Rat Vas Deferens Research Articles

Two isoforms of Trpm8 in rat vas deferens

Trpm8 is a nonselective Ca 2+ -permeable ion channel activated by temperatures below 28 °C and cooling chemical compounds such as menthol and icilin. Trpm8 is expressed in sensory neurons where it functions for temperature detection. However, Trpm8 is also expressed in various internal organs where temperature is stably higher than 36 °C, that is much higher than cool temperature needed for Trpm8 activation. This opens possible makes roles for Trpm8, such as nociception. Trpm8 mRNA is expressed in vas deferens (VD), a smooth muscle organ of male reproductive system. However, no Trpm8-mediated currents were previously registered in the myocytes. VD is located outside of testes, and it consists of smooth muscle tube covered by the epithelium. It actively contracts and transfers sperm from testes to ejaculatory ducts prior to ejaculation. Temperature in VD is stably high, thus, Trpm8 role there could be outside of cold detection. The objective of this study was to analyze Trpm8 mRNA and protein expression in rat VD, as well as splice variant analysis. Trpm8 mRNA expression in VD was confirmed with RT-PCR, and Trpm8 protein was detected by the Western-blot analysis. Additionally, we found that both mRNA and protein of shorter non-classical isoform, as well as canonical isoform of Trpm8 in VD. We isolated smooth muscle cells from VD and performed a multi-cell PCR. This technique makes possible non-myocytic mRNA detection that could be isolated from e.g. sensory neurons termini where Trpm8 is expressed at much higher levels. Interestingly, in the isolated smooth muscle cells, no canonical Trpm8 transcript was found, though the non-classical isoform was present. We propose the shorter isoform could be formed as a result of alternative splicing. This would account for a difference in Trpm8 function in VD, i.e. no Trpm8-mediated currents registered in the myocytes. A shorter isoform could have a truncated N-terminal domain, that is consistent with known human Trpm8 isoforms sM8-6 and sM8-18. Keywords : Trpm8, cold receptor, splice variant, vas deferens, myocytes, gene expression

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.

Immunolocalization of laminin during postnatal development of the testis, epididymis and vas deferens of albino rat.

Background: Disorders of testicular function may have their origins in fetal or early life as a result of abnormal development. Laminin-1 is emerging as the key molecule in early embryonic basement membrane assembly. Accumulating evidence supported the idea that extracellular matrix (ECM) molecules and mesenchymal cells might influence Sertoli and spermatogenic cell functions. Aim of the work: detecting the changes in the distribution and prevalence of laminin-1 assembly during postnatal development of the testis, epididymis and vas deferens in albino rats. Materials and methods: Thirty male albino rats were used and divided into six groups (n= 5 each) according to the age (postnatal day). These were one day, 1 week, 2 weeks, 3 weeks, 4 weeks, and 8 weeks postnatal. Specimens were fixed and processed, sectioned and stained by hematoxylin and eosin and immunohistochemical stain for laminin-1. The area percent of positive laminin immunostaining was measured and results were statistically analyzed. Results: at day one postnatal, thetestis was formed of solid un-canalized cords of seminiferous tubules with abundant laminin expression in the cells of the cords. With advancement of development the cords were luminized and the laminin expression declined to involve the basement membrane and the apical portions of the Sertoli cells at the 8th week postnatal. The epididymis at postnatal day one had a small diameter and narrow lumen and laminin expression involved the cytoplasm of the epithelial lining. As development proceeded the expression became confined to the apical portion, the site of stereocilia together with its presence in the basement membranes. The same pattern of changes in laminin expression together with morphological appearance was detected in the vas deferens. Conclusion: The present study was able to demonstrate a change in the distribution as well as the prevalence of laminin-1 immunoreactivity within the testis, epididymis and vas. During the period of postnatal development starting at postnatal day one up to 8 weeks postnatal. This would reflect an essential role for laminin in early postnatal period of development.

Immunolocalization of laminin during postnatal development of the testis, epididymis and vas deferens of albino rat.

Immunolocalization of laminin during postnatal development of the testis, epididymis and vas deferens of albino rat.

Testosterone enhances expression and functional activity of epithelial sodium channel (ENaC), cystic fibrosis transmembrane regulator (CFTR) and sodium hydrogen exchanger (NHE) in vas deferens of sex-steroid deficient male rats

Effects of testosterone on expression and functional activity of ENaC, CFTR and NHE in vas deferens were investigated. MethodsOrchidectomized, adult male rats were given 125 and 250 μg/kg/day testosterone subcutaneously, with or without flutamide and finasteride for seven consecutive days. At the end of the treatment, rats were anesthetized and vas deferens were perfused. Changes in vas deferens fluid secretion rate, pH, HCO3−, Cl− and Na+ concentrations were recorded in the presence of amiloride and Cftr inh-172. Rats were then sacrificed and vas deferens were harvested and subjected for molecular biological analysis. ResultsTestosterone treatment caused the fluid pH and HCO3− concentrations to decrease but secretion rate, Cl− and Na+ concentrations to increase, where upon amiloride administration, the pH and HCO3− concentration increased but Cl− and Na+ concentrations further increased. In testosterone-treated rats, administration of Cftr inh-172 caused all fluid parameters to decrease. In testosterone-treated rats co-administered with flutamide or finasteride, pH and HCO3− concentration increased but fluid secretion rate, Cl− and Na+ concentrations decreased and these parameters were not affected by amiloride or Cftr inh-172 administration. Under testosterone influence, CFTR and γ-ENaC were highly expressed at the apical membrane while NHE-1 and 4 were highly expressed at the basolateral membrane of vas deferens epithelium. Meanwhile, NHE-2 and 3 were highly expressed at the apical membrane. ConclusionsDifferential expression of ENaC, CFTR and NHE in vas deferens under testosterone influence indicated the important role of these transporters in creating optimal fluid microenvironment that is essential for preserving male fertility.

Testosterone up-regulates vacuolar ATPase expression and functional activities in vas deferens of orchidectomized rats

Precise regulation of vas deferens fluid pH is essential for sperm. However, the mechanisms underlying effect of testosterone on vas deferens fluid pH have never been identified, which could involve changes in expression and functional activity of vacoular (V)-ATPase. MethodsOrchidectomized, adult male Sprague-Dawley rats were treated subcutaneously with 125 μg/kg/day and 250 μg/kg/day testosterone with or without flutamide (androgen receptor blocker) and finasteride (5α-reductase inhibitor) for seven (7) days. Following treatment completion, in vivo perfusion of vas deferens lumen was performed and changes in fluid secretion rate, pH and HCO3− content were measured with and without bafilomycin, a V-ATPase inhibitor. Rats were then sacrificed and vas deferens were harvested and subjected for V-ATPase A1 and B1/2 protein expression and distribution analysis by western blotting and immunohistochemistry, respectively. ResultsIn sham-operated and testosterone-treated orchidectomized rats, higher fluid secretion rate, which was not antagonized by bafilomycin but lower HCO3− content and pH which were antagonized by bafilomycin were observed when compared to orchidectomized-only and orchidectomized, testosterone-treated rats receiving flutamide or finasteride, respectively. Bafilomycin had no effect on fluid secretion rate, HCO3− content and pH in orchidectomized and testosterone-treated orchidectomized rats receiving flutamide and finasteride. V-ATPase A1 and B1/2 proteins were expressed at high levels in vas deferens and were highly distributed at the apical membrane of luminal epithelium and in muscle layer of this organ, mainly in sham and testosterone-treated orchidectomized rats. ConclusionsV-ATPase is involved in acidification of vas deferens fluid under testosterone influence.

14-O-Methylmorphine: A Novel Selective Mu-Opioid Receptor Agonist with High Efficacy and Affinity

14-O-methyl (14-O-Me) group in morphine-6-O-sulfate (M6SU) or oxymorphone has been reported to be essential for enhanced affinity, potency and antinociceptive effect of these opioids. Herein we report on the pharmacological properties (potency, affinity and efficacy) of the new compound, 14-O-methylmorphine (14-O-MeM) in in vitro. Additionally, we also investigated the antinociceptive effect of the novel compound, as well as its inhibitory action on gastrointestinal transit in in vivo. The potency and efficacy of test compound were measured by [35S]GTPγS binding, isolated mouse vas deferens (MVD) and rat vas deferens (RVD) assays. The affinity of 14-O-MeM for opioid receptors was assessed by radioligand binding and MVD assays. The antinociceptive and gastrointestinal effects of the novel compound were evaluated in the rat tail-flick test and charcoal meal test, respectively. Morphine, DAMGO, Ile5,6 deltorphin II, deltorphin II and U-69593 were used as reference compounds.14-O-MeM showed higher efficacy (Emax) and potency (EC50) than morphine in MVD, RVD or [35S]GTPγS binding. In addition, 14-O-MeM compared to morphine showed higher affinity for μ-opioid receptor (MOR). In vivo, in rat tail-flick test 14-O-MeM proved to be stronger antinociceptive agent than morphine after peripheral or central administration. Additionally, both compounds inhibited the gastrointestinal peristalsis. However, when the antinociceptive and antitransit doses for each test compound are compared, 14-O-MeM proved to have slightly more favorable pharmacological profile.Our results affirm that 14-O-MeM, an opioid of high efficacy and affinity for MOR can be considered as a novel analgesic agent of potential clinical value.

The ethyl acetate fraction of a methanolic extract of unripe noni (Morinda citrifolia Linn.) fruit exhibits a biphasic effect on the dopaminergic system in mice.

In earlier ex vivo studies, we reported the biphasic effect of a methanolic extract of unripe Morinda citrifolia fruit (MMC) on dopamine-induced contractility in isolated rat vas deferens preparations. The present in vivo study was designed and undertaken to further explore our earlier ex vivo findings. This study examined the effect of the ethyl acetate fraction of a methanolic extract of unripe Morinda citrifolia Linn. fruit (EA-MMC; 5–100 mg/kg, p.o.) on the dopaminergic system using mouse models of apomorphine-induced climbing time and climbing behavior, methamphetamine-induced stereotypy (sniffing, biting, gnawing, and licking) and haloperidol-induced catalepsy using the bar test. Acute treatment with EA-MMC at a low dose (25 mg/kg, p.o.) significantly attenuated the apomorphine-induced climbing time and climbing behavior in mice. Similarly, EA-MMC (5 and 10 mg/kg, p.o.) significantly inhibited methamphetamine-induced stereotyped behavior in mice. These results demonstrated that the antidopaminergic effect of EA-MMC was observed at relatively lower doses (<25 mg/kg, p.o.). On the other hand, EA-MMC showed dopaminergic agonistic activity at a high dose (3,000 mg/kg, p.o.), which was evident from alleviation of haloperidol (a dopamine D2 blocker)-induced catalepsy in mice. Therefore, it is concluded that EA-MMC might possess a biphasic effect on the dopaminergic system, i.e., an antagonistic effect at lower doses (<25 mg/kg, p.o.) and an agonistic effect at higher doses (>1,000 mg/kg, p.o.). However, further receptor-ligand binding assays are necessary to confirm the biphasic effects of M. citrifolia fruit on the dopaminergic system.

Clonidine Inhibits Phenylephrine-Induced Contraction of Rat Thoracic Aortae by Competitive Antagonism of α&amp;lt;sub&amp;gt;1&amp;lt;/sub&amp;gt;-Adrenoceptors Independent of α&amp;lt;sub&amp;gt;2&amp;lt;/sub&amp;gt;-Adrenoceptor Stimulation

Clonidine is a classically categorized α2-adrenoceptor (α2-AR) agonist that produces vascular contractions by stimulating arterial smooth muscle α2-ARs. However, clonidine inhibits α1-AR-mediated arterial contractions. Recently, it was suggested that repeated stimulation with clonidine induces desensitization of α2-ARs, thus inhibiting noradrenaline-induced smooth muscle contractions. In the present study, we examined whether clonidine-mediated inhibition of α1-AR contractions involves interactions with α2-ARs in rat thoracic aortae. 1) Clonidine and guanfacine inhibited electrical field stimulation-induced contractions in a concentration-dependent, yohimbine-sensitive manner in isolated rat vas deferens preparations. 2) Clonidine almost completely suppressed phenylephrine-induced sustained contractions of rat thoracic aortae. 3) Clonidine competitively inhibited phenylephrine-induced contractions with a pA2 value of 6.77 at concentrations between 10-7 and 10-6 M. At 10-5 M, clonidine inhibited phenylephrine-induced contractions and dramatically reduced maximum contractions. 4) In contrast, clonidine did not inhibit contractions produced by high KCl or prostaglandin F2α. 5) Inhibition of phenylephrine-induced sustained contractions by clonidine was also produced in the presence of yohimbine. However, guanfacine did not inhibit phenylephrine-induced sustained contractions. These findings suggest that clonidine inhibits phenylephrine-induced contraction of rat thoracic aortae by competitive antagonism of α1-ARs, which is mediated through a mechanism independent of α2-AR stimulation.

Mitragyna speciosa Leaf Extract Exhibits Antipsychotic-Like Effect with the Potential to Alleviate Positive and Negative Symptoms of Psychosis in Mice.

In this study, we investigated the antipsychotic-like effect of methanolic extract of Mitragyna speciosa leaf (MMS) using in vivo and ex vivo studies. In vivo studies comprised of apomorphine-induced climbing behavior, haloperidol-induced catalepsy, and ketamine-induced social withdrawal tests in mice whereas the ex vivo study was conducted utilizing isolated rat vas deferens preparation. Acute oral administration of MMS (50–500 mg/kg) showed an inverted bell-shaped dose-response in apomorphine-induced cage climbing behavior in mice. The effective inhibitory doses of MMS (75 and 100 mg/kg, p.o.) obtained from the apomorphine study was further tested on haloperidol (subcataleptic dose; 0.1 mg/kg, i.p.)-induced catalepsy in the mouse bar test. MMS (75 and 100 mg/kg, p.o.) significantly potentiated the haloperidol-induced catalepsy in mice. Interestingly, MMS at the same effective doses (75 and 100 mg/kg, p.o.) significantly facilitated the social interaction in ketamine-induced social withdrawal mice. Furthermore, MMS inhibited the dopamine-induced contractile response dose-dependently in the isolated rat vas deferens preparations. In conclusion, this investigation provides first evidence that MMS exhibits antipsychotic-like activity with potential to alleviate positive as well as negative symptoms of psychosis in mice. This study also suggests the antidopaminergic activity of MMS that could be responsible for alleviating positive symptoms of psychosis.

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.

Synthesis and structure–activity relationships of novel arylpiperazines as potent antagonists of α1-adrenoceptor

Arylpiperazines 2–11 were synthesized, and their biological profiles at α1-adrenergic receptors (α1-ARs) assessed by binding assays in CHO cells expressing human cloned subtypes and by functional experiments in isolated rat vas deferens (α1A), spleen (α1B), and aorta (α1D). Modifications at the 1,3-benzodioxole and phenyl phamacophoric units resulted in the identification of a number of potent compounds (moderately selective with respect to the α1b-AR), in binding experiments. Notably, compound 7 (LDT451) showed a subnanomolar pKi of 9.41 towards α1a-AR. An encouragingly lower α1B-potency was a general trend for all the series of compounds, which showed α1A/D over α1B selectivity in functional assays. If adequately optimized, such peculiar selectivity could have relevance for a potential LUTS/BPH therapeutic application.

Stereoselectivity of butylidenephthalide on non-adrenergic prejunctional voltage-dependent Ca2+ 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 Ca2+ only partially reversed the twitch inhibition by Z-Bdph, but effectively reversed those by Ca2+ channel blockers, such as verapamil, diltiazem and aspaminol, suggesting that the action mechanisms may be different from those of Ca2+ channel blockers. K+ channel blockers, such as tetraethylammonium (TEA) and 4-aminopyridine (4-AP), may prolong duration of action potential to allow greater Ca2+ 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.

Modulation of Smooth Muscle Relaxation by Short and Long Carbon Based Chemicals

Introduction: Short and long-chained based carbon chemicals derived from plants are found in many household products and are widely used in complementary medicine. It has been suggested anecdotally that some of these chemicals have the ability to relax smooth muscle tissues, and to test this concept, this study examines the effects of 1,8-cineole, 1-hepta-nol, α-pinene, cis-3-hexen-1-ol and trans-2-hexenal on neurotransmission and contraction of mouse and rat vas deferens. Methods: Focal extracel-lular recordings from the surface of mouse vas deferens of electrically-evoked nerve terminal impulses (NTIs) and electrically-evoked excitatory junctional currents (EJCs) were examined in both the presence and the absence of each chemical. Likewise, noradrenaline-evoked contractile forces of rat vas deferens were examined in both the presence and the absence of each chemical. Results: Relative to the respective controls, 1,8-cineole, 1-heptanol and trans-2-hexenal each statistically significantly decreased both smooth muscle contractile forces and EJC amplitudes, with 1-heptanol and trans-2-hexenal also statistically significantly decreasing NTI amplitudes. On NTI and EJC amplitudes the effects of 1,8-cineole and 1-heptanol were reversible whereas the effects of trans-2-hexenal were irreversible. Conclusion: Our results demonstrate that 1,8-cineole and trans-2-hexenal decrease smooth muscle neurotransmission and contraction and thereby cause smooth muscle relaxation, thus suggesting that these chemicals may have clinical applications and health benefits.

Slowing Sinus Tachycardia in Heart Transplant Recipients: Is It Time?

We report the synthesis and the biological activity of new analogues of Ac-RFMWMK-NH2 and Ac-RYYRWK-NH2, modified in position 4 and 5, respectively, with incorporation of newly synthesized β2-tryptophan analogues. Trp was substituted by the (S)-2-(1-methyl-1H-indol-3-yl)propionic residue or by (S)-2-(5-methoxy-1H-indol-3-yl)propionic residue. The biological activity (pEC50 and Emax) of these compounds was tested on electrically stimulated preparations of rat vas deferens. The 5-methoxy β-tryptophan group reverses the affinity of the compounds.

Rats undernourished in utero have altered Ca2+ signaling and reduced fertility in adulthood.

Epidemiological and animal studies have shown that placental undernutrition impairs reproduction in adult offspring, but the underlying molecular mechanisms within the male genital tract remain unknown. Due to its special physiological characteristics in transport and the modulation of the environment to which its luminal content is exposed, we hypothesized that the vas deferens would be a highly sensitive target. The goals were to investigate whether intrauterine malnutrition affects molecular mechanisms related to Ca2+- and oxidative stress-modulated processes and causes structural alterations in the adult rat vas deferens that could attenuate fecundity and fertility. Male adult rats malnourished in utero had increased vas deferens weight associated with thickening of the muscular coat, a decrease in the total and haploid germ cells, a marked increase in the immature cells, and a decline in the numbers of pregnant females and total offspring per male rat. The ex vivo response of vas deferens from malnourished rats demonstrated an accentuated decrease in the contractile response to phenylephrine. The vas deferens had a marked decrease in Ca2+ transport due to the uncoupling of Ca2+-stimulated ATP hydrolysis and ATP-driven Ca2+ flux, and the downregulation of both sarco-endoplasmic reticulum Ca2+-ATPase 2 and the coupling factor 12-kDa FK506-binding protein. An increase in protein carbonylation (a marker of oxidative damage) and an imbalance between protein kinases C and A were observed as a legacy of undernutrition in early life. These results provide the structural and molecular basis to explain at least in part how maternal undernutrition affects fecundity and fertility in adult male rats.

Effects of long-term fluoxetine treatment on adrenergic plasticity in rat vas deferens

Chronic antidepressant administration increases neurotrophin levels in the central and peripheral nervous system, leading to an increase of neuronal sprouting, reestablishment of neural networks and neurotransmitter levels. Injured peripheral nerves regenerate at very slow rates. However, the recovery of the hypogastric nerve in rodents after injury is significantly improved with neurotrophin administration. Accordingly, our goal was to determine whether treatment with the antidepressant fluoxetine affects catecholamine levels and neuronal function, after surgical denervation of the rat vas deferens. Noradrenaline levels in the denervated vas deferens were higher in fluoxetine-treated animals than in the vehicle-treated group, as measured by high performance liquid chromatography. In functional studies of smooth muscle contraction, the responses induced by phenylephrine or ATP, as well as pre-synaptic α2-adrenoceptor reactivity, were not modified by chronic treatment with the antidepressant. However, the contraction mediated by neuronal release of noradrenaline induced by tyramine was increased on days 7 and 21 after denervation in rats treated with fluoxetine. These data indicate that fluoxetine can improve functional recovery after rat vas deferens denervation.

Effects of desipramine on prazosin potency at α1A- and α1D-adrenoceptors in rat vas deferens: Implications for the α1L-adrenoceptor subclassification

This study investigates the interaction between cocaine, desipramine and prazosin at α1-adrenoceptor subtypes mediating contractions to noradrenaline in epididymal portions of rat vas deferens. Noradrenaline potency was not significantly affected by desipramine (0.1–1.0μM) and reduced by desipramine (10μM), but was increased by the presence of cocaine (3.0–30μM), particularly in terms of phasic contractions to low concentrations of noradrenaline. In vehicle experiments, prazosin exhibited relatively low potency as an antagonist against the predominantly α1A-adrenoceptor mediated response (pKB 8.50). In the presence of cocaine, prazosin exhibited higher potency against the revealed α1D-adrenoceptor mediated component (e.g. pKB 9.12). In the presence of desipramine, the potency of prazosin was either unchanged or indeed decreased. Cocaine (0.3–30μM) significantly increased the single pulse nerve-stimulation-evoked contraction, with a maximum increase to 156±12% of control (n=9). In contrast, desipramine in low concentrations (0.1–0.3μM) produced a small but significant increase to 126.6±5.5% (n=11), but higher concentrations failed to increase the response. In conclusion, desipramine fails to produce sufficient noradrenaline transporter block in low concentrations (0.1μM) and produces α1-adrenoceptor antagonism in slightly higher concentrations (0.3–1μM), and so is unsuitable for use in α1-adrenoceptor subclassification studies. Contractions of rat vas deferens are mediated by α1A- and α1D-adrenoceptors, and prazosin has selectivity for α1D- over α1A-adrenoceptors. The α1L-adrenoceptor previously identified in rat vas deferens is the native α1A-adrenoceptor. The range of prazosin potencies and receptor subtypes previously reported in rat vas deferens may be explained by the choice of cocaine or desipramine as noradrenaline transporter blocker.

Biofunctional studies of new 2-methoxyphenylpiperazine xanthone derivatives with α1-adrenolytic properties

BackgroundThe aim of this study was to assess the selectivity of the studied xanthone derivatives for α1-adrenoceptor subtypes (α1A, α1B, α1D, α1L) in functional experiments in order to verify if they possess any selectivity for a distinct subtype of α1-adrenoceptor. Moreover, several pharmacological tests were carried out to assess whether they reveal other than α1-adrenoceptor blocking properties such as: antagonistic for 5-HT2 receptors, vasorelaxant or spasmolytic. MethodsThe influence on α1A-adrenoceptors was examined in biofunctional studies employing isolated rat vas deferens, on α1B-adrenoceptors in guinea-pig spleen, on α1D-adrenoceptors in rat aorta, and on α1L-adrenoceptors in rabbit spleen. Affinity for 5-HT2 receptors was measured in radioligand binding assay, whereas antagonistic potency for 5-HT2 receptors was studied on isolated rat aorta. Vasorelaxant effect of tested compounds was assessed in functional study employing rat aorta, whereas direct spasmolytic activity was investigated using the isolated rabbit small intestine. ResultsThe present study provides evidences that the tested 2-methoxyphenylpiperazine xanthone derivatives are non-selective α1-adrenoceptor blockers. However, at higher concentrations the direct spasmolytic effect could enhance their hypotensive activity. The obtained results indicate that the studied xanthones possessed weak calcium entry blocking activity, as well as antagonistic properties for 5-HT2A receptors. ConclusionsThe results of the present study support the idea that the hypotensive activity of the studied compounds is related to their α1-adrenolytic properties.

Ivermectin as an inhibitor of the plasma membrane and the sarcoplasmic/endoplasmic reticulum Ca2+-ATPases in rat vas deferens

ObjectiveThe present work investigated the effect of ivermectin on Ca2+ content and on the Ca2+-ATPase activity (represented by the plasma membrane Ca2+-ATPase and the sarcoplasmic/endoplasmic reticulum Ca2+-ATPase present in rat vas deferens.MethodsThe assays were carried out using ultracentrifuged homogenate preparations from rat vas deferens in the presence or absence of the 12-kDa FK506-binding protein-Ca2+ release channel complex. Measures of Ca2+ content and Ca2+ ATPase activity were then carried out in function of different concentrations of ivermectin. ResultsThe data show that ivermectin (10 μM) reduces the sarcoplasmic reticulum Ca2+ content in FK506-binding protein (+) and FK506-binding protein (-) fractions of ultracentrifuged homogenate from rat vas deferens (inhibition of 50% and 40%, respectively, p&lt;0.05) and inhibits both the activities of sarcoplasmic/endoplasmic reticulum Ca2+-ATPase and plasma membrane Ca2+-ATPases pumps (33% and 16%, respectively, p&lt;0.05).ConclusionThese data suggest that ivermectin effects Ca2+ handling in the rat vas deferens, indicating that this drug could alter the contractility of this smooth muscle. Therefore, ivermectin could be an interesting pharmacological tool to alter the physiological function of vas deferens and to manipulate the fertility status of male rats.Indexing terms: Calcium. Ivermectin. Rats.