Stimulation-evoked Overflow Of Tritium Research Articles

No effect of pertussis toxin on peripheral prejunctional α2‐adrenoceptor‐mediated responses and on endothelium‐dependent relaxations in the rat

1. We have investigated the effects of pertussis toxin treatment on a variety of peripheral tissues in the rat. 2. Incubation with pertussis toxin (1 microgram ml-1) in vitro failed to alter the negative inotropic actions of acetylcholine in rat left atria. 3. Pretreatment with pertussis toxin (6 micrograms kg-1, i.v., 3-4 days) abolished the negative inotropic actions of acetylcholine in rat left atria. 4. Pretreatment with pertussis toxin (40 micrograms kg-1, i.v., 3-4 days) failed to alter the prejunctional inhibitory actions of the alpha 2-adrenoceptor agonist xylazine, either in terms of the isometric contraction to a single stimulus in rat vas deferens or in terms of stimulation-evoked overflow of tritium in atria pre-incubated with [3H]-noradrenaline. 5. Pretreatment with pertussis toxin (6 micrograms kg-1, i.v., 3-4 days) failed to affect, and pertussis toxin (40 micrograms kg-1, i.v., 3-4 days) potentiated endothelium-dependent relaxations of rat aorta to histamine and acetylcholine. 6. It seems unlikely that peripheral prejunctional actions of alpha 2-adrenoceptor agonists or endothelium-dependent relaxations of rat aorta involve pertussis toxin-sensitive G proteins.

Evidence for β2-Adrenoceptor-Mediated Facilitation of 3H-Noradrenaline Release from Isolated Guinea Pig Papillary Muscles

The effects of beta-adrenoceptor agonists and antagonists on field-stimulated release of radioactivity from superfused guinea pig papillary muscles preincubated with 3H-noradrenaline were studied. Stimulation-evoked overflow of tritium was abolished in the absence of Ca2+ or the presence of tetrodotoxin. Isoprenaline (1 mumol/L) caused a slight facilitation of evoked overflow, whereas phentolamine (1 mumol/L) exerted a strong facilitatory action. However, when phentolamine (1 mumol/L) was present throughout superfusion, isoprenaline and the selective beta 2-adrenoceptor agonist, zinterol, caused concentration-dependent increases (half-maximal effects at 1 nmol/L). The effects of the agonists were inversely related to stimulation frequency. Furthermore, the concentration-response curve of isoprenaline was shifted to the right by the selective beta 2-adrenoceptor antagonist, ICI 118,551, but not by the selective beta 1-adrenoceptor antagonist, ICI 89,406. Schild-plot analysis revealed competitive antagonism and a pA2 value of 9.04 for ICI 118,551. Both ICI 118,551 and ICI 89,406, as well as beta-adrenoceptor antagonists with intrinsic sympathomimetic activity (pindolol and celiprolol; 1 mumol/L), had no effect on stimulation-evoked overflow of tritium (phentolamine present). It is concluded that guinea pig papillary muscles are endowed with prejunctional beta 2 adrenoceptors facilitating impulse-evoked noradrenaline release. The facilitation is markedly promoted by blockade of prejunctional alpha adrenoceptors.

Estimation ofpA2 values at presynaptic ?2-autoreceptors in rabbit and rat brain cortex in the absence of autoinhibition

An attempt was made to determine pA2 values of antagonists at the presynaptic, release-inhibiting alpha 2-autoreceptors of rabbit and rat brain cortex under conditions when there was very little released noradrenaline in the autoreceptor biophase and, hence, pA2 values were not distorted by endogenous autoinhibition. Cortex slices were preincubated with 3H-noradrenaline and then superfused and stimulated by trains of 4 pulses delivered at 100 Hz or, in a few cases, by trains of 36 pulses at 3 Hz. The alpha-adrenoceptor agonists clonidine, noradrenaline, and alpha-methylnoradrenaline concentration-dependently decreased the stimulation-evoked overflow of tritium. The alpha-adrenoceptor antagonists yohimbine, rauwolscine and idazoxan did not increase the overflow of tritium elicited by 4 pulses/100 Hz in rabbit brain slices and increased it only slightly in rat brain slices. In contrast, the antagonists increased markedly the overflow at 36 pulses/3 Hz. All antagonists caused parallel shifts to the right of the concentration-response curves of clonidine, noradrenaline, and alpha-methylnoradrenaline. pA2 values were calculated either from linear regression of log [agonist concentration ratio - 1] on log [antagonist concentration] or from sigmoid curve fitting. The slopes of the linear regression lines were close to unity, and the pA2 values calculated by the two methods agreed well. There was no consistent preferential antagonism of any antagonist to any agonist. pA2 values determined with stimulation by 4 pulses/100 Hz were by 0.53-0.80 log units higher than those determined with stimulation by 36 pulses/3 Hz.(ABSTRACT TRUNCATED AT 250 WORDS)

Presynaptic alpha 2-adrenoceptor, opioid kappa-receptor and adenosine A1-receptor interactions on noradrenaline release in rabbit brain cortex.

The interaction of presynaptic, release-inhibiting alpha 2-adrenoceptors, opioid kappa-receptors and adenosine A1-receptors was studied in slices of the occipito-parietal cortex of the rabbit. The slices were preincubated with 3H-noradrenaline and then superfused and stimulated electrically twice for 2 min each (S1, S2). The stimulation-evoked overflow of tritium was taken to reflect action potential-evoked release of noradrenaline. One of two release-modulating compounds to be examined for interaction was kept in the medium throughout superfusion, the other one was added before S2. In many experiments, the stimulation parameters were adjusted (frequency 0.5-7 Hz; voltage drop 2-5 V/cm) in order to obtain similar reference release (S1) values despite the presence of the first release-modulating compound. The selective kappa-receptor agonist ethylketocyclazocine (EK) attenuated markedly the release-inhibiting effects of the alpha 2-adrenoceptor-selective agonists clonidine and alpha-methylnoradrenaline as well as the release-facilitating effect of the alpha 2-adrenoceptor-selective antagonist yohimbine. The attenuation occurred both when the parameters of electrical stimulation were kept constant and when they were adjusted to obtain similar S1 release values. The selective A1-receptor agonist R-N6-phenylisopropyladenosine (PIA) also attenuated the effects of clonidine and yohimbine. Conversely, clonidine attenuated and yohimbine enhanced the release-inhibiting effect of PIA. Yohimbine also enhanced the release-facilitating effect of the adenosine receptor antagonist 8-phenyltheophylline. Again, these changes occurred both at constant stimulation parameters and when stimulation parameters were adjusted.(ABSTRACT TRUNCATED AT 250 WORDS)

Protein kinase C and presynaptic modulation of acetylcholine release in rabbit hippocampus.

1. The involvement of protein kinase C in the presynaptic modulation of stimulated acetylcholine release was investigated in rabbit hippocampus. 2. Slices of the rabbit hippocampus, labelled with [3H]-acetylcholine, were superfused with medium and stimulated electrically during superfusion. 3. The protein kinase C activating phorbol ester 4 beta-phorbol 12,13-dibutyrate (4 beta-PDB) enhanced the electrically evoked tritium overflow in a concentration-dependent manner. Its biologically inactive 4 alpha-isomer was without any effect on transmitter release. 4. The protein kinase C inhibitor polymyxin B decreased the stimulation-evoked tritium overflow and counteracted the enhancement of release caused by 4 beta-PDB. 5. The stimulation-evoked tritium overflow was facilitated when the muscarine receptor antagonist atropine was present. The effects of both atropine and 4 beta-PDB, given in combination, were additive. 6. The net inhibition of the evoked tritium overflow caused by the muscarine receptor agonists carbachol and oxotremorine was similar, irrespective of whether 4 beta-PDB was present or not. 7. Similar results to those for muscarine autoreceptor-mediated inhibition, were obtained for inhibition of the stimulated tritium overflow caused by the adenosine receptor agonist (-)-N6-(R-phenylisopropyl)-adenosine ((-)-PIA) and the opioid receptor agonist ethylketocyclazocine (EKC). The net inhibition of both agonists was independent of the presence of the phorbol ester. 8. The above results provide further evidence for participation of a presynaptically located protein kinase C in the modulation of acetylcholine release. However, the modulatory mechanisms which are coupled to presynaptic receptors and mediate inhibition of release seem not to be directly affected by protein kinase C.

Leucine-enkephalin- and neuropeptide Y-modulation of [ 3H]noradrenaline release in the oviduct of mature and juvenile rabbits

1. The effects of leucine-enkephalin and neuropeptide Y (NPY) on [ 3H]noradrenaline release induced by electrical field stimulation were studied in the isthmic part of the oviduct of juvenile and mature rabbits. 2. [ 3H]noradrenaline and total tritium overflow in the presence of cocaine, corticosterone and hyoscine were determined by liquid scintillation spectrometry. 3. Tritium overflow evoked by electrical stimulation (1 or 4 Hz, 1 msec) was calcium dependent. [ 3H]noradrenaline content (measured by ion exchange chromatography) accounted for 85% of the total tritium overflow. 4. Leucine-enkephalin (1 μM) in the presence of the peptidase inhibitor bacitracin reduced the stimulation-evoked tritium overflow in mature rabbits by 26.1 ± 1.6% and in juvenile rabbits by 11.9 ± 1.9%. Naloxone (1 μM) antagonized the effect of leucine-enkephalin. 5. NPY (0.2 μM) reduced the evoked tritium overflow in mature rabbits by 23.4 ± 2.4% and in juvenile rabbits by 17.2 ± 4.3%. 6. It is concluded that leucine-enkephalin and NPY inhibited [ 3H]noradrenaline release in rabbit oviduct and the modulatory effect of leucine-enkephalin depends on maturity while NPY modulation is a more independent system.

Facilitation of serotonin (5-HT) release in the rat brain cortex by cAMP and probable inhibition of adenylate cyclase in 5-HT nerve terminals by presynaptic alpha 2-adrenoceptors.

Stimulation-evoked tritium overflow was examined in superfused rat brain cortex slices (stimulus: electrical impulses; 3 Hz) and synaptosomes (stimulus: potassium 12 mmol/l) preincubated with 3H-5-HT. 1. In slices and synaptosomes, the evoked 3H overflow was facilitated by forskolin and 8-Br-cAMP, but was not affected by AH 21-132 (an inhibitor of cAMP phosphodiesterase; cis-6-(p-acetamidophenyl)-1,2,3,4,4a,10b-hexahydro-8,9-dimethoxy-2-methylbenzo [c] [1,6]-naphthyridine). In the presence of AH 21-132, the facilitatory effect of forskolin on evoked overflow was increased. 2. In slices, AH 21-132 or combined administration of forskolin plus AH 21-132 did not change the percentage of basal or evoked 3H overflow represented by unmetabolized 3H-serotonin (about 30% and 60%, respectively). 3. In slices, cocaine or 6-nitroquipazine, an inhibitor of serotonin uptake, did not influence the increase in evoked overflow produced by forskolin plus AH-21-132. Forskolin plus AH 21-132 did not alter the inhibitory effect of serotonin (examined in the presence of 6-nitroquipazine) and the facilitatory effect of metitepin (a serotonin receptor antagonist) on evoked 3H overflow, but considerably decreased the inhibitory effect of clonidine or B-HT 920 (2-amino-6-allyl-5,6,7,8-tetrahydro-4H-thiazolo-[5,4-d]-azepine). The present results suggest that the serotoninergic nerve terminals in the rat brain cortex are endowed with an adenylate cyclase, which is negatively coupled to the presynaptic alpha 2-adrenoceptors, but is not linked to the presynaptic autoreceptors.

The serotonin (5-HT) autoreceptor in the hippocampus of the rabbit: Role of 5-HT biophase concentration

Slices of hippocampus from the rabbit were preincubated with [ 3H]serotonin ([ 3H]5-HT), then superfused continuously and twice stimulated electrically. The stimulation-evoked overflow of tritium was inhibited by the 5-HT autoreceptor ligands 5-carboxamido-tryptamine (5-COHT), 5-HT, 5-methoxy- N, N-dimethyl-tryptamine (5-MeOMT), (±)-8-hydroxy-2-(di- n-propylamino)tetralin (8-OH-DPAT), methysergide and (±)-cyanopindolol in a concentration-dependent manner. These effects were competitively inhibited by the 5-HT autoreceptor antagonists, metitepin and metergoline. (±)-Cyanopindolol also reduced the evoked release of 5-HT from slices of cortex from the rat. The inhibitor of the uptake of 5-HT, 6-nitroquipazine diminished the autoreceptor-mediated depression of release of 5-HT. In cortex tissue from the rat, 6-nitroquipazine reversed the decreased release of 5-HT, due to (±)-cyanopindolol, to a facilitation. The disinhibition of the release of 5-HT by autoreceptor antagonists was further enhanced by 6-nitroquipazine. Non-linear regression analysis of concentration-response curves for 5-COHT yielded the following pK d of endogenous 5-HT at the autoreceptor: 7.753 ± 0.116. This value corresponds to the p K d of 5-HT at the 5-HT 1B binding site. The 5-HT biophase concentration at the autoreceptor of 10 8.220 ± 0.132M was markedly enhanced by 6-nitroquipazine (10 ∂M) to 10 -7476 ± 0.102M. It is concluded that the 5-HT autoreceptor belongs to the 5-HT 1B subtype of receptor; the corresponding 5-HT biophase concentration can be estimated quantitatively; 8-OH-DPAT decreased the evoked release of 5-HT through both 5-HT autoreceptors and α 2-heteroreceptors and (±)-cyanopindolol acts as partial agonist at the 5-HT autoreceptor.

Facilitation of Noradrenaline Release by Gallamine in the Rat Salivary Gland

The effect of gallamine on spontaneous and stimulation-evoked overflow of tritium was studied in the submandibular gland of the rat. The gland was perfused retrogradely and labeled with3H-noradrenaline. The stimulation-evoked (1 Hz for 60 s) overflow of tritium was facilitated by increasing concentrations of gallamine (0.3–20 mM). None of the concentrations of gallamine increased the spontaneous overflow of the tritium. The facilitatory effect of gallamine was observed in 0.3 to 5 mM calcium medium; the maximum facilitation was observed at the normal concentration of calcium (2.5 mM). The facilitatory effect of gallamine was inversely related to the frequency of stimulation (10-fold facilitation at 1 Hz and 3-fold at 10 Hz).

N-ethylmaleimide (NEM) diminishes alpha 2-adrenoceptor mediated effects on noradrenaline release.

The effect of N-ethylmaleimide (NEM), which has been shown to abolish rather selectively inhibition of adenylate cyclase, on the alpha 2-adrenoceptor-mediated modulation of noradrenaline release was studied. Slices of the rabbit hippocampus were loaded with 3H-noradrenaline, superfused continuously and stimulated twice electrically. NEM (30 mumol/l) applied for 30 min enhanced both basal and stimulation-evoked tritium overflow significantly. Occupation of the receptor by the alpha 2-adrenoceptor agonist clonidine prior to and during NEM treatment did not protect the alpha 2-adrenoceptor-mediated autoinhibitory feedback system from being affected by NEM. Preincubation of the hippocampal slices with NEM was without any influence on 3H-noradrenaline uptake. The inhibitory effect of clonidine on 3H-noradrenaline release was attenuated in a non-competitive manner. In addition, the facilitatory effect of the alpha 2-adrenoceptor antagonist yohimbine on the stimulus-evoked tritium overflow was reduced. The facilitation of the evoked noradrenaline release by yohimbine or yohimbine or yohimbine and NEM converged with increasing concentrations of yohimbine, suggesting that yohimbine and NEM were acting at the same signal-transduction system. These results are compatible with the idea that NEM, by alkylating the Ni-unit of a presynaptically located adenylate cyclase, prevents the alpha 2-adrenoceptor-mediated modulation of noradrenaline release.

Effects of clonidine on the stimulation-evoked release of 3H-noradrenaline from superfused rat brain slices as a function of the biophase concentration. Temperature dependent widening of extracellular space.

The influence of clonidine on the stimulation-evoked overflow of tritium was studied in brain slices preincubated with 3H-noradrenaline. The slices were prepared from parietal cortex (Cx), nucleus anterior hypothalami (nah) and nucleus tractus solitarii (nts). After preincubation, the tissues were superfused at 23 degrees C or 37 degrees C with a medium containing the noradrenaline uptake inhibitor desipramine. Electrical field stimulation was applied using stimulation frequencies of 0.3-10 Hz. At 23 degrees C/0.3 Hz, clonidine concentration-dependently inhibited the evoked overflow of tritium in all three brain regions. In contrast, at 23 degrees C/3 Hz the inhibitory effect of the drug in the Cx was abolished and a facilitation was observed in the nah and nts. When tested at increasing frequencies of stimulation in the nts at 23 degrees C, clonidine exerted a dual action, characterized by a reduction of electrically evoked responses at frequencies below 1 Hz and a facilitation at frequencies above 1 Hz. At 37 degrees C, clonidine concentration-dependently decreased the evoked overflow in all brain regions studied, this effect being more pronounced at 0.3 Hz than at 3 Hz. The apparent lack of an effect of clonidine on the stimulation-evoked overflow of tritium in the Cx at 23 degrees C/3 Hz was turned to a facilitation when noradrenaline (0.01 mumol/l) was included in the superfusion medium. Conversely, an inhibitory effect of clonidine was seen when the uptake blocker desipramine (as well as noradrenaline) was omitted from the superfusion medium.(ABSTRACT TRUNCATED AT 250 WORDS)

Modulation of [3H]-dopamine released by different frequencies of stimulation from rabbit retina.

Rabbit retinal pieces were incubated with [3H]-dopamine and superfused with Krebs solution. Calcium-dependent tritium release was elicited twice within each experiment by electrical field stimulation (360 pulses, 2 ms, 20 mA) at frequencies of 1 Hz, 3 Hz, and 6 Hz. The evoked release of [3H]-dopamine for the first period of stimulation (S1) was 2.09 +/- 0.23% (n = 5) at 1 Hz and was of similar magnitude at all other frequencies of stimulation employed. The D2-dopamine receptor agonist, LY 171555 (quinpirole HCl; 0.01-1 microM) added to the superfusion medium before the second period of stimulation, inhibited the calcium-dependent release of [3H]-dopamine in a concentration-dependent manner, and was more potent the lower the stimulation frequency. The isomer of quinpirole, LY 181990 (0.01-1 microM) did not inhibit the stimulation-evoked overflow of tritium, regardless of concentration or stimulation frequency. The stereoisomers of the D2-dopamine receptor antagonist sulpiride (0.01-3 microM) increased the calcium-dependent release of [3H]-dopamine in a concentration-dependent manner, being more potent the higher the frequency of stimulation. S-sulpiride was more potent than R-sulpiride at all stimulation frequencies. The inhibitory effect of quinpirole was stereoselectively antagonized by sulpiride, but not by LY 181990. The alpha-adrenoceptor antagonist phentolamine (1 microM) did not modify the quinpirole-induced inhibition of [3H]-dopamine release. When the synaptic concentration of dopamine was increased by the presence of the dopamine uptake inhibitor nomifensine (3 microM), the potency of the agonist quinpirole in inhibiting the release of [3H]-dopamine elicited by field stimulation at 1 Hz (180 pulses) was decreased. In contrast, the potency of S-sulpiride in enhancing the evoked release of [3H]-dopamine was increased when nomifensine was present in the superfusion medium. Picomolar concentrations of the hormone melatonin (0.1-10 nM) inhibited the calcium-dependent release of [3H]-dopamine from rabbit retina with the same potency regardless of the frequency of stimulation applied (1 Hz, 3 Hz or 6 Hz). The potency of D2-dopamine receptor agonists and antagonists in modifying [3H]-dopamine release from rabbit retina appears to depend on the synaptic concentration of dopamine which is altered by the frequency of stimulation, and by the dopamine uptake inhibitor nomifensine.(ABSTRACT TRUNCATED AT 400 WORDS)

Autoreceptors and alpha 2-adrenoceptors at the serotonergic axons of rabbit brain cortex.

Slices of the rabbit occipito-parietal cortex were preincubated with 3H-serotonin and then superfused and stimulated electrically (2 min at 3 Hz). In the absence of drugs, the stimulation-evoked overflow of tritium was approximately 3% of the tritium content of the tissue. Unlabelled serotonin and 5-carboxamido-tryptamine, when administered in the presence of 6-nitroquipazine, reduced the evoked overflow of tritium. Their effects were antagonized by metitepin (apparent pA2 value 8.1) and (+/-)-cyanopindolol (apparent pA2 value 6.4). Metitepin, but not cyanopindolol, increased evoked tritium overflow; the effect of metitepin was greater in the presence than in the absence of nitroquipazine. The evoked overflow of tritium was also depressed by clonidine, an effect antagonized by idazoxan (apparent pA2 value 7.0) but not by prazosin. Phenylephrine caused a decrease only at high concentrations that simultaneously accelerated basal tritium efflux. Prazosin and idazoxan did not change evoked tritium overflow, and phentolamine increased it significantly only when administered in the presence of (+)-oxaprotiline. Rauwolscine produced an inhibition that was prevented by metitepin. It is concluded that the serotonergic axons of the rabbit occipitoparietal cortex possess presynaptic, release-inhibiting serotonin autoreceptors and alpha 2-adrenoceptors. The receptors appear to receive an input of endogenous serotonin and, to a lesser extent, noradrenaline, under the conditions of these in vitro experiments.

Facilitation of noradrenaline release by gallamine in the rat salivary gland.

The effect of gallamine on spontaneous and stimulation-evoked overflow of tritium was studied in the submandibular gland of the rat. The gland was perfused retrogradely and labeled with 3H-noradrenaline. The stimulation-evoked (1 Hz for 60 s) overflow of tritium was facilitated by increasing concentrations of gallamine (0.3-20 mM). None of the concentrations of gallamine increased the spontaneous overflow of the tritium. The facilitatory effect of gallamine was observed in 0.3 to 5 mM calcium medium; the maximum facilitation was observed at the normal concentration of calcium (2.5 mM). The facilitatory effect of gallamine was inversely related to the frequency of stimulation (10-fold facilitation at 1 Hz and 3-fold at 10 Hz). Stimulation of the salivary gland by a single pulse (1 ms duration) in the normal medium did not evoke an overflow of tritium; however, the same stimulus produced a marked increase in the overflow in the presence of gallamine. The facilitatory action of gallamine on the release of sympathetic transmitter is ascribed to the enhanced availability of calcium ions to the secretory process resulting from blockade of potassium conductance during nerve activity.

An investigation of age-related changes in pre- and postjunctional α-adrenoceptors in human saphenous vein

The responsiveness of prejunctional α 1-, and postjunctional α 2-adrenoceptors was examined in human isolated saphenous veins from male patients in the age range 37–70. There was no age-related alteration in the prejunctional potency of the α 2-adrenoceptors agonist xylazine for inhibiting the stimulation-evoked overflow of tritium in tissues preincubated with [ 3H]noradrenaline. The α 2-adrenoceptor antagonist yohimbine (0.01–1 μM) and the α 1-adrenoceptor antagonist prazosin (0.1–1 μM) signifcantly reduced stimulation-evoked contractions in a concentration-dependent manner. There was no significant age-related correlation for the potency of prazosin but there was a significant negative correlation between the potency of yohimbine and age (r = 0.70, n = 11, P < 0.05), i.e. the potency of yohimbine decreased with increasing age. The decreased postjunctional potency of yohimbine may reflect a loss of α 2-adrenoceptors with increasing age.

Evidence for neuro-effector transmission through postjunctional alpha 2-adrenoceptors in human saphenous vein.

The effects of the alpha 1-adrenoceptor antagonist prazosin and the alpha 2-adrenoceptor antagonist yohimbine were examined against stimulation-evoked contractions in human isolated saphenous veins. The concentration of yohimbine producing 30% inhibition of stimulation-evoked contractions (IC30) was 13.2 nM, whereas the IC30 of prazosin was greater than 250 nM. The inhibition of stimulation-evoked contractions by yohimbine was not prejunctionally mediated since yohimbine (0.01-0.1 microM) significantly potentiated the stimulation-evoked overflow of tritium in tissues pre-incubated with [3H]-noradrenaline. The high potency of yohimbine and the low potency of prazosin indicate that neuro-effector transmission in human saphenous vein is mediated predominantly by postjunctional alpha 2-adrenoceptors.

Modulation of hippocampal serotonin (5-HT) release by endogenous adenosine

Slices of rabbit hippocampus were preincubated with [ 3H]serotonin then superfused continuously and stimulated twice electrically. The stimulation-evoked overflow of tritium was Ca 2+-dependent, tetrodotoxin-sensitive and subject to modulation by serotonin autoreceptors. It was decreased by various adenosine receptor agonists in an order of potency that was typical for A 1 − (R i − ) receptors: N 6-cyclohexyladenosine > (−)N 6-phenylisopropyladenosine >5′-N-ethylcarboxamideadenosine> (+)N 6-phenylisopropyladenosine = adenosine.The effects of the agonists were antagonized by 8-phenyltheophylline. The hypothesis that endogenous adenosine influences hippocampal serotonin release is supported by the following findings: both the adenosine receptor antagonists (theophylline or 8-phenyltheophylline (10 μM, each)) and the enzyme adenosine deaminase (10 μg/ml) increased, whereas R-E 244 (3 μM), an inhibitor of adenosine uptake, significantly decreased the evoked tritium overflow. When 8-phenyltheophylline was present throughout superfusion the effects of both R-E 244 and exogenous adenosine deaminase were abolished. It is concluded that serotonin release in the rabbit hippocampus is depressed by endogenous adenosine via A 1 − (R i − ) receptors.

Role of cAMP for regulation of impulse-evoked noradrenaline release from the rabbit pulmonary artery and its possible relationship to presynaptic ACTH receptors.

Strips of the rabbit pulmonary artery preincubated with 3H-noradrenaline were superfused with physiological salt solution containing cocaine, corticosterone and propranolol. Basal tritium efflux and electrically evoked tritium overflow were determined. The basal efflux of tritium was not affected by forskolin 0.01-10 mumol/l, 8-Br-cAMP and dibutyryl-cAMP 10-330 mumol/l, or the phosphodiesterase inhibitors rolipram 1-10 mumol/l and AH 21-132 l mumol/l; it was increased by AH 21-132 10-100 mumol/l. Forskolin concentration-dependently increased the evoked 3H overflow, and this effect was not attenuated by omission of cocaine. The facilitatory effect of forskolin was more pronounced at 0.66 Hz than at 2 Hz. Rolipram, AH 21-132, 8-Br-cAMP or dibutyryl-cAMP also produced a concentration-dependent increase in evoked 3H overflow (8-Br-cAMP was more effective than dibutyryl-cAMP in this respect). Except for the highest concentration investigated, AH 21-132 was more effective in facilitating evoked overflow than in increasing basal efflux. Forskolin, AH 21-132 or 8-Br-cAMP did not alter the percentages of 3H-noradrenaline and 3H-metabolites contained in basal tritium efflux or in stimulation-evoked tritium overflow. When a combination of AH 21-132 plus 8-Br-cAMP or AH 21-132 plus forskolin was administered, the facilitatory effect on evoked tritium overflow was more pronounced than with the single compounds alone. ACTH1-24 also facilitated the evoked tritium overflow. Combined exposure to ACTH1-24 plus forskolin, ACTH1-24 plus AH 21-132 or ACTH1-24 plus forskolin plus AH 21-132 resulted in a clearly more pronounced increase in evoked tritium overflow than exposure to the single compounds alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Alpha 2-adrenoceptor antagonism and other pharmacological antagonist properties of some substituted benzoquinolizines and yohimbine in vitro.

Comparison of pA2 values for antagonism of clonidine induced inhibition of the electrically evoked contraction of the rat isolated vas deferens (alpha 2-adrenoceptor) and antagonism of contractions to methoxamine on the rat isolated anococcygeus (alpha 1-adrenoceptor) showed a group of substituted benzoquinolizines (Wy 25309, 26392 and 26703) to be more potent and more selective alpha 2-adrenoceptor antagonists than yohimbine. The benzoquinolizines and yohimbine enhanced stimulation-evoked overflow of tritium from rabbit isolated pulmonary arteries preloaded with [3H]-noradrenaline, as expected for alpha 2-adrenoceptor antagonists. In contrast to the results on the rat vas deferens, yohimbine was more potent than the benzoquinolizines. At higher concentrations all the alpha-adrenoceptor antagonists reduced the stimulation-evoked contraction of the pulmonary artery. The benzoquinolizines were competitive antagonists of 5-hydroxytryptamine on the rat isolated ileum. Wy 25309 showed only weak activity (pA2 = 5.21) whereas Wy 26703 was more potent (pA2 = 7.25). Yohimbine was a potent antagonist of 5-hydroxytryptamine. Wy 26703 was the only compound to have histamine antagonist effects in the guinea pig isolated ileum and to antagonise the chronotropic effect of isoprenaline on the isolated atria of the guinea pig and in both instances activity was weak (pA2 values 5.3 and 5.5 respectively). Yohimbine reduced the spontaneous beating of the atria at 3 X 10(-6) M. No compound at 10(-5) M antagonised acetylcholine on the guinea pig ileum. These novel substituted benzoquinolizines should be useful experimental compounds for the study of alpha 2-adrenoceptor mediated responses.

Inhibition by GABA, baclofen and gabapentin of dopamine release from rabbit caudate nucleus: Are there common or different sites of action?

Slices of rabbit caudate nucleus were preincubated with [ 3H]dopamine then superfused and stimulated electrically. Baclofen, GABA and gabapentin 10 −4 and 10 −3 mol/l reduced the stimulation-evoked overflow of tritium in a similar manner. The effects of the substances were reinvestigated while one of them was present in a high concentration throughout the superfusion. These interaction experiments showed that baclofen and GABA have a common presynaptic site of action which is different from that of gabapentin.