In vivo release of catecholamines in the locus coeruleus

To investigate the release of endogenous dopamine, noradrenaline and adrenaline in the locus coeruleus, this brain area was superfused with artificial cerebrospinal fluid (CSF) through push-pull cannulae and the release of catecholamines was determined in the superfusate radioenzymatically.Collection of superfusates in time periods of 10 min revealed that release rates of the three catecholamines fluctuated, thus pointing to the existence of ultradian rhythms with following mean periods (minutes per cycle): noradrenaline 52±4, dopamine 37±2, adrenaline 36±2. The rhythm frequency of noradrenaline was significantly lower than the frequencies of dopamine and adrenaline.When the locus coeruleus was superfused with neuroactive drugs, superfusates were collected in time periods of 3 min. Superfusion with tetrodotoxin (1 mol 1^–1) for 12 min elicited a prompt and sustained decrease (–70%) in the release rates of dopamine and adrenaline. The release rate of noradrenaline was also reduced, although to a lesser extent (–40%). Superfusion with veratridine (50 mol 1^–1) led to an immediate and very pronounced enhancement in the release rates of dopamine, noradrenaline and adrenaline. The veratridine-induced increase in catecholamine outflow was decreased strongly by simultaneous superfusion with tetrodotoxin.The findings suggest that the release of endogenous catecholamines in the locus coeruleus fluctuates according to ultradian rhythms. Changes in the release on superfusion with veratridine and tetrodotoxin demonstrate the neuronal origin of the three catecholamines. The observed differences in the release characteristics between noradrenaline on the one hand and dopamine and adrenaline on the other might indicate that noradrenaline is partly released from somatodendritic sites of the noradrenergic cell bodies in the locus coeruleus.This work was supported by the Fonds zur Förderung der wissenschaftlichen Forschung Correspondence to: N. Singewald at the above address     

Catecholamine release in the locus coeruleus is modified by experimentally induced changes in haemodynamics

Summary The involvement of catecholaminergic neurons of the locus coeruleus in central cardiovascular control was investigated in the anaesthetized cat. Push-pull cannulae were bilaterally inserted into the LC and the release of noradrenaline and dopamine was determined radioenzymatically in the superfusate. The effects of experimentally induced changes in blood volume and vascular resistance on catecholamine release in the locus coeruleus were studied.Hypervolaemia strongly inhibited the release of noradrenaline in the locus coeruleus. Intravenous infusion of noradrenaline (5 g·kg^–1·min^–1) elicited a pronounced pressor response which was also associated with a decrease in the release of noradrenaline in the locus coeruleus. Conversely, a fall of blood pressure caused by a controlled haemorrhage enhanced the release of noradrenaline. A profound fall in blood pressure caused by infusion of nitroprusside (8 g·kg^–1·min^–1) did not modify the release rate of noradrenaline. Dopamine release rate was not significantly influenced by these cardiovascular alterations.The results demonstrate that increases in blood pressure elicited by vascular constriction or hypervolaemia inhibit the release of noradrenaline in the locus coeruleus. Decreases in blood pressure elicited by hypovolaemia enhance the release of noradrenaline, but lowering blood pressure by vasodilatation is ineffective. Hence, the release of endogenous noradrenaline in the locus coeruleus is responsive to haemodynamic signals, thus supporting the suggested integrative role of the locus coeruleus in central cardiovascular control.Correspondence to N. Singewald at the above address     

In vivo effect of tramadol on locus coeruleus neurons is mediated by alpha2-adrenoceptors and modulated by serotonin

Tramadol is a centrally-acting analgesic endowed with opioid, noradrenergic and serotonergic properties. Various data suggest that, in addition to its analgesic effect, tramadol may have antidepressant and anxiolytic-like effects. This study investigates, through single-unit extracellular recording techniques, the in vivo effects of tramadol on locus coeruleus (LC) neurons and its possible effects on alpha(2)-adrenoceptors, opioid receptors and the 5-HT system. Tramadol produced a dose-dependent and complete inhibition of LC activity (ED(50)=2.1mg/kg). This inhibitory effect was prevented and reversed by the selective alpha(2)-adrenoceptor antagonist, idazoxan, but not by the opioid receptor antagonist, naloxone. The inhibition of the synthesis of 5-HT by p-chlorophenylalanine and the pre-administration of the 5-HT(1A) receptor agonist, 8-OH-DPAT at 40microg/kg, caused a significant potentiation of the tramadol effect decreasing the ED(50) by 53% and 67% respectively. Lower doses of 8-OH-DPAT, of 1 and 4microg/kg, did not significantly modify the tramadol effect. In summary, the results indicate that tramadol elicits an inhibitory effect on LC neurons in vivo through alpha(2)-adrenoceptors. Moreover, this effect is modulated by the 5-HT system and particularly by 5-HT(1A) receptors.     

The stimulatory effect of clonidine through imidazoline receptors on locus coeruleus noradrenergic neurones is mediated by excitatory amino acids and modulated by serotonin

Clonidine and other imidazoline/oxazoline drugs, such as cirazoline and rilmenidine, have been shown to stimulate the activity of noradrenergic neurones in the locus coeruleus (NA-LC) by an ₂-adrenoceptor-independent mechanism through the activation of I-imidazoline receptors. The endogenous modulation of the stimulatory effect of clonidine on NA-LC neurones was further investigated after inactivation of ₂-adrenoceptors with N-ethoxycarbonyl-2-ethoxy-1, 2-dihydroquinoline (EEDQ). In EEDQ-pretreated rats (6 mg/kg, i.p., 6h), clonidine caused a rapid and dose-dependent (320–5120 g/kg, i.v.) increase in the firing rate of NA-LC neurones (ED₅₀ = 809 g/kg, E_max = 90%). The stimulatory effect of clonidine on NA-LC neurones was completely blocked by pretreatment of rats with the excitatory amino acid receptor antagonist kynurenic acid (1–3 mol in 10–30 mol i.c.v., 2–5 min before clonidine). In contrast, the stimulatory effect of clonidine on NA-LC neurones was potentiated by pretreatment with reserpine (5 mg/kg, s.c., 18 h) (E_max increased by 63%). Pretreatment with -methyl-p-tyrosine (250 mg/kg, i.p., 24 h) did not alter the stimulatory effect of clonidine, but pretreatment with p-chloro-phenylalanine (400 mg/kg, i.p., 24 h) markedly enhanced the stimulatory effect of clonidine on NA-LC neurones (E_max increased by 139%). The present results indicate that the imidazoline receptor-mediated stimulatory effect of clonidine on NA-LC neurones is an indirect effect dependent on an excitatory amino acid pathway and modulated by an inhibitory serotonin mechanism.     

In vivo release of endogenous catecholamines in the hypothalamus

Summary The posterior hypothalamus of anaesthetized cats was superfused with a push-pull cannula and the release of the endogenous catecholamines noradrenaline, adrenaline and dopamine was determined in the superfusate. The rate of release of the three catecholamines followed an ultradian rhythm, the time interval between two adjacent phases of high rate of release being about 70 min. Pretreatment of the animals with reserpine decreased the levels of catecholamines in the hypothalamus and rest of the brain and reduced their rate of release into the superfusate. Hypothalamic superfusion with superfusing fluid of high concentration of potassium and low concentration of sodium enhanced the rate of release of noradrenaline and adrenaline; this effect was abolished when the hypothalamus was superfused with calcium-free solution. Electrical stimulation of the locus coeruleus ipsilateral to the superfused hypothalamus increased the release of noradrenaline and adrenaline, stimulation of the contralateral locus coeruleus enhanced the release of noradrenaline, adrenaline and dopamine. In both cases, the rate of release of adrenaline was enhanced to a lesser extent than the rate of release of noradrenaline. The release of noradrenaline and adrenaline was increased to a higher extent on stimulation of the ipsilateral locus coeruleus than on stimulation of the contralateral one.Part of the results was presented at the Spring Meeting of the German Pharmacological Society, Mainz, March 1978 and at the IV International Catecholamine Symposium, Asilomar, September 1978This work was supported by the Deutsche Forschungsgemeinschaft     

Noradrenaline release in rat locus coeruleus is regulated by both opioid and alpha(2) -adrenoceptors.

Extracellular fluid levels of noradrenaline (NA) in the locus coeruleus (LC) during naloxone-precipitated morphine withdrawal with pretreatment of yohimbine (1 mg kg (-1), s.c.) or clonidine (1 mg kg (-1), s.c.) were measured in rats. There was a significant increase in the NA level after the injection of naloxone (2 mg kg (-1), i.p.) in the morphine-dependent rats. Moreover, the NA levels in the LC markedly increased during the 30-60 min following the naloxone (i.p.) challenge in the morphine-dependent rats pretreated with yohimbine. In contrast, the naloxone challenge in morphine-dependent rats pretreated with clonidine notably decreased the levels of NA in the LC. Behavioral signs of withdrawal were observed following the naloxone challenge in the morphine-dependent rats pretreated with yohimbine, with minimal signs in the morphine-infused rats pretreated with clonidine, and none in the saline-infused controls. These results directly suggest that NA increased within the LC after the naloxone challenge in morphine-dependent animals pretreated with yohimbine may be, at least in part, regulated by alpha(2) -adrenoceptors in the LC.     

Pulsatile release of catecholamines in the hypothalamus of conscious rats

Summary To investigate the patterns of catecholamine release in the brain, the hypothalamus of conscious, freely moving rats was superfused through a push-pull cannula with artificial cerebrospinal fluid and the catecholamines dopamine, noradrenaline and adrenaline were determined in the superfusate radioenzymatically. Superfusate was continuously collected in time periods of 20 min for at least 20h. Dopamine, noradrenaline and adrenaline release rates fluctuated according to an ultradian rhythm with a frequency of 1 cycle/92 min (dopamine and noradrenaline) or 99 min (adrenaline). Additionally, the three catecholamines were released according to an ultradian rhythm with the following frequencies: noradrenaline and adrenaline 1 cycle/ 12 h, dopamine 1 cycle/8 h. The release rates of dopamine and adrenaline were similar during light and dark periods, while the release rate of noradrenaline in the dark period was slightly lower than that during the light period. It is concluded that in the hypothalamus of the conscious rat the release rates of dopamine, noradrenaline and adrenaline fluctuate according to two ultradian rhythms with different frequencies.This work was supported by the Deutsche Forschungsgemeinschaft and the Fonds zur Förderung der wissenschaftlichen ForschungParts of this work have been presented at the 29th Spring Meeting of the German Society of Pharmacology and Toxicology, 1988 and at the 12th Annual Meeting of the European Neuroscience Association, 1989Correspondence to H. Prast at the above address     

In vivo release of newly synthesized catecholamines from the hypothalamus by amphetamine

Summary The posterior hypothalamus of cats immobilized with gallamine was superfused through a push-pull cannula with artificial cerebrospinal fluid. Addition of³H-tyrosine into the superfusing fluid led to synthesis of³H-catecholamines which were released spontaneously. Separation of the³H-catecholamines by column chromatography or their acetylation and separation by paper chromatography revealed that both³H-noradrenaline and³H-dopamine were released. In most experiments³H-noradrenaline represented about 10 to 25% of total³H-catecholamines. Superfusion of the hypothalamus with amphetamine (1×10^–5 M) enhanced the release of total³H-catecholamines, the release of³H-noradrenaline being relatively more enhanced than that of³H-dopamine. Determination of the readioactive compounds in the hypothalamus at the end of the experiments showed that total³H-catecholamines represented 3% of³H-tyrosine. About 15% of the total³H-catecholamines were due to³H-noradrenaline and 85% to³H-dopamine.     

Stress-induced release of substance P in the locus coeruleus modulates cortical noradrenaline release

Several lines of evidence implicate the neuropeptide substance P (SP) in the modulation of emotional behavior. Interaction between SP and noradrenergic systems has been proposed to be important in the regulation of stress, depression, and anxiety mechanisms; however, most evidence so far is based on studies in unchallenged and/or anesthetized animals. Thus, by using a dual-probe microdialysis approach in freely moving animals, the aim of the present study was to investigate whether a relevant stressor can trigger the release of SP in the locus coeruleus (LC) and whether and how this response modulates noradrenaline (NA) transmission both in the LC and in the medial prefrontal cortex (mPFC), an important LC terminal region involved in emotional processing. While confirming previous reports that neurokinin 1 receptor (NK1R) antagonists activate cortical noradrenergic transmission under resting conditions, we present evidence that this interaction is opposite during stress challenge. Our results show that exposure to forced swimming considerably enhanced the release of SP and NA in the LC. Administration of a selective NK1R antagonist into the LC potentiated this NA response within the LC but abolished the stress-induced increase in NA release within the mPFC. These findings demonstrate stress-induced increase in endogenous extracellular SP levels within the LC exerting a facilitatory effect on the noradrenergic pathway to the mPFC. The attenuation of stress-induced hyperactivation of this pathway by NK1R antagonists, presumably via enhancing NA and autoinhibition in the LC, may contribute to the therapeutic efficacy of these drugs known to ameliorate symptoms of stress-related disorders.     

Inhibition of catecholamine (noradrenaline,dopamine) release in the locus coeruleus and the hypothalamus by baroreceptor activation: identification of the involved baroreceptors

We have previously shown that experimentally induced blood pressure changes modify the release rates of catecholamines in the hypothalamus and the locus coeruleus. The aim of the present investigation was to identify the peripheral baroreceptors and the centripetal pathways responsible for the changes of catecholamine release in these brain areas.In anaesthetized cats, push-pull cannulae were bilaterally inserted into the locus coeruleus and the posterior hypothalamus. The two brain areas were superfused simultaneously with artificial cerebrospinal fluid. Baroreceptor activation by phenylephrine-induced blood pressure elevation decreased the release rate of noradrenaline in the locus coeruleus and the release rates of noradrenaline and dopamine in the posterior hypothalamus. Similar effects were elicited by electrical stimulation of the central trunk of the transected vagus and aortic depressor nerves (vagus-ADN). Transection of the nerves abolished the effect of phenylephrine on the release of noradrenaline in the locus coeruleus. Nerve transections attenuated slightly the decreased release of noradrenaline elicited by phenylephrine in the posterior hypothalamus, while the reduced dopamine release rate was not influenced. The selective stimulation of baroreceptors in the carotid sinus by an inflatable catheter did not influence the release of catecholamines in the locus coeruleus, while release rates of noradrenaline and dopamine in the posterior hypothalamus were decreased.The simultaneous superfusion of locus coeruleus and hypothalamus revealed that, in both areas, noradrenaline release is inhibited by baroreceptor activation. Noradrenergic neurons of the posterior hypothalamus are inhibited by baroreceptor impulses conducted by the carotid sinus nerve and vagus-ADN, while the noradrenergic neurons of the locus coeruleus seem to respond to impulses transmitted by vagus-ADN. Furthermore, baroreceptor activation inhibits dopaminergic neurons in the hypothalamus but not in the locus coeruleus.     

In vivo release by histamine agonists and antagonists of endogenous catecholamines in the cat hypothalamus

Summary The posterior hypothalamus of anaesthetized cats was superfused through a push-pull cannula with histamine agonists and antagonists and the release of endogenous catecholamines was determined in the superfusate. Hypothalamic superfusion with histamine, 2-methylhistamine (H₁-agonist), dimaprit (H₂-agonist) or metiamide (H₂-antagonist) enhanced the release of the catecholamines dopamine, noradrenaline and adrenaline. The releasing effects of these substances depended on the presence of calcium ions. Superfusion with 2-pyridylethylamine (H₁-agonist) was virtually ineffective, while superfusion with 2-thiazolethylamine (H₁-agonist) enhanced the rate of release of noradrenaline and adrenaline without influencing the release of dopamine. Superfusion with mepyramine (H₁-antagonist) inhibited the release of noradrenaline and adrenaline without affecting the release of dopamine. Hypothalamic superfusion with a concentration of procaine which was equi-anaesthetic to that of mepyramine was ineffective. Ranitidine (H₂-antagonist) did not alter the rates of release of the catecholamines. The releasing effect of histamine was inhibited on hypothalamic superfusion with mepyramine and ranitidine. Ranitidine also inhibited the releasing effects of dimaprit and 2-methylhistamine thus indicating that the releasing action of the latter compound was mainly due to stimulation of H₂-receptors. These data suggest that blockade of H₁-receptors of the posterior hypothalamus reduces the release of noradrenaline and adrenaline, while stimulation of H₁-receptors seems to increase the rates of release of these two catecholamines. Stimulation of H₂-receptors enhances the release of all three catecholamines. Thus, dopaminergic neurones of the hypothalamus seem to possess H₂-receptors, while noradrenergic and adrenergic neurones possess H₁- and H₂-receptors.This work was supported by the Deutsche Forschungsgemeinschaft     

Serotonin release is modulated by presynaptic autoreceptors.

The existence of presynaptic autoreceptors controlling the release of 5-hydroxytryptamine (5HT) from serotonergic nerve endings was investigated utilizing superfused hypothalamic synaptosomes. Extracellular 5HT reduced the high K+-induced release of previously accumulated 3H-5HT. The central 5HT receptor blocker methiothepin counteracted the inhibitory effect of 5HT. Other 5HT antagonists (cyproheptadine, methysergide and mianserin) were inactive and may therefore act preferentially at the postsynaptic receptors.     

Dependence of serotonin release in the locus coeruleus on dorsal raphe neuronal activity

The serotonergic innervation of the locus coeruleus paetly derives from the dorsal raphe nucleus (DRN). Using the push-pull superfusion technique, we investigated whether and to what extent the release of serotonin and the extracellular concentration of its metabolite 5-hydroxyindoleacetic acid (5-HIAA) in the locus coeruleus are influenced by the neuronal activity of the DRN. In anaesthetized rats, a push-pull cannula was inserted into the locus coeruleus, which was continuously superfused with artificial cerebrospinal fluid (aCSF). Serotonin and 5-HIAA levels in the superfusate were determined by HPLC combined with electrochemical detection. Electrical stimulation (5 Hz, 300 μA, 1 ms) of the DRN for 5 min, or its chemical stimulation by microinjection of glutamate (3.5 nmol, 50 nl), led to an increased release of serotonin in the locus coeruleus and to a slight (2 mmHg) decrease in blood pressure. Superfusion of the locus coeruleus with tetrodotoxin (1 μM) abolished the increase in the release rate of serotonin evoked by electrical stimulation of the DRN, while the slight fall in blood pressure was not influenced. Thermic lesion (75 °C, 1 min) of the DRN elicited a pronounced decline in serotonin release rate within the locus coeruleus, the maximum decrease being 52%. The decrease in the release of serotonin was associated with a long-lasting rise in blood pressure. Microinjection of the serotonin neurotoxin 5,7-dihydroxytryptamine (5 μg, 250 nl) into the DRN led to an initial increase in the serotonin release rate that coincided with a short-lasting fall in blood pressure. Subsequently, the release of serotonin was permanently reduced and was associated with hypertension. Microinjection of the 5-HT_1A receptor agonist (±)-8-hydroxy-dipropylaminotetralin (8-OH-DPAT; 7.5 nmol, 50 nl) into the DRN led to a long-lasting reduction of the release rate of serotonin in the locus coeruleus. Microinjection of 8-OH-DPAT into the DRN also slightly lowered blood pressure (3 mmHg). Neither stimulations nor lesion of the DRN, nor microinjection of 8-OH-DPAT into this raphe nucleus, altered the extracellular concentration of 5-HIAA. Judging from the present biochemical results it appears that the serotonergic afferents to the locus coeruleus originate to more than 50% from cell bodies located in the DRN. The neuronal serotonin release in the locus coeruleus is modulated by 5-HT_1A receptors lying within the DRN. Changes in blood pressure and release of serotonin elicited by stimulating or lesioning the DRN point to the importance of serotonergic neurons extending between this raphe nucleus and the locus coeruleus in central cardiovascular control. Received: 5 November 1998 / Accepted: 21 February 1999     

Characterization of noradrenaline release in the locus coeruleus of freely moving awake rats by in vivo microdialysis

Rationale The origin and regulation of noradrenaline (NA) in the locus coeruleus (LC) is unknown.Objectives The neurochemical features of NA overflow (nerve impulse dependence, neurotransmitter synthesis, vesicle storage, reuptake, ₂-adrenoceptor-mediated regulation) were characterized in the LC.Methods Brain microdialysis was performed in awake rats. Dialysates were analyzed for NA.Results NA in the LC decreased via local infusion of Ca²⁺-free medium (–42±5%) or the sodium channel blocker tetrodotoxine (TTX) (–47±8%) but increased (333±40%) via KCl-induced depolarization. The tyrosine hydroxylase (TH) inhibitor -methyl-p-tyrosine (250 mg kg^–1, i.p.) and the vesicle depletory drug reserpine (5 mg kg^–1, i.p.) decreased NA. Therefore, extracellular NA in the LC satisfies the criteria for an impulse flow-dependent vesicular exocytosis of neuronal origin. Local perfusion of the ₂-adrenoceptor agonist clonidine (0.1–100 M) decreased NA (E_max=–79±5%) in the LC, whereas the opposite effect (E_max=268±53%) was observed with the _2A-adrenoceptor antagonist BRL44408 (0.1–100 M). This suggests a tonic modulation of NA release through local _2A-adrenoceptors. The selective NA reuptake inhibitor desipramine (DMI) (0.1–100 M) administered into the LC increased NA in the LC (E_max=223±40%) and simultaneously decreased NA in the cingulate cortex, confirming the modulation exerted by NA in the LC on firing activity of noradrenergic cells and on the subsequent NA release in noradrenergic terminals.Conclusion Synaptic processes underlying NA release in the LC are similar to those in noradrenergic terminal areas. NA in the LC could represent local somatodendritic release, but also the presence of neurotransmitter release from collateral axon terminals.     

Role of nitric oxide in the stress-induced release of serotonin in the locus coeruleus

Serotonergic mechanisms within the locus coeruleus (LC) are thought to be important in various functions including the stress response. In this study we investigated a possible role of nitric oxide (NO) as an intermediary messenger in the regulation of the serotonin (5-HT) neurotransmission within the LC. Using the push-pull superfusion technique coupled with HPLC and electrochemical detection, the in vivo release of 5-HT was determined in time periods of 10 min in the LC of freely moving rats. Superfusion with three different NO donors, SIN-1 (linsidomine), S-nitroso-N-penicillamine (SNAP) or 3-(2-hydroxy-2-nitroso-1-propylhydrazino)-1-propanamine (PAPANO) increased 5-HT release in the LC. Superfusion with the precursor of NO, L-arginine, for 1 h led to a sustained increase in 5-HT release. On the other hand, the NOS inhibitor N-methyl-L-arginine methyl ester (L-NAME) did not significantly change the release of 5-HT. Infusion of N-methyl-D-aspartate (NMDA) or kainic acid, as well as exposure of rats to noise stress or tail pinch increased the release of 5-HT in the LC. Superfusion with L-NAME prevented the increase in 5-HT outflow by all these procedures, while the inactive isomer D-NAME had no effect. Taken together, the results of this study suggest that the release of 5-HT in the LC is facilitated by NO. Under resting conditions inhibition of NOS does not appear to substantially influence the release of 5-HT in the LC. However, there seems to be a facilitatory nitrergic influence on serotonergic responses evoked by excitatory amino acid receptor stimulation or various stress stimuli.     

Reduced pressor responses to stimulation of the locus coeruleus after lesion of the posterior hypothalamus

Summary Electrical stimulation of the locus coeruleus or of the hypothalamic posterior area of cats under pentobarbitone anaesthesia elicited a rise of the arterial blood pressure. Electrocoagulation of the hypothalamic posterior area or its lesion by the injection of ethanol significantly diminished the pressor response to electrical stimulation of the locus coeruleus. The pressor response to electrical stimulation of the area posterior was almost completely abolished after electro-coagulation and strongly inhibited after injection of ethanol. It is suggested that adrenergic neurones ascending from the locus coeruleus to the posterior hypothalamus may be involved in the hypothalamic regulation of the arterial blood pressure.This work was supported by the Deutsche Forschungsgemeinschaft and the Stiftung Volkswagenwerk.     

Influence of catecholamines on the in vivo release of histamine in the hypothalamus.

The hypothalamus of conscious, freely moving rats was superfused with artificial CSF through push-pull cannulae and the release of endogenous histamine was determined radioenzymatically in the superfusate. Superfusion with potassium chloride enhanced the release rate of histamine. The effect of potassium chloride was abolished by alpha-fluoromethylhistidine. Noradrenaline (alpha 1- and alpha 2-agonist) and clonidine (alpha 2-agonist) decreased the release rate of histamine and inhibited the potassium-induced histamine release. In preliminary experiments, yohimbine (alpha 2-antagonist) seemed to increase the release rate of histamine in the superfusate. beta-Agonists and antagonists (isoprenaline, salbutamol, propranolol) did not influence the release of histamine.     

Blockade of the release of LH induced by pharmacological suppression of adrenergic inflow into the locus coeruleus

Previous studies have indicated that stimulation of the locus coeruleus (LC) produced inhibition of the release of LH induced by stimulation of the medial preoptic area (mPOA) in ovariectomized, estrogen-primed rats. A similar response following the application of drugs into the LC was taken as an index of activation of the LC neurons. The injection of the phenylethanolamine N-methyltransferase (PNMT) blocking-agent, 2,3- dichloromethylbenzylamine ( DCMB , 0.5 micrograms) into the LC, 2 and 1 h before starting the mPOA stimulation greatly attenuated the induced release of LH. Injection of saline had no effect. The reduced release of LH in DCMB -treated rats was restored to normal by injecting clonidine (0.5 micrograms) into the LC immediately before the mPOA stimulus started, but not by injecting phenylephrine (0.5 micrograms) or saline (0.5 microliter). The release of LH induced by mPOA stimulation was also blocked in rats in which the alpha 2-antagonist, piperoxane (0.5 micrograms) was injected into the LC but not in those injected with the alpha 1-antagonist, phenoxybenzamine (0.5 micrograms) or the beta-antagonist, propranolol (0.5 micrograms). It is concluded that adrenergic afferents into the LC tonically inhibited the activity of LC neurons and that this effect is mediated by alpha 2-adrenoceptors. Interference with the synthesis of epinephrine or blockade of alpha 2-adrenoceptors resulted in activation of LC neurons and thereby in inhibition of LH release.     

The production of reactive oxygen species in peripheral blood neutrophils is modulated by airway mucous

Neutrophils are a major source of reactive oxygen species (ROS). The role of airway mucous on ROS production is unknown. The aim of our study was to investigate the direct influence of bronchoalveolar lavage fluid (BALF) and induced sputum (IS) alone or in combination with chemical/biological stimulus on ROS production in peripheral blood neutrophils during chronic obstructive pulmonary disease (COPD). Neutrophils were isolated from peripheral blood of 47 patients with moderate COPD and 14 healthy individuals (HI). BALF/RPMI (1:1) or IS/RPMI (1:1) from COPD patients were used to stimulate neutrophils alone or in combination with phorbolmyristate- acetate (PMA) (0.1–30 nM) or Staphylococcus aureus bacteria (0.7–500 bact/neutrophil). Relative generation of ROS was measured flow cytometrically. BALF/RPMI and in combination with relatively low PMA or all bacteria concentrations stimulated ROS; while, combination with relatively high PMA concentrations suppressed ROS in of COPD patients and HI. IS/RPMI and its combination with PMA inhibited ROS generation in both groups; whereas, IS stimulated or had a tendency to stimulate ROS production with relatively high bacteria concentrations. In conclusion, BALF and IS directly or in combination with chemical/biological factors modulated ROS production. This effect was stronger in neutrophils from COPD patients and depended on chemical/biological stimulus intensity.