Prostaglandin synthesis elicited by adrenergic stimuli is mediated via alpha-2C and alpha-1A adrenergic receptors in cultured smooth muscle cells of rabbit aorta.

This study was performed to characterize the subtype of adrenergic receptor(s) (AR) involved in prostacyclin synthesis [measured as 6-keto-prostaglandin (PG)F1 alpha] elicited by AR agonists in cultured vascular smooth muscle cells of rabbit aorta. Both alpha-1 and alpha-2 AR agonists enhanced 6-keto-PGF1 alpha synthesis in a dose-dependent manner with the following order of potency: norepinephrine greater than BHT 933 greater than UK 14304 greater than xylazine greater than phenylephrine greater than or equal to methoxamine greater than cirazoline. Isoproterenol and oxymetazoline did not alter 6-keto-PGF1 alpha synthesis. Methoxamine-induced 6-keto-PGF1 alpha synthesis was not reduced by the alpha-2 AR antagonist rauwolscine. The affinities of AR antagonists (PA2 value) in inhibiting methoxamine-induced 6-keto-PGF1 alpha synthesis were of the following order: prazosin greater than WB 4101 greater than corynanthine greater than yohimbine. Administration of WB 4101 and the irreversible alpha-1B AR antagonist chloroethylclonidine reduced norepinephrine (in the presence of rauwolscine, 10(-8) M)- or methoxamine-induced 6-keto-PGF1 alpha synthesis; WB 4101 was more potent than chloroethylclonidine. UK 14304-induced 6-keto-PGF1 alpha synthesis was not reduced by chloroethylclonidine or BRL 44408, a selective alpha-2A AR antagonist, but it was inhibited by other alpha AR antagonists. The affinities of AR antagonists (PA2 values) in inhibiting UK 14304-induced 6-keto-PGF1 alpha synthesis were of the following order: rauwolscine greater than yohimbine greater than BAM 1303 greater than BRL 41992 greater than WB 4101 greater than ARC 239 greater than or equal to prazosin greater than SKF 104078 greater than or equal to corynanthine. The order of affinity of alpha-2 AR antagonists in inhibiting UK 14304-induced 6-keto-PGF1 alpha synthesis in vascular smooth muscle cells was similar to that derived from radioligand binding studies in opossum kidney cell line receptors classified as alpha-2C receptors. These data suggest that 6-keto-PGF1 alpha synthesis elicited by adrenergic stimuli in cultured vascular smooth muscle cells of rabbit aorta is mediated primarily via alpha-2C and to a lesser extent alpha-1A receptors.     

Prostaglandin synthesis elicited by adrenergic stimuli in rabbit aorta is mediated via alpha-1 and alpha-2 adrenergic receptors

The purpose of this study was to characterize the type of adrenergic receptor(s) involved in both prostaglandin (PG) synthesis and the contractile response elicited by adrenergic receptor agonists in the rabbit aorta. The synthesis of prostacyclin as measured by the production of 6-keto-PGF1 alpha was assessed in vitro after exposing the aortic rings to different adrenergic agonists. Norepinephrine (NE), selective alpha 1 adrenergic receptor agonists methoxamine (MET), phenylephrine (PHE) and cirazoline (CIR) and the alpha 2 adrenergic receptor agonists UK 14304 (UK) and xylazine (XYL), but not the beta adrenergic receptor agonist isoproterenol (ISP), enhanced 6-keto-PGF1 alpha synthesis in a concentration-dependent manner with following order of potency: NE greater than UK 14304 greater than XYL greater than PHE greater than MET greater than CIR. The NE-induced increased in 6-keto-PGF1 alpha synthesis was attenuated by the alpha 1 adrenergic receptor antagonists prazosin (PZ) and corynanthine (COR) and by the alpha 2 adrenergic receptor antagonists rauwolscine (RW) and yohimbine (YOH). MET-induced 6-keto-PGF1 alpha synthesis was reduced by PZ and COR but not by RW. UK-induced 6-keto-PGF1 alpha synthesis was reduced by RW, YOH, and PZ, which also acts as alpha-2B receptor antagonist, but not by COR. In rabbit aortic rings, adrenergic agonists produced contraction with the following order of potency: NE greater than PHE greater than MET greater than CIR greater than UK greater than XYL.(ABSTRACT TRUNCATED AT 250 WORDS)     

Activation of alpha-2 adrenergic receptors augments neurotransmitter-stimulated cyclic AMP accumulation in rat brain cerebral cortical slices

Norepinephrine-stimulated cyclic AMP (cAMP) accumulation in brain is mediated by both alpha and beta adrenergic receptors. The functional activity of these receptors can be differentiated by examining the response to isoproterenol alone and isoproterenol + 6-fluoronorepinephrine, and alpha adrenergic agonist. The present study was undertaken to define the pharmacological characteristics of the alpha adrenergic component associated with cAMP accumulation in brain. Using a prelabeling technique it was found that norepinephrine- or isoproterenol plus 6-fluoronorepinephrine-stimulated cAMP accumulation was only inhibited partially by an alpha-1 adrenergic receptor antagonist. In contrast, alpha-2 adrenergic receptor antagonists inhibited completely that portion of the response exceeding that produced by isoproterenol alone (alpha adrenergic augmentation). Furthermore, selective alpha-1 adrenergic agonists were incapable of enhancing the cAMP response to isoproterenol, whereas alpha-2 adrenergic agonists were active in this regard. Partial agonists for the alpha-2 adrenergic receptor were ineffective as augmentors. The data suggest that the alpha adrenergic component of the norepinephrine response in rat brain slices has characteristics of both alpha-1 and alpha-2 receptors, with the alpha-2 adrenergic component being a major contributor in this regard.     

Estimation of agonist dissociation constants at central presynaptic alpha-2 autoreceptors in the absence of autoinhibition.

The agonist dissociation constants (KA) and relative efficacies of UK-14304, norepinephrine (NE) and clonidine at presynaptic release-modulating alpha-2 adrenoceptors were determined in rat cerebral cortex slices using the irreversible alpha-2 adrenoceptor antagonist N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) (0.8 mg/kg i.p.) to reduce the receptor pool. Eighteen hours after treatment, the slices were incubated with [3H]NE, superfused in the presence of reuptake inhibitor and stimulated electrically with short bursts of 4 pulses delivered at 100 Hz (pseudo-one-pulse; POP) or trains of 36 or 72 pulses delivered at 3 Hz. EEDQ treatment did not affect the overflow of radioactivity evoked by POP, but greatly facilitated overflow at 36 or 72 pulses/3 Hz indicating that the autoinhibition seen under the latter conditions is totally lacking with POP stimulation. KA values determined using 4 pulses/100 Hz were 136, 50 and 625 nM for UK-14304, clonidine and NE, respectively. At 36 or 72 pulses/3 Hz the values were higher by a factor of up to 3. The percentage of receptors active after EEDQ treatment was found to be 5.5 to 8.2% and was not influenced by conditions of stimulation. Receptor reserves were estimated to be about 65% for UK-14304 and NE and 40% for clonidine. The efficacies of UK-14304 and clonidine relative to NE were 1 and 0.5, respectively. The data indicate that KA values for agonists at presynaptic alpha-2 autoreceptors are inevitably underestimated if the released transmitter causes inhibition of release in addition to the drug under investigation.     

Desensitization of the alpha-2 adrenergic receptor in HT29 and opossum kidney cell lines

Preincubation of HT29 cells with an alpha-2 adrenergic agonist resulted in a parallel rightward shift in the subsequent dose-response curve to 5-bromo-6-[2-imidazoline-2-yl-amino] quinoxaline (an alpha-2 adrenergic agonist) in inhibiting vasoactive intestinal peptide-stimulated cyclic AMP production. This rightward shift in the dose-response curve, which was concentration and time dependent, was interpreted as desensitization of the alpha-2 adrenergic receptor-mediated inhibition of cyclic AMP production. The fact that no decrease in efficacy was observed appears to result from a receptor reserve. Agonist preincubation effects on subsequent p-aminoclonidine (an alpha-2 adrenergic partial agonist) inhibition and partial irreversible inactivation of receptors confirmed the presence of an alpha-2 adrenergic receptor reserve in HT29 cells. Desensitization appeared to have a heterologous component since inhibition of vasoactive intestinal peptide-stimulated cyclic AMP production by somatostatin was also attenuated. We also assessed the effect of alpha-2 agonist preincubation on subsequent 5-bromo-6-[2-imidazoline-2-yl-amino] quinoxaline inhibition of parathyroid hormone-stimulated cyclic AMP production in OK cells. As with HT29 cells, agonist preincubation resulted in a concentration- and time-dependent shift in the dose-response curve. In both cell lines, long-term preincubation with an alpha-2 adrenergic agonist resulted in a 40% decrease in subsequent [3H]yohimbine binding, indicating down-regulation of the alpha-2 adrenergic receptor.     

Pharmacological characterization of alpha adrenoceptors in the rat gastric fundus

Alpha receptors on the intramural cholinergic neurons and on the smooth muscle cells are involved in the inhibitory effect of catecholamines on rat gastric fundus motility. The pharmacological characteristics of these alpha receptors were assessed using longitudinal muscle strips of the rat gastric fundus, contracted to a similar degree by electrical stimulation and by methacholine. All alpha agonists studied (norepinephrine, phenylephrine, methoxamine, clonidine, UK-14,304 and B-HT 920) concentration-dependently inhibited the stimulation-induced contractions. Norepinephrine, phenylephrine, methoxamine and clonidine also inhibited the methacholine-induced contractions, but for the same concentration of agonist, the inhibition was less pronounced than during electrical stimulation-induced contractions; UK-14,304 and B-HT 920 inhibited the methacholine-induced contractions only in a concentration of 10(-4) M. The effect of clonidine and UK-14,304 on electrical stimulation-induced contractions was antagonized competitively by the alpha antagonists rauwolscine and yohimbine (slope in the Schild plot not different from 1). The effect of norepinephrine and phenylephrine on methacholine-induced contractions was antagonized by the alpha antagonists prazosin, corynanthine and yohimbine; against phenylephrine, the antagonism was competitive (slope in the Schild plot not different from 1). It is concluded that the muscular alpha receptors in the rat gastric fundus are of the alpha-1-type. On the postganglionic cholinergic neurons, alpha-2-like receptors are present; it is not yet clear whether the pronounced effect of alpha-1 agonists on the cholinergic neuron activity is due to interaction with these receptors.     

Receptor reserve at the alpha-2 adrenergic receptor in the rat cerebral cortex

N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ), an irreversible alpha-2 antagonist, was used to establish and quantitate the receptor reserve at the alpha-2 adrenergic autoreceptor mediating inhibition of [3H]norepinephrine ([3H]NE) release in rat cerebral cortical slices. EEDQ treatment had no effect on [3H]NE uptake or base-line release. Four hours after EEDQ treatment (0.8 mg/kg i.p.), the EC50 was shifted 7-fold to the right and there was a 21.5% decrease in the maximal response to the full alpha-2 agonist UK-14304. Using the double-reciprocal plot analysis, the equilibrium activation constant (KA) was calculated to be 1.41 +/- 0.8 microM. Similar analysis of alpha-2 autoreceptor response at various times after 1.6 mg/kg of EEDQ gave similar values for the KA. Therefore, evaluation of either the response of the remaining native receptors after partial irreversible inactivation or the response of newly synthesized receptors after nearly complete irreversible inactivation can be used to determine the KA of the receptor. Comparison of repopulation kinetics analyses for alpha-2 receptor response and estimated receptor number revealed that recovery of maximal response was much faster than actual receptor recovery. By examining the relationship between alpha-2 autoreceptor occupancy and response it was possible to determine that there is approximately a 60 to 70% receptor reserve; only 1.5% of the receptors need to be occupied by UK-14304 in order to obtain 50% of the maximal inhibition of [3H]NE release. The presence of a large receptor reserve must be taken into account when evaluating alpha-2 adrenergic autoreceptor regulation in the rat cerebral cortex.     

Systemic and regional hemodynamic characterization of alpha-1 and alpha-2 adrenoceptor agonists in pithed rats

In pithed rats, blood pressure dose-response curves to i.v. cirazoline, methoxamine and phenylephrine (full alpha-1 adrenoceptor agonists) exhibited higher maxima than those to B-HT 920, M-7, UK-14,304 (full alpha-2 adrenoceptor agonists) and indanidine (Sgd 101/75: partial alpha-1 adrenoceptor agonist). For an 80 mm Hg increase in blood pressure, full alpha-1 adrenoceptor agonists enhanced total peripheral, renal and mesenteric vascular resistances significantly more than alpha-2 adrenoceptor stimulants or indanidine. In contrast, all compounds produced a similar degree of hindquarter vasoconstriction, suggesting that both types of alpha adrenoceptors have the same functional importance in this skeletal muscle vascular bed. Application of a multivariate discriminant analysis to the drug-induced changes in the total peripheral and mesenteric vascular resistances associated with a pressor effect of 80 mm Hg allowed their assignment to two distinct groups corresponding to the full alpha-1 and the full alpha-2 adrenoceptor agonists plus indanidine. All investigated compounds in low doses increased cardiac output, which returned to base-line values after high doses of alpha-1 but plateaued after high doses of alpha-2 adrenoceptor agonists or indanidine. alpha-1 adrenoceptor agonists decreased whereas alpha-2 stimulants and indanidine successively increased and then decreased renal blood flow. Finally, all investigated compounds increased hindquarter blood flow at low doses but decreased it at high doses. The ratios of the doses of cirazoline required to produce a 100% rise in systemic and local vascular resistances in the presence or in the absence of prazosin were of similar magnitude. This was also true for M-7 when studied in the presence or in the absence of yohimbine. These findings suggest pharmacological identity within alpha-1 as well as within alpha-2 adrenoceptor populations in all investigated vascular beds. Finally, the calcium entry blocker diltiazem did not affect the increases in systemic and regional resistances evoked by cirazoline but depressed profoundly the effects of M-7 and indanidine. In conclusion, full alpha-1 and alpha-2 adrenoceptor agonists can be discriminated easily on the basis of their systemic and regional hemodynamics in the pithed rat. That the hemodynamic effects of the partial alpha-1 adrenoceptor agonist indanidine are similar to those of alpha-2 adrenoceptor agonists and susceptible to calcium channel blockade suggests that the alpha-1 adrenoceptors stimulated by this drug have the same coupling modality as alpha-2 adrenoceptors and share with the latter the same functional expression when stimulated.     

Stimulation of adenosine 3',5'-monophosphate formation in rat cerebral cortical slices by methoxamine: interaction with an alpha adrenergic receptor.

Methoxamine elicits a rapid accumulation of adenosine 3',5'-monophosphate (cyclic AMP) in rat cerebral cortical slices with maximal effects at 100 muM. The accumulations of cyclic AMP elicited by this amine are completely blocked by the alpha adrenergic antagonists, phenoxybenzamine and dihydroergokryptine, partially blocked by the alpha antagonist, phentolamine, and unaffected by the beta blocking agent, propranolol, or by the local anesthetic, tetracaine. The magnitude of the accumulations of cyclic AMP elicited by methoxamine in cerebral cortical slices of four rat strains (F-344, ACI, BUF, and Sprague-Dawley) exhibit a strong negative correlation with spontaneous motor activity and a positive correlation with the magnitude of norepinephrine-elicited accumulations of cyclic AMP. The stimulatory interaction of methoxamine with alpha adrenergically regulated cyclic AMP-generating systems differs from the interaction of norepinephrine with alpha receptors as evidenced by the following observations: 1) the stimulatory effects of methoxamine and norepinephrine are nearly additive; 2) the stimulatory effects of methoxamine and adenosine are nearly additive, whereas the effects of norepinephrine and adenosine are much more than additive. Methoxamine, however, does not increase further the magnitude of accumulation of cyclic AMP elicited by a combination of norepinephrine and adenosine. The results are consonant with the interaction of methoxamine with alpha adrenergic receptors which are normally activated by norepinephrine only to a marginal extent. However, in the presence of adenosine, these receptors are now sensitive to activation by norepinephrine.     

Enhancement of alpha-1 and alpha-2 adrenergic agonist-induced vasoconstriction by removal of endothelium in rat aorta

It is well known that the vascular endothelial cell layer plays an essential role in the vasodilatory response of several agents. In this study we have investigated the possibility that the endothelium may also modulate alpha adrenergic agonist-induced vasoconstriction. The responses of rat aortae to selective alpha-1 and alpha-2 adrenergic agonists were studied. Removal of the endothelium did not significantly alter the maximum contractile response to norepinephrine. However, the maximum responses to selective alpha-1 agonists (phenylephrine and methoxamine) were increased 2-fold. The vasoconstrictor effects of both clonidine and B-HT920 (selective alpha-2 agonists) were enhanced 5- to 7-fold after removal of the endothelial cell layer. The sensitivity of the tissue, as reflected by the EC50 value, to each alpha adrenergic agonist was enhanced in the absence of endothelium. An explanation for the present results is that alpha-1 and alpha-2 adrenergic agonists activate adrenoceptors in the endothelial cells and thereby may promote the release of a relaxing factor to inhibit vascular smooth muscle contraction. Removal of the endothelium would abolish release of this putative inhibitory substance and adrenergic agonist would activate only adrenoceptors in the muscle to cause vasoconstriction. On the other hand, endothelial cells may function as an uptake site for the various adrenergic agonists. Ablation of this uptake process could conceivably result in a greater effective concentration of the agonist in the receptor area and thus promote a stronger vasoconstrictor effect.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibition of vasopressin-stimulated cyclic AMP accumulation by alpha-2 adrenoceptor agonists in isolated papillary collecting ducts

The effects of selective alpha adrenoceptor agonists and antagonists on vasopressin (VP)-sensitive cyclic AMP (cAMP) formation in microdissected rat papillary collecting ducts were examined. In the presence of 10(-10) M VP, norepinephrine and the selective alpha-2 adrenoceptor agonist, B-HT 933, produced almost total inhibition of VP-stimulated cAMP accumulation. Half-maximal inhibition occurred at 1 x 10(-8) M and 6 x 10(-7) M for norepinephrine and B-HT 933, respectively. Cirazoline, a selective alpha-1 adrenoceptor agonist, had no significant effect on VP-stimulated cAMP accumulation. The inhibitory effects of norepinephrine and B-HT 933 were antagonized by rauwolscine but not by prazosin. The antagonism of B-HT 933-induced inhibition of VP-stimulated cAMP accumulation was competitive with an antagonist dissociation constant (KB) of 10.9 x 10(-9) M. Preincubation of papillary collecting ducts with pertussis toxin (1 microgram/ml for 1 hr at 37 degrees C) attenuated, by 65%, the inhibitory effect of B-HT 933 on VP-stimulated cAMP levels. These results demonstrate that alpha-2 adrenoceptors capable of inhibiting VP action are present on the papillary collecting duct. Furthermore, the alpha-2 adrenoceptor-induced inhibition of VP-stimulated cAMP accumulation is pertussis-toxin sensitive. This suggests that alpha-2 adrenoceptors are coupled negatively to adenylate cyclase, via the guanine nucleotide binding protein, in the collecting tubule.     

Ablation of the myenteric plexus impairs alpha but not beta adrenergic receptor-mediated mechanical responses of rat jejunal longitudinal muscle

The mechanical responses produced by alpha and beta adrenergic receptor agonists were evaluated in control and myenteric neuron-ablated rat jejunal longitudinal muscle. The myenteric plexus of the jejunum was destroyed by serosal application of benzalkonium chloride (BAC). The beta adrenergic receptor agonists isoproterenol and sulfonterol produced a concentration-dependent relaxation of both control and BAC-treated jejunum. Dose-response curves obtained in control and BAC-treated jejunum were nearly superimposable regardless of the beta agonist used. Isoproterenol-induced relaxation was antagonized by the beta receptor antagonists propranolol and practolol but not by butoxamine. The alpha-1 selective agonists phenylephrine and methoxamine were more potent and efficacious in producing relaxation of control than BAC-treated jejunum. The relaxant responses of methoxamine and phenylephrine in control jejunum were blocked by prazosin but not by yohimbine. The supposed alpha-2 selective agonist clonidine also produced a concentration-dependent, prazosin-sensitive, yohimbine-resistant relaxation which was markedly greater in control than BAC-treated jejunum, consistent with alpha-1 receptor stimulation. Clonidine tested in the presence of prazosin and the alpha-2 selective receptor agonists UK-14,304, M-7 and B-HT 920 all produced a concentration-dependent contraction of control but not BAC-treated jejunum. The contractile response produced by UK-14,304 was antagonized by yohimbine but not by atropine. Our results suggest that in rat jejunal longitudinal muscle: beta adrenergic receptors mediate relaxation and are located on the smooth muscle; alpha-1 adrenergic receptors mediate relaxation and are located on both the smooth muscle and myenteric plexus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evaluation of alpha-1 and alpha-2 adrenoceptor-mediated vasoconstriction in the in situ, autoperfused, pulmonary circulation of the anesthetized dog

Alpha-1 and alpha-2 adrenoceptor-mediated vasoconstriction was studied in the in situ, autoperfused pulmonary circulation of the open-chest anesthetized dog utilizing selective alpha adrenoceptor agonists and antagonists. Animals were pretreated with propranolol (1 mg/kg i.v.) to eliminate beta adrenoceptor-mediated effects in the pulmonary circulation. Blood was withdrawn from the right femoral artery and transferred, via a peristaltic pump, to the pulmonary arterial branch supplying the left diaphragmatic lobe of the lung. The flow rate of the pump was set so that the perfusion pressure in the lobe was equal to resting diastolic pulmonary artery pressure (10 +/- 1 mm Hg). Under conditions of constant left atrial pressure and pulmonary blood flow, intralobar administration of alpha adrenoceptor agonists elicited increases in perfusion pressure of the lobe, reflecting changes in pulmonary vascular resistance. Intralobar administration of the selective alpha-1 adrenoceptor agonist methoxamine and the selective alpha-2 adrenoceptor agonist B-HT 933 elicited dose-dependent increases in lobar perfusion pressure, as did the nonselective alpha adrenoceptor agonist norepinephrine. Prazosin (100 micrograms/kg i.v.), a selective alpha-1 adrenoceptor antagonist, inhibited pulmonary vasopressor responses to methoxamine and norepinephrine without altering significantly the response to B-HT 933. Rauwolscine (100 micrograms/kg i.v.), a selective alpha-2 adrenoceptor antagonist, inhibited the response to B-HT 933 and norepinephrine with little effect on methoxamine. Intralobar administration of tyramine to evoke the release of endogenous norepinephrine resulted in dose-dependent increases in lobar perfusion pressure. The response to tyramine was inhibited selectively by prazosin with little effect of rauwolscine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization of alpha-2 adrenergic receptors in the OK cell, an opossum kidney cell line

We have characterized alpha-2 adrenergic receptors in OK cells, an opossum kidney-derived cell line. In membrane saturation binding experiments, [3H]rauwolscine (Kd = 74 pM) was 3-fold more potent than [3H]yohimbine (Kd = 230 pM). Each labeled a single class of binding sites with densities of 135 and 124 fmol/mg of protein for [3H]rauwolscine and [3H]yohimbine, respectively. Inhibition of [3H]rauwolscine and [3H]yohimbine binding by several alpha adrenergic agonists and antagonists demonstrated the radioligands labeled an alpha-2 type adrenergic receptor with a pharmacological profile similar to the alpha-2B receptor subtype. The rank order of potency for antagonist inhibition of binding was yohimbine greater than prazosin = phentolamine greater than chlorpromazine = corynanthine, whereas the rank order of agonist potency was oxymetazoline = clonidine greater than or equal to UK-14,304 greater than or equal to (-)-epinephrine greater than (-)-norepinephrine. The oxymetazoline, clonidine and antagonist inhibition curves were routinely monophasic and modeled best as a single class of binding sites. For the other agonists, inhibition binding curves were biphasic with approximately 35% of the binding sites existing in a high affinity state. These curves were shifted to the right in the presence of 0.1 mM GTP, and in general modeled as a single class of binding sites. UK-14,304, (-)-epinephrine, (-)-norepinephrine and oxymetazoline attenuated parathyroid hormone-stimulated cyclic AMP production by up to 70% in whole cell monolayers in a dose-dependent manner via a pertussis toxin-sensitive mechanism. With the exception of oxymetazoline, this inhibition could be reversed with alpha adrenergic antagonists.(ABSTRACT TRUNCATED AT 250 WORDS)     

Differences in responsiveness of intrapulmonary artery and vein to arachidonic acid: mechanism of arterial relaxation involves cyclic guanosine 3':5'-monophosphate and cyclic adenosine 3':5'-monophosphate

The objective of this study was to examine the relationship between responses of bovine intrapulmonary artery and vein to arachidonic acid and cyclic nucleotide levels in order to better understand the mechanism of relaxation elicited by arachidonic acid and acetylcholine. Arachidonic acid relaxed phenylephrine-precontracted arterial rings and elevated both cyclic GMP and cyclic AMP levels in arteries with intact endothelium. In contrast, endothelium-damaged arterial rings contracted to arachidonic acid without demonstrating significant changes in cyclic nucleotide levels. Indomethacin partially inhibited endothelium-dependent relaxation and abolished cyclic AMP accumulation whereas methylene blue, a guanylate cyclase inhibitor, partially inhibited relaxation and abolished cyclic GMP accumulation in response to arachidonic acid. All vessel responses were blocked by a combination of the two inhibitors. Prostaglandin (PG) I2 relaxed arterial rings and elevated cyclic AMP levels whereas PGE2 and PGF2 alpha caused contraction, suggesting that the indomethacin-sensitive component of arachidonic acid-elicited relaxation is due to PGI2 formation and cyclic AMP accumulation. The methylene blue-sensitive component is attributed to an endothelium-dependent but cyclooxygenase-independent generation of a substance causing cyclic GMP accumulation. Intrapulmonary veins contracted to arachidonic acid with no changes in cyclic nucleotide levels and PGI2 was without effect. Homogenates of intrapulmonary artery and vein formed 6-keto-PGF1 alpha, PGF2 alpha and PGE2 from [14C]arachidonic acid, which was inhibited by indomethacin. Thus, bovine intrapulmonary vein may not possess receptors for PGI2. The failure of endothelium-intact vein to relax to acetylcholine may be related to the lack of a relaxant effect by arachidonic acid, perhaps attributed to the absence of generation of an endothelium-derived relaxing factor.     

Enhanced pulmonary alpha-2 adrenoceptor responsiveness under conditions of elevated pulmonary vascular tone

Alpha-1 and alpha-2 adrenoceptor-mediated vasoconstriction was studied in the in situ, autoperfused pulmonary circulation of the open-chest anesthetized dog under conditions of normal and elevated pulmonary vascular tone. Under conditions of normal pulmonary vascular tone (10 +/- 1 mm Hg), methoxamine, a selective alpha-1 adrenoceptor agonist, and B-HT 933, a selective alpha-2 adrenoceptor agonist, elicited maximal increases in lobar perfusion pressure of 5 and 2 mm Hg above resting pulmonary tone, respectively. When pulmonary vascular tone was elevated progressively with the thromboxane mimetic, U-46619, serotonin or PGF2 alpha, alpha-1 adrenoceptor-mediated pulmonary vasoconstrictor responses to methoxamine were unaffected, whereas alpha-2 adrenoceptor-mediated pulmonary pressor responses to B-HT 933 were enhanced. Overall the response to B-HT 933 was enhanced 4-fold when pulmonary perfusion pressure was elevated to 19.8 +/- 0.8 mm Hg with U-46619 and almost 5-fold when elevated to 27.0 +/- 1.2 mm Hg. Pulmonary vasoconstrictor responses to angiotensin II were unaffected by elevated pulmonary vascular tone. Enhanced responsiveness of B-HT 933 to elevated pulmonary vascular tone was antagonized by the selective alpha-2 adrenoceptor antagonist, rauwolscine (100 micrograms/kg i.v.), and unaffected by the selective alpha-1 adrenoceptor antagonist, prazosin (100 micrograms/kg i.v.). When canine intralobar pulmonary veins were studied in vitro they contracted to B-HT 933 whereas intralobar pulmonary arteries did not respond. These data indicate that alpha-2 adrenoceptor responsiveness is enhanced markedly and selectively under conditions in which pulmonary vascular tone is elevated.     

The role of adrenergic receptors in the initiation of vomiting and its gastrointestinal motor correlates.

We investigated the role of adrenergic receptors in the mechanisms of initiation of vomiting and its gastrointestinal (GI) motor correlates. The effects of clonidine, UK-14304, St-91, naphazoline, phenylephrine and isoproterenol were examined for their ability to initiate vomiting and its GI motor correlates. Only the alpha-2 adrenoceptor agonists UK-14304, clonidine, St-91 and naphazoline activated vomiting and its GI motor correlates. Tolerance of vomiting, but not its GI motor correlates, readily developed to all alpha-2 adrenergic receptor agonists but St-91. The responses to UK-14304 or clonidine were blocked by idazoxan, yohimbine, clonidine tolerance or high doses of phenoxybenzamine, but not by propranolol or prazosin. The responses to UK-14304 or clonidine were also blocked by fentanyl, 1-(1-naphthyl) piperazine, methysergide, SCH 22390 or scopolamine, but not by haloperidol, sulpiride, domperidone or naloxone. Adrenoceptor antagonists, clonidine tolerance or sympathetic blockade did not block vomiting or its GI motor correlates activated by apomorphine, CuSO4 or cholecystokinin-octapeptide. We concluded that alpha-2 adrenergic receptors of the chemoreceptive trigger zone can initiate vomiting and its GI motor correlates, but these receptors do not mediate vomiting induced by another chemoreceptive trigger zone stimulant, apomorphine, or stimulation of the GI tract using CuSO4. However, 5-hydroxytryptamine-2 serotonergic, muscarinic cholinergic and opiate receptors within the central nervous system participate in controlling emesis activated by alpha-2 adrenergic agonists. Peripheral adrenergic receptors do not mediate the GI motor correlates of vomiting.     

Alpha adrenoceptor-mediated vasoconstriction in rat hindlimb: innervated alpha-2 adrenoceptors in the saphenous arterial bed

The alpha adrenoceptor subtypes mediating vasoconstriction to exogenous agonists and to spinal sympathetic nerve stimulation have been characterized in the autoperfused (constant flow) femoral (predominantly skeletal musculature) and saphenous (predominantly cutaneous) vascular beds of the pithed rat. Intra-arterial infusion of the alpha-1 adrenoceptor agonist, methoxamine, increased perfusion pressure in both vascular beds over the same range of infusion rates, and the maximum responses were similar. The selective alpha-2 adrenoceptor agonist, B-HT 933, also increased perfusion pressure in both beds, although the maximum response to B-HT 933 in the saphenous bed was approximately twice that observed in the femoral bed. Responses to methoxamine were blocked by the alpha-1 adrenoceptor antagonist, prazosin (0.1 mg/kg), but not the alpha-2 adrenoceptor antagonist, rauwolscine (1 mg/kg), or by the selective postjunctional alpha-2 adrenoceptor antagonist, SK&F 104078 (1 mg/kg). Conversely, responses to B-HT 933 were blocked by rauwolscine and by SK&F 104078, but not by prazosin. Vasopressor responses to B-HT 933 in both vascular beds of the rat hindlimb also were reduced markedly by the calcium channel blocker, nifedipine (1 mg/kg), whereas responses to methoxamine were relatively resistant to inhibition by nifedipine. In the femoral bed, as in the systemic arterial circulation, responses to sympathetic nerve stimulation were strongly inhibited by prazosin, were potentiated by rauwolscine and were unaffected by SK&F 104078. In contrast, in the saphenous arterial bed, the responses to sympathetic nerve stimulation were inhibited by all three antagonists.(ABSTRACT TRUNCATED AT 250 WORDS)     

Simultaneous measurements of cytosolic free calcium level and prostaglandin synthesis reveal a correlation between them in perfused monolayer of cultured rat vascular smooth muscle cells: effects of bradykinin and angiotensin II.

The level of cytosolic free calcium ([Ca2+]i) and the production rate of prostacyclin were simultaneously measured in perfused monolayers of cultured vascular smooth muscle (VSM) cells. After loading of fura-2 (a fluorescent calcium indicator), the monolayer of VSM cells (cultured on a cover glass) was fixed in the perfusion cuvette and the cuvette was placed in a fluorometer to monitor the change in [Ca2+]i. The monolayer was perfused and the fractionated perfusion solution was collected to determine 6-keto-PGF1 alpha (a metabolite of prostacyclin) production found in the solution. Afterwards, the time-dependent changes in [Ca2+]i and 6-keto-PGF1 alpha synthesis were compared. Bradykinin (BK, 10(-6) M), angiotensin (Ang) II (10(-7) M) as well as ionomycin (10(-6) M) induced simultaneous increases in [Ca2+]i and 6-keto-PGF1 alpha production. An inhibitor against prostaglandin synthesis, acetylsalicylic acid (ASA, 10(-6) M) abolished BK-induced 6-keto-PGF1 alpha synthesis, whereas ASA did not affect the increase in [Ca2+]i. BK-induced increases in [Ca2+]i and 6-keto-PGF1 alpha production occurred in a dose-dependent manner and the half-maximal response was observed at the same concentration of BK (10(-7) M). These results indicate that an increase in [Ca2+]i is closely associated with BK as well as AngII-induced prostacyclin synthesis. It is suggested that an increase in [Ca2+]i plays a prior role in prostacyclin synthesis. Thus, an interaction between phospholipase A2 (prostaglandin synthesis) and phospholipase C (inositol trisphosphate-Ca2+ mobilization) is suggested.     

Smooth muscle alpha-2 adrenoceptors mediate vasoconstrictor responses to exogenous norepinephrine and to sympathetic stimulation to a greater extent in spontaneously hypertensive than in Wistar Kyoto rat tail arteries

The alpha adrenoceptor-mediated vasoconstriction in isolated perfused tail arteries from spontaneously hypertensive (SHR) and age matched Wistar Kyoto (WKY) normotensive rats has been examined. Responses induced by periarterial field stimulation, exogenous norepinephrine or the selective alpha-1 adrenoceptor agonist methoxamine were preferentially antagonized by prazosin in both SHR or WKY tail arteries. However, in SHR only, the alpha-2 adrenoceptor antagonist idazoxan (RX 781094) at low concentrations, significantly antagonized responses to periarterial field stimulation and to exogenous norepinephrine. Except at rather high concentrations, idazoxan was inactive as an antagonist of responses induced by methoxamine. The alpha-1 adrenoceptor blocking agent prazosin was a very potent antagonist of the responses induced by periarterial field stimulation and by methoxamine. These results indicate that alpha-2 adrenoceptors predominate in both SHR and WKY tail arteries, but a significant subpopulation of smooth muscle alpha-2 adrenoceptors is present in tail arteries of SHR but not of WKY rats. In contrast to WKY normotensive rats, postjunctional alpha-2 adrenoceptors may also be involved in the vasoconstrictor responses to sympathetic nerve stimulation in tail arteries of SHR.