Frequency-Dependence of 3H-Noradrenaline Release from Rabbit Pulmonary Artery: Effect of α-Adrenoceptor Antagonists and Inhibitors of Transmitter Inactivation

Abstract: The aim of this study was to examine the modulating role of presynaptic α₂-adrenoceptors on transmitter release from vascular sympathetic neurones. This was done by examining the influence of removal of inactivation pathways on the effect of α-adrenoceptor antagonists on the release of transmitter from noradrenergic neurones. The rabbit main pulmonary artery preloaded with ³H-noradrenaline (³H-NA) was used. The artery was stimulated with 300 pulses at various frequencies (1, 3, 10 and 30 Hz). Pargyline (3 × 10^?4M) increased the stimulation-evoked ³H-overflow at 1 and 3 Hz and decreased it at 30 Hz. U-0521 (3′,4′-dihydroxy-2-methylpropiophenone; 3 × 10^?6M) enhanced the overflow at 1 Hz and had no effect at 3–30 Hz. Corticosterone (4 × 10^?5M) did not alter the stimulation-evoked ³H-overflow at 1–30 Hz. Cocaine (3 × 10^?6M) enhanced the ³H-overflow slightly at 1–30 Hz. At 3 × 10^?5M, cocaine enhanced ³H-overflow at 1 Hz and reduced it at 30 Hz. Neither corticosterone (4 × 10^?5M) nor propranolol (10^?7M) modified this effect of cocaine. Propranolol (10^?7M) alone decreased the ³H-overflow at 30 Hz and had no effect at 1–10 Hz. Phenoxybenzamine (10^?6M) and chlorpromazine (3 × 10^?6M) potentiated the stimulation-evoked ³H-overflow at 1–30 Hz. Phentolamine (10^?6M) decreased the ³H-overflow at 1 Hz and enhanced it at 3–30 Hz. Rauwolscine (10^?6M) enhanced the stimulation-evoked ³H-overflow maximally at 10 Hz and had no effect at 1 and 30 Hz. Cocaine (3 × 10^?5M), cocaine (3 × 10^?5M) + corticosterone (4 × 10^?5M), pargyline (3 × 10^?4M) and U-0521 (3 × 10^?6M), but not corticosterone (4 × 10^?5M), increased the rauwolscine-induced enhancement at 1 and 3 Hz, but had no influence at 10 and 30 Hz. It is concluded that at a low frequency (1 Hz) of nerve stimulation, inhibition of either neuronal uptake, extraneuronal uptake, monoamine oxidase (MAO) or catechol-O-methyltransferase (COMT) causes an enhancement in transmitter overflow that is due to removal of inactivation pathways. The enhancement is not mediated via facilitatory β-adrenoceptors. At higher frequencies (3–30 Hz) inhibition of either neuronal uptake or MAO causes a sufficient amount of transmitter in the neuromuscular gap to activate the presynaptic α-adrenoceptor mediated negative feed-back system causing a decrease in transmitter release. α- Adrenoceptor antagonists enhance the stimulation-evoked release of ³H-NA only when the junctional gap concentration of transmitter is sufficient to trigger the presynaptic α-adrenoceptor auto-inhibitory system.     

Inhibition of adrenergic neurotransmission by prostaglandin E1 (PGE1) in the rabbit ear artery.

Isolated central ear arteries of rabbits were perfused with Krebs solution at a constant flow rate of 2–7 ml/min. Pressure changes to periarterial nerve stimulation (1, 3, 10 and 30 Hz; 5–32 pulses) and to noradrenaline were measured in the presence and absence of prostaglandin E₁ (PGE₁). This substance in concentrations of 7 × 10^?9– 4.5 × 10^?7 produced a dose-dependent inhibition of constrictor responses to nerve stimulation. The degree of inhibitory action of PGE₁ became progressively less as the frequency or length of stimulation was increased. Prostaglandin E₁ (1.1 × 10^?7 M) failed to affect the constrictor responses to noradrenaline and it markedly reduced the stimulation-evoked release of radioactivity from arteries incubated with [³]noradrenaline. The results indicate that PGE₁ inhibits the vasoconstrictor responses of rabbit ear artery to nerve stimulation by reducing the release of the adrenergic transmitter.     

Dual Effect of the Muscarinic Agonist McN-A-343 on Vascular Neuroeffector Transmission

Abstract: The site and mechanism of action of McN-A-343 (4-m-chlorophenylcarbamoyloxy)-2-butynyltrime-thylammonium chloride) on sympathetic neuroeffector transmission in the rabbit isolated pulmonary artery was studied. Low concentrations (10^?6 — 3 × 10^?5 M) of McN-A-343 and cocaine enhanced (up to 210 and 236%, respectively) the contractions evoked by electrical-field stimulation, while higher concentrations (10^?4 — 3 × 10^?4 M) inhibited them. McN-A-343 (10^?4 M) caused an initial transitory potentiation (222% of control) of the stimulation-evoked contractions followed by an inhibition. In the presence of cocaine (3 × 10^?5 M), the potentiation caused by McN-A-343 was prolonged and the secondary inhibitory phase was thus abolished. Physostigmine (10^?5 — 10^?4 M), hexamethonium (10^?5 M), atropine (3 × 10^?7 M), suprofen (10^?5 M), and 4-aminopyridine did not alter the effect of McN-A-343 (10^?4 M). Cocaine (3 × 10^?5 M)and(+)-amphetamine (10^?5 M) reversed the McN-A-343-evoked block, while they did not alter the inhibition caused by tetracaine (3.2 × 10^?5 M). Atropine (3 × 10^?7 M) had no effect on the McN-A-343-induced block, while 4-aminopyridine (10^?4 M) caused a partial and transitory reversal. On the aorta McN-A-343 (10^?4 M) did not alter the contractile concentration-response curve of (—)-noradrenaline (10^?9 — 3 × 10^?4 M), while that of serotonin (10^?8 — 3 × 10^?5 M) was moved to the right in a competitive manner. McN-A-343 (10^?4 M) did not alter the contractions evoked by noradrenaline (10^?7 M) during the period corresponding to the stimulation-evoked enhancement and subsequent inhibition. McN-A-343 (10^?4 M) slightly antagonized the contractions caused by tyramine (10^?6 — 10^?3 M) and carbachol (10^?6 — 10^?3 M). It is concluded that McN-A-343 enhances stimulation-evoked transmitter release by a presynaptic facilitatory action mediated via receptors localized on the outer surface of adrenergic neurones and to a lesser extent by inhibition of noradrenaline re-uptake. The enhancement does not involve presynaptic nicotine or muscarine receptors. Furthermore, McN-A-343 inhibits transmitter release by acting as an adrenergic neurone blocking agent at an intraneuronal site. The inhibition does not involve presynaptic muscarine inhibitory receptors and is prostaglandin-independent.     

Dual Inhibitory Action of ATP on Adrenergic Neuroeffector Transmission in Rabbit Pulmonary Artery

The aim of this study was to determine whether the inhibitory action of ATP on sympathetic neuroeffector transmission in the isolated pulmonary artery is due to ATP itself or one of its dephosphorylated breakdown products, ADP, AMP or adenosine. Furthermore, the mechanism of the inhibitory action was investigated. ATP (10^?6?3 × 10^?4 M), the degradation-resistant ATP-analogue, β, γ-methylene-5′-triphosphate (10^?5?3 × 10^?4 M), ADP (10^?6?3 × 10^?4 M), AMP (10^?5?3 × 10^?4 M), adenosine (10^?5?3 × 10^?4 M) and 2-chloroadenosine (10^?7?3 × 10^?4 M) reduced the contractions evoked by field-stimulation. This was also the case for prostaglandin E₂ (3 × 10^?9?3 × 10^?7 M), while prostaglandin F_2α(1.4 × 10^?8 M) slightly augmented the neurogenic response. The time course of the inhibitory effect of purinergic compounds on the stimulation evoked contractions was studied. In the case of ATP and ADP the inhibition was biphasic: an initial marked block (1 min. after drug addition) which in the continued presence of either compound recovered partially 10 min. later and then remained almost constant for another 90 min. The other purinergic agents caused a monophasic reduction. In the presence of indomethacin (5 × 10^?5 M), ATP and ADP also reduced the neurogenic contractions in a monophasic manner. Indomethacin did not alter the β,γ-methylene-5′-triphosphate-induced inhibition. Dilazep (3 × 10^?6 M) plus deoxycoformycin (3.6 × 10^?6 M), augmented the inhibitory effect of ATP. In contrast, theophylline (5 × 10^?5 M) did not alter the effect of ATP. The inhibitory effect of ATP (10^?4 M) on stimulation-evoked contractions was inversely proportional to the extracellular Ca²⁺ concentration (0.3–5.2 mM) and to frequency of stimulation (3–15 Hz). These results suggest that ATP initially causes a presynaptic inhibition of noradrenaline release evoked by field-stimulation. This phase I block is probably mainly due to an ADP-mediated short-lasting release of prostaglandins of the E type. The continuous inhibition (phase II) is probably due to ATP and its metabolites, possibly mainly adenosine. The phase II inhibition may possibly involve a decreased entry of Ca²⁺ into adrenergic nerve terminals during depolarization.     

A pre-junctional action of 5-hydroxytryptamine and methysergide on noradrenergic nerves in dog isolated saphenous vein

Electrical stimulation (2 Hz for 2 min) of dog isolated saphenous vein strips pre-incubated with tritiated noradrenaline increased the overflow of tritium of which about 80% was noradrenaline. 5-Hydroxytryptamine (5-HT; 1·0 × 10^?9-1·0 × 10^?7 mol litre^?1) and methysergide (3·0 × 10^?8-3·0 × 10^?8 mol litre^?1) inhibited the induced overflow of total tritium by a maximum of 78 ± 4% and 47 ± 7% respectively (mean ± s.e. mean, n = 6 for each). Methysergide was about 30 times less potent than 5-HT and the maximum inhibition obtained was less than with 5-HT. Both compounds inhibited electrically-induced contractions and overflow of tritiated noradrenaline. Their inhibitory actions on tritium overflow were little affected by phentolamine (1·0 × 10^?8 mol litre^?1) or cyproheptadine (1·0 × 10^?8 mol litre^?1), nor was the inhibitory effect of methysergide on electrically induced contractions antagonized by atropine, mepyramine, cimetidine or propranolol. The findings suggest that the prejunctional inhibitory effect of methysergide may be mediated via stimulation of a 5-HT receptor which, unlike the D-receptor, is not blocked by cyproheptadine. The possibility that the pre-junctional 5-HT receptor in the dog saphenous vein is the same as the post-junctional receptor in this preparation is discussed.     

Involvement of alpha-receptors in clonidine-induced inhibition of transmitter release from central monoamine neurones

Slices of rat cerebral cortex preincubated with (?)-³H-noradrenaline or ³H-5-hydroxytryptamine were stimulated by an electrical field, and the stimulation-induced overflow of tritium was determined. (1) Clonidine diminished the stimulation-evoked tritium overflow from slices preincubated with ³H-noradrenaline. The degree of this inhibition was greater at a low than at a high frequency of stimulation. (2) A high concentration of clonidine (10^?5 M) did not antagonize the increase of the stimulation-induced overflow caused by 10^?6 M or 10^?5 M cocaine, but abolished the increase caused by 10^?7 M of phentolamine or phenoxybenzamine. In the presence of cocaine, the inhibitory effect of clonidine was reduced. (3) 10^?5 M clonidine diminished the stimulation-evoked overflow of tritium from slices preincubated with ³H-5-hydroxytryptamine. (4) It is concluded that clonidine decreases, and phentolamine and phenoxybenzamine increase, the stimulation-induced release of noradrenaline from cerebral neurones by an activation and a blockade of α-receptors, respectively. A variety of secretory cells (secreting catecholamines, acetylcholine, 5-hydroxytryptamine, insulin or renin) seem to be endowed with structures similar to α-adrenergic receptors, which can modulate the secretion process.     

The inhibitory action of pgf2α on release of [3H]noradrenaline enhanced by α2-adrenoceptor blockade,sodium-pump inhibition and 4-aminopyridine in the main pulmonary artery of the rabbit

Large concentrations of prostaglandin PGF_2α inhibited the stimulation (2 Hz) evoked release of [³H]noradrenaline from the isolated main pulmonary artery of the rabbit (the inhibition caused by 3 × 10^?5 M PGF_2α was 62%). Furthermore, PGF_2α inhibited the release evoked by stimulation when it was enhanced by different procedures. During blockade of presynaptic α₂-adrenoceptors by 3 × 10^?7 M yohimbine, which by itself enhanced the overflow of [³H]NA in response to stimulation, the inhibitory action of PGF_2α was more pronounced (78.2%). In tissue in which the Na⁺-pump was inhibited (K⁺-free treatment) where the overflow of ³H was markedly increased, PGF_2α exerted nearly equal inhibition of transmitter release to that observed in control experiments (64.3%). The inhibitory effect of PGF_2α on the stimulation-evoked release of [³H]NA was less pronounced (32.1%) in the presence of 10^?4M 4-aminopyridine (a blocker of K⁺ -channels).     

Effect of ω‐Conotoxin GVIA on Noradrenaline Release from Postganglionic Sympathetic Neurones in Rabbit Aorta

Abstract: The aim of the present work was to examine the effect of the selective N‐type calcium blocking agent ω‐conotoxin GVIA on stimulation‐evoked release of noradrenaline from sympathetic nerves in rabbit isolated aorta with regard to stimulation frequency, extracellular Ca²⁺ concentration, and transmitter uptake. Rings of rabbit isolated aorta were preloaded with (‐)‐³H‐noradrenaline and the fractional ³H‐overflow evoked by electrical‐field stimulation was determined by liquid scintillation spectrometry. ω‐Conotoxin GVIA (3×10^?10– 3×10^?8 M) did not alter the spontaneous ³H‐outflow. ω‐Conotoxin GVIA (3×10^?10– 3×10^?8 M) caused a slowly developing reduction of stimulation‐evoked ³H‐overflow at 1 and 30 Hz. The Emax for the ω‐conotoxin‐induced inhibition was less (70%) at 30 Hz than that (96%) seen at 1 Hz. Short‐term incubation with ω‐conotoxin GVIA caused a subsequent steady‐state inhibition. The inhibitory action of ω‐conotoxin GVIA (3×10^?10– 3×10^?9 M) was inversely related to the extracellular Ca²⁺ concentration (6.5×10^?4– 2.7×10^?3 M). Cocaine (3×10^?5 M) plus corticosterone (4×10^?5 M), neuronal and extraneuronal uptake inhibitors, respectively, did not alter the inhibitory effect of ω‐conotoxin GVIA (3×10^?9 M) on ³H‐overflow evoked by stimulation at a frequency of either 1 or 30 Hz. It is concluded that ω‐conotoxin GVIA acts on prejunctional N‐type calcium channels to inhibit stimulation‐evoked noradrenaline release from sympathetic neurone terminals in rabbit aorta. At a high frequency, another subtype calcium channel may possibly be involved. The action of ω‐conotoxin GVIA is independent of neuronal and extraneuronal uptake mechanisms for noradrenaline, but dependent on the amount of Ca²⁺ to be transported across the neurilemma from the extracellular space into the neurone.     

Calcium Channels Involved in Noradrenaline Release from Sympathetic Neurones in Rabbit Carotid Artery*

Abstract: Transmitter release from nerve terminals is dependent on the entry of Ca²⁺ through neuronal voltage‐gated calcium channels. In sympathetic neurones both N‐ and L‐type calcium channels are present. Potassium channel blockade increases Ca²⁺ entry into sympathetic neurones. We examined the participation of N‐ and L‐type calcium channels in the stimulation‐evoked release of noradrenaline from vascular sympathetic neurones. Rings of rabbit carotid artery were preincubated with [³H]‐noradrenaline. Electrical field stimulation was used to evoke ³H overflow. The selective N‐type calcium channel blocking agent ω‐conotoxin GVIA (single concentrations: 3×10^?10–10^?8 M) caused a slowly developing reduction of the stimulation‐evoked ³H overflow. At 3×10^?8 M, ω‐conotoxin GVIA caused an equilibrium block with a rapid (15 min.) onset. After 2 hr exposure to ω‐conotoxin the inhibition was steady (pIC₅₀ (‐log M): 9.43; E_max: 91%). The selective L‐type calcium blocking agents nifedipine (10^?7–10^?5 M) and nimodipine (10^?8–10^?5 M) had no effect on the stimulation‐evoked ³H overflow. The calcium channel opener Bay K 8644 (10^?6 M) likewise had no effect. The potassium channel blocking agent 4‐aminopyridine (10^?5–10^?3 M) enhanced the stimulation‐evoked ³H overflow up to 5 times. 4‐Aminopyridine (10^?4 M) did not alter the inhibitory effect of ω‐conotoxin GVIA (3×10^?8 M). In the presence of 4‐aminopyridine (10^?4 M), nifedipine (10^?5 M) and nimodipine (10^?6 M) enhanced the ³H overflow. We conclude that the stimulation‐evoked release of noradrenaline from sympathetic neurones in rabbit carotid artery is mediated by N‐type calcium channels and that L‐type channels are not involved even when potassium channels are blocked by 4‐aminopyridine.     

Reassessment of the blocking activity of prazosin at low and high concentrations on sympathetic neurotransmission in the isolated mesenteric vasculature of rats

1 The overflow of endogenous noradrenaline from the rat mesenteric arterial vasculature was determined along with the perfusion pressure response to periarterial nerve stimulation (PNS) (4–12 Hz). 2 The PNS-induced pressor responses were blocked by prazosin at 3x 10⁸ M but not by prazosin at 10^?6 M or the combination of prazosin at 3 × 10^?8 M and yohimbine (10^?7 M). 3 The PNS-induced overflow of endogenous noradrenaline was significantly augmented by prazosin at 10^?6 M or the combination of prazosin at 3 × 10^?8 M and yohimbine (10^?7 M) but not by prazosin at 3 × 10^?8 M alone. 4 The PNS-induced pressor response and endogenous noradrenaline overflow, in the absence of prazosin, were not significantly influenced by α, β-methylene-adenosine triphosphate (α, β-methylene ATP) (3x10^?6 M). The pressor responses to PNS which remained after prazosin at 10^?6 M were abolished by pretreatment with α, β-methylene ATP and therefore attributable to coreleased ATP. 5 The pressor responses to exogenous noradrenaline were abolished by prazosin at both 3 × 10^?8 M and 10^?6 M. 6 These results suggest that no functionally significant amount of ATP was coreleased with noradrenaline by PNS in the presence of prazosin at low concentrations which blocked only postjunctional α₁-adrenoreceptors. 7 Thus, these results indicate that the effects of prazosin at high concentrations, such as 10^?6 M, on sympathetic neurotransmission should not be ignored when considering the mechanisms of vasoconstrictor responses to PNS following high concentrations of prazosin.     

Influence of fentanyl,levorphanol and pethidine on the release of noradrenaline from rat brain cortex slices

Summary In slices of rat brain cortex preincubated with (–)-³H-noradrenaline, the influence of fentanyl, levorphanol and pethidine on the efflux of tritium was investigated. The spontaneous outflow of tritium was not changed by low, and was accelerated by high concentrations of the drugs. The overflow of tritium evoked by electrical stimulation at 3 Hz was diminished by 10^–8–10^–7 M fentanyl and by 10^–7–10^–6 M levorphanol, but was augmented by 10^–5 M levorphanol. Naloxone prevented the inhibitory effect of fentanyl and levorphanol. In contrast to fentanyl and levorphanol, pethidine did not decrease, but at concentrations of 10^–6–10^–5 M greatly increased the stimulation-induced overflow of tritium. However, the increase was abolished, and the stimulation-evoked overflow slightly reduced, after the re-uptake of noradrenaline had been blocked by cocaine. It is concluded that fentanyl, levorphanol and pethidine share with morphine the ability to inhibit the release of transmitter from cerebrocortical noradrenaline neurones evoked by nerve impulses.     

Prejunctional Muscarinic Receptor Modulation of Noradrenaline Release from Sympathetic Neurones in Rabbit Aorta *

The prejunctional muscarinic modulation of stimulation‐evoked release of ³H‐noradrenaline from sympathetic neurones in rabbit aorta was examined. The role of transmitter uptake, α‐adrenoceptor blockade, stimulation frequency and endothelium on the modulation was investigated. Rings of aorta were incubated with (‐)‐³H‐noradrenaline and subsequently subjected to electrical‐field stimulation. Fractional ³H‐overflow was determined by liquid scintillation counting. Acetylcholine (10^?8–3×10^?6 M) added cumulatively, reduced the stimulation‐evoked ³H‐overflow up to 80%. The effect of acetylcholine was the same in intact and endothelium‐free aorta. The inhibitory effect of acetylcholine was inversely related to the frequency of stimulation (1–10 Hz). The maximal inhibition (%) was 80 (1 Hz), 53 (3 Hz) and 14 (10 Hz). The inhibitory effect of acetylcholine (10^?6 M) and carbachol (10^?5 M) reached a maximum 15 min. after addition and then remained almost constant. Cocaine (3×10^?5 M) did not alter the effect of acetylcholine. Desipramine (10^?6 M) and corticosterone (4×10^?5 M) attenuated the inhibition seen with low concentrations (10^?8–10^?7 M) of acetylcholine. The acetylcholine‐induced inhibition was antagonized by desipramine. Cocaine plus corticosterone attenuated the inhibition seen with high concentrations (10^?6–3×10^?6 M) of acetylcholine. Rauwolscine (10^?6 M) enhanced the maximal inhibitory effect of acetylcholine. We conclude that the inhibitory effect of acetylcholine on ³H‐overflow from rabbit aorta preloaded with ³H‐noradrenaline is (1) inversely related to stimulation frequency; (2) independent of endothelium; (3) unaffected by neuronal and extraneuronal transmitter uptake; (4) that cocaine is not a prejunctional muscarinic antagonist; (5) that cocaine, but not desipramine, is suited as a neuronal uptake inhibitor in studies of prejunctional muscarinic receptor subtypes; and (6) and that there is an inverse interaction between prejunctional α₂‐adrenoceptors and muscarinic receptors.     

Influence of morphine and naloxone on the release of noradrenaline from rat brain cortex slices

Summary In slices of rat brain cortex preincubated with (–)-³H-noradrenaline, the influence of morphine and naloxone on the efflux of tritium was investigated. The spontaneous outflow of tritium was not changed by 10^–7–10^–5 M morphine and by 10^–6–10^–4 M naloxone, but was accelerated by 10^–4 M morphine. Electrical field stimulation augmented tritium outflow. The overflow evoked per ppulse decreased as the frequency of stimulation was increased from 0.3 to 3 Hz, but remained approximately constant when it was further increased to 10 Hz. At frequencies of 0.3, 1, and 3 Hz, but not at 10 Hz, morphine in concentrations of 10^–7–10^–5 M depressed the stimulation-induced overflow of tritium. 10^–4 M morphine did not influence the overflow induced by stimulation at 0.3 and 1 Hz and increased that evoked by stimulation at 10 Hz. Naloxone (10^–6–10^–4 M) did not change the response to stimulation. In the presence of 10^–4 M naloxone, 10^–6 M morphine did not diminish, and 10^–5 M morphine even enhanced the stimulation-induced overflow of tritium. The inhibitory effect of 10^–6 M morphine was not reduced, after tyrosine hydroxylase had been blocked by -methyltyrosine-methylester. It is concluded that morphine through an action on specific opiate receptors inhibits the release of transmitter from cerebrocortical noradrenergic neurones evoked by nerve impulses. By an action unrelated to opiate receptors, morphine at high concentrations increases the stimulation-induced overflow of noradrenaline, presumably by inhibiting its re-uptake into nerve endings.     

Facilitation of noradrenaline release by gallamine in the rat salivary gland

Summary 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).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.     

The influence of uptake2 on the inhibition by unlabelled noradrenaline of the stimulation-evoked overflow of 3H-noradrenaline in rabbit aorta with regard to surface of amine entry

The release by electrical field stimulation of 3H-noradrenaline from the adrenergic nerve endings in the rabbit aorta was studied in a special double-chambered organ bath. Independently of the frequency (1-10 Hz) and the number of pulses used (300-3,000 pulses), only 10-20% of the stimulation-evoked overflow of tritium (total evoked overflow) left the aortic wall via the intimal surface (intimal evoked overflow). The corresponding percentage value for the basal efflux was twice that for the evoked overflow, thus indicating that part of the radioactivity in the basal efflux originated from an extraneuronal compartment. The radioactivity in the total evoked overflow (in response to stimulation at 10 Hz) originated from at least two compartments (compartment I and II) with half times for efflux of about 2 and 6 min, respectively. In the intimal evoked overflow, compartment II (but not compartment I) was involved. When uptake1 was inhibited, 70% of the radioactivity in the intimal evoked overflow (stimulation at 1 Hz) consisted of metabolites, while unchanged amine prevailed by far in the total and adventitial evoked overflow, respectively. Additional inhibition of uptake2 thus had a striking effect only on the composition of the radioactivity in the intimal evoked overflow. The intimal surface was exposed to unlabelled noradrenaline in order to inhibit the evoked overflow of tritium (stimulation at 1 Hz; uptake1 inhibited). When uptake2 was inhibited additionally, the dose-response curve for the inhibitory effect of noradrenaline was shifted to the left by a factor of 4.(ABSTRACT TRUNCATED AT 250 WORDS)     

Interactions between tetraethylammonium and methohexitone at the chick neuromuscular junction: Experiments with the moving fluid electrode technique

The effect of methohexitone on the depolarization and contracture responses produced by tetraethylammonium (TEA), acetylcholine (ACh) and repetitive indirect stimulation were investigated, using the moving fluid electrode technique, in the chick biventer cervicis (BVC) nerve muscle preparation. TEA (4.8 × 10^?4?4.8 × 10^?2 M) produced contracture and depolarization responses which were concn-dependent. These responses were potentiated by methohexitone (8.8 × 10^?5 M). The mean ED₅₀S for the contracture responses in the control Krebs solution and with methohexitone were (mean ± S.E.M.) 6.5 ± 0.03 × 10^?3 M and 1.3 ± 0.04 × 10^?3 M (N = 6) respectively. The mean ED₅₀S for the depolarizations were (mean ± S.E.M.) 5.9 ± 0.1 × 10^?3 and 1.5 ± 0.06 × 10^?3 M (N = 6) respectively. ACh (5.5 × 10^?6?1.1 × 10^?2 M) produced contracture and depolarization responses which were concn-dependent. These responses were reduced by methohexitone (8.8 × 10^?5 M). The mean ED₅₀S for the contracture responses in the control Krebs solution and with methohexitone were (mean ± S.E.M. 2.4 ± 0.21 × 10^?4 and 2.3 ± 0.1 × 10^?3 M (N = 6) respectively. The mean (± S.E.M.) ED₅₀S for the depolarizations were 8.4 ± 0.33 × 10^?4 and 3.7 ± 0.14 × 10^?3 M (N = 6), respectively. Repetitive indirect stimulation, at 1–20 Hz, produced contraction and depolarization responses which were frequency-dependent. These responses were slightly potentiated by methohexitone (8.8 × 10^?5 M). The mean (± S.E.M.) frequency₅₀S for the contractions produced in the control Krebs solution and with methohexitone were 9.2 ± 0.1 and 8.5 ± 0.2 Hz (N = 6) respectively. The mean frequency₅₀S for the depolarizations were (mean ± S.E.M.) 7.2 ± 0.1 and 5.8 ± 0.19 Hz (N = 6) respectively. It is concluded that TEA may have a direct post-synaptic action, in addition to releasing ACh from the presynaptic nerve terminals. TEA produces more contracture tension than does ACh for a given level of membrane depolarization. Methohexitone, non-competitively, reduces the responses produced by applied ACh whereas it potentiates those produced by TEA and repetitive nerve stimulation.     

1-methyl substituent and stereochemical effects of 2-phenylcyclopropylamines on the inhibition of rat brain mitochondrial monoamine oxidase A and B

(E)-2-Phenylcyclopropylamine ((E)-TCP), (Z)-2-phenylcyclopropylamine ((Z)-TCP), (E)-1-methyl-2-phenylcyclopropylamine ((E)-MTCP), and (Z)-1-methyl-2-phenylcyclopropylamine ((Z)-MTCP) were synthesized and used to determine to what extent 1-methyl substitution and stereochemistry of 2-phenylcyclopropylamines affect inhibition of monoamine oxidase (MAO). Inhibition of rat brain mitochondrial MAO-A and B by the compounds were measured using serotonin and benzylamine as the substrate, respectively and IC₅₀ values obtained with 95% confidence limits by the method of computation. For the inhibition of MAO-A, (E)-MTCP (IC₅₀=6.2×10^?8M) was found to be 37 times more potent than (Z)-MTCP (IC₅₀=2.3×10^?6 M), whereas the activity of (E)-TCP (IC₅₀=2.9×10^?7 M) was slightly less than that of (Z)-TCP (IC₅₀=2.3×10^?7 M). Similarly, for the inhibition of MAO-B, (E)-MTCP (IC₅₀=6.3×10^?8 M) was 7 times more potent than (Z)-MTCP (IC₅₀=4.7×10^?7 M) and (E)-TCP (IC₅₀=7.8×10^?8 M), 0.6 times as potent as (Z)-TCP (IC₅₀=4.4×10^?8 M). The results suggested that while without 1-methyl group, potency of a (Z)-isomer was comparable to that of (E)-isomer, the methyl group in its (Z)-position was very unfavorable to the inhibition of MAO and that in its (E)-position, the methyl group contributed positively to the potency as found by the fact that (E)-MTCP was 1–5 times more potent than (E)-TCP. In view of the selective inhibition of MAO-A or B, all compounds elicited 4–10 times higher preference for the inhibition of MAO-B over MAO-A and 1-methyl substitution as well as the stereochemical factors did not significantly influence the selectivity.     

Facilitation of noradrenaline release from rat brain slices by \-adrenoceptors

The present study examined whether stimulation of \-adrenoceptors facilitated noradrenaline release in the rat brain. Electrical stimulation-evoked overflow of tritium from rat cerebral cortical, hypothalamic and hippocampal slices labelled with ³H-noradrenaline was measured during superfusion for 100 min. Tissue slices were electrically simulated (1 Hz, 20 mA, 2 ms, 2 min), at 20(S1) and 70(S2) min after the onset of superfusion. The nonselective \-adrenoceptor agonist isoproterenol (0.1 – 10 nM) enhanced stimulation-evoked overflow of tritium from slices of cerebral cortex, hypothalamus and hippocampus in a concentration-dependent manner; mean S2/S1 ratios with 10 nM isoproterenol were 161 +- 11%, 142 +- 15% and 143 - 12% of control, respectively, in the three brain regions. The facilitatory effect of isoproterenol in cerebral cortical slices was antagonized by propranolol (50 nM), a nonselective \sb-adrenoceptor antagonist, and by the \sb₁- and \sb₂-selective adrenoceptor antagonists ICI 89,406 (1 nM) and ICI 118,551(1 nM), respectively. The \sb₁- and \sb₂-selective adrenoceptor agonists prenalterol and albuterol (0.1 \2- 10 nM), respectively, also increased stimulation-evoked overflow of tritium from cerebral cortical slices; these effects were antagonized by \sb-adrenoceptor antagonists. These findings suggest that stimulation of \sb-adrenoceptors enhance noradrenaline release from rat cerebral cortical, hypothalamic and hippocampal slices; this release mechanism appears to involve both \sb₁- and \sb₂-adrenoceptor subtypes. These facilitating presynaptic receptors may be involved in mediating the antidepressant-like behavioral effects of \sb₂-adrenoceptor agonists.     

No evidence for reverse trans-synaptic regulation of neuronal uptake by beta-adrenoceptors in kitten atria

Kitten atria incubated with [³H]noradrenaline, 1.18 × 10^?7 M for 10 min or 3 × 10^?9 M for 30 min, actively accumulated the amine. Final tissue tritium concentrations were 2–4-fold and 8–12-fold higher, respectively, than those of the incubation fluid. Uptake was consistently greater in right than in left atria. The β₁-adrenoceptor antagonist, practolol, 10^?6 or 10^?5 M, and the β₁- and β₂-adrenoceptor antagonist, propranolol, 5 × 10^?8, 5 × 10^?7or 5 × 10^?6 M, did not affect noradrenaline uptake. Reverse trans-synaptic regulation of neuronal noradrenaline uptake by β-adrenoceptors therefore does not appear to operate in kitten atria as has been reported for rat atria and other tissues.     

Effect of Cocaine and Corticosterone on the Metabolism of 3H-Noradrenaline Released from Rabbit Isolated Aorta and Adventitia

Abstract: The effect of cocaine and corticosterone (neuronal and extraneuronal uptake inhibitors, respectively) on the metabolism of ³H-noradrenaline (³H-NA) released spontaneuosly or by electrical-field stimulation was studied in the rabbit isolated aorta and tunica adventitia preloaded with ³H-NA. The spontaneous outflow of tritium from untreated aorta consisted of ³H-NA (25%); ³H-3,4-dihydroxyphenylglycol (³H-DOPEG; 24%); ³H-3,4-dihydroxymandelic acid (³H-DOMA; 8%); ³H-O-methylated plus deaminated compounds (³H-OMDA; 45%); and ³H-normetanephrine (³H-NMN; 1%). The percentage distribution of ³H-NA and its ³H-metabolites in this outflow was not altered by either cocaine (3 × 10^?5 M), corticosterone (4 × 10^?5 M), or cocaine (3 × 10^?5 M) + corticosterone (4 × 10^?5 M). The spontaneous ³H-outflow from untreated tunica adventitia did not differ from that of aorta. Cocaine (3 × 10^?5 M) + corticosterone (4 × 10^?5 M) did not alter the composition of ³H-NA and its ³H-metabolites in the spontaneous outflow from adventitia. The stimulation-evoked ³H-overflow from untreated rabbit aorta preloaded with ³H-NA consisted of ³H-NA (40%); ³H-DOPEG (15%); ³H-DOMA (2%); ³H-OMDA (50%); and ³H-NMN (3%). Cocaine (3 × 10^?5 M) decreased the percentage recovered as ³H-DOPEG, increased ³H-NMN and had no effect on the other ³H-metabolites. Corticosterone (4 × 10^?5 M) had no effect on the percentage distribution of ³H-NA and its ³H-metabolites. Cocaine (3 × 10^?5 M) + corticosterone (4 × 10^?5 M) increased ³H-NA, decreased ³H-DOPEG and had no effect on the percentage distribution of the other ³H-metabolites. In the case of adventitia, cocaine (3 × 10^?5 M) + corticosterone (4 × 10^?5 M) only decreased the percentage recovered as ³H-DOPEG, without any significant effect on ³H-NA and its other ³H-metabolites. It is concluded that inhibition of neuronal plus extraneuronal uptake of H-NA by cocaine plus corticosterone, respectively does not fully prevent metabolism of ³H-NA released either spontaneously or by electrical-field stimulation.