Lymphocytes sensitize rat isolated atria to the inotropic and chronotropic effects of sodium arachidonate

Normal human lymphocytes (4 X 10(5) ml-1) incubated with sodium arachidonate (8 X 10(-7)M) (NaA-L) induced a strong enhancement of the tension and frequency of spontaneously beating rat atria. Normal human lymphocytes (L) or NaA alone at 8 X 10(-7)M did not modify this contractile activity. Between 2 X 10(-6)M to 1 X 10(-5)M NaA alone increased the tension of the atria without effect on the rate. In the presence of L (4 X 10(5) ml-1) the dose-response curve to NaA shifted to the left, the potency and the efficiency of NaA were enhanced and the chronotropic action was triggered. Inhibitors of cyclo-oxygenase (indomethacin 1 X 10(-6)M or acetylsalicylic acid (ASA) 1.8 X 10(-4)M) completely blocked the positive inotropic effect induced by NaA alone. Inhibitors of lipoxygenase/s (nordihydroguaiaretic acid (NDGA) 1 X 10(-5)M or 5,8,11,14-eicosatetraynoic acid (ETYA) 1 X 10(-7)M did not modify this effect. Indomethacin and ASA did not block the positive inotropic and chronotropic effects of the lower concentration of NaA-L and significantly reduced the inotropic effect of the higher ones. NDGA and ETYA shifted to the right the inotropic and chronotropic dose-response curve to NaA-L. FPL-55712 (1 X 10(-7)M), the slow reacting substance of anaphylaxis (SRS-A) antagonist, significantly reduced the overall inotropic and chronotropic effect of NaA-L. Direct contact of NaA-L with the atria was not necessary. Cell-free supernatants of L exposed to NaA increased the tension and the frequency of beating rat atria.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of St-587 on the alpha-adrenoceptors in the bisected rat vas deferens

The effects of the alpha-adrenoceptor agonist St-587 have been studied on the twitch responses induced by field stimulation in the prostatic portion of rat vas deferens. Moreover the drug's influence on the unstimulated prostatic and epididymal halves of rat vas deferens has also been determined. Alone and after addition of yohimbine (0.3 microM) it enhanced in a concentration-dependent manner the twitch responses in the prostatic half. Prazosin competitively antagonized (pA2 = 8.41 +/- 0.03) this effect. The enhancing effect of St-587 was not reduced in reserpinized animals. These results suggest that post-synaptic alpha 1-adrenoceptors are involved in the potentiation of twitch responses induced by St-587. When alpha 1-adrenoceptors were blocked by prazosin (0.1 microM), St-587 partially inhibited the twitch responses of the prostatic portion of rat vas deferens (Emax = 49.5 +/- 3.5%). Yohimbine completely reversed the inhibitory effects of both St-587 and clonidine. Furthermore St-587 antagonized the inhibitory effects of clonidine on twitch responses. Thus it appears that St-587 also behaves as a partial agonist of presynaptic alpha 2-adrenoceptors in this portion of rat vas deferens, but it did not induce contractions in the unstimulated prostatic half of the vas deferens. However, it competitively antagonized the alpha 1-adrenoceptor agonist phenylephrine by acting as an antagonist of prostatic postsynaptic alpha 1 adrenoceptors. These alpha 1-adrenoceptors are probably different from those that mediate the twitch enhancing response to St-587 in that portion. On the other hand, St-587 was a partial agonist of alpha 1-adrenoceptors in the epididymal half.(ABSTRACT TRUNCATED AT 250 WORDS)     

CHANGES IN PREJUNCTIONAL POTENCY OF B-HT 933 DURING AGING IN THE RAT VAS DEFERENS

1. Age-related changes in prejunctional alpha 2-adrenoceptors were examined in the rat vas deferens using pharmacological techniques. 2. B-HT 933 (1 x 10(-8) - 1 x 10(-6) mol/L) caused a concentration-dependent inhibition of isometric contractions (tetrodotoxin-sensitive) induced by stimulation with single field-stimulus pulses, in both the epididymal and prostatic regions of rat vas deferens. The concentration-response curve to B-HT 933 was shifted to the right with age in the prostatic regions of the vas deferens. 3. In high concentrations (10(-6) - 3 x 10(-4) mol/L), B-HT 933 caused concentration-dependent enhancement of the contractile response to stimulation and evoked spontaneous contractile activity. No significant difference in this postjunctional activity occurred with age in either the prostatic or epididymal regions of the vas deferens. 4. Schild analysis revealed no significant differences in pA2 values for the antagonisms of the prejunctional inhibitory effect of B-HT 933 by rauwolscine in either the prostatic or epididymal regions of vas deferens between young and old rats. 5. These results could be interpreted as a decrease in alpha 2-adrenoceptor number with age. The more marked decrease in the prejunctional inhibitor potency of B-HT 933 in prostatic regions of vas deferens with aging may be due to a smaller receptor reserve in this region of the vas deferens.     

Activation of alpha 1-adrenoceptors increases [3H]inositol metabolism in rat vas deferens and caudal artery

Rings of rat vas deferens and caudal artery were incubated with [3H]inositol in Krebs-Ringer bicarbonate buffer containing 10 mM lithium chloride, and the production of water-soluble [3H]inositol phosphates was monitored. Norepinephrine increased [3H]inositol phosphate accumulation 7-fold in rings from vas deferens and 3-fold in rings from caudal artery. Epinephrine, phenylephrine and methoxamine were as effective as norepinephrine, suggesting that these drugs are full agonists in causing this response. Prazosin, phentolamine and yohimbine completely blocked the stimulation by norepinephrine in both tissues with potencies typical of blockade of alpha 1-adrenoceptors. Despite a substantial receptor reserve for alpha 1-adrenoceptor mediated contractile responses, clonidine, p-amino-clonidine, phenylpropanolamine and ephedrine can only cause a partial contractile response in rat vas deferens. However, all of these partial agonists were either as effective or more effective in increasing [3H]inositol phosphate accumulation in rat vas deferens as they were in activating a contractile response. These data suggest that alpha 1-adrenoceptors increase phosphatidylinositol turnover in rat vas deferens and caudal artery, and that there may be a receptor reserve for alpha 1-adrenoceptor mediated increases in [3H]inositol phosphate accumulation in these smooth muscles.     

Effects of alpha,beta-methylene ATP on biphasic responses of rat vas deferens

The epididymal region of isolated vas deferens of the rat was stimulated locally with field electrodes. Continuous perfusion with alpha,beta-methylene-adenosine-5'-triphosphate (mATP, 10(-5) M) desensitized the P2-purinoceptors and the effect of this on the two components of tetanic responses and the 'non-adrenergic' and adrenergic phases of single shock responses were examined. Exposure to mATP selectively prevented the contractions to adenosine-5'-triphosphate (ATP, 10(-4)-5 X 10(-4) M) but not noradrenaline (NA, 10(-5) M) and preferentially blocked the secondary but not the primary tetanic component. It also depressed markedly the non-adrenergic phase of single shocks and either reduced or abolished the adrenergic phase. Thus the secondary tension development depends in the rat vas deferens on both the activation of alpha 1-adrenoceptors and the non-adrenergic twitch mechanism. The action of mATP involves more processes than the desensitization of the P2-purinoceptors so does not positively identify purinergic transmission. Direct NA-induced contraction contributes more to the primary than to the secondary tetanic component.     

Characterization of alpha 1-adrenoceptor subtypes in tension response of human prostate to electrical field stimulation.

1. The effects of various alpha 1-adrenoceptor antagonists and nifedipine on tension responses of human prostate to electrical field stimulation were evaluated in this study. 2. Prazosin (3 x 10(-10) to 10(-8) M) and 5-methyl-urapidil (10(-9) to 3 x 10(-8) M) blocked concentration-dependently the tension responses to electrical field stimulation and completely abolished them in the maximal concentrations (10(-8) M and 3 x 10(-8) M, respectively); in contrast, chloroethylclonidine (CEC), in the maximal concentration of 100 microM, blocked these effects by only 50%. 3. The contractile responses of rat vas deferens and spleen to exogenously-applied alpha 1-adrenoceptor agonists were competitively inhibited by prazosin and 5-methyl-urapidil; in addition, the pA2 values were calculated and the relative potencies with reference to prazosin were obtained. The relative potency of 5-methyl-urapidil in human prostate (0.105) was close to that in rat vas deferens (0.257), which contains primarily putative alpha 1A-adrenoceptors. However, it was much more than that in rat spleen (0.011), which contains primarily putative alpha 1B-adrenoceptors. 4. Nifedipine (10(-8) to 10(-6) M) inhibited concentration-dependently the contractile responses to electrical field stimulation in human prostate; in addition, the inhibition percentages were similar to those to exogenously-applied noradrenaline in rat vas deferens. In contrast, CEC (10 microM), which almost flattened the concentration-response curve of the rat spleen to phenylephrine, only partially inhibited (by 33.1%) the nerve-mediated contraction of human prostate. 5. The involvement of prejunctional alpha 2-adrenoceptors situated on the sympathetic nerve terminals of human prostate was also examined.(ABSTRACT TRUNCATED AT 250 WORDS)     

Harman and 3-methylharman--blockaders of presynaptic alpha 2-adrenoreceptors

It has been shown in experiments on rat vas deferens that melipramine, harmane and 3-methylharman in low concentrations of the order of 10(-8)--10(-6) M increase the contraction of the vas deference induced by noradrenaline and transmural stimulation. Meanwhile in higher concentrations (10(-6)--4 X 10(-5) M) they have been discovered to decrease this effect (except for harmane). In low concentrations harmane and 3-methylharman competitively remove the inhibitory effect of clonidine on the transmural stimulation-induced contractions. It is inferred that harmane and methylharman are competitive blockers of presynaptic alpha 2-adrenoceptors in adrenergic synapses.     

Failure of AH11110A to functionally discriminate between alpha(1)-adrenoceptor subtypes A, B and D or between alpha(1)- and alpha(2)-adrenoceptors

The potency of the putatively alpha(1B)-adrenoceptor selective drug, 1-[biphenyl-2-yloxy]-4-imino-4-piperidin-1-yl-butan-2-ol (AH11110A), to antagonize contraction upon stimulation of alpha(1A)-adrenoceptors in rat vas deferens and rat perfused kidney, alpha(1B)-adrenoceptors in guinea-pig spleen, mouse spleen and rabbit aorta, and alpha(1D)-adrenoceptors in rat aorta and pulmonary artery was evaluated and compared to that of a number of subtype-discriminating antagonists. N-[3-[4-(2-Methoxyphenyl)-1-piperazinyl]propyl]-3-methyl-4-oxo-2-phenyl-4H-1-benzopyran-8-carboxamide (Rec 15/2739) and (+/-)-1,3,5-trimethyl-6-[[3-[4-((2,3-dihydro-2-hydroxymethyl)-1,4-benzodioxin-5-yl)-1-piperazinyl]propyl]amino]-2,4(1H,3H)-pyrimidinedione (B8805-033) were confirmed as selective for alpha(1A)-adrenoceptors, 8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspiro[4.5]decane-7,9-dione (BMY 7378), 8-[2-(1,4-benzodioxan-2-ylmethylamino)ethyl]-8-azaspiro[4.5]decane-7,9-dione (MDL 73005EF), and cystazosin were found to be selective for alpha(1D)-adrenoceptors, whereas spiperone was weakly selective for alpha(1B)-over alpha(1A)-adrenoceptors. However, from the functional affinity profile obtained for AH11110A at alpha(1A)-adrenoceptors (pA(2)=6.41 in rat vas deferens), alpha(1B)-adrenoceptors (pA(2)=5.40-6.54) and alpha(1D)-adrenoceptors (pA(2)=5.47-5.48), the affinity and presumed selectivity previously obtained for AH11110A in radioligand binding studies at native alpha(1B)- and cloned alpha(1b)-adrenoceptors (pK(i)=7.10-7.73) could not be confirmed. Additionally, AH11110A enhanced the general contractility of rat vas deferens, produced a bell-shaped dose-response curve of vasodilation in perfused rat kidney, and its antagonism in most other tissues was not simply competitive. The affinity of AH11110A for prejunctional alpha(2)-adrenoceptors in rabbit vas deferens (pA(2)=5.44) was not much lower than that displayed for alpha(1)-adrenoceptor subtypes, revealing that AH11110A, besides alpha(1)-adrenoceptors, also interacts with alpha(2)-adrenoceptors, and thus may be unsuitable for alpha-adrenoceptor subtype characterization, at least in smooth muscle containing functional studies.     

Investigation of neurotransmission in vas deferens from alpha(2A/D)-adrenoceptor knockout mice

1. We have investigated pre- and post-junctional responsiveness in vas deferens from wild-type and alpha(2A/D)-adrenoceptor knockout mice. The response to a single stimulus was not significantly different between wild-type and knock-out mice. The isometric contraction to 10 Hz stimulation for 4 s was significantly larger in vas deferens from knockout as compared with wild-type. 2. The maximum potentiation of 10 Hz stimulation-evoked contractions by yohimbine was to 206.2+/-38.0% of control in wild-type but to 135.8+/-13.6% of control in knockout. The alpha(2A/D)-adrenoceptor selective antagonist BRL 44408 significantly increased the 10 Hz stimulation-evoked contraction in wild-type but not knockout, and the reverse was true for the alpha(2C)-adrenoceptor selective antagonist spiroxatrine. The alpha(2B)-adrenoceptor antagonist imiloxan had no effect on the evoked contraction except at high concentrations, and only in wild-type. Following cocaine (3 microM) and BRL 44408 (1 microM), 10 Hz responses were similar in shape and maximum between wild-type and knock-out. 3. The alpha(2)-adrenoceptor agonist xylazine virtually abolished the early component of the contraction to 10 Hz stimulation in the presence of nifedipine (10 microM) in vas deferens from knockout mice in a way consistent with a change of receptor subtype but without clear evidence for a reduced receptor number. However, the late component of the contraction to 10 Hz stimulation was significantly potentiated by xylazine in tissues from knock-out mice. 4. It is concluded that, although non-alpha(2A/D)-adrenoceptors replace alpha(2D)-adrenoceptors in this knockout, the alpha(2)-adrenoceptor agonist and antagonist data are contradictory. The antagonist data suggest a major loss of prejunctional alpha(2)-adrenoceptors, but this is not necessarily supported by the agonist data.     

Acute and long-term administration of citalopram desensitizes alpha2-adrenoceptors in the rat vas deferens

The aim of this study was to investigate the effect of citalopram, a selective serotonin reuptake inhibitor, on the sensitivity of rat vas deferens alpha2-adrenoceptors and to compare it with the effects of serotonin and the dual noradrenaline-serotonin uptake inhibitor duloxetine. To this end, we studied the inhibitory effect of the alpha2-adrenoceptor agonist bromoxidine on the electrically induced contraction of the vas deferens. Citalopram (1, 3 x 10(3) and 3 x 10(4) nM) applied in-vitro significantly attenuated the concentration-response inhibition induced by activation of alpha2-adrenoceptors on the electrically evoked contraction of the vas deferens (concentration of the agonist required to promote 50% of the maximal effect, EC50, for bromoxidine increased by 232%, 421% and 818%, respectively). Similarly, serotonin also attenuated the concentration-response inhibition mediated by presynaptic alpha2-adrenoceptors (96% increase in EC50). Acute and long-term systemic administration of citalopram and duloxetine also produced a loss in the sensitivity of alpha2-adrenoceptors to bromoxidine (EC50 for bromoxidine increased by 97% and 144%, respectively, after citalopram, and by 214% and 167% after duloxetine). In addition, we observed that an increased fraction of receptors was required to be occupied to yield 50% of the inhibitory effect of bromoxidine after long-term administration of citalopram and duloxetine (KE increased by 142% and 83%). These results are indicative of early-onset and persistent down-regulation of peripheral alpha2-adrenoceptors by citalopram, which may account for some of its side effects.     

Effects of alpha,beta-methylene ATP on potentiation of contractions to field stimulation of rat vas deferens by carbachol

Carbachol (0.1-300 mumol/L) potentiated contractions to field stimulation (0.1 Hz, 1 ms, supramaximal V) in the rat epididymal and prostatic vas deferens. Desensitization of P2-purinoceptors by exposure to alpha,beta-methylene ATP (30 mumol/L) markedly reduced (greater than 80%) the potentiating effect of carbachol in the prostatic vas deferens but only moderately reduced (about 20%) the maximal stimulated response to carbachol in the epididymal segment. The presence of prazosin (10 mumol/L) and yohimbine (10 mumol/L), being selective alpha 1- and alpha 2-adrenoceptor antagonists, did not modify the attenuation of carbachol potentiation caused by alpha,beta-methylene ATP treatment. At 0.1 mmol/L, alpha,beta-methylene ATP had no significant effect on the binding of [3H]QNB to muscarinic cholinergic receptors. It is concluded that carbachol may potentiate the contractions to field stimulation in the prostatic vas deferens via an enhancement of purinergic neurotransmission. The molecular mechanism of carbachol potentiation in the epididymal vas deferens remains to be established.     

Alpha-adrenoceptor occupancy by N,N-dimethyl-2-bromo-2-phenethylamine hydrobromide (DMPEA) in rat vas deferens

The effects of N,N-dimethyl-2-bromo-2-phenethylamine hydrobromide (DMPEA) on alpha 1- and alpha 2-adrenoceptors were examined in the isolated rat vas deferens. The active species of DMPEA was the corresponding ethyleniminium ion, which forms in the biophase, since pretreatment of tissues with sodium thiosulphate completely prevented the DMPEA-induced inhibition at alpha 1- and alpha 2-adrenoceptors. DMPEA was approximately 42-fold more potent in inhibiting alpha 2-adrenoceptors than it was in inhibiting alpha 1-adrenoceptors. The antagonism of both receptor types was reversible since washing of the tissues after incubation with DMPEA brought the agonist dose-response curve back to the control value. At alpha 1-adrenoceptors, DMPEA displaced the noradrenaline dose-response curve to the right and concomitantly depressed the maximum response, effects which are consistent with a non-competitive mechanism of action. At alpha 2-adrenoceptors, DMPEA caused a parallel shift of the clonidine or noradrenaline dose-response curve to the right in field-stimulated prostatic portions of the rat vas deferens. The antagonism appeared to be competitive at low concentrations, whereas the shift of the clonidine dose-response curve at higher concentrations became overproportional to the DMPEA concentration. A combination of DMPEA with idazoxan produced a less-than-additive shift of the dose-response curve for clonidine, indicating that these antagonists do not bind to the same site.     

Alpha(1D)-adrenoceptors mediate nerve and agonist-evoked contractions in mouse vas deferens: evidence obtained from knockout technology

1 It has been demonstrated that nerve-evoked contractions of the rat vas deferens involve alpha(1D)-adrenoceptors. Definitive evidence for a similar alpha(1D)-adrenoceptor-mediated response in mouse vas deferens has been more difficult to obtain. In this study, we have used alpha(1D)-adrenoceptor knockout (alpha(1D)-KO) mice to aid in the pharmacological characterization. 2 Mouse whole vas deferens was stimulated with a single pulse every 5 min. Once a stable response had been obtained, vehicle or antagonist was administered cumulatively at 5-min intervals and a response to stimulation obtained 5 min later. Cumulative concentration-response curves were also obtained for noradrenaline. 3 In vas deferens from alpha(1D)-KO mice, the contractile response to low concentrations of noradrenaline and the contractile response to a single stimulus were significantly reduced as compared to wild type (WT). 4 The alpha(1D)-adrenoceptor selective antagonist, BMY 7378, produced a concentration-dependent inhibition of single pulse-evoked contractions of vas deferens from WT and alpha(1D)-KO mice. BMY 7378 was significantly less potent in inhibiting stimulation-evoked contractions in vas deferens from alpha(1D)-KO mice. 5 It is concluded that alpha(1D)-adrenoceptors mediate a component of nerve- and agonist-evoked contractions of the vas deferens of WT mice.     

Effects of castration and of testosterone replacement on alpha(1)-adrenoceptor subtypes in the rat vas deferens

The contractions of the rat vas deferens in response to noradrenaline are mediated through alpha(1A)-adrenoceptors. We observed participation of alpha(1B)-adrenoceptors in these contractions after castration. We now investigated the time course of this plasticity and the effects of testosterone by determining the actions of competitive antagonists on noradrenaline-induced contractions after 7, 14, 21 and 30 days of castration. BMY 7378 (8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspiro[4.5]decane-7,9-dione dihydrochloride) antagonised noradrenaline-induced contractions in control and castrated rats with low pA(2) values (approximately = 6.8). In control vas deferens, WB 4101 (2-(2,6-dimethoxyphenoxyethyl)aminomethyl-1,4-benzodioxane hydrochloride) had a slope in the Schild plot no different from 1.0, while slopes lower than 1.0 (approximately 0.6) were observed for vas deferens from castrated rats. Chloroethylclonidine was ineffective in the control vas while it inhibited noradrenaline-induced contractions in vasa from castrated rats and converted the complex antagonism by WB 4101 into simple competitive antagonism. Treatment of castrated rats with testosterone prevented the effects of castration. The results suggest that alpha(1B)-adrenoceptors are detectable in vas deferens from at least the 7th through the 30th day after castration and that testosterone prevents this plasticity.     

Sympathectomy reveals alpha 1A- and alpha 1D-adrenoceptor components to contractions to noradrenaline in rat vas deferens

We have previously demonstrated that contractions of rat vas deferens to exogenous noradrenaline involve predominantly alpha(1A)-adrenoceptors, but that contractions to endogenous noradrenaline involve predominantly alpha(1D)-adrenoceptors. In this study, we have examined the effects of sympathectomy on the subtypes of alpha(1)-adrenoceptor in rat vas deferens in radioligand binding and functional studies. In vehicle-treated tissues, antagonist displacement of [(3)H]prazosin binding to alpha(1)-adrenoceptors was consistent with a single population of alpha(1)-adrenoceptors. Binding affinities for a range of alpha(1)-adrenoceptor antagonists were expressed as pK(i) values and correlated with known affinities for alpha(1)-adrenoceptor subtypes. The correlation was significant only with alpha(1A)-adrenoceptors. In tissues from rats sympathectomised with 6-hydroxy-dopamine (2 x 100 mg kg(-1) i.p.), binding affinity for the alpha(1D)-adrenoceptor antagonist BMY 7378 fitted best with a two-site model. In functional studies, the potency of noradrenaline at producing total (phasic plus tonic) but not tonic contractions was increased in tissues from sympathectomised rats. Results obtained from sympathectomised rats suggest that phasic contractions are mainly alpha(1D)-adrenoceptor mediated, whereas tonic contractions are mainly alpha(1A)-adrenoceptor mediated, based on the effects of BMY 7378 and the alpha(1A)-adrenoceptor antagonist RS 100329. It is concluded that the predominant alpha(1)-adrenoceptor in vehicle-treated rat vas deferens is the alpha(1A)-adrenoceptor, both in terms of ligand binding and contractions to exogenous agonists. The alpha(1D)-adrenoceptor is only detectable by ligand binding following chemical sympathectomy, but is involved in noradrenaline-evoked contractions, particularly phasic contractions, of rat vas deferens.     

Receptor interactions of a series of imidazolines: comparison of the alpha 2-adrenoceptors between the rabbit vas deferens and guinea pig ileum

A series of imidazolines and norepinephrine were used to characterize and differentiate the presynaptic alpha 2-adrenoceptors in the rabbit vas deferens and the guinea pig ileal longitudinal muscle using pharmacological procedures. Based on pEC50-values (the negative log of the 50% effective concentration) for each imidazoline, a rank order of potency of p-aminoclonidine greater than oxymetazoline greater than or equal to clonidine greater than naphazoline greater than phentolamine was obtained in the rabbit vas deferens and an order of p-aminoclonidine greater than clonidine greater than naphazoline greater than oxymetazoline was obtained in the guinea pig ileum. In the rabbit vas deferens, phentolamine, which is generally considered to be a competitive alpha 1- and alpha 2-adrenoceptor antagonist, acted as a full alpha 2-adrenoceptor agonist. The dissociation constants of oxymetazoline and yohimbine were significantly lower in the rabbit vas deferens than in the guinea pig ileum. These results suggest that the presynaptic alpha 2- adrenoceptors in these tissues are different. Furthermore, the pKB-value of yohimbine against norepinephrine was significantly one log unit lower than those obtained using a series of imidazolines. Data from our studies add to increasing evidence of the existence of high and low affinity binding sites on the alpha 2-adrenoceptors in the rabbit vas deferens.     

Discrepancy between alpha 1-adrenoceptor-mediated contraction and the occupation theory in rat vas deferens--possible existence of a 'silent' receptor

The relation of the amount of alpha 1-adrenoceptors (alpha 1-R) with contraction to norepinephrine (NE) through alpha 1-R in rat vas deferens was examined by means of radiobinding assays. Treatment with dibenamine decreased the maximal contraction to NE with a decrease in the amount of alpha 1-R but the relation was not linear. The contractile response disappeared completely when 20% of the alpha 1-R still remained. Moreover, culture of dibenamine-pretreated muscle restored the contraction to NE without a significant increase in the amount of alpha 1-R in the muscle. These findings suggest that some alpha 1-R are 'silent' in the contraction of rat vas deferens in response to NE under physiological conditions.     

Calcium mobilization and phosphatidylinositol turnover in vas deferens smooth muscle of diabetic rats

The study concerned Ca2+ channels that are receptor-operated by norepinephrine (NE) and mediate hyper-reactivity of vas deferens smooth muscle from rats with streptozotocin (STZ)-induced diabetes, and the mediatory responses of these channels, such as tension development, Ca2+ uptake and phosphatidylinositol (PI) turnover. The contractile responses induced by adrenoceptor agonists were significantly greater in diabetic rat vas deferens than in the controls. A greater Ca2+ uptake was induced by 10(-5) M NE in strips from diabetic rats than in the controls. The uptake of Ca2+ was completely inhibited by 10(-6) M prazosin but not by 10(-5) M verapamil. Enhancement of Ca2+ release by 10(-5) M NE was faster and greater in diabetic muscles than in the controls. The accumulation of [3H]inositol phosphates was increased 4-fold in the controls and 7-fold in diabetic muscles by 10(-5) M NE. This increase was completely inhibited by 10(-6) M prazosin but not by 10(-6) M yohimbine. The data suggest that vas deferens smooth muscle hyper-reactivity in diabetic rats is due to increased PI turnover mediated by alpha 1-adrenoceptors, to the release of intracellular bound Ca2+ and to an increase of Ca2+ uptake through receptor-operated Ca2+ channels.     

Coupling to protein kinases A and C of adenosine A2B receptors involved in the facilitation of noradrenaline release in the prostatic portion of rat vas deferens

In the prostatic portion of rat vas deferens, the non-selective adenosine receptor agonist NECA (0.1-30 microM), but not the A(2A) agonist CGS 21680 (0.001-10 microM), caused a facilitation of electrically evoked noradrenaline release (up to 43 +/- 4%), when inhibitory adenosine A(1) receptors were blocked. NECA-elicited facilitation of noradrenaline release was prevented by the A(2B) receptor-antagonist MRS 1754, enhanced by preventing cyclic-AMP degradation with rolipram, abolished by the protein kinase A inhibitors H-89, KT 5720 and cyclic-AMPS-Rp and attenuated by the protein kinase C inhibitors Ro 32-0432 and calphostin C. The adenosine uptake inhibitor NBTI also elicited a facilitation of noradrenaline release; an effect that was abolished by adenosine deaminase and attenuated by MRS 1754, by inhibitors of the extracellular nucleotide metabolism and by blockade of alpha(1)-adrenoceptors and P2X receptors with prazosin and NF023, respectively. It was concluded that adenosine A(2B) receptors are involved in a facilitation of noradrenaline release in the prostatic portion of rat vas deferens that can be activated by adenosine formed by extracellular catabolism of nucleotides. The receptors seem to be coupled to the adenylyl cyclase-protein kinase A pathway but activation of the protein kinase C by protein kinase A, may also contribute to the adenosine A(2B) receptor-mediated facilitation of noradrenaline release.