unknown. Therefore, BPS/IC management is directed to pain relief, as bladder pain is believed to drive both voiding frequency and nocturia. Botulinum toxin (BoNT-A) has been shown to decrease noxious input (60, 61).
The analgesic effect of BoNT-A presumably results from decreased neuropeptide release at peripheral extremities (62). In the first case, neurogenic inflammation is prevented, and in the second, nociceptive transmission becomes inhibited at the spinal cord.
Most nociceptive bladder afferents are concentrated in the trigone (63). This may suggest that previous studies that injected the entire bladder of BPS/IC patients with BoNT-A (64, 65) placed most of the neurotoxins far from the receptive fibers and increased the risk of decreasing detrusor contractility. The hazard of vesicoureteral reflux after trigonal BoNT-A is nonexistent (66). The treatment of the BPS/IC is to inject 10 sites in the trigone with 100 I.U. BoNT-A diluted with 10 mL saline. These trigonal injections improved BPS/IC symptoms without significant complications. Reinjections after 3 and 6 months remain effective, because BoNT-A prevents exocytosis of Acetylcholine (Ach) vesicles at the nerve terminal, thereby inhibiting neurotransmission and muscle contraction. The action of BoNT is not permanent because neuronal death does not occur and eventually the toxin is inactivated and removed. BoNT subtype A (BoNT-A) is the most relevant clinically. Onabotulinum toxin A (BoNT-ONA), abobotulinum toxin A, and incobotulinum toxin A are available BoNTAs (67, 68).
Recently, the use of BoNT-ONA in LUTS was comprehensively reviewed. Its efficacy in treatment of idiopathic Detrusor Overactivity was mentioned above. It seems that the dose of 100 i.U. may be the one that appropriately balances the symptoms benefits with the safety profile (67). Improvement of lower urinary tract symptoms associated with Benign Prostato Enlargement (BPE) has been reported with the use of intraprostatic injection of BoNT-ONA in clinical studies. Recently, intraprostatic injection of BoNT-ONA 100 I.U., 200 I.U., and 300 I.U. was tested versus placebo injection in a phase 2 dose ranging study. IPSS, Q-max and prostate volume significantly improved in all groups, owing to a large placebo effect from the injectable therapy. A post hoc analysis revealed a significant reduction with BoNT-ONA 100 I.U. in prior to a-blocker users at week 12, which will be explored in further studies (69, 70).
 van Ermengem E. Classics in infectious diseases. A new anaerobic bacillus and its relation to botulism. E. van Ermengem. Originally published as ‘Ueber einen neuen ana�roben Bacillus und seine Beziehungen zum Botulismus’. Z Hyg Infektionskr 1897; 26: 1-56, Rev Infect Dis 1979; 1: 701-19
 Schantz EJ, Johnson EA. Botulinum toxin. The story of its development for the treatment of human disease. Perspect Biol Med 1997; 40: 317-27
 Schantz EJ, Johnson EA. Properties and use of botulinum toxin and other microbial neurotoxins in medicine. Microbiol Rev 1992; 56: 80-99
 Scott AB. Botulinum toxin injection into extraocular muscles as an alternative to strabismus surgery. Ophthalmology 1980; 87: 1044-9
 Carruthers A, Kiene K, Carruthers J. Botulinum A exotoxin use in clinical dermatology. J Am Acad Dermatol 1996; 34: 788-97
 Naumann M, Flachenecker P, Br?cker EB et al. Botulinum toxin for palmar hyperhydrosis. Lancet 1997; 349: 252
 Schurch B, Schmid DM, Stohrer M. Treatment of neurogenic incontinence with botulinum toxin A. N Engl Med 2000; 342: 665
 Schmid DM, Schurch B, John H, Hauri D. Botuline toxin injection to treat overactive bladder. Eur Urol 2004 (Suppl.): Abstract 516
 Cheng CL, de Groat WC. The role of capsaicin-sensitive afferent fibers in the lower urinary tract dysfunction induced by chronic spinal cord injury in rats. Exp Neurol 2004; 187: 445-54
 Mills IW, Drake MJ, Noble JG, Brading AF. Are unstable detrusor contractions dependant on efferent excitatory innervation? Neurourol Urodyn 1998; 17: 352-4
 Brading AF, Turner WH. The unstable bladder: towards a common mechanism. Br J Urol 1994; 73:3-8
 German K, Bedwani J, Davies J, Brading AF, Stephenson TP. Physiological and morphometric studies into the pathophysiology of detrusor hyperreflexia in neuropathic patients. J Urol 1995; 153: 1678-83
 Harrison SC, Hunnam GR, Farman P, Ferguson DR, Doyle PT. Bladder instability and denervation in patients with bladder outflow obstruction. Br J Urol 1987; 60: 519-22
 Sethia KK, Brading AF, Smith JC. An animal model of non-obstructive bladder instability. J Urol 1990; 143: 1243-6
 Sibley GN. Developments in our understanding of detrusor instability. Br J Urol 1997; 80 (Suppl. 1): 54-61
 Speakman MJ, Brading AF, Gilpin CJ, Dixon JS, Gilpin SA, Gosling JA. Bladder outflow obstruction � a cause of denervation supersensitivity. J Urol 1987; 138: 1461-6
 Mills IW, Greenland JE, McMurray G et al. Studies of the pathophysiology of idiopathic detrusor instability: the physiological properties of the detrusor smooth muscle and its pattern of innervations. J Urol 2000; 163: 646-51
 Drake MJ, Hedlund P, Mills IW et al. Structural and functional denervation of human detrusor after cord injury. Laboratory Invest 2000; 80: 1491-9
 Fry CH, Sui GP, Severs NJ, WU C. Spontaneous activity and electrical coupling in human detrusor smooth muscle; implications for detrusor overactivity? Urology 2004; 63 (Suppl. 1): 3-10
 Sui GP, Coppen SR, Dupont E et al. Impedance measurements and connexin expression in human detrusor muscle from stable and unstable bladders. BJU Int 2003; 92: 297-305
 Haferkamp A, Dorsam J, Elbadawi A. Ultrastructural diagnosis of neuropathic detrusor overactivity: validation of a common myogenic mechanism. Adv Exp Medical Biol 2003; 539: 281-91
 Sui GP,Rothery S, Dupont E, Fry CH, Severs NJ. Gap junctions and connexin expression in human suburothelial interstitial cells. BJU Int 2002; 90: 118-29  Birder LA. Role of the urothelium in urinary bladder dysfunction following spinal cord injury. Prog Brain Res 2006; 152: 135-46
 Birder L. Role of the urothelium in bladder function. Scand J Urol Nephrol Suppl 2004; 215: 48-53
 Beckel JM, Kanai A, Lee SJ, de Groat WC, Birder LA. Expression of functional nicotinic acetylcholine receptors in rat urinary bladder epithelial cells. Am J Physiol Renal Physiol 2006; 290: F103-10
 Gevaert T, Vriens J, Segal A et al. Deletion of the transient receptor potential cation channel TRPV4 impairs murine bladder voiding. J Clin Invest 2007; 117: 3453-62
 Birder L, Kullmann FA, Lee H et al. Activation of urothelial transient receptor potential vanilloid 4 by 4alpha-phorbol 12, 13-didecanoate contributes to altered bladder reflexes in the rat. J Pharmacol Exp Ther 2007; 323: 227-35
 Smet PJ, Jonavicius J, Marshall VR, de Vente J. Distribution of nitric oxide synthase-immunoreactive nerves and identification of the cellular targets of nitric oxide in guinea-pig and human urinary bladder by cGMP immunohistochemistry. Neuroscience 1996; 71: 337-48
 Powell DW, Mifflin RC, Valentich JD et al. Paracrine cells important in health and disease. Am J Physiol 1999; 277: C1-9
 Drake MJ, Fry CH, Eyden B. Structural characterization of myofibroblasts in the bladder. BJU Int 2006; 97: 29-32
 Drake MJ, Hedlund P, Andersson KE et al. Morphology, phenotype and ultrastructure of fibroblastic cells from normal and neuropathic human detrusor: absence of myofibroblast characteristics. J Urol 2003; 169: 1573-6
 Hashitani H, Yanai Y, Suzuki H. Role of interstitial cells and gap junctions in the transmission of spontaneous Ca2+ signals in detrusor smooth muscles of the guinea-pig urinary bladder. J Physiol 2004; 559: 567-81
 Wiseman OJ, Fowler CJ, Landon DN. The role of the human bladder lamina propria myofibroblast. BJU Int 2003; 91: 89-93
 Gillespie JI, Harvey IJ, Drake MJ. Agonist- and nerve-induced phasic activity in the isolated whole bladder of the guinea pig: evidence for two types of bladder activity. Exp Physiol 2003; 88: 343-57
 van AF, Roskams T, Blyweert W, Ost D, Bogaert G, De Ridder D. Identification of kit positive cells in the human urinary tract. J Urol 2004; 171: 2492-6
 Sergeant GP, Hollywood MA, McCloskey KD, Thornbury KD, McHale NG. Specialised pacemaking cells in the rabbit urethra. J Physiol 2000; 526: 359-66
 Sergeant GP, Thornbury KD, McHale NG, Hollywood MA. Interstitial cells of Cajal in the urethra. J Cell Mol Med 2006; 10: 280-91
 Sjogren C, Andersson KE, Husted S, Mattiasson A, Moller Madsen B. Atropine resistance of transmurally stimulated isolated human bladder muscle. J Urol 1982; 128: 1368-71
 Kinder RB, Mundy AR. Pathophysiology of idiopathic detrusor instability and detrusor hyper-reflexia. An in vitro study of human detrusor muscle. Br J Urol 1987; 60: 509-15
 Bayliss M, Wu C, Newgreen D, Mundy AR, Fry CH. A quantitative study of atropine-resistant contractile responses in human detrusor smooth muscle, from stable, unstable and obstructed bladders. J Urol 1999; 162: 1833-9
 Skehan AM, Downie JW, Awad SA. The pathophysiology of contractile activity in the chronic decentralized feline bladder. J Urol 1993; 149: 1156-64
 Andersson KE. Mechanisms of disease: central nervous system involvement in overactive bladder syndrome. Nat Clin Pract Urol 2004; 1: 103-8
 Downie JW, Armour JA. Mechanoreceptor afferent activity compared with receptor field dimensions and pressure changes in feline urinary bladder. Can J Physiol Pharmacol 1992; 70: 1457-67
 Drake MJ, Mills IW, Gillespie JI. Model of peripheral autonomous modules and a myovesical plexus in normal and overactive bladder function. Lancet 2001; 358: 401-3
 Drake MJ. The integrative physiology of the bladder. Ann R Coll Surg Engl 2007; 89: 580-5
 Black JD, Dolly JO. Interaction of 125I-labeled botulinum neuro-toxins with nerve terminals. II. Autoradiographic evidence for its uptake into motor nerves by acceptor-mediated endocytosis. J Cell Biol 1986; 103: 535-44
 Dong M, Yeh F, Tepp WH et al. SV2 is the protein receptor for botulinum neurotoxin A. Science 2006; 312: 592-6
 Tang-Liu DD, Aoki KR, Dolly JO et al. Intramuscular injection of 125I-botulinum neurotoxin-complex versus 125I-botulinum-free neurotoxin: time course of tissue distribution. Toxicon 2003; 42: 461-9
 Aoki KR, Ranoux D, Wissel J. Using translational medicine to understand clinical differences between botulinum toxin formulations. Eur J Neurol 2006; 13 (Suppl. 4): 10-9
 Aoki KR. Botulinum neurotoxin serotypes A and B preparations have different safety margins in pre-clinical models of muscle weakening efficacy and systemic safety. Toxicon 2002; 40: 923-8
 Guiliani S, Santicioli P, Lippi A, Lecci A, Tramontana M, Maggi CA. The role of sensory neuropeptides in motor innervation of the hamster isolated urinary bladder. Naunyn Schmiedebergs Arch Pharmacol 2001; 364: 242-8
 Smith CP, Franks ME, McNeil BK et al. Effect of botulinum toxin A on the autonomic nervous system of the rat lower urinary tract. J Urol 2003; 169: 1896-900
 Khera M, Somogyi GT, Kiss S, Boone TB, Smith CP. Botulinum toxin A inhibits ATP release from bladder urothelium after chronic spinal cord injury. Neurochem Int 2004; 45: 987-93
 Smith CP, Vemulakonda VM, Kiss S, Boone TB, Somogyi GT. Enhanced ATP release from rat bladder urothelium during chronic bladder inflammation: effect of botulinum toxin A. Neurochem Int 2005; 47: 291-7
 Khera M, Somogyi GT, Salas NA, Kiss S, Boone TB, Smith CP. In vivo effects of botulinum toxin A on visceral sensory function in chronic spinal cord-injured rats. Urology 2005; 66: 208-12
 Burton TJ, Edwardson JM, Ingham J, Tempest HV, Ferguson DR. Regulation of Na+ channel density at the apical surface of rabbit urinary bladder epithelium. Eur J Pharmacol 2002; 448: 215-23
 Lagou M, Gillespie J, Kirkwood T, Harvey I, Drake MJ. Muscarinic stimulation of the mouse isolated whole bladder: physiological responses in young and ageing mice. Auton Autacoid Pharmacol 2006; 26: 253-60
 Abrams P, Cardozo L, Fall M, et al. The standardisation of terminology of lower urinary tract function: report from the Standardisation Sub-committee of the International Continence Society. Neurourol Urodyn 2002; 21: 167-78.
 Van de Merwe JP, Nordling J, Bouchelouche P, et al. Diagnostic criteria, classification, and nomenclature for painful bladder syndrome/interstitial cystitis: an ESSIC proposal. Eur Urol 2008; 53: 60-7
 Apostolidis A, Dasgupta P, Denys P, et al. Recommendations on the use of botulinum toxin in the treatment of lower urinary tract disorders and pelvic floor dysfunctions: a European consensus report. Eur Urol 2009; 55: 100-20.
 Chuang YC, Yoshimura N, Huang CC, et al. Intravesical botulinum toxin A administration produces analgesia against acetic acid induced bladder pain response in rats. J Urol 2004; 172: 1529-32.
 Rapp DE, Turk KW, Bales GT, et al. Botulinum toxin type A inhibits calcitonin gene-related peptide release from isolated rat bladder. J Urol 2006; 175: 1138-42.
 Avelino A, Cruz C, Nagy I, et al. Vanilloid Receptor 1 expression in the rat urinary tract. Neuroscience 2002; 109: 787-98.
 Giannantoni A, Porena M, Costantini E, et al. Botulinum A toxin intravesical injection in patients with painful bladder syndrome: 1-yr follow-up. J Urol 2008; 179: 1031-4.
 Kuo HC, Chancellor MB. Comparison of intravesical botulinum toxin type A injections plus hydrodistension with hydrodistension alone for the treatment of refractory interstitial cystitis/painful bladder syndrome. BJU Int 2009; 1: 1-5.
 Karsenty G, Elzayat E, Delapparent T, et al. Botulinum toxin type A injections into the trigone to treat idiopathic overactive bladder do not induce vesicoureteral reflux. J Urol 2007; 177: 1011-4.
 Mangera A, Andersson KE, Apostolidis A, et al. Contemporary management of lower urinary tract disease with botulinum toxin A: a systemic review of botox (onabotulinumtoxinA) and dysport (abobotulinumtoxinA). Eur Urol 2011; 60: 784-95.
 Chancellor MB, Fowler CJ, Apostolidis A, et al. Drug insight: biological effects botulinum toxin A in the lower urinary tract. Nat Clin Pract Urol 2008; 5: 319-28.
 Marberger M, Chartier-Kastler E, Egerdie B, et al. A randomized double-blind placebo-controlled phase 2 dose-ranging study of onabotulinumtoxinA in men with benign prostatic hyperplasia. Eur Urol 2013; 63: 496-503.
 Rui Pinto, Tiago Lopes, B?rbara Frias, et al. Trigonal Injection of Botulinum Toxin A in Patients with Refractory Bladder Pain Syndrome/Interstitial Cystitis. Eur Urol 2010; 58: 360-365.
Prof. Dr. SEMIR AHMED SALIM AL SAMARRAI
Professor Doctor of Medicine-Urosurgery, Andrology, and Male Infertility
Dubai Healthcare City, Dubai, United Arab Emirates.
Mailing Address: Dubai Healthcare City, Bldg. No. 64, Al Razi building, Block D,
2nd floor, Dubai, United Arab Emirates, PO box 13576