The pivotal role of neuronal nitric oxide synthase in the release of 6-nitrodopamine from mouse isolated vas deferens.

6-Nitrodopamine (6-ND) is released from rat and human vas deferens and is considered a major mediator of both tissues contractility. The contractions induced by 6-ND are selectively blocked by both tricyclic antidepressants and α1-adrenoceptor antagonists. Endothelial nitric oxide synthase (eNOS) is the major isoform responsible for 6-ND release in mouse isolated heart, however the origin of 6-ND in the vas deferens is unknown. Here it was investigated by LC-MS/MS the basal release of 6-ND from isolated vas deferens obtained from control, eNOS-/-, nNOS-/-, and iNOS-/- mice. In addition, it was evaluated in vitro vas deferens contractility following electric field stimulation (EFS). Basal release of 6-ND was significantly reduced in nNOS-/- mice compared to control mice, but not decreased when the vas deferens were obtained from either eNOS-/- or iNOS-/- mice. Pre-incubation of the vas deferens with tetrodotoxin (1 µM) significantly reduced the basal release of 6-ND from control, eNOS-/-, and iNOS-/- mice but had no effect on the basal release of 6-ND from nNOS-/- mice. EFS-induced frequency-dependent contractions of the vas deferens, which were significantly reduced when the tissues obtained from control, eNOS-/- and iNOS-/- mice, were pre-incubated with l-NAME, but unaltered when the vas deferens was obtained from nNOS-/- mice. In addition, the EFS-induced contractions were significantly smaller when the vas deferens were obtained from nNOS-/- mice. The results clearly demonstrate that nNOS is the main NO isoform responsible for 6-ND release in mouse vas deferens and reinforces the concept of 6-ND as a major modulator of vas deferens contractility.

Effects of RS17053 on α1 -adrenoceptors in rat vas deferens and aorta.

BACKGROUND: RS17053 is classed as an α1A -adrenoceptor selective antagonist. OBJECTIVES: We have examined its profile of action at all subtypes of α1 -adrenoceptor. METHODS: Noradrenaline (NA) evoked contractions of rat vas deferens involve α1D -adrenoceptors in phasic contractions and α1A -adrenoceptors in tonic contractions. Contractions of rat aorta to NA involve α1D - and α1B -adrenoceptors. RESULTS: RS17053 (10-5  M) shifted NA potency and virtually abolished tonic contractions to NA, with little or limited effect on phasic contractions. The α1D -adrenoceptor antagonist BMY7378 (3 × 10-7 M) significantly inhibited the remaining phasic component of the contractions, and the α1A -adrenoceptor antagonist RS100329 (10-7  M) inhibited further the residual tonic contraction. Hence, RS17053 shows high selectivity for α1A -adrenoceptors over α1D -adrenoceptors in rat vas deferens. However, RS17053 (10-5  M) produced a large shift in the potency of NA in rat aorta, with a pKB of 6.82. Large shifts of NA potency in rat aorta involve α1B -adrenoceptor blockade. CONCLUSION: Results in rat vas deferens demonstrate low potency of RS17053 at α1D -adrenoceptors, but results from rat aorta can only be explained as demonstrating α1B -adrenoceptor antagonism by RS17053. RS17053 may be a useful pharmacological tool when reclassified as a mainly α1A - and to a lesser extent α1B -adrenoceptor antagonist with little effect at α1D -adrenoceptors.

The P2X1 receptor as a therapeutic target.

Within the family of purinergic receptors, the P2X1 receptor is a ligand-gated ion channel that plays a role in urogenital, immune and cardiovascular function. Specifically, the P2X1 receptor has been implicated in controlling smooth muscle contractions of the vas deferens and therefore has emerged as an exciting drug target for male contraception. In addition, the P2X1 receptor contributes to smooth muscle contractions of the bladder and is a target to treat bladder dysfunction. Finally, platelets and neutrophils have populations of P2X1 receptors that could be targeted for thrombosis and inflammatory conditions. Drugs that specifically target the P2X1 receptor have been challenging to develop, and only recently have small molecule antagonists of the P2X1 receptor been available. However, these ligands need further biological validation for appropriate selectivity and drug-like properties before they will be suitable for use in preclinical models of disease. Although the atomic structure of the P2X1 receptor has yet to be determined, the recent discovery of several other P2X receptor structures and improvements in the field of structural biology suggests that this is now a distinct possibility. Such efforts may significantly improve drug discovery efforts at the P2X1 receptor.

Existence and distribution of Niemann-Pick type 2C (NPC2) in prawn reproductive tract and its putative role as a cholesterol modulator during sperm transit in the vas deferens.

Sequestering of cholesterol (CHO) is a hallmark molecular event that is known to be associated with sperm gaining their fertilizing ability in a broad array of animals. We have shown previously that the level of CHO declines in the Macrobrachium rosenbergii sperm membrane when they are migrating into the vas deferens, prompting us to search for CHO transporters, one of which is Niemann-Pick type 2C (NPC2), within the prawn male reproductive tract. Sequence comparison of MrNPC2 with other NPC2, from crustaceans to mammals, revealed its conserved features in the hydrophobic cavity with 3 amino acids forming a CHO lid that is identical in all species analyzed. Expressions of MrNPC2 transcript and protein were detected in testicular supporting and interstitial cells and along the epithelial cells of the vas deferens. As confirmed by live cell staining, the testicular sperm (Tsp) surface was devoid of MrNPC2 but it first existed on the vas deferens sperm, suggesting its acquisition from the luminal fluid, possibly through trafficking of multi-lamellar vesicles during sperm transit in the vas deferens. We further showed that recombinant MrNPC2 had a high affinity towards CHO in the lipid extracts, either from Tsp or from lipid vesicles in the vas deferens. Together, our results indicated the presence of MrNPC2 in the male reproductive tract, which may play an important role as a CHO modulator between the sperm membrane and vas deferens epithelial communication.

Neurotoxicity of Tityus bahiensis (brown scorpion) venom in sympathetic vas deferens preparations and neuronal cells.

Systemic scorpion envenomation is characterized by massive neurotransmitter release from peripheral nerves mediated primarily by scorpion venoms neurotoxins. Tityus bahiensis is one of the medically most important species in Brazil, but its venom pharmacology, especially regarding to peripheral nervous system, is poorly understood. Here, we evaluated the T. bahiensis venom activity on autonomic (sympathetic) neurotransmission by using a variety of approaches, including vas deferens twitch-tension recordings, electrophysiological measurements (resting membrane potentials, spontaneous excitatory junctional potentials and whole-cell patch-clamp), calcium imaging and histomorphological analysis. Low concentrations of venom (≤ 3 µg/mL) facilitated the electrically stimulated vas deferens contractions without affecting postsynaptic receptors or damaging the smooth muscle cells. Transient TTX-sensitive sustained contractions and resting membrane depolarization were mediated mainly by massive spontaneous ATP release. High venom concentrations (≥ 10 µg/mL) blocked the muscle contractions and induced membrane depolarization. In neuronal cells (ND7-23wt), the venom increased the peak sodium current, modified the current-voltage relationship by left-shifting the Nav-channel activation curve, thereby facilitating the opening of these channels. The venom also caused a time-dependent increase in neuronal calcium influx. These results indicate that the sympathetic hyperstimulation observed in systemic envenomation is presynaptically driven, probably through the interaction of α- and ß-toxins with neuronal sodium channels.

Localization of sperm intracellular Ca2+ keeps fertilizability in the newt vas deferens.

Sperm intracellular Ca2+ is crucial for the induction of sperm-egg interaction, but little is known about the significance of Ca2+ maintenance prior to induction. In sperm of the newt Cynops pyrrhogaster, intracellular Ca2+ is localized to the midpiece during storage in the vas deferens, while extracellular Ca2+ is influxed in modified Steinberg's salt solution to promote a spontaneous acrosome reaction related to the decline of sperm quality. In the present study, sperm from the vas deferens were loaded with the Ca2+ indicator Fluo8H, and changes in Ca2+ localization in modified Steinberg's salt solution were examined. Calcium ions expanded from the cytoplasmic area of the midpiece to the entire tail in most sperm during a 1-h incubation and localized to the principal piece in some sperm within 24 h. Similar changes in Ca2+ localization were observed in reconstructed vas deferens solution that included ions and pH at equivalent levels to those in the vas deferens fluid. Sperm with Ca2+ localization in the entire tail or the principal piece weakened or lost responsiveness to sperm motility-initiating substances, which trigger sperm motility for fertilization, but responded to a trigger for acrosome reaction. The change in Ca2+ localization was delayed and transiently reversed by ethylene glycol tetraacetic acid or a mixture of Ca2+ channel blockers including Ni2+ and diltiazem. These results suggest that C. pyrrhogaster sperm localize intracellular Ca2+ to the midpiece through Ca2+ transport in the vas deferens to allow for responses to sperm motility-initiating substances.

The effects of varying Mg2+ ion concentrations on contractions to the cotransmitters ATP and noradrenaline in the rat vas deferens.

The vas deferens responds to a single electrical pulse with a biphasic contraction caused by cotransmitters ATP and noradrenaline. Removing Mg2+ (normally 1.2 mM) from the physiological salt solution (PSS) enhances the contraction. This study aimed to determine the effect of Mg2+ concentration on nerve cotransmitter-mediated contractions. Rat vasa deferentia were sequentially bathed in increasing (0, 1.2, 3 mM) or decreasing (3, 1.2, 0 mM) Mg2+ concentrations. At each concentration a single field pulse was applied, and the biphasic contraction recorded. Contractions to exogenous noradrenaline 10 µM and ATP 100 µM were also determined. The biphasic nerve-mediated contraction was elicited by ATP and noradrenaline as NF449 (10 µM) and prazosin (100 nM) completely prevented the respective peaks. Taking the contractions in normal PSS (Mg2+ 1.2 mM) as 100%, lowering Mg2+ to 0 mM enhanced the ATP peak to 170 ±â€¯7% and raising Mg2+ to 3 mM decreased it to 39 ±â€¯3%; the noradrenaline peak was not affected by lowering Mg2+ to 0 mM (97 ±â€¯3%) but was decreased to 63 ±â€¯4% in high Mg2+ (3 mM). Contractions to exogenous ATP, but not noradrenaline, were increased in Mg2+ 0 mM and both were inhibited with Mg2+ 3 mM. Changing Mg2+ concentration affects the contractions elicited by the cotransmitters ATP and noradrenaline. The greatest effects were to potentiate the contraction to ATP in Mg2+ 0 mM and to inhibit the contraction to both ATP and noradrenaline in high Mg2+. Future publications should clearly justify any decision to vary the magnesium concentration from normal (1.2 mM) values.

Effects of mu opioid receptors in paraventricular nucleus on ejaculation through mediating sympathetic nerve system activity.

To investigate the roles of mu opioid receptors (MORs) in paraventricular nucleus of the hypothalamus (PVN) on ejaculation and its underlying mechanism in the rats, we performed copulation behavioral testing and acute experiments. During the acute experiments, mean arterial pressure (MAP), heart rate (HR), bulbospongiosus muscle-electromyogram (BSM-EMG) and pressure of vas deferens (PVD) were all recorded. The expression levels and distributions of opioid receptors were also assessed in PVN of male rats. Moreover, adeno-associated virus type 1 (AAV1) was microinjected into PVN to demonstrate whether there are direct projections from PVN to lumbar spinothalamic (LSt) cells. We found that microinjection of MOR agonist, D-A1a2-NM9-Phe4-Gly(ol)5enkephalin (DAGO), into the PVN prolonged the intromission latency and inhibited ejaculation (P = 0.0241, P = 0.0473, respectively), while the opposed results appeared in CTAP (D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2, MOR antagonist) group (P = 0.0021, P = 0.0286, respectively). Moreover, DAGO caused a significant decrease in MAP and HR (P = 0.0065, P = 0.0030, respectively), and PVD decreased significantly after DAGO microinjection in PVN (P = 0.0383). CTAP not only blocked the effect of DAGO but also significantly increased MAP, HR and PVD (P = 0.0003, P = 0.0010, P = 0.0074, respectively). Meanwhile, a significant increase was observed in BSM-EMG activity after microinjecting of CTAP (P = 0.0022), accompanied by visible BSM contraction. Additionally, anterograde monosynaptic transneuronal tracer AAV1 labeling revealed that neurons in PVN projected directly to LSt cells in L3-4 spinal cord. These results indicate that MORs in PVN centrally mediate ejaculation by regulating the sympathetic outflow, which may be treated as a therapeutic target for ejaculation disorders in the future.

Ultrastructure of the vasa deferentia of Terrobittacus implicatus and Cerapanorpa nanwutaina (Insecta: Mecoptera).

The fine structures of vasa deferentia and postvesicular vasa deferentia were investigated in the hangingfly Terrobittacus implicatus (Cai et al. 2006) and the scorpionfly Cerapanorpa nanwutaina (Chou 1981) using light and transmission electron microscopy, and schematic diagrams were drawn accordingly. The vasa deferentia of both species comprise muscular layers, a basal lamina, and a mono-layered epithelium, but the postvesicular vasa deferentia contain muscular layers, a basal lamina, a single-layered epithelium, a subcuticular cavity, and an inner cuticle respectively. The vas deferens releases secretions into the lumen directly, probably by means of merocrine production. On the contrary, the cells of the postvesicular vas deferens correspond to class I glandular cells, discharging secretions into the subcuticular cavity first, and then into the lumen through an inner cuticle. The epithelium in both structures of Bittacidae is well developed and contains more microvilli, organelles, and more types of secretions than in Panorpidae. In Panorpidae, the spine of the postvesicular vas deferens may serve as a barricade for the reflow of the sperm and to protect the extraordinarily long structure from being collapsed or injured.

Methylhexaneamine causes tachycardia and pressor responses indirectly by releasing noradrenaline in the rat.

We have investigated the mode of cardiovascular action of the stimulant methylhexaneamine (MHA) in terms of direct or indirect adrenergic actions in anaesthetised rats. Male and female rats were anaesthetised with pentobarbitone and pressor (changes in diastolic blood pressure) and cardioaccelerator responses to MHA were examined in vehicle treated or chemically sympathectomised rats. MHA produced pressor and cardioaccelerator responses over the same dose range in vehicle treated animals, with significant cardioaccelerator and pressor responses occurring at MHA (0.1 mg/kg). However, tachycardia was more marked than pressor responses. In sympathectomised rats, cardiac and pressor actions of MHA were greatly attenuated. MHA was also studied in isolated tissues. In rat vas deferens, MHA produced small tonic contractions, but these were virtually abolished by sympathectomy In rat aorta, MHA produced almost no contractions. These results are also consistent with largely indirect actions. There were no differences between male and female rats. It is concluded that MHA acts predominantly indirectly in both male and female rats causing noradrenaline release to produce cardiovascular actions and that as a result pressor and cardiac responses occur at similar doses. This propensity for MHA to cause tachycardia and rises in blood pressure at similar doses range may have implications for adverse cardiovascular actions.

Mechanisms of butylidenephthalide for twitch facilitation in electrically stimulated mouse vas deferens.

CONTEXT: The rhizome of Ligusticum chuaxiong Hort. (Umbelliferae) has been used by Chinese for several thousand years. Its main constituent, butylidenephthalide (Bdph), was proved to be active in inhibiting rat uterine contractions induced by prostaglandin F2α and was reported to be a nonspecific antispamodic and a blocker of voltage-dependent Ca2+ channels (VDCCs). OBJECTIVES: The present study investigates the mechanisms of Bdph for twitch facilitation in ICR mouse vas deferens (MVD). MATERIALS AND METHODS: Electrical field stimulation (EFS, supramaximal voltage ranging from 60-90 V, 1 ms, 0.2 Hz) was applied to the isolated MVD in Krebs solution. Interactions between Bdph (50 µM) and calcium antagonist (verapamil, diltiazem or aspaminol) on the EFS-evoked twitch responses were determined. The number of experiments was 3-18. RESULTS: Bdph (50 µM)-induced twitch facilitations from 100 to 391.9% were unrelated to activation of postjunctional cholinergic or adrenergic receptors. Verapamil and Bdph unabolished the twitch facilitation each other. Diltiazem unabolished the Bdph-induced twitch facilitation. In contrast, Bdph abolished those induced by diltiazem. Aspaminol at 20 µM abolished the Bdph-induced twitch facilitation. In contrast, Bdph abolished those induced by aspaminol. Tetraethylammonium and 4-aminopyridine, the K+ channel blockers, significantly augmented the Bdph-induced twitch facilitation. DISCUSSION AND CONCLUSIONS: Bdph may bind to the different, more and same subtypes of VDCCs from verapamil, than diltiazem, and as aspaminol does on prejunctional membrane, respectively. Besides a blocker of VDCCs, Bdph may be a blocker of K+ channels on prejunctional membrane. Thus, Bdph depolarized the membrane and facilitated the cumulative Ca2+-induced twitch responses.

Big conductance calcium-activated potassium channel openers control spasticity without sedation.

BACKGROUND AND PURPOSE: Our initial aim was to generate cannabinoid agents that control spasticity, occurring as a consequence of multiple sclerosis (MS), whilst avoiding the sedative side effects associated with cannabis. VSN16R was synthesized as an anandamide (endocannabinoid) analogue in an anti-metabolite approach to identify drugs that target spasticity. EXPERIMENTAL APPROACH: Following the initial chemistry, a variety of biochemical, pharmacological and electrophysiological approaches, using isolated cells, tissue-based assays and in vivo animal models, were used to demonstrate the activity, efficacy, pharmacokinetics and mechanism of action of VSN16R. Toxicological and safety studies were performed in animals and humans. KEY RESULTS: VSN16R had nanomolar activity in tissue-based, functional assays and dose-dependently inhibited spasticity in a mouse experimental encephalomyelitis model of MS. This effect occurred with over 1000-fold therapeutic window, without affecting normal muscle tone. Efficacy was achieved at plasma levels that are feasible and safe in humans. VSN16R did not bind to known CB1 /CB2 /GPPR55 cannabinoid-related receptors in receptor-based assays but acted on a vascular cannabinoid target. This was identified as the major neuronal form of the big conductance, calcium-activated potassium (BKCa ) channel. Drug-induced opening of neuronal BKCa channels induced membrane hyperpolarization, limiting excessive neural-excitability and controlling spasticity. CONCLUSIONS AND IMPLICATIONS: We identified the neuronal form of the BKCa channel as the target for VSN16R and demonstrated that its activation alleviates neuronal excitability and spasticity in an experimental model of MS, revealing a novel mechanism to control spasticity. VSN16R is a potential, safe and selective ligand for controlling neural hyper-excitability in spasticity.

Effects of in vitro, acute and chronic treatment with fluoxetine on the sympathetic neurotransmission of rat vas deferens.

It is described that fluoxetine treatment is able to induce ejaculatory disorders. However, the exact mechanism is still not fully understood. Therefore, this study was carried out to further evaluate the anti-ejaculatory effects of fluoxetine, using different approaches (in vitro or in vivo treatments), on the sympathetic neurotransmission of the rat vas deferens. Vas deferens from male Wistar rats were used to check the in vitro effects of fluoxetine 10-6M, 3.10-6M or 10-5M. Animals were also acutely (20mg/kg, i.p. 4h or 24h) or chronically (10mg/kg, i.p., 30days) treated with fluoxetine or drug-free vehicle. The vas deferens from non-treated and treated animals were isolated and mounted in an isolated organ bath for the study of the contractions induced by adrenergic agonists, tyramine, 5-HT, Ca2+ or electrical field stimulation. In vitro or acute treatment with fluoxetine decreased the contraction induced by agonists, Ca2+ or electrical field stimulation. The chronic treatment with fluoxetine decreased the contractions induced agonists, tyramine or Ca2+, but did not modify the contractions induced by electrical field stimulation. We have shown that in vitro or in vivo fluoxetine treatment is able to alter the sympathetic neurotransmission of the rat vas deferens which could be related to alterations in the calcium signalling.

The human epididymis: its function in sperm maturation.

BACKGROUND: Spermatozoa acquire their fertilizing ability and forward motility properties during epididymal transit. Our knowledge of gamete physiology is based on studies conducted in laboratory and domestic species; our knowledge of these processes in humans is limited. Medical indications for assisted reproductive technologies (ART) have progressed to include male infertility. Surgical procedures allow collection of spermatozoa from all along the human excurrent ducts, and the former have been used with some success in reproductive medicine. This has raised questions over the role of the epididymis in human sperm physiology. OBJECTIVE AND RATIONALE: To reanalyze what we now know about epididymal physiology in humans and to assess the relevance of laboratory animal models for understanding human physiology and the pathophysiology of the epididymis. SEARCH METHODS: A systematic bibliographic search of PubMed for articles published in English before May 2015 was carried out using the search terms 'epididymis' and 'sperm maturation'. Literature on the consequences of vasectomy on the epididymis was also searched. OUTCOMES: Whereas the proximal epididymis is almost exclusively occupied by efferent ducts, the sperm reservoir capacity is poorly developed in humans. At the molecular level, the human transcriptome and proteome show some segment specificity; conflicting results persist with regard to secretome variation along the tubule. The number of genes regulated along the excurrent ducts in men is lower when compared to rodent species, but remains significant. It is challenging to reconcile biochemical and physiological studies with clinical data obtained from men undergoing reanastomosis of the vas deferens at different points along the excurrent duct. We propose that vasectomy/vasovasostomy is a model to understand the consequences of obstruction on epididymis function in humans. WIDER IMPLICATIONS: Despite the scarcity of biological material available, the interspecies variability of the male reproductive tract urges us to use modern molecular and cellular biology tools to better understand human epididymis physiology in order to apply ART in a more responsible manner.

Stereoselectivity of butylidenephthalide on non-adrenergic prejunctional voltage-dependent Ca(2+) channels in prostatic portion of rat vas deferens.

The naturally occurring and synthetic butylinenephthalide (Bdph) has two geometric isomers. Z- and E-Bdph were reported to have geometric stereoselectivity for voltage-dependent calcium channels (VDCCs) in guinea-pig ileum. The aim of this study was to investigate whether the binding of Z- and E-Bdph on prejunctional VDCCs of rat vas deferens (RVD) is stereoselective. The twitch responses to electrical field stimulation (EFS, supramaximal voltage, 1 ms, 0.2Hz) were recorded on a polygraph. Z- and E-Bdph concentration-dependently inhibited the twitch responses to EFS in full tissue, prostatic portion and epididymal portion of RVD. The pIC50 value of Z-Bdph was greater than that of E-Bdph in the electrically stimulated prostatic portion of RVD, suggesting that the binding of Bdph on the non-adrenergic prejunctional VDCCs of cell membrane is stereoselective. In the prostatic portion, exogenous Ca(2+) only partially reversed the twitch inhibition by Z-Bdph, but effectively reversed those by Ca(2+) channel blockers, such as verapamil, diltiazem and aspaminol, suggesting that the action mechanisms may be different from those of Ca(2+) channel blockers. K(+) channel blockers, such as tetraethylammonium (TEA) and 4-aminopyridine (4-AP), may prolong duration of action potential to allow greater Ca(2+) entry and induced more release of transmitters. Therefore both blockers via their prejunctional actions reversed the twitch inhibition induced by Z-Bdph in all preparations of RVD by a non-specific antagonism.

Mirabegron relaxes urethral smooth muscle by a dual mechanism involving ß3 -adrenoceptor activation and α1 -adrenoceptor blockade.

LINKED ARTICLE: This article is commented on by Michel, M. C., pp. 429-430 of this issue. To view this commentary visit http://dx.doi.org/10.1111/bph.13379. BACKGROUND AND PURPOSE: Mirabegron is the first ß3 -adrenoceptor agonist approved for treatment of overactive bladder syndrome. This study aimed to investigate the effects of ß3 -adrenoceptor agonist mirabegron in mouse urethra. The possibility that mirabegron also exerts α1 -adrenoceptor antagonism was also tested in rat smooth muscle preparations presenting α1A - (vas deferens and prostate), α1D - (aorta) and α1B -adrenoceptors (spleen). EXPERIMENTAL APPROACH: Functional assays were carried out in mouse and rat isolated tissues. Competition assays for the specific binding of [(3) H]prazosin to membrane preparations of HEK-293 cells expressing each of the human α1 -adrenoceptors, as well as ß-adrenoceptor mRNA expression and cyclic AMP measurements in mouse urethra, were performed. KEY RESULTS: Mirabegron produced concentration-dependent urethral relaxations that were shifted to the right by the selective ß3 -adrenoceptor antagonist L-748,337 but unaffected by ß1 - and ß2 -adrenoceptor antagonists (atenolol and ICI-118,551 respectively). Mirabegron-induced relaxations were enhanced by the PDE4 inhibitor rolipram, and the agonist stimulated cAMP synthesis. Mirabegron also produced rightward shifts in urethral contractions induced by the α1 -adrenoceptor agonist phenylephrine. Schild regression analysis revealed that mirabegron behaves as a competitive antagonist of α1 -adrenoceptors in urethra, vas deferens and prostate (α1A -adrenoceptor, pA2  â‰… 5.6) and aorta (α1D -adrenoceptor, pA2  â‰… 5.4) but not in spleen (α1B -adrenoceptor). The affinities estimated for mirabegron in functional assays were consistent with those estimated in radioligand binding with human recombinant α1A - and α1D -adrenoceptors (pKi  â‰… 6.0). CONCLUSION AND IMPLICATIONS: The effects of mirabegron in urethral smooth muscle are the result of ß3 -adrenoceptor agonism together with α1A and α1D -adrenoceptor antagonism.

[Phosphodiesterase type 5 inhibitors for premature ejaculation: advances in studies].

Premature ejaculation (PE) is a common male sexual disorder with an incidence rate of 20-30%. Recent clinical trials have demonstrated that phosphodiesterase type 5 inhibitors (PDE5i), as the first-line drug for erectile dysfunction (ED), can improve ejaculatory function probably by acting on the peripheral and central adrenergic nerves. The possible action mechanisms of PDE5i may involve lessening of the central sympathetic output, modulation of the contractile responses from the vas deferens, seminal vesicles, prostate and urethra, induction of peripheral analgesia, and prolonging of the total erectile duration, increasing the confidence of ejaculation control, and reducing the post-ejaculation refractory time. This review discusses the possible mechanisms and clinical application of PDE5i in the treatment of PE.

Vas deferens neuro-effector junction: from kymographic tracings to structural biology principles.

The vas deferens is a simple bioassay widely used to study the physiology of sympathetic neurotransmission and the pharmacodynamics of adrenergic drugs. The role of ATP as a sympathetic co-transmitter has gained increasing attention and furthered our understanding of its role in sympathetic reflexes. In addition, new information has emerged on the mechanisms underlying the storage and release of ATP. Both noradrenaline and ATP concur to elicit the tissue smooth muscle contractions following sympathetic reflexes or electrical field stimulation of the sympathetic nerve terminals. ATP and adenosine (its metabolic byproduct) are powerful presynaptic regulators of co-transmitter actions. In addition, neuropeptide Y, the third member of the sympathetic triad, is an endogenous modulator. The peptide plus ATP and/or adenosine play a significant role as sympathetic modulators of transmitter's release. This review focuses on the physiological principles that govern sympathetic co-transmitter activity, with special interest in defining the motor role of ATP. In addition, we intended to review the recent structural biology findings related to the topology of the P2X1R based on the crystallized P2X4 receptor from Danio rerio, or the crystallized adenosine A2A receptor as a member of the G protein coupled family of receptors as prototype neuro modulators. This review also covers structural elements of ectonucleotidases, since some members are found in the vas deferens neuro-effector junction. The allosteric principles that apply to purinoceptors are also reviewed highlighting concepts derived from receptor theory at the light of the current available structural elements. Finally, we discuss clinical applications of these concepts.

Influence of acute treatment with sibutramine on the sympathetic neurotransmission of the young rat vas deferens.

The effects of acute treatment with sibutramine on the peripheral sympathetic neurotransmission in vas deferens of young rats were still not evaluated. Therefore, we carried out this study in order to verify the effects of acute sibutramine treatment on the neuronal- and exogenous agonist-induced contractions of the young rat vas deferens. Young 45-day-old male Wistar rats were pretreated with sibutramine 6 mg/kg and after 4h the vas deferens was used for experiment. The acute treatment with sibutramine was able to increase the potency (pD2) of noradrenaline and phenylephrine. Moreover, the efficacy (Emax) of noradrenaline was increased while the efficacy of serotonin and nicotine were decreased. The maximum effect induced by a single concentration of tyramine was diminished in the vas deferens from treated group. Moreover, the leftward shift of the noradrenaline curves promoted by uptake blockers (cocaine and corticosterone) and ß-adrenoceptor antagonist (propranolol) was reduced in the vas deferens of treated group. The initial phasic and secondary tonic components of the neuronal-evoked contractions of vas deferens from treated group at the frequencies of 2 Hz were decreased. Moreover, only the initial phasic component at 5 Hz was diminished by the acute treatment with sibutramine. In conclusion, we showed that the acute treatment with sibutramine in young rats was able to affect the peripheral sympathetic nervous system by inhibition of noradrenaline uptake and reduction of the neuronal content of this neurotransmitter, leading to an enhancement of vas deferens sensitivity to noradrenaline.

Cardiovascular stimulant actions of bupropion in comparison to cocaine in the rat.

Stimulants are banned in competition by the World Anti-Doping Agency, except for a small number of therapeutic agents subject to monitoring, including bupropion. We have examined the potency of bupropion in comparison with two agents banned in competition, adrafinil and modafinil, and with cocaine and desipramine as blockers of the noradrenaline re-uptake transporter in peripheral tissues of the rat. For studies in vivo, the pressor response to noradrenaline in the anaesthetized rat was studied. Cocaine, desipramine and bupropion at doses of 0.1, 0.3 and 1mg/kg, respectively, significantly increased the pressor response to noradrenaline. Overall, cocaine and desipramine were approximately 2-5 times more potent than bupropion in vivo in the rat. Adrafinil and modafinil (both 3mg/kg) did not significantly affect the pressor response. Bupropion was chosen for further study. In 1Hz paced rat right ventricular strips, bupropion (30µM) significantly increased the potency of noradrenaline at increasing the force of contraction. In rat vas deferens, bupropion and cocaine produced concentration-dependent increases in the contractile response to nerve stimulation, and cocaine was 11 times more potent than bupropion. Since bupropion is used clinically in doses of up to 300mg, it is likely that bupropion has actions at the noradrenaline transporter, and thus cardiovascular stimulant actions, in clinical doses. This may explain findings of increased exercise performance with bupropion.