Urethral Smooth Muscle Research Articles

Evidence of Gap Junctions in the Stable Nonobstructed Human Bladder

Evidence of Gap Junctions in the Stable Nonobstructed Human Bladder

Expression Of Estrogen Receptors α And β By Sympathetic Ganglion Neurons Projecting To The Proximal Urethra Of Female Rats

Urinary incontinence is prevalent in postmenopausal women and estrogen is commonly administered therapeutically. In animal models estrogen increases urethral smooth muscle agonist induced contraction but a consistent clinical benefit in humans has not been confirmed. A reason may be that estrogen affects tissues other than the urethra that are involved in continence. We determined if sympathetic nerves projecting to the urethra may also be a target for estrogen. Sympathetic neurons innervating proximal urethra smooth muscle were identified by injection of the retrograde tracer Fast Blue (Dr. Illing GmbH and Co. KG, Gross-Umstadt, Germany) in 10 ovariectomized adult female rats. Rats received a single injection of 10 microg./kg. estradiol benzoate or vehicle 24 hours before tissue harvest. Retrograde labeled sympathetic neurons expressing estrogen receptors alpha and beta in prevertebral and paravertebral ganglia were identified by immunostaining. Approximately 80% of Fast Blue labeled neurons were located in the T11 to L5 paravertebral ganglia. The remainder was located predominantly in the prevertebral suprarenal ganglia with fewer in celiac and superior mesenteric ganglia. Estrogen receptor beta was detected in more than 90% of urethra projecting neurons, while approximately 30% expressed estrogen receptor alpha. No significant change occurred after estrogen administration. Almost all examined sympathetic neurons projecting to the proximal urethra express estrogen receptor beta and a substantial subset expresses estrogen receptor alpha irrespective of estrogen titer. Therefore, estrogen may influence continence by acting not only on the urethral target, but also on its excitatory sympathetic innervation.

Long-Term Effect of Ovariectomy and Simulated Birth Trauma on the Lower Urinary Tract of Female Rats

Using an animal model we studied the long-term effects of ovariectomy and simulated birth trauma in the development of apoptosis as well as the urodynamic, histological and ultrastructural findings 9 months after such procedures. A total of 24 pregnant Sprague-Dawley female rats were used. Immediately after delivery 14 animals underwent vaginal ballooning and ovariectomy, while the remaining 10 served as controls. At 9 months the animals underwent urodynamic evaluation, which included the urethral pressure profile. The rats were then sacrificed and urogenital tissue was obtained for immunostaining using terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate nick end-labeling, histomorphometry evaluation and electron microscopy. Immunostaining demonstrated a significant increase in the apoptotic index in the urethra of castrated/ballooning rats with a predominance in the submucosa layer. Maximum urethral closure pressure was significantly lower in that group, although there was no correlation of apoptosis with maximum urethral closure pressure measurement. Urodynamic evaluation revealed only discrete alterations in cystometric parameters. Morphometric evaluation showed increased connective tissue in the vagina. Electron microscopy of urethral smooth muscle demonstrated altered cellular shape, increased intercellular space with collagen deposition and some degeneration of the mitochondria. Apoptosis in the urethra occurs 9 months after castration and simulated birth trauma. However, this finding was not seen in the muscle layers or in other urogenital tissues. Some ultrastructural changes also occurred that may explain some symptoms that women have after vaginal childbirth and menopause.

An investigation of the effects of zaprinast, a PDE inhibitor, on the nitrergic control of the urethra in anaesthetized female rats

1. The effects of L-NAME and zaprinast were investigated (i.v.) on reflex-evoked changes in bladder and urethral pressures in urethane-anaesthetized female rats. 2. L-NAME attenuated reflex-evoked urethral relaxations (65+/-10%), while zaprinast potentiated these responses (68+/-24%). L-NAME and zaprinast also increased baseline urethral pressure and urethral striated muscle (EUS-EMG) activity. These drugs had little effect on the bladder. 3. Following pre-treatment with alpha-bungarotoxin (i.v.) to block urethral striated muscle, L-NAME and zaprinast failed to increase baseline urethral pressure. Further zaprinast failed to alter the size of reflex-evoked urethral relaxations. 4. Intra-urethral zaprinast caused a significant increase while sodium nitroprusside (SNP) and isoprenaline caused decreases in urethral pressure (+14+/-3%, -25+/-5%, -29+/-7%, respectively). These changes were associated with increases in EUS-EMG activity. After chlorisondamine (i.v.), zaprinast caused a significant fall in urethral pressure, while the decrease in urethral pressure caused by SNP and isoprenaline was potentiated. No changes in EUS-EMG activity occurred. 5. These results indicate that a nitrergic pathway mediates reflex-evoked urethral smooth muscle relaxations. The data also indicates that there is a background release of NO, which reduces sphincter skeletal muscle activity. Further, the ability of zaprinast to potentiate nitrergic evoked urethral relaxations involves an increase in striated muscle tone. This appears to be an indirect result of smooth muscle relaxation and is mediated, at least in part, by a chlorisondamine-sensitive mechanism.

Localization and Role of Nitric Oxide Synthase and Endogenous Nitric Oxide Synthase Inhibitors in the Rabbit Lower Urinary Tract

We investigated the possible role of endogenous methylarginine derivatives, such as NG-monomethyl-L-arginine (L-NMMA), asymmetrical NG, NG-dimethyl-L-arginine (ADMA) and symmetrical NG, N'G-dimethyl-L-arginine (SDMA), for regulating nitric oxide synthase in the rabbit lower urinary tract. Strips of detrusor, trigone and proximal urethra were processed for the determination of endogenous methylarginines and L-arginine by automated high performance liquid chromatography. We also compared nitric oxide synthase activity and nitric oxide mediated functional responses to electrical field stimulation in the 3 regions. Nitric oxide synthase activity was measured by determining the conversion of [3H] L-arginine to [3H] L-citrulline. L-NMMA, ADMA and SDMA were detectable by high performance liquid chromatography in the detrusor, trigone and proximal urethra. Electrical field stimulation induced nitric oxide mediated prominent relaxation in the trigone and proximal urethra, while no relaxation response was observed in the detrusor. Exogenously applied 1 to 100 microM. L-NMMA and 1 to 100 microM. ADMA but not 100 microM. SDMA dependently inhibited Ca2+ dependent nitric oxide synthase activity in all regions, and electrical field stimulation induced relaxation in the trigone and proximal urethra. Inhibition with L-NMMA and ADMA was blocked in the presence of 3 mM. L-arginine but not by 3 mM. D-arginine. Three methylarginines and nitric oxide synthase are localized throughout the rabbit lower urinary tract. Endogenous L-NMMA and ADMA may be involved in regulating nitric oxide biosynthesis of the micturition reflex in the normal or disease state.

Carbon Monoxide Relaxes The Female Pig Urethra As Effectively As Nitric Oxide In The Presence Of Yc-1

Nitric oxide (NO) and carbon monoxide (CO) have been suggested to relax smooth muscle by activating soluble guanylate cyclase (sGC), binding to the same site of the enzyme. 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole (YC-1) (Cayman Co., Malmö, Sweden) increases the catalytic rate of sGC by binding to an allosteric site. We investigated whether YC-1 can modulate the relaxant responses of isolated urethral smooth muscle to exogenous CO, (NO) and electrical field stimulation. In spontaneously active and noradrenaline (Sigma-Aldrich Chemie GmbH, Steinheim, Germany) pre-contracted preparations of circular urethral smooth muscle from female pigs relaxant responses were evoked by electrical field stimulation before and after incubation with 10(-5) M. YC-1. The concentration-response curves for CO and NO were investigated in noradrenaline pre-contracted strips before and after incubation with YC-1. The tissue contents of cyclic 3',5'-guanosine monophosphate (cGMP) and cyclic adenosine monophosphate after electrical field stimulation, and the administration of CO or NO was investigated in the absence and presence of YC-1. YC-1 significantly increased the amplitude of the relaxations evoked by electrical field stimulation, CO and NO, and simultaneously caused significant increases in the cGMP content in all preparations. The effect on CO induced relaxant responses was conspicuous. In the presence of YC-1 the potency and maximal relaxant effect of CO were similar to those of NO in the absence of YC-1. YC-1 enhances cGMP dependent relaxant responses of the female pig urethra in vitro. The finding that the response to CO was greatly increased after sensitizing sGC suggests a potential for CO as a relaxant mediator in urethral smooth muscle.

Bladder and urethral responses to pelvic nerve stimulation in the pig.

The aim of our study of the pig was to investigate the responses of smooth muscle of the bladder and the urethral sphincter to preganglionic parasympathetic stimulation, and to assess the effect on the lower urinary tract of IV administration of alpha,beta-methylene-ATP. In seven anaesthetised female pigs, the responses to repeated 20 s pelvic nerve stimulations before and after IV administration of 0.02 mg/kg alpha-beta-methylene-ATP, and the responses to the drug itself, were recorded in the bladder and the urethra separately. In the urethral high-pressure zone, pre-stimulation pressure was a mean of 61+/-11 cmH2O. During pelvic nerve stimulation, urethral pressure declined by 48+/-9 cmH2O, while the bladder pressure increased to 30+/-18 cmH2O. The rate of pressure changes during the first 3 s of stimulation (initiation of voiding) was larger in the urethra than in the bladder (urethral pressure decrease: 13.0+/-3.1 cmH2O/s, bladder pressure increase: 3.2+/-2.5 cmH2O/s). Administration of alpha,beta-methylene-ATP was followed by a significant but temporary enlargement in the bladder response to pelvic nerve stimulation to 36+/-20 cmH2O, p = 0.028, n = 7, but no change in urethral response. At least 80% of the urethral pre-stimulation pressure was exerted by the smooth muscle. The synergic activation of the detrusor and the urethral smooth muscle in response to preganglionic parasympathetic nerve stimulation was controlled by the peripheral nerves or by the neuromuscular transmission. Administration of alpha,beta-methylene-ATP increased the bladder response to pelvic nerve stimulation without changing the urethral response.

Pharmacological Effects of Berberine and Palmatine on the Prostatic and Urethral Smooth Muscle of the Rabbit

Pharmacological Effects of Berberine and Palmatine on the Prostatic and Urethral Smooth Muscle of the Rabbit

CONTRIBUTION OF CEREBRAL NITRIC OXIDE TO BLADDER OVERACTIVITY AFTER CEREBRAL INFARCTION IN RATS

We investigated the contribution of cerebral nitric oxide to neurogenic voiding dysfunction after cerebral infarction. The left mid cerebral artery in female Sprague-Dawley rats was occluded with 4-zero monofilament nylon thread. Bladder activity was monitored during infusion cystometrography. Time or dose dependent effects of intracerebral ventricular administration of the nonselective nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME), were investigated in conscious, sham operated and cerebral infarcted rats. The selective neuronal nitric oxide synthase inhibitor 1-(2-trifluoromethylphenyl) imidazole was also administered to determine the participation of nitric oxide synthase subtypes. Cross-sectional infarct area was measured and infarct volume was calculated 12 hours after mid cerebral artery occlusion. Bladder capacity was reduced by 54% 30 minutes after mid cerebral artery occlusion. L-NAME significantly increased bladder capacity in a dose and time dependent manner in cerebral infarcted rats but had no effect on sham operated rats. L-NAME (50 microg./kg.) administered 3 or 5 hours after occlusion significantly increased bladder capacity. This effect of L-NAME was reversed by injecting 250 microg. L-arginine per rat, which alone did not produce any significant change in bladder capacity in cerebral infarcted rats. Administration of 1-(2-trifluoromethylphenyl) imidazole also significantly increased bladder capacity in these rats. On the other hand, 5 microg. of the nitric oxide donor FK-409 per rat reduced bladder capacity for 10 to 15 minutes. None of the drugs affected infarct volume. These results indicate that supraspinal nitric oxide has an important role in bladder overactivity after cerebral infarction but it does not affect normal micturition in rats. This finding suggests a central mechanism sensitive to nitric oxide for bladder overactivity after cerebral infarction.

The involvement of L-type Ca(2+) channels in the relaxant effects of the ATP-sensitive K(+) channel opener ZD6169 on pig urethral smooth muscle.

1. The effects of ZD6169, a novel ATP-sensitive K(+) channel (K(ATP) channel) opener, were investigated on membrane currents in isolated myocytes using patch-clamp techniques. Tension measurement was also performed to study the effects of ZD6169 on the resting tone of pig urethral smooth muscle. 2. Levcromakalim was more potent than ZD6169 in lowering the resting urethral tone. Relaxation induced by low concentrations of ZD6169 (< or =3 microM) was completely suppressed by additional application of glibenclamide (1 microM). In contrast, glibenclamide (1-10 microM) only partially inhibited the relaxation induced by higher concentrations of ZD6169 (> or = microM). 3. Bay K8644 (1 microM) reduced the maximum relaxation produced by ZD6169 (> or =10 microM). 4. In whole-cell configuration, ZD6169 suppressed the peak amplitude of voltage-dependent Ba(2+) currents in a concentration- and voltage-dependent manner, and at 100 microM, shifted the steady-state inactivation curve of the voltage-dependent Ba(2+) currents to the left at a holding potential of -90 mV. 5. In cell-attached configuration, open probability of unitary voltage-dependent Ba(2+) channels (27 pS, 90 mM Ba(2+)) was inhibited by 100 microM ZD6169 and by 10 microM nifedipine. 6. Reverse transcriptase-polymerase chain reaction (RT - PCR) analysis revealed the presence of the transcript of the alpha(1C) subunit of L-type Ca(2+) channels in pig urethra. 7. These results demonstrate that ZD6169 causes urethral relaxation through two distinct mechanisms, activation of K(ATP) channels at lower concentrations and inhibition of voltage-dependent Ca(2+) channels at higher concentrations (about 10 microM).

URETHRA IS MORE SENSITIVE TO ISCHEMIA THAN BLADDER: EVIDENCE FROM AN IN VITRO RAT STUDY

URETHRA IS MORE SENSITIVE TO ISCHEMIA THAN BLADDER: EVIDENCE FROM AN IN VITRO RAT STUDY

Neurotransmission and drug effects in urethral smooth muscle.

Urethral smooth muscle tension is one of the factors contributing to the maintenance of intraurethral pressure and to continence. This tension is controlled by nervous mechanisms involving a complex pattern of efferent and afferent signalling in parasympathetic, sympathetic, and somatic nerves, with resulting release of contraction- and relaxation-producing transmitters and mediators. Sympathetic activity, via release of NA and stimulation of urethral smooth muscle alpha1-ARs, is the main contraction-producing factor. Among the three high affinity alpha1-AR subtypes identified in molecular cloning and functional studies, the alpha1A subtype seems to predominate in both the male and female urethra. The physiological role of the urethral beta-adrenoceptors has not been established, and the importance of muscarinic receptors (and subtypes) for urethral function is presently unclear. Nitric oxide, produced by nitric oxide synthase within cholinergic nerves, seems to be the predominant inhibitory neurotransmitter, but there is good evidence for the existence of other, as yet unidentified, non-adrenergic, non-cholinergic inhibitory messengers. Vasoactive intestinal polypeptide, adenosine 5'-triphosphate, and carbon monoxide (CO) all relax urethral smooth muscle, and are potential candidates for the nonadrenergic, noncholinergic relaxation, but their roles for urethral function remain to be defined.

Regional variation in myosin isoforms and phosphorylation at the resting tone in urinary bladder smooth muscle.

Urinary bladder filling and emptying requires coordinated control of bladder body and urethral smooth muscles. Bladder dome, midbladder, base, and urethra showed significant differences in the percentage of 20-kDa myosin light chain (LC20) phosphorylation (35.45 +/- 4.6, 24.7 +/- 2.2, 13.6+/- 2.1, and 12.8 +/- 2.7%, respectively) in resting muscle. Agonist-mediated force was associated with a rise in LC20 phosphorylation, but the extent of phosphorylation at all levels of force was less for urethral than for bladder body smooth muscle. RT-PCR and quantitative competitive RT-PCR analyses of total RNA from bladder body and urethral smooth muscles revealed only a slight difference in myosin heavy chain mRNA copy number per total RNA, whereas mRNA copy numbers for NH2-terminal isoforms SM-B (inserted) and SM-A (noninserted) in these muscles showed a significant difference (2.28 x 10(8) vs. 1.68 x 10(8) for SM-B and 0.12 x 10(8) vs. 0.42 x 10(8) for SM-A, respectively), which was also evident at the protein level. The ratio of COOH-terminal isoforms SM2:SM1 in the urethra was moderately but significantly lower than that in other regions of the bladder body. A high degree of LC20 phosphorylation and SM-B in the bladder body may help to facilitate fast cross-bridge cycling and force generation required for rapid emptying, whereas a lower level of LC20 phosphorylation and the presence of a higher amount of SM-A in urethral smooth muscle may help to maintain the high basal tone of urethra, required for urinary continence.

Neural control of the urethra.

Coordination between the urinary bladder and the urethra is mediated by multiple reflex pathways organized in the brain and spinal cord. Some reflexes promote urine storage; whereas other reflexes facilitate voiding. During bladder filling, activation of mechanoreceptor afferent nerves in the bladder wall triggers firing in the cholinergic efferent pathways to the external urethral sphincter (EUS) and in sympathetic adrenergic pathways to the urethral smooth muscle. These storage reflexes are dependent upon interneuronal circuitry in the spinal cord. During voiding the spinal storage reflexes are inhibited by supraspinal mechanisms which originate in the pontine micturition center. Glutamatergic, serotonergic and alpha, adrenergic excitatory transmission as well as GABAergic/glycinergic inhibitory transmission have been implicated in the central control of sphincter reflexes. During voiding, a parasympathetic nitrergic inhibitory input to the urethral smooth is activated. This reflex mechanism which is triggered by bladder afferents persists in paraplegic rats and therefore must be mediated at least in part by spinal interneuronal circuitry. In female rats, the parasympathetic nitrergic pathway is prominent; but in male rats it is obscured by a dominant parasympathetic cholinergic excitatory input to the urethral smooth muscle. The function of the cholinergic pathway in voiding is uncertain. Stimulation of urethral afferents can also influence bladder activity. Contraction of the external urethral sphincter activates afferents that inhibit reflex bladder contractions; whereas infusion of fluid through the urethra facilitates bladder contractions. These reflexes are also organized in the spinal cord and presumably play a role in urine storage and elimination. Alterations in primitive bladder-to-urethra and urethra-to-bladder reflex mechanisms may contribute to neurogenic bladder dysfunction.

Functional characteristics of urinary tract smooth muscles in mice lacking cGMP protein kinase type I.

Nitric oxide (NO)-mediated smooth muscle relaxation is mediated by cGMP through activation of cGMP-dependent protein kinase I (cGKI). We studied the importance of cGKI for lower urinary tract function in mice lacking the gene for cGKI (cGKI-/-) and in litter-matched wild-type mice (cGKI+/+) in vitro and in vivo. cGKI deficiency did not result in any changes in bladder gross morphology or weight. Urethral strips from cGKI-/- mice showed an impaired relaxant response to nerve-derived NO. The cGMP analog 8-bromo-cGMP (8-BrcGMP) and the NO-donor SIN-1 relaxed the wild-type urethra (50-60%) but had only marginal effects in the cGKI-deficient urethra. Bladder strips from cGKI-/- mice responded normally to electrical field stimulation and to carbachol but not to 8-BrcGMP. In vivo, the cGKI-deficient mice showed bladder hyperactivity characterized by decreased intercontraction intervals and nonvoiding bladder contractions. Loss of cGKI abolishes NO-cGMP-dependent relaxations of urethral smooth muscle and results in hyperactive voiding. These data suggest that certain voiding disturbances may be associated with impaired NO-cGKI signaling.

Contractile properties of urethral smooth muscles of young and aged female dogs: morphological and pharmacological aspects.

Alpha1-adrenoceptors are highly concentrated in the urethral smooth muscles and may play an important role in the contraction of this area. However, detailed examinations of age-related changes of the properties of urethral smooth muscle have rarely been undertaken. The contractile properties of urethras from young non-parous and old parous female beagles were determined with a urethral function study, macroscopic autoradiography for urethras using [3H]-labeled tamsulosin and morphometry of the urethral muscles. The antagonistic effect (pA2) of prazosin for norepinephrine was 7.76+/-0.13 in young dogs and 7.62+/-0.06 in aged dogs. The specific binding of [3H]-tamsulosin (a relatively selective alpha1A-adrenoceptor antagonist) was recognized diffusely in proximal urethras with in vitro autoradiography. The density of binding in smooth muscles was approximately 60 and 40% in circular longitudinal layers, respectively, for both dogs. The female canine urethra had alpha1A, and alpha1L-adrenoceptors. No age-related changes were seen in the function of the proximal urethra, distribution of alpha1-adrenoceptor binding sites and smooth muscle densities.

Intraurethral ultrasonography: correlation of urethral anatomy with functional urodynamic parameters in stress incontinent women.

To determine whether differences in functional urodynamic parameters can be explained by changes in urethral anatomy, 39 patients underwent intraurethral ultrasonography to obtain a 360 degree view of the urethra. The point of maximal rhabdosphincter thickness was identified in all patients. The thickness, circumference and area of the urethral smooth and skeletal muscle layers were calculated. Data from patient histories and urodynamic evaluations were compared with this anatomical survey. The urodynamic diagnoses were as follows: 10 patients were normal, 24 had genuine stress incontinence and 5 had intrinsic sphincter deficiency. These patients had decreasing rhabdosphincter thicknesses of 3.91, 3.35 and 2.70 mm (P = 0.048). A weak linear relationship was found between maximal urethral closure pressure and rhabdosphincter (r = 0.40, P = 0.013) and longitudinal smooth muscle (r = 0.35, P=0.027) thickness. It was concluded that a loss of urethral resistance as measured by maximal urethral closure pressure is associated with changes in urethral anatomy identified by intraurethral ultrasonography.

Prejunctional α-adrenoceptors regulate nitrergic neurotransmission in the rabbit urethra

We evaluated the effects of prejunctional α-adrenoceptors on nitric oxide (NO)-mediated urethral relaxation in rabbits using a muscle bath technique and high-performance liquid chromatography coupled with a microdialysis procedure. The amount of NO 2 −/NO 3 − released during electrical field stimulation was measured by an NO 2 −/NO 3 − analyzer based on the Griess method. Pretreatment with phenylephrine (0.01 μM) and yohimbine (0.1–10 μM) significantly reduced the relaxation responses induced by electrical field stimulation. In contrast, pretreatment with clonidine (0.01 μM) and prazosin (0.01–1 μM) enhanced the relaxation responses. Cys-NO-induced relaxations of rabbit urethral smooth muscle were not affected by pretreatment with α-adrenoceptor agonists and antagonists. The amount of NO 2 −/NO 3 − released by electrical field stimulation increased after pretreatment with clonidine (0.01 μM) and prazosin (0.01–1 μM), but decreased after pretreatment with phenylephrine (0.01 μM) and yohimbine (0.1–10 μM). The results suggest that the release of NO from nitrergic nerves in the rabbit urethra is reduced and increased by stimulation of prejunctional α 1- and α 2-adrenoceptors, respectively.

Ultrastructure of detrusor and urethral smooth muscle in women with urinary incontinence

Objective: We performed a quantitative study to determine whether mixed urinary incontinence was associated with any ultrastructural changes in detrusor and urethral smooth muscle. Study Design: Detrusor and urethral smooth muscle biopsy specimens were obtained at the time of laparotomy from 5 women aged 35 to 65 years with mixed urinary incontinence and from a control group of 5 continent women. Smooth muscle morphologic characteristics were assessed from a systematic random sample of electron micrographs. A further 16 urethral biopsy specimens were similarly analyzed to confirm the findings of the initial study. Results: The electron-dense portion of the sarcolemma was smaller in urethral biopsy specimens taken from patients with intrinsic sphincter deficiency than in those from control subjects (χ 2 (1) = 4.9; P = .027). No other morphologic characteristics were unique to patients with incontinence. Conclusions: Our study suggests that focal adhesion architecture is decreased in urethral smooth muscle of patients with intrinsic sphincter deficiency. (Am J Obstet Gynecol 2000;182:879-84.)

Spontaneous photo-relaxation of urethral smooth muscle from sheep, pig and rat and its relationship with nitrergic neurotransmission.

1. In the present work we have characterized the relaxant response induced by light stimulation (LS) in the lower urinary tract from sheep, pig and rat, establishing its relationship with nitrergic neurotransmission. 2. Urethral, but not detrusor, preparations showed pronounced photo-relaxation (PR) which declined progressively following repetitive LS. Sheep urethral PR was again restored either spontaneously or (to a greater extent) by exogenous nitric oxide (NO) addition and by electrical field stimulation (EFS) of intrinsic nitrergic nerves. 3. Greater NO generation was detected from sheep urethral than from detrusor homogenates following illumination. 4. Sheep urethral PR was inhibited by oxyhaemoglobin, but not by methaemoglobin, carboxy-PTIO, extracellular superoxide anion generators or superoxide dismutase. Guanylyl cyclase but not adenylyl cyclase activation mediates urethral relaxation to LS. 5. Urethral PR was more resistant to inhibition by L-thiocitrulline than EFS-induced responses, although this agent prevented PR restoration by high-frequency EFS. 6. Urethral PR was TTX insensitive and partially modified in high-K+ solutions. Cold storage for 24 h greatly impaired urethral PR, although it was restored by high-frequency EFS. 7. Repetitive exposure to LS, EFS or exogenous NO induced changes in the shape of the EFS-induced nitrergic relaxation, possibly by pre-synaptic mechanisms. 8. In conclusion, we suggest the presence of an endogenous, photo-labile, nitro-compound store in the urethra, which seems to be replenished by neural nitric oxide synthase activity, indicating a close functional relationship with the nitrergic neurotransmitter.