1-, 
-adrenoceptors in the urinary bladder, urethra and prostateIntroduction
The lower urinary tract is responsible for urine storage and voiding (see Andersson & Wein, 2004). During the storage phase of the micturition cycle, the bladder relaxes to accommodate increasing volumes of urine at acceptable pressure, and the bladder neck and urethra contract to provide resistance to prevent involuntary leakage. During the micturition phase, the bladder neck and urethral muscles relax to allow the detrusor to contract and expel urine without major resistance. While the prostate does not appear to play a major physiological role in continence, its enlargement in patients with benign prostatic hyperplasia (BPH) can increase bladder outlet resistance and thereby disturb physiological voiding.
Diseases of the lower urinary tract are frequent in the general population. They include the syndrome of the overactive bladder (OAB), which is defined as urgency, with or without incontinence, usually accompanied by frequency and nocturia (Abrams et al., 2002), and is present in about 16% of the population aged 40 years and over (Milsom et al., 2001). Another frequent condition is stress urinary incontinence, a condition largely affecting the female population. Its reported prevalence in the general female population ranges between 5 and 37% (Hampel et al., 2004). The reasons for this remarkable heterogeneity include differences between study populations and the use of varying definitions of the condition. More consistently, stress incontinence accounts for approximately 80% of incontinence in women (Hampel et al., 2004). While BPH is a very frequent condition in elderly males, its prevalence estimates depend on whether the histological diagnosis of BPH or the associated bothersome symptoms are assessed, the latter being reported in about 30% of men aged 50–80 in population-based studies (Berges et al., 2001).
The autonomic nervous system plays a key role in the regulation of lower urinary tract function (see Bannowsky & Juenemann, 2003; Michel et al., 2005c). Its sympathetic innervation occurs via the hypogastric nerve arising from the nucleus intermediolateralis of spinal cord segments Th12–L2. Noradrenaline released from these nerves can act on all three classes of adrenoceptors, that is, 
1-, 2- and 
-adrenoceptors. Three receptor subtypes have been cloned within each of these classes and are designated as 1A (in earlier papers, sometimes also referred to as 1A/D or 1C), 1B, 1D, 2A (its rodent analogue sometimes referred to as 2D), 2B, 2C, 1, 2 and 3 (see Bylund et al., 1994; Hieble et al., 1995). Multiple splice variants of the 1A-adrenoceptor have been reported, but they have a very similar ligand recognition profile and hence their pharmacological relevance remains unclear (Hirasawa et al., 1995; Chang et al., 1998; Daniels et al., 1999). Moreover, the 1A-adrenoceptor gene product can exhibit low affinity for prazosin and several other drugs upon expression in some cell types, and this phenotype is often referred to as '1L' (Ford et al., 1997; Daniels et al., 1999). Similarly, the 1-adrenoceptor gene product can exhibit low affinity for propranolol and several other drugs upon expression in some cell types, and this phenotype is sometimes referred to as '4' and sometimes as 'atypical -adrenoceptor' (Joseph et al., 2003; 2004). Moreover, it should be considered that single-nucleotide polymorphisms exist for most of the nine cloned human adrenoceptor subtypes, which could lead to altered tissue responses (see Leineweber et al., 2004; Lei et al., 2005). The present manuscript reviews the expression, functional responses and regulation of each of these adrenoceptor subtypes in the bladder, urethra and prostate, and discusses their therapeutic implications and potential value as drug targets.
Bladder
The bladder stores and expels urine. The force needed to expel it during the voiding phase of the micturition cycle is generated by the detrusor smooth muscle (sometimes with the help of increasing intra-abdominal pressure), which is anatomically largely found in the bladder dome. In contrast, the bladder neck is involved in generating resistance during the filling phase of the micturition cycle to help prevent involuntary urine leakage. Therefore, the bladder neck is functionally more closely related to the urethra than to the detrusor. The trigone and bladder base are anatomically located close to the bladder neck. Interestingly, the bladder neck appears to have a much denser sympathetic innervation than the detrusor, and the role of neuronally released noradrenaline in activating adrenoceptors expressed in the detrusor has not been well established.
1-Adrenoceptors
mRNA expression
The presence of 1-adrenoceptor subtype mRNA in the urinary bladder has been assessed in rats, mice, monkeys and humans, with rats and humans apparently differing considerably. Using competitive RT–PCR, 1A-, 1B- and 1D-adrenoceptors were found to account for 95, 1 and 4%, respectively, of total 1-adrenoceptor mRNA in rat bladder (Scofield et al., 1995). Another study based upon RNase protection assays has reported a roughly similar abundance of all three subtypes in whole rat bladder, but did not provide quantitative information (Malloy et al., 1998), whereas a later, more quantitative report from those investigators based upon competitive RT–PCR has shown the presence of the three subtypes in a ratio of 70 : 5 : 25% (Hampel et al., 2002). Microarray analysis detected hybridization signals for the 1A-adrenoceptor, but not for any other -adrenoceptor subtype (Lluel et al., 2003b). For each of the three subtypes, expression in the rat bladder base was shown to be markedly greater than in the detrusor (Yono et al., 2004). A predominant expression of the 1A-subtype was qualitatively confirmed using in situ hybridization studies, which found a strong expression of this subtype in the urothelium, a moderate expression in smooth muscle (quantitatively similar to that in prostate smooth muscle), but no presence in connective tissue; bladder dome and bladder base were similar in this regard (Walden et al., 1997). The same study also found a very similar situation in the rhesus monkey bladder (Walden et al., 1997). In contrast, that study detected a moderate expression of 1A-adrenoceptors in the human bladder dome smooth muscle (quantitatively similar to that in prostate smooth muscle), but not in bladder dome connective tissue or urothelium or in prostate epithelium (Walden et al., 1997). Using real-time PCR, other investigators confirmed a moderate expression of 1-adrenoceptor mRNA in the human bladder (corresponding to only 3% of -adrenoceptor mRNA abundance), to which 1A-, 1B- and 1D-adrenoceptors contributed 33, 53 and 14%, respectively (Nomiya & Yamaguchi, 2003). RT–PCR studies reported a dominant abundance of 1A- and 1D-adrenoceptor mRNA with less, if any, 1B-adrenoceptor mRNA in the human bladder (Sigala et al., 2004). Other investigators, using RNase protection assays, detected 1A-, 1B- and 1D-adrenoceptors in a 34 : 0 : 66% ratio (Malloy et al., 1998). The predominance of 1D-adrenoceptor mRNA has been confirmed in a recent study from the same group using two independent sets of samples using quantitative real-time PCR confirmed by RNase protection assays (Schwinn, personal communication). Real-time PCR studies in mice reported 1A-, 1B- and 1D-adrenoceptors in a 42 : 8 : 50% ratio (Chen et al., 2005), that is, in a roughly similar ratio as in the human bladder. Taken together, the total quantity of 1-adrenoceptor mRNA expression in the detrusor appears low. While the 1A-adrenoceptor is the most abundant subtype in the rat bladder, the relative contributions of 1-adrenoceptor subtypes in the human bladder remain controversial.
Protein expression
The presence of 1-adrenoceptors in the detrusor of rats, rabbits, guinea-pigs, pigs, cats, monkeys and humans has been examined at the protein level using radioligand-binding studies in tissue homogenates and, in some cases, receptor autoradiography. Using [125I]BE 2254 (also known as [125I]HEAT) as the radioligand, a low density of 1-adrenoceptors (
7 fmol mg-1 protein) was found in rat bladder, which was shown to represent a homogeneous population of 1A-adrenoceptors (Hampel et al., 2002). A low density of 1-adrenoceptors was confirmed in autoradiography studies using [3H]prazosin (Monneron et al., 2000). Saturation-binding experiments with the 1A-selective radioligand [3H]L-771,688 also detected a relatively low density of this subtype in the rat bladder as compared to several other tissues (Chang et al., 2000). In rabbit bladder, a slightly greater but still only moderate 1-adrenoceptor density (14–18 fmol mg-1 protein) was reported using [125I]BE 2254 (Tsujimoto et al., 1986) or [3H]prazosin (Latifpour et al., 1990). Autoradiography studies using [3H]prazosin also detected only few, if any, 1-adrenoceptors in the guinea-pig, cat and female pig bladder (Monneron et al., 2000). Using the same radioligand in membrane preparations, no quantifiable amounts of 1-adrenoceptors were detected in male or female porcine detrusor (Goepel et al., 1997). Autoradiography studies with [3H]prazosin found very little 1-adrenoceptor expression at the protein level in the urothelial or smooth muscle layers of the rhesus monkey detrusor (Walden et al., 1997). Studies in the human detrusor using [125I]BE 2254 as the radioligand reported a low 1-adrenoceptor density (6 fmol mg-1 protein); based upon competition studies with BMY 7378, 66% of these were described as 1D-adrenoceptors (Malloy et al., 1998). A low level of 1-adrenoceptor expression at the protein level in the detrusor was confirmed using [125I]BE 2254 by other investigators (Sigala et al., 2004). Using Western blots with subtype-selective antibodies, the latter study demonstrated the presence of all three subtypes at the protein level in the human detrusor, but did not provide subtype-specific quantification (Sigala et al., 2004). Another group, however, using [3H]prazosin as the radioligand, has not detected quantifiable numbers of 1-adrenoceptors in the human detrusor (Goepel et al., 1997). Thus, the overall density of 1-adrenoceptors in the detrusor of various species, including humans, is low.
The presence of 1-adrenoceptors has also been investigated in the trigone, bladder base and/or bladder neck of several species. In this regard, pigs appear to be the only species where 1-adrenoceptors have not been detected in the bladder neck (Goepel et al., 1997). In the rabbit bladder base, early studies had found 1-adrenoceptors of an unspecified subtype (Andersson et al., 1984; Larsson et al., 1986; Levin et al., 1988). Direct comparative studies in rats (Monneron et al., 2000) and rabbits (Latifpour et al., 1990) reported greater 1-adrenoceptor binding in the trigone and bladder base, respectively, than in the dome. Similar autoradiography studies with [3H]prazosin found greater 1-adrenoceptor expression in the monkey bladder base than detrusor; based upon competition by the highly 1A-selective SNAP 5272, the latter appeared to predominantly represent 1A-adrenoceptors (Figure 1) (Walden et al., 1997). 1-Adrenoceptors have also been found in the human bladder base (Levin et al., 1988). This was confirmed by other investigators, using not only radioligand binding but also Western blotting with subtype-selective antibodies, which detected all three subtypes (Sigala et al., 2004). Thus, in agreement with the mRNA measurements, radioligand binding and receptor autoradiography studies have detected only low densities of 1-adrenoceptors in the detrusor of several species, including humans; in this regard, detection by [125I]BE 2254 appears to be more sensitive than that by [3H]prazosin. A more consistent, and in some direct comparative studies greater, 1-adrenoceptor expression was seen in the trigone/bladder base/bladder neck region. While the 1A-adrenoceptor appears to be the most abundant subtype in healthy rats, the 1D-adrenoceptor appears to be the most abundant subtype in humans.
Figure 1.
Presence of 1A-adrenoceptor protein in the lower urinary tract of the monkey. Receptors were localized by autoradiography, using [3H]praozsin and defining non-specific binding in the presence of SNAP 5272. Receptor autoradiograms were scanned into computer as a 16 grey scale image. The 16 grey levels corresponding to specific 1A-adrenoceptor receptor binding were each assigned colour (see scale) to allow subtle differences in film exposure to be easily visible. Sections show bladder dome and prostate (i), bladder trigone (ii), bladder base (iii), prostatic urethra (iv) and penile urethra (v). Key: sm (smooth muscle); ur (urothelium); lm (longitudinal muscle). Schematic representation of monkey urinary tract together with the orientation of sectioning planes (i)–(v) shown in (b). Taken with permission from Walden et al. (1997).
In vitro function
In vitro studies on the functional role of 1-adrenoceptors in the urinary bladder have focused not only on direct contractile effects but also on the modulation of neurotransmitter release. The 1-adrenoceptor agonists phenylephrine and methoxamine enhanced the field stimulation-induced release of both noradrenaline and acetylcholine in the isolated rat bladder (Somogyi et al., 1995). Phenylephrine also increased the basal release of noradrenaline, but not of acetylcholine. While the phenylephrine effect on acetylcholine release was blocked by the 1-adrenoceptor antagonist terazosin, that on noradrenaline release was not, indicating that the latter may have been -adrenoceptor-mediated. The increased acetylcholine release was accompanied by an enhancement of field stimulation-induced contraction, which was mainly seen at low-frequency nerve stimulation and at low extracellular Ca2+ concentrations. It was concluded that cholinergic nerve terminals in rat bladder express 1-adrenoceptors that facilitate acetylcholine release. Later studies from these investigators confirmed the initial observations and further demonstrated that the facilitation of acetylcholine release was largely, if not exclusively, mediated by 1A-adrenoceptors (Szell et al., 2000). An 1-adrenoceptor-mediated depolarization of parasympathetic nerves in the vesical ganglia has also been demonstrated in cats (Nakamura et al., 1984).
The possible direct contractile effects of 1-adrenoceptor stimulation have been investigated in rat, rabbit, guinea-pig and human bladder. In the rat detrusor, 1-adrenoceptor agonists such as phenylephrine produced only weak contractions, that is, in the range of 10–43% of those reached by muscarinic receptor stimulation or receptor-independently by KCl (Kolta et al., 1984; Somogyi et al., 1995; Lluel et al., 2000; 2003a, 2003b; Szell et al., 2000). The effect appears to be mediated by a subtype with low affinity for the 1D-selective BMY 7378, most likely the 1A-adrenoceptor (Lluel et al., 2003b). Interestingly, and in line with the data on 1-adrenoceptor expression at the protein level (see above), the phenylephrine-induced contraction was about three times as large in rat bladder neck as compared to the detrusor (Lluel et al., 2003a). One study demonstrates that the weak direct contractile effects of 1-adrenoceptor agonists in the rat detrusor occur via chloroethylclonidine-sensitive 1B- or 1D-adrenoceptors, that is, a different subtype than the one mediating enhanced acetylcholine release and hence indirect contractile effects (Szell et al., 2000). The direct contractile effects of 1-adrenoceptor agonists in the rabbit detrusor were also weak (Ueda et al., 1984; Tsujimoto et al., 1986; Latifpour et al., 1990). However, several direct comparative studies demonstrate greater 1-adrenoceptor-mediated contraction in rabbit trigone (Ueda et al., 1984) and bladder base (Latifpour et al., 1990). Interestingly, some data show that the 1-adrenoceptor subtype mediating the contraction of rabbit trigone, bladder base and/or bladder neck resembles the cloned 1A-adrenoceptor (Honda & Nakagawa, 1986; van der Graaf et al., 1997; Kava et al., 1998; Williams et al., 1999), but has relatively low affinity for prazosin (pA2 8.0–8.4), indicating that it may belong to the 1L-phenotype of the 1A-adrenoceptor (Lefevre-Borg et al., 1993; Deplanne & Galzin, 1996; van der Graaf et al., 1997; Kava et al., 1998; Williams et al., 1999) (a more detailed discussion of the 1L-phenotype is given in the prostate section). Based upon the high potency of the antagonist L-771,688 (also known as SNAP 6383) in inhibiting the contractile effects of the agonist A61603, the receptor mediating contraction of the monkey bladder neck was also classified as being 1A (Chang et al., 2000).
In analogy to rats and rabbits, studies in the human detrusor have found only very weak contraction (up to 5% of the maximum muscarinic response) by the 1-adrenoceptor agonist phenylephrine (Nomiya & Yamaguchi, 2003). A more robust contraction was observed in studies with the human bladder base and bladder neck (Caine et al., 1975). In the latter tissue, contraction was potently elicited by the 1A-selective agonist A-61603 and inhibited potently by the 1A-selective antagonist L-771,688 (Chang et al., 2000). In contrast to the rabbit bladder neck, however, responses in the human bladder base exhibited high potency for prazosin (pA2 8.9) (Kunisawa et al., 1985), indicating that the 1L-phenotype of the 1A-adrenoceptor was not involved.
In contrast to rats, rabbits and humans, 1-adrenoceptor stimulation in the isolate guinea-pig bladder did not enhance, but rather inhibited, the amplitude and frequency of phasic contractions (Gillespie, 2004), but the reasons for such species differences remain unclear. Thus, in most species, including humans, 1-adrenoceptor stimulation produces only weak detrusor contraction, whereas a stronger contraction is observed for the trigone, bladder base and/or bladder neck. The physiological relevance of this, however, remains unclear, since the bladder neck appears largely under the control of the parasympathetic (and perhaps nonadrenergic–noncholinergic) rather than the sympathetic nervous system (Deplanne et al., 1998).
In vivo function
The in vivo analysis of a role for 1-adrenoceptors in the regulation of bladder function is complicated by the fact that both central and peripheral receptors may be involved and may serve distinct functions. In anaesthetized rats, intra-thecal injections of prazosin inhibited bladder contraction evoked from the locus coeruleus (Yoshimura et al., 1988). Using continuous cystometry in conscious rats, doxazosin given intra-thecally was shown to reduce the size of the bladder pressure (Ishizuka et al., 1996b). Two studies have investigated the 1-adrenoceptor subtypes involved in the central stimulation of bladder contraction. Reductions in the height of isovolumetric contraction were reported for the 1A-adrenoceptor antagonist RS 100,329 given intra-thecally and for the moderately 1B-selective antagonist (+)-cyclazosin; however, the latter effect was not dose-related (Yoshiyama & De Groat, 2001). Further, both drugs increased the frequency of these contractions, while the 1D-adrenoceptor antagonist BMY 7378 had no effect. Naftopidil, which may have some selectivity for 1D-adrenoceptors, given intra-thecally, was reported to inhibit the appearance of regular isovolumetric bladder contractions and reduce their height (Sugaya et al., 2002). In addition, tamsulosin, which has high affinity for both 1A- and 1D-adrenoceptors, was also reported to inhibit the appearance of these contractions. These studies demonstrate that central, most likely spinal, 1-adrenoceptors are involved in stimulating bladder contractility, and that an 1A-adrenoceptor is the most likely candidate mediating such effects.
Studies with systemic administration of 1-adrenoceptor antagonists have yielded less consistent results. For example, i.v. 1-adrenoceptor antagonists inhibited the sympathetic control of the bladder by reducing hypogastric nerve activity (Danuser & Thor, 1995; Ramage & Wyllie, 1995) and somatic activity to the urethra (Danuser & Thor, 1995). However, spontaneous bladder contractions, presumably mediated by the parasympathetic nervous system, were unaffected (Ramage & Wyllie, 1995). Others compared the intra-thecal and intra-arterial effects of doxazosin, phentolamine, prazosin, tamsulosin and yohimbine upon cystometric parameters in anaesthetized rats (Jeong & Lee, 2000); based upon differences between drugs and modes of administration, these authors proposed that 1-adrenoceptors suppress the micturition effect via a peripheral mechanism, whereas 2-adrenoceptors do so via a central mechanism. Finally, antagonists selective for 1A-, 1B- and 1D-adrenoceptors, that is, RS-100,329, RS-51,385 and BMY 7378, given i.v. to anaesthetized rats, were found to have no effect on bladder contraction height induced by infusion of saline into the bladder, but the associated reflex urethral contractions were attenuated by blockade of 1A/D-adrenoceptors (Conley et al., 2001). Both RS-100,329 and BMY 7378 also decreased resting urethral pressure. The failure to see any changes in evoked bladder contraction may reflect a difference in the method being used to evoke it and/or the route of administration. However, the increase in frequency observed for intra-thecal 1A-adrenoceptor antagonists would be expected to be translated into a decrease in the volume threshold, but this was also not observed.
Thus, despite some ongoing controversies, the overall in vivo data suggest that at a spinal level 1-adrenoceptors are probably involved in mediating bladder contractions and decreasing the frequency of micturition. Therefore, systemically administered 1-adrenoceptor antagonists that penetrate into the central nervous system may predominantly inhibit bladder contractions. On the other hand, 1-adrenoceptors are also involved in the peripheral control of the sympathetic supply to the bladder and thus storage. In this respect, stimulation of the hypogastric nerve has also been shown to facilitate cholinergic transmission at the level of the pelvic ganglia via the action of 1-adrenoceptors (Keast et al., 1990) and thus also enhancing bladder contractions. Interestingly, it has been reported that in anaesthetized dogs the -adrenoceptor agonist midodrine did not affect bladder capacity in young animals, but reduced it in old animals (Takahashi et al., 1996).
Treatment with a very high dose of the 1-adrenoceptor antagonist doxazosin (30 mg kg-1 orally) was reported to attenuate obstruction-induced bladder hypertrophy (Das et al., 2002). However, these findings are difficult to interpret since another study in nonobstructed rats found that doxazosin (2 or 4 mg kg-1 s.c. plus 4 mg kg-1 orally) increased the weight of the bladder base and, in at least some dose groups, upregulated 1A-adrenoceptor mRNA in the bladder base (Yono et al., 2004), and also because the doxazosin doses in both studies are very high as compared to a therapeutic dose of 4–8 mg per patient. Such high doses of doxazosin may have growth-inhibiting or apoptotic effects on the lower urinary tract, which are independent of 1-adrenoceptors (Walden et al., 2004).
Regulation of receptor expression and function
Some studies have investigated a possible regulation of the role of 1-adrenoceptors in bladder function by gender, ageing and bladder outlet obstruction. Expression of 1A-adrenoceptor mRNA was similar in the detrusor and bladder base of male and female rats (Walden et al., 1997), and the number of 1-adrenoceptor-binding sites was also similar in the detrusor and bladder base of male and female rabbits (Latifpour et al., 1990). A study in humans confirmed a lack of gender effect on 1-adrenoceptor binding in detrusor and bladder neck, but found a significantly greater 1-adrenoceptor density in female than in male trigone; this study also reported on the quantity of 1A-, 1B- and 1D-adrenoceptor mRNA in all three regions of both genders, but did not provide a statistical analysis of the observed differences (Sigala et al., 2004).
Ageing studies on 1-adrenoceptors in the bladder have been reported for rats, rabbits and dogs. A comparison of 7-, 17- and 29-month-old Fischer rats did not detect significant alterations in the maximum effects or potency of phenylephrine (Kolta et al., 1984). This was confirmed in studies on 10- and 30-month-old female (Lluel et al., 2000) and male Wistar rats (Lluel et al., 2003b), as well as in 6- and 24-month-old male Sprague–Dawley rats (Lluel et al., 2003a). However, the former two studies surprisingly reported a markedly increased noradrenaline-induced contraction in aged animals, which was not explained by a possible 2-adrenoceptor stimulation (Lluel et al., 2000; 2003b). Since the same studies did not detect differential expression of any -adrenoceptor subtype in a microarray analysis, the phenylephrine findings appear somewhat more plausible than the noradrenaline findings. Ageing was also reported not to affect the number of 1-adrenoceptor-binding sites in detrusor and bladder base of 6 months vs 4.5–5-year-old male or female rabbits (Latifpour et al., 1990). On the other hand, studies in anaesthetized dogs found reductions of bladder capacity upon systemic administration of the 1-adrenoceptor agonists in 68-month-old, but not in 12-month-old, animals; the interpretation of these findings, however, is complicated by the fact that the old dogs had been parous, whereas the young ones were nonparous (Takahashi et al., 1996). Thus, the overall data suggest that neither gender nor ageing has a major effect on 1-adrenoceptor function in the bladder.
Due to the high prevalence of BPH, bladder outlet obstruction is frequent in elderly men. Since it has been speculated that the 1-adrenoceptor antagonist-induced symptom relief in BPH patients may involve effects on bladder function (see Michel, 2002; Roehrborn & Schwinn, 2004), potential alterations of 1-adrenoceptors have been investigated in the bladder of animal models of obstruction and in patients. Studies in a rat model of obstruction found an unchanged total 1-adrenoceptor mRNA and radioligand binding. However, this was accompanied by a reduction of 1A- and an increase of 1D-adrenoceptor mRNA; in competition-binding experiments, 1D-adrenoceptors had been undetectable in control rats, but represented approximately 40% of all 1-adrenoceptors in obstructed animals (Hampel et al., 2002). The contractile effects of phenylephrine were reported to remain unchanged in obstructed patients (Nomiya & Yamaguchi, 2003). Further studies are needed to define the role of 1-adrenoceptors in the detrusor in settings of bladder outlet obstruction.
Clinical implications
In conclusion, 1-adrenoceptors appear to play a small functional role if any in the detrusor of healthy animals and humans. Since some of the beneficial effects of 1-adrenoceptor antagonists in BPH patients cannot easily be explained solely based upon prostatic 1-adrenoceptors (see the section on prostate), it nevertheless appears plausible that those located in the detrusor may have therapeutic relevance. This hypothesis, however, remains to be tested. In a similar vein, it has been reported in a small group of patients with spinal cord injury that treatment with the 1-adrenoceptor antagonist terazosin increases bladder compliance and results in less incontinence and dysreflexia (Swierzewski et al., 1994), but it remains unclear whether this reflects a direct effect on the bladder or an indirect effect. On the other hand, 1-adrenoceptors may play a more prominent functional role in the bladder neck and hence the regulation of bladder outlet resistance. Antagonizing their function may contribute to the beneficial effects of 1-adrenoceptor antagonists in BPH patients (see the section on prostate), whereas their stimulation provides a potential target in the treatment of stress incontinence (see the section on urethra).
2-Adrenoceptors
mRNA and protein expression
To the best of our knowledge, the presence of 2-adrenoceptor subtype mRNA in the bladder has not been reported. At the protein level, however, radioligand-binding studies have detected 2-adrenoceptors in the detrusor and bladder base/bladder neck of rabbits (Andersson et al., 1984; Levin et al., 1988; Latifpour et al., 1990), pigs (Goepel et al., 1997) and humans (Levin et al., 1988; Goepel et al., 1997). Their density in the rabbit bladder base was reported to be smaller (Levin et al., 1988), larger (Andersson et al., 1984) and similar (Latifpour et al., 1990) to that of 1-adrenoceptors within the same tissue. In the porcine and human bladder, their density (15–25 and 40 fmol mg-1 protein, respectively) clearly exceeded that of 1-adrenoceptors, but was somewhat smaller than that of -adrenoceptors within the same study (Goepel et al., 1997). In competition-binding experiments in porcine and human bladder, a predominant, if not exclusive, population of 2A-adrenoceptors was found (Goepel et al., 1997).
In vitro function
Only few studies have assessed the functional role of 2-adrenoceptors in the bladder. The pre-junctional inhibition of neurotransmitter release from both post-ganglionic sympathetic and parasympathetic nerve terminals is the best-established function of 2-adrenoceptors in most tissues. Consistent with this concept, 2-adrenoceptor stimulation inhibited field stimulation-induced contraction of rat bladder in a tetrodotoxin, but not hexamethonium-sensitive, manner (Santicioli et al., 1983; Maggi et al., 1985). Moreover, 2-adrenoceptor stimulation also inhibits parasympathetic nerve activity in the bladder of rabbits (Tsurusaki et al., 1990) and cats (Nakamura et al., 1984; Keast et al., 1990) by an effect on the vesical parasympathetic ganglion. Despite the considerable abundance of 2-adrenoceptors in bladder homogenates, their post-junctional function has not been established as they do not mediate contractile effects in the rabbit detrusor (Ueda et al., 1984), whole guinea-pig bladder (Gillespie, 2004) or human bladder base (Kunisawa et al., 1985).
In vivo function
In vivo studies on 2-adrenoceptor function in the bladder are often difficult to interpret because central and peripheral 2-adrenoceptor stimulation may not have the same effects; they may even partly counteract each other and their relative roles may depend on the use of anaesthetized vs conscious animals. In anaesthetized rats 2-adrenoceptor stimulation reduced volume-induced bladder contraction (Maggi et al., 1985; Harada & Constantinou, 1993). Using somewhat different methods, an opposite conclusion was drawn from a more recent study (Jeong & Lee, 2000). However, all of these studies agree that the site of action of the 2-adrenoceptor agonists and antagonists is in the spinal cord rather than in the periphery. An initial report in conscious rats reported that intra-thecal and intra-arterial (close the bladder) administration of an 2-adrenoceptor agonist reduced micturition pressure, bladder capacity and micturition volume; while an 2-antagonist inhibited the effects of intra-thecal agonist, it mimicked those of the peripheral administration (Ishizuka et al., 1996a). In contrast, other studies found that 2-adrenoceptor agonists increase the frequency of bladder contractions (Durant et al., 1988; Kontani et al., 2000) and voiding (Harada & Constantinou, 1993), but the interpretation of this finding would be complicated by a diuretic effect of the agonist (Harada & Constantinou, 1993). Similar to the situation in anaesthetized rats, all of the above studies agree that 2-adrenoceptors in the spinal cord are likely to be the main source of modulation of bladder function.
Regulation of receptor expression and function
Studies in rabbit detrusor or bladder base reported a similar 2-adrenoceptor density in male and female animals as well as in young (6 months) and old (4.5–5 years) rabbits (Latifpour et al., 1990). A study in male and female pigs confirmed the lack of gender difference in detrusor and bladder neck, with 2A-adrenoceptors being the only detectable subtype in all groups (Goepel et al., 1997). Based upon all of the above data, 2-adrenoceptors are not considered a promising target for the treatment of voiding disorders.
-Adrenoceptors
mRNA expression
The presence of -adrenoceptors in the rat and human bladder at the mRNA level has been studied using Northern blots, PCR and in situ hybridization. Messenger RNA for all three -adrenoceptor subtypes has been detected in rats (Seguchi et al., 1998; Fujimura et al., 1999; Matsubara et al., 2002). It has been claimed that the 3-adrenoceptor may be the most abundant subtype (Fujimura et al., 1999), but no specific quantitative data were reported. Studies in the human bladder have also detected mRNA for all three -adrenoceptor subtypes (Fujimura et al., 1999; Igawa et al., 1999; Takeda et al., 1999; Li et al., 2003; Nomiya & Yamaguchi, 2003). Based upon quantitative PCR experiments, it appears that the 3-adrenoceptor accounts for more than 95% of all -adrenoceptor mRNA in the human bladder (Nomiya & Yamaguchi, 2003). The presence of 3-adrenoceptor mRNA in the human detrusor has also been confirmed in in situ hybridization studies (Takeda et al., 1999).
Protein expression
The identification of -adrenoceptors at the protein level is typically based upon binding studies with radioligands such as [125I]iodocyanopindolol, [125I]iodopindolol, [3H]CGP 12,177 or [3H]dihydroalprenolol. [125I]iodocyanopindolol and [3H]CGP 12,177 have much lower affinity for 3- than for 1- or 2-adrenoceptors (Hoffmann et al., 2004; Baker, 2005). Data from our lab confirm this and further demonstrate that [3H]dihydroalprenolol yields a similarly poor labelling of 3-adrenoceptors (Niclauss et al., unpublished observations), a finding that is entirely consistent with the low 3-adrenoceptor affinity of unlabelled alprenolol (Hoffmann et al., 2004). While high concentrations of [125I]iodocyanopindolol and [3H]CGP 12,177 have successfully been used to label 3-adrenoceptors in transfected cells, the use of similarly high concentrations in tissues yields very high nonspecific binding and will saturate 1- and 2-adrenoceptors. Both problems make the detection of 3-adrenoceptors in tissues expressing mixed -adrenoceptor subtype populations virtually impossible. A potential alternative would be the use of a 3-adrenoceptor-selective radioligand such as [3H]SB 206,606. However, this ligand has only high nanomolar affinity for 3-adrenoceptors (Kd values of 200–500 nM) (Muzzin et al., 1994; Klaus et al., 1995). Therefore, [3H]SB 206,606 is also a poor choice for the labelling of 3-adrenoceptors in tissues. These technical limitations must be considered when interpreting existing radioligand-binding data in the bladder and other tissues.
Radioligand-binding studies on bladder -adrenoceptors have been reported for rats, rabbits, pigs and humans. Saturation-binding studies with various radioligands have reported 6–42 fmol mg-1 protein in rats (Nishimoto et al., 1995; Ma et al., 2002), 60–92 fmol mg-1 protein in rabbits (Levin et al., 1988; Latifpour et al., 1990; Morita et al., 1998), 30–154 fmol mg-1 protein in pigs (Goepel et al., 1997; Yamanishi et al., 2002b, 2002c) and 22–60 fmol mg-1 protein in humans (Levin et al., 1988; Goepel et al., 1997; Morita et al., 2000; Li et al., 2003). Limited attempts have been made to identify the receptor subtypes in the bladder by radioligand binding. Based upon competition studies with the 2-selective antagonist ICI 118,551 and a 1-selective antagonist, sites in the rabbit (Latifpour et al., 1990) and human bladder (Goepel et al., 1997) were reported to largely belong to the 2-subtype. On the other hand, three studies in the porcine bladder detected few, if any, high-affinity sites for ICI 118,551, and the 1-selective antagonist CGP 20,712A recognized largely low-affinity sites in those studies (Goepel et al., 1997; Yamanishi et al., 2002b, 2002c). Two of the studies additionally report about 60% high-affinity sites for SR 59,230A (Yamanishi et al., 2002b, 2002c); the latter authors interpreted these findings as evidence in favour of the presence of a population of largely 3-adrenoceptors. However, three reasons argue against this interpretation: Firstly, ICI 118,551 may not be 2-selective in pigs (Goepel et al., 1996), which make the low affinity of this compound in the porcine bladder difficult to interpret. Secondly, while SR 59,230A can be used to functionally block 3-adrenoceptors, it is not selective for this subtype and, at least in humans, has even slightly lower affinity for 3- than for 1- and 2-adrenoceptors (Hoffmann et al., 2004). Thirdly, the radioligands used in all of the above studies are unlikely to label a major fraction of possibly present 3-adrenoceptors due to their low affinity for this subtype (at least in humans; see above). Therefore, we consider the presently available pig data to be inconclusive. This does not exclude the presence of 3-adrenoceptors at the protein level in any of these species, but the currently available radioligand-binding techniques are probably inadequate to detect their presence. Hence, the reported densities of -adrenoceptors in the bladder may represent an underestimation if the additional presence of 3-adrenoceptors is taken into account.
In vitro function
Since activation of adenylyl cyclase is the prototypical signalling pathway of -adrenoceptors, it is not surprising that an isoprenaline-stimulated, propranolol-sensitive elevation of cAMP content has also been reported in rat bladder (Derweesh et al., 2000; Ma et al., 2002; Uchida et al., 2005). However, various recent studies have questioned whether this can sufficiently explain -adrenoceptor-mediated smooth muscle relaxation (Horinouchi et al., 2003; Peters & Michel, 2003; Tanaka et al., 2003). One study in rat bladder demonstrated that the concentration–response relationships for isoprenaline, clenbuterol and FR 165,101 for relaxation and cAMP elevations were largely superimposable in noncontracted muscle; however, no such relationship was observed during KCl-induced contraction (Uchida et al., 2005). Accordingly, the adenylyl cyclase inhibitor SQ 22,536, in a concentration where it fully suppressed cAMP formation, inhibited rat bladder relaxation by all three agonists in the absence of pre-contraction, but not in its presence (Uchida et al., 2005). Similarly, SQ 22,536 and the protein kinase A inhibitors H7, H89 and Rp-cAMPs, if anything, inhibited isoprenaline-induced relaxation of rat bladder only against passive tension, but not against KCl-induced tension in another study (Frazier et al., 2005a). These data demonstrate that, at least in rats, elevation of cAMP is relevant for the regulation of bladder smooth muscle tone against passive tension, but not in the presence of a depolarizing stimulus such as KCl. Interestingly, a combination of adenylyl and guanylyl cyclase inhibitors (SQ 22,536 and ODQ) caused the strongest inhibition of relaxation against passive tension, but was also inactive against KCl-induced tension (Frazier et al., 2005a).
A possible modulation of membrane potential, ion-channel activity and intracellular ion concentrations has been studied as an alternative means of -adrenoceptor control of bladder function. In guinea-pig bladder smooth muscle bundles exhibiting spontaneous action potentials, isoprenaline was found to hyperpolarize the cells, prevent action potentials and reduce the associated Ca2+ transients; the elevation of membrane potential was blocked by protein kinase A inhibitors and by high extracellular K+ concentrations, but not by K+ channel inhibitors (Nakahira et al., 2001). In other studies, both isoprenaline and the receptor-independent adenylyl cyclase activator forskolin were shown to increase iberiotoxin-sensitive K+ currents in guinea-pig bladder smooth muscle cells, and such stimulation was sensitive to a peptidergic inhibitor of protein kinase A (Kobayashi et al., 2000). In a later study, these investigators also demonstrated propranolol-sensitive isoprenaline inhibition of Ba2+ current through L-type Ca2+ channels due to a shift of steady-state for inactivation by 11 mV; this effect was apparently mediated by protein kinase A, but did not involve protein kinase G (Kobayashi et al., 2003). Other investigators reported that isoprenaline caused marginal increases in Ca2+ currents after large conditioning depolarizations (but not in their absence) in the guinea-pig bladder, and that this effect was not mimicked by forskolin (Smith et al., 1999). On the other hand, a third group found that isoprenaline causes intracellular Ca2+ sparks and activates voltage-dependent Ca2+ channels in guinea-pig bladder, and proposed that this may underlie the activation of large-conductance, iberiotoxin-sensitive K+ channel (Petkov & Nelson, 2005). Differences in the electrophysiological procedures used by the two groups may have contributed to this apparent controversy. Activation of iberiotoxin-sensitive K+ channels can relax the urinary bladder (Malysz et al., 2004). Several studies have assessed the functional relevance of ion channel modulation by -adrenoceptor stimulation. Studies using KCl-precontracted bladder strips from guinea-pigs (Kobayashi et al., 2000) or rats (Frazier et al., 2005a; Uchida et al., 2005) have consistently found that K+ channel blockers such as iberiotoxin or charybdotoxin inhibit isoprenaline-induced bladder relaxation. Interestingly, the latter two studies also report that relaxation against passive tension is not sensitive to those toxins.
Prostaglandins may play a role in bladder contraction by several agents such as protease-activated receptors or bradykinin (Nakahara et al., 2003; 2004; Chopra et al., 2005). Therefore, it is surprising that prostaglandins were also postulated to play a permissive role for -adrenoceptor-mediated relaxation of the urinary bladder (Bolle et al., 1999).
The key function of -adrenoceptors in the bladder is smooth muscle relaxation and an increase in bladder compliance during the filling phase of the micturition cycle. The interpretation of in vitro bladder relaxation experiments has to take into account that the results are sensitive to the experimental conditions. Thus, it has been found that the -adrenoceptor agonist isoprenaline was approximately six times more potent when tested against passive tension than when tested against KCl-induced bladder tone in rats (Frazier et al., 2005a; Uchida et al., 2005). This is consistent with indirect comparisons in the published literature, where a pEC50 for isoprenaline of 8.3 (Yamazaki et al., 1998) vs 7.2 (Longhurst & Levendusky, 1999) and of 9.1 (Yamazaki et al., 1998) vs 7.3 (Oshita et al., 1997) were reported in rats and rabbits, respectively, for passive tension vs pre-contraction. In a comparison between KCl-induced and carbachol-induced tension in rat isolated detrusor, isoprenaline was significantly less potent and effective against the latter (Longhurst & Levendusky, 1999). Moreover, the choice of passive tension vs pre-contraction for relaxation experiments may also affect the underlying signal transduction of the -adrenoceptor response (Frazier et al., 2005a; Uchida et al., 2005). A second methodological consideration relates to the use of muscarinic receptor agonists to induce bladder pre-contraction in combination with -adrenoceptor agonists such as BRL 37,344 to induce relaxation. This drug has affinity for muscarinic acetylcholine receptors in the same concentration range where it acts as a -adrenoceptor agonist (Kubota et al., 2002); hence, data using this combination may at least partly reflect direct muscarinic receptor antagonism rather than -adrenoceptor agonism (see below).
A relaxation of bladder smooth muscle by -adrenoceptor agonists has been demonstrated against passive tension (Igawa et al., 2001; Takeda et al., 2002a), endothelin receptor-mediated (Takeda et al., 2003), muscarinic receptor-mediated (Seguchi et al., 1998; Nomiya & Yamaguchi, 2003) and KCl-induced pre-contraction (Nishimoto et al., 1995; Yamanishi et al., 2003a) or against field stimulation-induced tone (Nishimoto et al., 1995; Hudman et al., 2001). Moreover, relaxation responses have been demonstrated in the detrusor of various species, including rats (Kolta et al., 1984; Nishimoto et al., 1995; Oshita et al., 1997; Seguchi et al., 1998; Yamazaki et al., 1998; Fujimura et al., 1999; Longhurst & Levendusky, 1999; Lluel et al., 2000; Morita et al., 2000; Woods et al., 2001; Matsubara et al., 2002; Inci et al., 2003; Malysz et al., 2004; Uchida et al., 2005; Frazier et al., 2005a), mouse (Matsui et al., 2003), rabbits (Oshita et al., 1997; Morita et al., 1998; 2000; Yamazaki et al., 1998; Bing et al., 2003), guinea-pigs (Li et al., 1992; Gopalakrishnan et al., 1999; Kobayashi et al., 2000; Malysz et al., 2004), ferrets (Takeda et al., 2000a), cats (Nergardh et al., 1977), dogs (Yamazaki et al., 1998), pigs (Yamanishi et al., 2002b, 2002c; 2003a), monkeys (Takeda et al., 2002a) and humans (Nergardh et al., 1977; Fujimura et al., 1999; Igawa et al., 1999; 2001; Takeda et al., 1999; Morita et al., 2000; Nomiya & Yamaguchi, 2003). In contrast, -adrenoceptor stimulation did not consistently relax the basal tone of the human bladder neck (Caine et al., 1975).
Some studies have performed direct inter-species comparisons regarding the ability of -adrenoceptor agonists to induce bladder relaxation. Such comparisons of, for example, rat vs dog (Takeda et al., 2003), rat vs rabbit (Oshita et al., 1997) or rat vs rabbit vs dog (Yamazaki et al., 1998) have consistently reported that the maximum effects of an agonist without subtype selectivity, such as isoprenaline, were similar in various species. However, within the same study, the rank order of isoprenaline potency consistently was rabbit>rat>dog, suggesting that rabbits may have the largest and dogs the smallest receptor reserve for this response, respectively. Similar inter-species comparisons with subtype-selective -adrenoceptor agonists are more difficult to interpret, since the subtype being involved may differ between species.
Functional studies into the -adrenoceptor subtypes mediating bladder relaxation have been hampered by several problems. Firstly, some drugs proposed to be 3-adrenoceptor-selective agonists may have effects independent of -adrenoceptors. For example, it was reported that both BRL 37,344 and SR 58,611 can cause vasodilatation, which is insensitive to -adrenoceptor antagonists (Brahmadevara et al., 2003). Moreover, BRL 37,344 was reported to be a direct muscarinic receptor antagonist (Kubota et al., 2002) and 1-adrenoceptor antagonist (Leblais et al., 2005). Secondly, no truly 3-adrenoceptor-selective antagonist has been described. Thus, SR 59,230, the most frequently used drug to antagonize 3-adrenoceptors, does not discriminate human -adrenoceptor subtypes (Hoffmann et al., 2004) and, similarly to the chemically related bupranolol, may also be an 1-adrenoceptor antagonist (Leblais et al., 2005). When binding to 3-adrenoceptors, SR 59,230 may exhibit agonist rather than antagonist properties in some tissues (Horinouchi & Koike, 2001). Such limitations should be taken into account when interpreting the functional data presented below.
Studies in various species have used agonist and antagonist potency to identify the functional involvement of -adrenoceptor subtypes in bladder relaxation. Since absolute agonist potency may differ between species even for nonsubtype-selective agonists (see above), the former approach has used either rank orders of potency of various agonists or the potency of highly subtype-selective agonists to classify the receptor subtype being involved. Most studies have been reported from rats. Based upon a high potency of 3-selective agonists such as CL 316,243 (Woods et al., 2001) and FK175 (Fujimura et al., 1999), it has been proposed that rat bladder relaxation predominantly occurs via this subtype. However, studies assessing the rank order of potency of multiple subtype-selective agonists have proposed a mixed involvement of 2- and 3-adrenoceptors in rat bladder relaxation in most cases. These were based upon rank orders such as isoprenaline=procaterol (2-selective)>CL 316,243>dobutamine (1-selective) (Takeda et al., 2003), CL 316,243
isoprenalineprocaterol (Takeda et al., 2000b), isoprenalineCL 316,243procaterol>dobutamine (Yamazaki et al., 1998), BRL 37,344isoprenaline (Oshita et al., 1997), isoprenaline=GS-332 (3-selective)clenbuterol (2-selective) (Morita et al., 2000) or isoprenaline>FR 165101 (3-selective)clenbuterol
dobutamine (Uchida et al., 2005). One study, based upon a rank order of agonist potency of isoprenaline>BRL 37,344T-0509 (1-selective)>terbutaline (2-selective)SR 58,611 (3-selective), has even proposed a mixed involvement of 1-, 2- and 3-adrenoceptors in rat bladder relaxation (Longhurst & Levendusky, 1999). Antagonist studies have reported that ICI 118,551 inhibits the effects of clenbuterol against low-, but not high-frequency field stimulation (Hudman et al., 2000). Relaxant effects of the 3-agonist FK175 were moderately inhibited by the nonselective bupranolol, but not by even high concentrations of the 1-selective CGP 20,712 or the 2-selective ICI 118,551 (Fujimura et al., 1999). Similarly, relaxation induced by BRL 37,344 was not inhibited by low propranolol concentrations, but by CGP 12,177 or SR 59,230 when added atop of propranolol; in the same study, relaxation by CGP 12,177 was not affected even by high propranolol concentrations (Longhurst & Levendusky, 1999). These data indicate that 2- and 3-selective agonists may indeed cause rat bladder relaxation via their cognate receptor subtypes. With regard to nonsubtype-selective agonists such as isoprenaline or noradrenaline, several studies report relatively poor antagonism by propranolol, metoprolol, butoxamine or ICI 118,551 (Oshita et al., 1997; Seguchi et al., 1998; Longhurst & Levendusky, 1999). However, SR 59,230, which should inhibit the cloned 3-adrenoceptor, also caused only poor isoprenaline antagonism (Longhurst & Levendusky, 1999). Taken together, these data argue against a strong involvement of 1- and 2-adrenoceptors, but also fail to provide clear evidence for a 3-adrenoceptor. Interestingly, the isoprenaline-induced cAMP response in rat bladder was fully sensitive to propranolol (Ma et al., 2002), which is in line with the proposal that -adrenoceptor-mediated bladder relaxation occurs largely cAMP-independent (Frazier et al., 2005a; Uchida et al., 2005).
In vitro relaxation studies in rabbit bladder have reported agonist rank orders of potency of isoprenalineadrenaline>noradrenalineBRL 37,344 (Oshita et al., 1997), procaterol>isoprenaline>adrenalineCGP 12,177>noradrenalinedobutamine>CL 316,243 (Yamazaki et al., 1998) or clenbuterolGS-332 (Morita et al., 2000). Propranolol, bupranolol and ICI 118,551 antagonized the isoprenaline-induced relaxation with high potency, whereas CGP 20,712, in concentrations up to 100 nM, had no effect (Oshita et al., 1997; Yamazaki et al., 1998). Taken together, these data demonstrate that relaxation of the rabbit detrusor is predominantly mediated by a 2-adrenoceptor.
In the porcine detrusor, there was a rank order of potency of salbutamol (2-agonist)>noradrenaline>BRL 37,344>CGP 12,177 (the latter two being partial agonists only); while the BRL 37,344 response was antagonized by SR 59,230, the corresponding Schild slope was significantly smaller than unity (Yamanishi et al., 2002a). The same investigators also reported a low potency of BRL 37,344 sensitive to SR 59,233 in the porcine bladder base (Yamanishi et al., 2002c). More recently, these authors also reported porcine bladder base relaxation by isoprenaline and salbutamol (Yamanishi et al., 2003a). CGP 20,712 did not inhibit the isoprenaline responses, whereas propranolol and ICI 118,551 caused inhibition, but with a Schild slope of less than unity; in contrast, ICI 118,551 inhibited the salbutamol responses with high potency and a Schild slope close to unity. Another group of investigators found an order of potency of isoprenaline=adrenalineprocaterolBRL 37,344>CGP 12,177salbutamol>CL 316,243noradrenaline; in this regard, BRL 37,344, CL 316,243 and, surprisingly, noradrenaline were reported to be partial agonists and CGP 12,177 was found to be a weak partial agonist (Badawi et al., 2005). Taken together, these findings suggest that both 2-adrenoceptors and an additional subtype, possibly 3-adrenoceptors, mediate porcine bladder relaxation.
Data from several other animal species are too limited or controversial to allow definitive conclusions. In guinea-pigs, a predominant role of 1-adrenoceptors was proposed based upon relaxation by dobutamine, but not by BRL 37,344, salbutamol or clenbuterol, and antagonism of the isoprenaline, noradrenaline and adrenaline responses by atenolol (Yamamoto et al., 1998). Another study also proposed an involvement of 1-adrenoceptors based upon partial antagonism of the isoprenaline response by metoprolol, but reported an even greater role of 2-adrenoceptors based upon partial agonism by salbutamol and terbutaline and antagonism of the isoprenaline response by ICI 118,551 (Li et al., 1992). A more recent study based upon whole bladder contraction reported relaxation by noradrenaline and BRL 37,344, but not by formoterol (2-selective) (Gillespie, 2004). Limited data from one study in cats have suggested a predominant involvement of 1-adrenoceptors (Nergardh et al., 1977). One study in ferrets has proposed a primary involvement of 3-adrenoceptors based upon an agonist rank order of potency of BRL 37,344>CGP 12,177isoprenalineCL 316,243>dobutamineprocaterol and upon antagonism of the isoprenaline response by SR 58,894, but not by CGP 20,712 or ICI 118,551 (Takeda et al., 2000a). One study in dogs reported an agonist rank order of potency of CL 316,243>isoprenalineCGP 12,177>noradrenalinedobutamineprocateroladrenaline, and that the isoprenaline-induced relaxation was inhibited with high potency by bupranolol, but not by CGP 20,712 or ICI 118,551 (Yamazaki et al., 1998); the same group later confirmed the rank order of CL 316,243>dobutamineprocaterol (Takeda et al., 2003), suggesting predominantly an involvement of 3-adrenoceptors. A study in Cynomolgus monkeys found an agonist rank order of potency of isoprenaline>noradrenalineCGP 12,177>BRL 37,344adrenaline>dobutaminesalbutamolprocaterol, with the 1-selective xamoterol being a very weak partial agonist; the effects of isoprenaline were inhibited by bupranolol, but not by CGP 20,712 or ICI 118,551 (Takeda et al., 2002a), suggesting a predominant involvement of a 3-adrenoceptor.
Early reports on human bladder relaxation already proposed that this does not occur via a 1- or 2-adrenoceptor (Nergardh et al., 1977). Several more recent studies suggest that it indeed occurs via a 3-adrenoceptor. Igawa et al. (1998) originally reported relaxation of the human bladder (inhibited by bupranolol), whereas dobutamine, procaterol and CGP 12,177 caused much smaller if any relaxation. Thereafter, they reported an agonist order of potency of BRL 37,344isoprenalinenoradrenalineadrenalineCGP 12,177CL 316,243; in that study, isoprenaline responses were inhibited by SR 58,894, but only poorly by ICI 118,551 and not at all by CGP 20,712 (Figure 2) (Igawa et al., 1999). Another study from the same group reported an order of BRL 37,344isoprenaline>CGP 12,177CL 316,243, with all but isoprenaline being partial agonists (Igawa et al., 2001). Another study reported a rank order of potency of BRL 37,344>CGP 12,177>isoprenaline, with the former two being partial agonists only, and the 3-adrenoceptor agonist ZD 7114 being a very poor partial agonist; the isoprenaline responses were inhibited by SR 59,230, but not by butoxamine and atenolol (Takeda et al., 1999). In another study, isoprenaline and the 3-adrenoceptor selective agonist L 755,507, but not dobutamine or clenbuterol, relaxed carbachol-contracted human bladder strips (Nomiya & Yamaguchi, 2003). A very recent study reported a rank order of potency of isoprenaline>procaterol=CL 316,243=salbutamol, with the latter three compounds being considerably less effective than isoprenaline (Badawi et al., 2005). Finally, GS 332 was found to be more potent in the human bladder than clenbuterol in another study (Morita et al., 2000). In agreement with the predominant expression of 3-adrenoceptor mRNA in the human bladder (see above), these data demonstrate that this subtype is also most important for bladder relaxation in vitro. With the possible exception of ferrets and monkeys, the role of this subtype in other animal species is less prominent.
Figure 2.
Inhibition of isoprenaline-induced relaxation of human bladder detrusor by the 1-antagonist CGP 20,712, the 2-antagonist ICI 118,551 and the nonselective antagonist SR 58,894. Taken with permission from Igawa et al. (1999).
In vivo function
Functional in vivo effects on bladder function can be assessed in several ways. Noninvasive studies frequently look at micturition frequency, which is a key symptom of OAB (see Abrams et al., 2002). Invasive studies are based upon the insertion of a catheter coupled to a pressure transducer into the bladder and subsequent filling of the bladder endogenously or by installation of fluid. This allows various types of measurements, including the frequency of bladder contractions, maximum detrusor pressure, filling volume at first contraction or bladder compliance, all of which are typically also assessed in urodynamic studies in humans (see Abrams et al., 2002). Moreover, it should be considered that the effects of systemically administered drugs on bladder function are not necessarily mediated by drug targets located in the bladder (see the above section on bladder 1-adrenoceptors). Finally, the use of anaesthetized vs conscious animals may differentially affect the endogenous sympathetic tone.
Studies in rats (Lecci et al., 1998; Takeda et al., 2000b; 2003; Kaidoh et al., 2002; Tucci et al., 2002), ferrets (Takeda et al., 2000a) and monkeys (Takeda et al., 2002a) demonstrate that -adrenoceptor agonists such as isoprenaline can reduce intra-vesical pressure, indicating that this is a consistent feature in biology. On the other hand, propranolol had little, if any, effects on bladder function on its own (Durant et al., 1988), indicating that either there is little endogenous