Modulation of the release of [3H]norepinephrine from the base and body of the rat urinary bladder by endogenous adrenergic and cholinergic mechanisms

Modulation of [3H]NE release was studied in rat urinary bladder strips prelabeled with [3H]NE. [3H]NE uptake occurred in strips from the bladder base and body, but was very prominent in the base where the noradrenergic innervation is most dense. Electrical field stimulation markedly increased [3H]NE outflow from the superfused tissue. The quantity of [3H]NE release was approximately equal during three consecutive periods of stimulation. Activation of presynaptic muscarinic receptors by 1.0 microM oxotremorine reduced [3H]NE release to 46% of the control. Atropine (1 microM) blocked the effect of oxotremorine and increased the release to 147% of predrug control levels. Activation of presynaptic alpha-2 adrenoceptors by 1 microM clonidine reduced [3H]NE release to 55% of control. Yohimbine blocked the action of clonidine and increased the release to 148% of control. The release of [3H]NE from the bladder base and body was increased by both 1 microM atropine (to 167% and 174% of control, respectively) and 1 microM yohimbine (to 286% and 425% of control, respectively). Atropine and yohimbine administered in combination had similar facilitatory effects as when administered alone. We conclude that the release of [3H]NE from adrenergic nerve endings in electrically stimulated bladder strips is modulated via endogenous transmitters acting on both muscarinic and alpha-2 adrenergic presynaptic receptors and that the latter provide the most prominent control.     

Neurogenic cholinergic prejunctional inhibition of sympathetic beta adrenergic relaxation in the canine coronary artery

Electrical stimulation of isolated canine coronary arteries causes release of norepinephrine and subsequent relaxation by activation of beta adrenoceptors. The purpose of the present study was to determine if this beta adrenergic relaxation was influenced by a concomitant release of acetylcholine. Rings of epicardial coronary arteries of the dog were studied in organ chambers filled with physiological salt solution. The tetrodotoxin-sensitive, beta adrenergically mediated relaxation induced by electrical stimulation was studied during contractions evoked by prostaglandin F2 alpha. The relaxation to low-frequency stimulation was inhibited and augmented, respectively, by acetylcholine and atropine, suggesting that release of acetylcholine may modulate the beta adrenergic response to sympathetic nerve stimulation. The relaxation caused by high-frequency stimulation was not affected by atropine or removal of the endothelium, indicating that endogenously released acetylcholine does not act directly on the smooth muscle or initiate an endothelium-dependent vasodilator response. In superfused strips of coronary artery preincubated in [3H]norepinephrine, acetylcholine depressed the stimulated overflow of [3H]norepinephrine, indicating prejunctional cholinergic receptors on adrenergic nerve endings. Atropine augmented the overflow, suggesting that endogenous acetylcholine, released during stimulation, inhibits the release of norepinephrine. These observations suggest that prejunctional inhibition of norepinephrine release, which limits the sympathetic beta adrenergic relaxation of the smooth muscle, is the primary neurogenic cholinergic effect in canine epicardial coronary arteries.     

Interaction of the tricyclic antidepressant amitriptyline with prejunctional alpha and muscarinic receptors in the dog saphenous vein

Amitriptyline can cause tachycardia and arrhythmia associated with an excessive release of cardiac catecholamines. We have investigated its effects on norepinephrine release from adrenergic nerves by using the dog saphenous vein as a model of the sympathetic neuroeffector junction. Isolated strips of vein were mounted for isometric tension recording or incubated with [3H]norepinephrine and mounted for superfusion, tension recording and the superfusate. Amitriptyline (10(-6); 5 x 10(-6) M) increased the overflow of [3H]norepinephrine but decreased that of [3,4-3H]dihydroxyphenylglycol from electrically stimulated strips. The selective decreased in the overflow of this metabolite indicates that amitriptyline inhibits neuronal uptake. However, the increased overflow of [3H]norepinephrine caused by amitriptyline also occurred when neuronal uptake was blocked by cocaine (3 x 10(-5) M) but was abolished when prejunctional alpha receptors were blockade by phentolamine (10(-5) M). Amitriptyline attenuated the prejunctional inhibitory action of exogenous norepinephrine, this indicates that the drug interacts with prejunctional alpha receptors. Amitriptyline also antagonized the prejunctional inhibitory action of acetylcholine, both in the absence and presence of cocaine and phentolamine. These effects were not due to a nonspecific action of the drug as it did not reduce the prejunctional inhibitory effect of histamine. Thus, amitriptyline can increase the concentration of norepinephrine at the neuroeffector junction by blockade of neuronal uptake and by interacting with prejunctional alpha and muscarinic receptors. Since the cardiac adrenergic nerves also possess these receptors, the results could help to explain the cardiotoxic effects of the drug.     

Modulatory role of alpha adrenoceptors on the release of [3H]norepinephrine elicited by preganglionic stimulation of the cat superior cervical ganglion

In the isolated cat superior cervical ganglion labeled in vitro with [3H]norepinephrine ([3H]NE), the overflow of radioactivity evoked by preganglionic stimulation at 10 Hz (80 V, 2 msec duration for 5 min) was reduced to 50% of control values by the alpha adrenoceptor agonists clonidine (0.001 microM) and methoxamine (12.0 microM). The alpha adrenoceptor antagonist phenoxybenzamine (2.9 microM) produced a 2-fold increase in the overflow of [3H]NE elicited by nerve stimulation. Preincubation with drugs that reduce the neuronal uptake of norepinephrine in the isolated ganglion (8.8 microM cocaine and 0.33 microM desipramine) did not modify the release of [3H]NE by preganglionic stimulation. However, a higher concentration of desipramine (3.3 microM) produced a 4-fold increase in the overflow of tritium evoked by stimulation. As this concentration of desipramine produced a shift to the right in the concentration-response curve to methoxamine in the isolated nictitating membrane of the cat, the conclusion is drawn that a feedback mechanism mediated through presynaptic alpha adrenoceptors regulates the release of [3H]NE induced by preganglionic stimulation of the cat superior cervical ganglion. In addition, it is suggested that regulatory mechanisms for norepinephrine release by nerve stimulation are not restricted to nerve terminals but are also present in dendrites of the postganglionic adrenergic neurons.     

Interaction between serotonin uptake inhibitors and alpha-2 adrenergic heteroreceptors in the rat hypothalamus

The effectiveness of presynaptic receptor agonists to inhibit the electrically evoked release of [3H]monoamines from brain slices is attenuated in the presence of blockade of neuronal uptake for the serotonin (5-HT) and the norepinephrine (NE) systems. There is controversy, however, as to the existence of a functional link between the presynaptic receptors and the neuronal uptake carriers. An alternative hypothesis involves competition for the presynaptic receptor sites between the exogenous agonist and the released neurotransmitter. In order to examine the proposed functional interaction, we studied the alpha-2 adrenoceptor-mediated inhibition of the electrically evoked release of [3H]-5-HT from slices of the rat hypothalamus, a model in which endogenous NE does not activate the alpha-2 heteroreceptors located on 5-HT terminals. The inhibitors of 5-HT uptake, citalopram (0.01-1 microM) and paroxetine (1 microM), which by themselves did not modify [3H]-5-HT release, antagonized the inhibition of [3H]-5-HT overflow produced by UK 14.304, an alpha-2 adrenoceptor agonist. The inhibition of the electrically evoked release of [3H]-5-HT by exogenous NE (0.1-1 microM) was also attenuated in the presence of citalopram. In contrast, citalopram did not modify the electrically evoked release of [3H]-NE or the inhibition of [3H]-NE release mediated by UK 14.304. When the 5-HT autoreceptor was blocked by cyanopindolol, the inhibitory effect of UK 14.304 on [3H]-5-HT release was unaltered in the presence of citalopram.(ABSTRACT TRUNCATED AT 250 WORDS)     

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.     

Ethanol inhibits N-methyl-D-aspartate-stimulated [3H]norepinephrine release from rat cortical slices

The effects of ethanol on N-methyl-D-aspartate (NMDA)-stimulated [3H]norepinephrine (NE) release from rat cortical slices was studied. NMDA-stimulated [3H]NE release was inhibited by tetrodotoxin, Mg++ and 2-amino-5-phosphonopentanoic acid, indicating that NMDA receptors in the cortex have characteristics similar to those observed using electrophysiological studies. Ethanol (60-200 mM) decreased the release of [3H]NE evoked by 100 microM NMDA in a concentration-dependent manner (32-52% inhibition), but it did not significantly alter the basal release. The inhibitory effect of 100 mM ethanol was due to a reduction in the maximal response with no significant change in the EC50 for NMDA. Pretreatment of the slices with 100 mM ethanol up to 6 min did not alter the magnitude of inhibition. The inhibition of NMDA-stimulated [3H]NE release due to ethanol was reversible after a 13-min recovery period. The presence of ethanol did not significantly affect the IC50 for Mg++ inhibition of NMDA-stimulated [3H]NE release (23 +/- 3 microM). Glycine (10-300 microM) potentiated the release of [3H]NE stimulated by 250 microM NMDA, and 60 mM ethanol did not alter this effect of glycine. Ethanol (100 mM) inhibited the release of [3H]NE evoked by 18.9 mM KCl in the presence or absence of 2-amino-5-phosphonopentanoic acid, but had no effect on release induced by 49.1 mM KCl. Tetrodotoxin (0.3 mM) significantly decreased the release of [3H] NE evoked by 23.2 mM KCl, and 60 to 200 mM ethanol did not alter this release. These results suggest that NMDA receptors in rat cortical slices are located on nerve cell bodies.(ABSTRACT TRUNCATED AT 250 WORDS)     

Autonomic receptors in urinary tract: sex and age differences

As age and sex affect the function of the lower urinary tract, we studied the characteristics of adrenergic and cholinergic receptors in various parts of lower urinary tract smooth muscle of young (6 months) and old (4 1/2-5 years) male and female rabbits. Saturation experiments performed with [3H]prazosin, [3H]yohimbine, [3H]dihydroalprenolol and [3H]quinuclidinyl benzylate in rabbit bladder base, bladder dome and urethra indicate the presence of regional, sex- and age-related differences in the density of alpha-1, alpha-2, and beta adrenergic and muscarinic cholinergic receptors. Alpha-2 adrenergic receptor density is considerably higher in the female than in the male urethra of both age groups, whereas the higher density of beta adrenergic receptors in the female than in the male bladder base is observed only in the younger animals. The density of muscarinic receptors is higher in bladder dome than in bladder base or urethra in young rabbits of both sexes. In the old animals, the density of muscarinic receptors in bladder base increases to the level observed in bladder dome. Inhibition experiments with selective adrenergic agonists and antagonists indicate that the pharmacological profiles of alpha-2 adrenergic receptors in the urethra and beta adrenergic receptors in the bladder dome and bladder base are similar in both sexes and at both ages. Beta-2 adrenergic receptors are shown to be predominant in bladder base and bladder dome of rabbits. Parallel studies in rabbit urethra, adult rat cortex and neonatal rat lung show that the urethral alpha-2 adrenergic receptors are of the alpha-2A subtype.     

Inhibition by 5,6-dihydroxy-2-dimethylaminotetralin (M7) of noradrenergic neurotransmission in the rabbit hypothalamus: role of alpha-2 adrenoceptors and of dopamine receptors

Rabbit hypothalamic slices were prelabeled with [3H]norepinephrine and transmitter release elicited by electrical stimulation. In the presence of 10 microM cocaine and in a low Ca++ medium (0.65 mM), exposure for 8 min to exogenous dopamine (0.01-1 microM) inhibited, in a concentration-dependent manner, the electrically evoked release of [3H]norepinephrine. This inhibitory effect of dopamine on [3H]norepinephrine release was antagonized by the dopamine receptor antagonist S-sulpiride (1 microM), but remained unchanged in the presence of the alpha-2 adrenoceptor antagonists idazoxan (1 microM) or yohimbine (0.1 microM). These results indicate that, in a low Ca++ medium, exposure to dopamine decreased [3H]norepinephrine overflow in rabbit hypothalamic slices through the exclusive activation of presynaptic inhibitory dopamine receptors. M7 (5,6-dihydroxy-2-dimethylaminotetralin) is a potent agonist at central presynaptic dopamine autoreceptors and at peripheral alpha-2 adrenoceptors. Exposure to M7 in a normal Ca++ medium, inhibited in a concentration-dependent manner the electrically evoked release of [3H]norepinephrine without affecting the spontaneous outflow of radioactivity. The slope of the concentration-effect curve for these inhibitory effects of M7 was rather flat and the maximal inhibition obtained was 80%. The selective D2 receptor antagonist S-sulpiride (1 microM) failed to produce a significant shift to the right in the concentration-effect curve for the inhibitory effects of M7 on [3H]norepinephrine release. The preferential alpha-2 adrenoceptor antagonist yohimbine (0.1 microM) significantly antagonized the inhibition of [3H]norepinephrine release elicited by 0.01 microM M7, but not for higher concentrations of this aminotetraline.(ABSTRACT TRUNCATED AT 250 WORDS)     

Differential inhibition by ergonovine of norepinephrine release in the coronary artery and saphenous vein of the dog

Experiments were performed to determine the effect of ergonovine on adrenergic neurotransmission in the blood vessel wall. Strips of canine left circumflex coronary arteries and saphenous veins were incubated with [3H]norepinephrine and suspended for superfusion. Ergonovine (10(-6) M) decreased the overflow of [3H] norepinephrine to a modest degree in the coronary artery, whereas a profound inhibition of overflow was observed in the saphenous vein. Methiothepin, but not rauwolscine, reversed the inhibitory effect of ergonovine in both blood vessels. Treatment with atropine or droperidol did not alter the response to ergonovine in the coronary artery. The results demonstrate that ergonovine, by activating prejunctional serotonergic receptors, inhibits adrenergic neurotransmission to a greater extent in the saphenous vein than in the coronary artery. The differential effect of ergonovine on norepinephrine release in these vessels may provide evidence for heterogeneity in the prejunctional modulation of adrenergic neurotransmission.     

Mechanism and metabolic disposition of verapamil-evoked overflow of radioactivity from isolated atria preloaded with [3H]norepinephrine

In this study the mechanism and metabolic profile of verapamil-evoked release of radioactivity was investigated in the rat isolated atria preloaded with [3H]norepinephrine [( 3H]NE). Verapamil (10(-7) to 10(-3) M) caused a dose-related increase in the outflow (or the fractional release) of 3H. The fractional 3H-release produced by verapamil was reduced markedly in tissues which had been preloaded with [3H]NE in the presence of cocaine (10 microM) or after pretreatment of animals with reserpine (5 mg/kg i.p., 24 hr before sacrifice). Verapamil-evoked fractional 3H-release was unchanged in the presence of tetrodotoxin (5 X 10(-6) M) or in Ca++-free Krebs' medium containing 2 mM ethylene glycol bis(beta-aminoethyl ether)N,N'-tetraacetic acid. Whereas greater than 90% of tissue 3H-content consisted of unchanged [3H]NE, 60 to 75% of the spontaneous outflow and the verapamil-evoked overflow consisted of [3H]-3,4-dihydroxyphenylglycol and 2 to 10% was unchanged [3H]NE. When both cocaine (10 microM) and hydrocortisone (28 microM) (uptake-1 and uptake-2 blockers, respectively) were present, although the spontaneous outflow, as well as verapamil-evoked overflow, of radioactivity was increased, the metabolic profiles remained essentially unchanged. The addition of pargyline (10 microM), a monoamine oxidase inhibitor, in addition to the uptake-1 and uptake-2 blockers to the Krebs' solution significantly depressed both the spontaneous outflow and verapamil-evoked overflow of 3H; the verapamil-evoked overflow under this condition, however, consisted of unchanged [3H]NE (greater than 90%).(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization of alpha-1 adrenergic receptors linked to [3H]inositol metabolism in rat cerebral cortex

The properties of alpha-1 adrenergic receptors in rat cerebral cortex were examined by measuring increases in [3H]inositol metabolism in brain slices. Slices of rat cerebral cortex were incubated in the presence of 0.23 microM [3H]inositol in Krebs-Ringer-bicarbonate buffer containing 10 mM lithium chloride, and the production of water-soluble [3H]inositol phosphates was monitored after extraction and anion-exchange chromatography. Norepinephrine caused a 4- to 6-fold increase in [3H]inositol metabolism in cerebral cortical slices, and this response was blocked much more potently by the alpha-1-selective antagonist prazosin than by the alpha-2-selective antagonist yohimbine. Epinephrine and norepinephrine were both full agonists and stimulated [3H]inositol metabolism to the same extent in this system. The synthetic drugs phenylephrine and methoxamine were both partial agonists at these receptors, with intrinsic activities only 56 to 58% of epinephrine and norepinephrine. A variety of imidazoline and other partial agonists caused no measurable stimulation of [3H]inositol metabolism in this preparation. The response to norepinephrine was completely blocked by alpha adrenergic receptor antagonists with the potency order prazosin greater than BE 2254 greater than indoramin = phentolamine greater than azapetine greater than piperoxan greater than yohimbine. In the absence of calcium, basal [3H]inositol metabolism was increased, but norepinephrine caused the same 5-fold stimulation as in the presence of 2.5 mM CaCl2. The potencies of both antagonists and agonists in inhibiting or activating [3H]inositol metabolism in slices of rat cerebral cortex were highly correlated with their ability to displace the alpha-1 adrenergic receptor selective radioligand [125I]BE 2254 from specific binding sites in membrane preparations of rat cerebral cortex.(ABSTRACT TRUNCATED AT 250 WORDS)     

Epinephrine and norepinephrine enhance p-aminohippurate transport into basolateral membrane vesicles

The effect of exogenous l-norepinephrine (NE) and l-epinephrine (EP) on transmembrane transport of p-aminohippurate (PAH) was studied in rat proximal tubular basolateral membrane vesicles. A gradient of 50 mM Na+ (out greater than in) and preloading of vesicles with unlabeled PAH were utilized to promote the influx of [3H]PAH into the vesicles. At final concentrations of 1 microM, NE and EP each produced significant elevations in vesicle uptake of [3H]PAH. The enhancement of PAH transport by NE or EP was inhibited by either phentolamine (100 microM) or yohimbine (100 microM). Prazosin (100 microM) or atenolol (100 microM) were unable to inhibit the response to NE. Similarly, prazosin or propranolol (100 microM) were unable to inhibit the response to EP. Clonidine (1 microM) also produced a significant elevation of PAH uptake, an effect inhibited by both phentolamine and yohimbine. Basolateral Na+-K+-adenosine triphosphatase activity also was increased significantly by either NE or EP (1 microM). Both agonists produced significant elevations of PAH uptake into vesicles preloaded with ATP. However, in the absence of NE or EP, PAH uptake into ATP-loaded vesicles was not significantly greater than into control vesicles. It was concluded that NE and EP enhance Na+-coupled PAH transport and that this effect may be mediated by alpha-2 adrenergic receptors. Activation of Na+-K+-adenosine triphosphatase is a possible mechanism whereby adrenergic agonists may exert effects on Na+-coupled transport across the basolateral membrane.     

Effects of amiloride and 5-(N,N-hexamethylene)amiloride on dopaminergic and cholinergic neurotransmission

In order to establish the role of the Na+/H+ exchange transport on neurotransmission, we investigated the effects of amiloride and of 5-(N,N-hexamethylene)amiloride (HMA) on dopamine (DA) and acetylcholine (ACh) release and on receptor-mediated modulation of DA and ACh release. Superfused rabbit striatal slices prelabeled with [3H]DA and [14C]choline were stimulated electrically in the presence and absence of several concentrations of these agents. Amiloride (3-10 microM) and HMA (0.3-10 microM) reduced the basal efflux and the stimulation evoked overflow of total 3H and of [3H]-3,4-dihydroxyphenylacetic acid and inhibited monoamine oxidase activity. The inhibition of stimulation evoked overflow of total 3H was blocked by pretreatment with nomifensine but not by sulpiride. Amiloride had no effect on the basal efflux and the stimulation evoked overflow of ACh or it did modify apomorphine-induced inhibition of DA and ACh release. However, at 3 to 10 microM, HMA enhanced the basal efflux of 3H; this effect was not prevented either by uptake inhibition with nomifensine or by low extracellular calcium. These results suggest that amiloride-sensitive Na+ transport and the amiloride and HMA-sensitive Na+/H+ antiporter play no role on the secretion of DA and ACh, or on the mechanisms by which activation of pre- and postsynaptic DA receptors lead to inhibition of neurotransmitter release. Amiloride- and HMA-induced monoamine oxidase inhibition accounts for the effects of amiloride and HMA on DA efflux and overflow. The guanidine moiety present in the amiloride and HMA molecules is most likely responsible for these effects.     

Alpha-2 adrenoceptors modulate [3H]dopamine release from rabbit retina

In the rabbit retina, preloaded in vitro with [3H]dopamine, calcium-dependent release of radioactivity was elicited by a 1-min period of field stimulation at 3 Hz (20 mA, 2 msec). In the presence of the catecholamine uptake inhibitor nomifensine (30 microM), unlabeled catecholamines (0.01-3 microM), namely, dopamine, norepinephrine and epinephrine, inhibited in a concentration-dependent manner the field stimulation-evoked release of [3H]dopamine from the retina. The concentrations of dopamine, norepinephrine or epinephrine which inhibited by 50% the release of [3H]dopamine (IC50) were 0.30, 0.25 and 0.25 microM, respectively. In the presence of 30 microM nomifensine, S-sulpiride (1 microM) significantly increased the calcium-dependent release of [3H]dopamine, suggesting that this dopamine antagonist blocks a receptor tonically activated by endogenous dopamine in the rabbit retina. In contrast, the alpha receptor antagonist phentolamine (1 microM) alone did not affect the release of [3H]dopamine from the retina. The inhibitory effect of norepinephrine and epinephrine on [3H]dopamine overflow was not modified by S-sulpiride which, on the contrary, selectively antagonized the effect of exogenous dopamine. Phentolamine (1 microM) competitively antagonized the inhibitory effect of norepinephrine and epinephrine on [3H]dopamine release, suggesting that these catecholamines activate alpha adrenoceptors in retina. In the absence of nomifensine, the selective alpha-2 agonist clonidine (IC50 = 0.056 microM) inhibited the stimulation-evoked release of [3H]dopamine from retina, whereas the alpha agonist methoxamine was without effect. The inhibitory effect of clonidine was antagonized by yohimbine (1 microM), but not prazosin, suggesting that the release modulating alpha receptors of the retina are of the alpha-2 subtype.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of d-amphetamine and dopamine synthesis inhibitors on dopamine and acetylcholine neurotransmission in the striatum. I. Release in the absence of vesicular transmitter stores

The release of endogenous dopamine (DA) elicited by electrical stimulation and by d-amphetamine (AMPH) from superfused striatal slices of reserpine-pretreated rabbits was examined. Although reserpine pretreatment reduced tissue DA levels by greater than 95%, the basal efflux of DA and the DA metabolite dihydroxyphenylacetic acid (DOPAC) was slightly greater than that observed in untreated slices. DOPAC constituted the large majority of the basal efflux of endogenous compounds. No overflow of endogenous compounds was evoked by electrical stimulation (3 Hz, 3 min) after reserpine pretreatment. Superfusion with alpha-methyl-p-tyrosine (100 microM) abolished the efflux of endogenous DA and DOPAC. AMPH (0.3-10 microM) produced a concentration-dependent increase in the basal efflux of endogenous DA and a concomitant decrease in endogenous DOPAC efflux. The total efflux of endogenous compounds (DA + DOPAC) tended to be decreased by AMPH. No electrically evoked overflow of endogenous compounds was observed in the presence of AMPH. The increase in synaptic DA produced by AMPH was reflected by a concentration-dependent reduction in the electrically evoked overflow of [3H]acetylcholine (ACh). The ability of AMPH to increase DA efflux and inhibit [3H]ACh release was blocked by inhibition of DA synthesis with alpha-methyl-p-tyrosine (100 microM) or by blockade of the DA neuronal uptake carrier with nomifensine (NOM) (10 microM) and was potentiated by inhibition of monoamine oxidase with pargyline (10 microM). NOM also blocked partially the ability of AMPH to reduce endogenous DOPAC efflux. NOM increased the basal efflux of endogenous DA and inhibited electrically evoked [3H]ACh release but these effects were quantitatively much less than those produced by AMPH. NOM had no effect on DOPAC efflux. Pargyline had little effect on endogenous DA efflux or electrically evoked [3H]ACh release but abolished DOPAC efflux and increased tissue DA levels measured at the end of superfusion. When given in combination, NOM and pargyline produced a similar degree of inhibition of [3H]ACh release as AMPH, although the increase in DA efflux produced by this drug combination was less than that produced by AMPH. These results suggest that in the absence of vesicular transmitter stores (reserpine-pretreatment): synthesis provides a continuous supply of DA which is metabolized rapidly within the neuron and is lost as DOPAC; AMPH facilitates the synthesis-dependent efflux of extravesicular DA probably by an accelerated exchange diffusion mechanism.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of clinically effective concentrations of halothane on adrenergic and cholinergic synapses in rat brain in vitro

The effects of clinically relevant concentrations of halothane on various synaptic events in rat brain were examined in vitro. Slices or membranes prepared from various brain regions were equilibrated with 1.25% halothane at 37 degrees C, resulting in tissue concentrations greater than those found in brains of completely anesthetized animals (greater than 40 nmol/mg of lipid). Under these conditions, no effects of halothane were observed on agonist or antagonist binding to alpha-1 adrenergic, beta adrenergic, alpha-2 adrenergic or muscarinic cholinergic receptors, or on the ability of 5'-guanylylimidodiphosphate to reduce agonist binding affinity. This concentration of halothane also had no effect on norepinephrine-stimulated cyclic AMP accumulation, or inositol phosphate accumulation in response to norepinephrine, carbachol, histamine or 5-hydroxytryptamine. Halothane did reduce potassium-evoked release of [3H]norepinephrine from slices of rat cerebral cortex although it did not affect potassium-evoked release of [3H]acetylcholine. These results suggest that halothane may be relatively specific in its synaptic actions and may affect depolarization-evoked release of norepinephrine at concentrations achieved clinically.     

Prejunctional alpha adrenoceptor and angiotensin receptor function in isolated human, monkey and dog arteries

Transmural electrical stimulation (2-20 Hz) produced a frequency-dependent contraction in dog mesenteric, monkey mesenteric and human gastroepiploic arterial strips, which was abolished by tetrodotoxin and suppressed by phentolamine. Treatment with yohimbine (10(-9) and 10(-8) M) potentiated the response to nerve stimulation dose-dependently in the dog arteries, but rather attenuated the response of the primate arteries. Yohimbine (10(-8) M) attenuated the contraction caused by exogenous norepinephrine in the dog and monkey arteries. Angiotensin (ANG) II (2 X 10(-10) M) and ANG I (10(-9) M) potentiated the response to transmural stimulation in the dog and monkey arteries, whereas the response to norepinephrine was unaffected. The ANG I-induced potentiation was suppressed by treatment with ANG converting enzyme inhibitors. 3H-Overflow evoked by transmural stimulation in tissues prelabeled with [3H] norepinephrine was increased by yohimbine in the superfused dog arteries but was not increased significantly in the monkey arteries. The overflow was increased significantly by ANG II in the dog and monkey arteries. It may be concluded that prejunctional alpha-2 adrenoceptors mediating the inhibition of transmitter release do not function significantly in human gastroepiploic and monkey mesenteric arteries, and postjunctional alpha-2 receptors are involved partly in contractions of the monkey arteries due to adrenergic nerve stimulation. ANG II appears to be synthesized from ANG I via ANG converting enzyme in the primate arteries; the octapeptide potentiates the contraction caused by adrenergic nerve stimulation, possibly due to prejunctional ANG receptor activation and increased norepinephrine release.     

Effects of naftidrofuryl on adrenergic nerves, endothelium and smooth muscle in isolated canine blood vessels

Experiments were designed to determine the effects of the vasoactive drug naftidrofuryl on vascular smooth muscle, endothelial cells and adrenergic nerves in isolated canine blood vessels. Naftidrofuryl inhibited contractions of basilar arteries (in a decreasing order of potency), evoked by 5-hydroxytryptamine greater than KCl = anoxia (in rings with endothelium) greater than prostaglandin F2 alpha = uridine-5'-triphosphate. Naftidrofuryl antagonized competitively the contractions evoked by 5-hydroxytryptamine in the femoral artery and the saphenous vein. Naftidrofuryl caused the release of an endothelium-derived relaxing factor(s) from the endothelium of femoral arteries. The compound depressed contractions of saphenous veins evoked by electrical stimulation of the adrenergic nerve endings, but not those caused by the indirect sympathomimetic amine tyramine or exogenous norepinephrine. In saphenous veins incubated previously with [3H]norepinephrine, the drug inhibited the contractions and the release of transmitter evoked by electrical stimulation. Thus, naftidrofuryl acts at different levels in the blood vessel wall to cause: release of endothelium-derived relaxing factor(s); inhibition of S2-serotonergic receptors on vascular smooth muscle; prejunctional inhibition of adrenergic neurotransmission; and nonselective inhibition of the contractile process in vascular smooth muscle, which is particularly pronounced in cerebral arteries.     

Beta2-adrenoceptor-mediated prejunctional facilitation and postjunctional inhibition of sympathetic neuroeffector transmission in the guinea pig vas deferens.

This study examines the role of prejunctional and postjunctional beta-adrenoceptors in the modulation of sympathetic cotransmission in the guinea pig vas deferens. The prejunctional involvement of beta-adrenoceptors was evaluated by testing the effects of several agonists and antagonists on the nerve stimulation-evoked overflow of ATP and norepinephrine (NE) from the "in vitro" vas deferens. The nonsubtype-selective beta-adrenoceptor agonist isoproterenol and the beta2-subtype-selective agonist clenbuterol increased, to a similar degree, the overflow of ATP and NE, while the beta1-subtype-selective agonist xamoterol and the beta3-subtype-selective agonist BRL 37 344 had no effect. Pretreatment with ICI 118, 551, a beta2-subtype-selective antagonist, abolished the facilitation of cotransmitter release by isoproterenol and clenbuterol, while the beta1-subtype-selective antagonist atenolol had no effect. Activation of beta-adrenoceptors by either isoproterenol or clenbuterol, but not by xamoterol and BRL 37 344, reduced the amplitude of contractions evoked by exogenously applied ATP. Pretreatment with propranolol or ICI 118, 551, but not atenolol, prevented these inhibitory effects. Isoproterenol in lower concentrations produced dose-dependent reduction of the purinergic but not the adrenergic phase of nerve stimulation-induced contraction of the guinea pig vas deferens. When applied in concentrations greater than 1 microM, isoproterenol, but not clenbuterol, actually produced a concentration-dependent facilitation of contractions evoked by both nerve stimulation and exogenously applied ATP. Antagonists of alpha-adrenoceptors blocked these facilitatory effects. Together, these results demonstrate that beta2-adrenoceptors can influence sympathetic neuroeffector transmission both prejunctionally, where they facilitate equally well the release of sympathetic cotransmitters and postjunctionally, where they inhibit smooth muscle contractions evoked by ATP.