Stimulation-evoked Noradrenaline Overflow Research Articles

N-type and P/Q-type calcium channels regulate differentially the release of noradrenaline, ATP and β-NAD in blood vessels

Using HPLC techniques we evaluated the electrical field stimulation-evoked overflow of noradrenaline (NA), adenosine 5′-triphosphate (ATP), and β-nicotinamide adenine dinucleotide (β-NAD) in the presence of low nanomolar concentrations of ω-conotoxin GVIA or ω-agatoxin IVA in the canine mesenteric arteries and veins. ω-conotoxin GVIA abolished the evoked overflow of NA and β-NAD in artery and vein, whereas the evoked overflow of ATP remained unchanged in the presence of ω-conotoxin GVIA. ω-agatoxin IVA significantly reduced the evoked overflow of ATP and β-NAD. The overflow of NA remained largely unaffected by ω-agatoxin IVA, except at 16 Hz in the vein where the overflow of NA was reduced by about 50%. Artery and vein exhibited similar expression levels of the α 1B (CaV2.2, N-type) subunit, whereas the vein showed greater levels of the α 1A (CaV2.1, P/Q-type) subunit than artery. Therefore, there are at least two release sites for NA, β-NAD and ATP in the canine mesenteric artery and vein: an N-type-associated site releasing primarily NA, β-NAD and some ATP, and a P/Q-type-associated site releasing ATP, β-NAD and some NA. The N-type-mediated mechanisms are equally expressed in artery and vein, whereas the P/Q-type-mediated mechanisms are more pronounced in the vein and may ensure additional neurotransmitter release at higher levels of neural activity. In artery, β-NAD caused a dual effect consisting of vasodilatation or vasoconstriction depending on concentrations, whereas vein responded with vasodilatation only. In contrast, ATP caused vasoconstriction in both vessels. β-NAD and ATP may mediate disparate functions in the canine mesenteric resistive and capacitative circulations.

Effects of the neuropeptide Y Y2 receptor antagonist BIIE0246 on sympathetic transmitter release in the pig in vivo.

This study investigated the effects of BIIE0246, a novel neuropeptide Y (NPY) Y(2) receptor antagonist, on renal sympathetic nerve-evoked release of noradrenaline and NPY-like immunoreactivity (-LI) in the pig in vivo. Stimulation (5 Hz, 60 s) of renal sympathetic nerves evoked vasoconstriction and overflow of noradrenaline and NPY-LI. Infusion of peptide YY (PYY; 1 microg/kg per min) strongly inhibited the stimulation-evoked overflow of noradrenaline and NPY-LI. BIIE0246 (100 nmol/kg) abolished the inhibitory effect of PYY. BIIE0246 did not however affect the stimulation-evoked overflow of noradrenaline and NPY-LI, or basal cardiovascular parameters, per se. The alpha(2)-adrenoceptor antagonist yohimbine (0.2 mg/kg) significantly elevated both basal plasma levels and the nerve stimulation-evoked overflow of noradrenaline and NPY-LI. Subsequent administration of BIIE0246 did not further alter these effects. However, BIIE0246 caused splenic vasodilatation per se when given after yohimbine. It is concluded that the renal sympathetic nerves in the pig possess NPY Y(2) receptors, which upon activation inhibit transmitter (noradrenaline and NPY) release. Furthermore, when circulating levels of NPY were enhanced after blockade of alpha(2)-adrenoceptors, an involvement of endogenous NPY, acting on the NPY Y(2) receptor, in regulation of basal splenic vascular tone was unveiled.

ORL1 receptor-mediated inhibition by nociceptin of noradrenaline release from perivascular sympathetic nerve endings of the rat tail artery.

In the present study the effect of the opioid heptadecapeptide nociceptin, also termed orphanin FQ, an endogenous ligand for the orphan receptor named ORL1 (opioid receptor-like 1) receptor, was investigated on [3H]noradrenaline release induced by electrical field stimulation (24 pulses at 0.4 Hz, 200 mA, 0.3 ms duration) in the rat tail artery in the absence and presence of an alpha2-adrenoceptor antagonist, rauwolscine 3 microM. Nociceptin inhibited the electrically-evoked tritiated noradrenaline release in a concentration-dependent manner from rat tail arteries. This inhibitory effect of nociceptin was enhanced in the presence of the alpha2-adrenoceptor antagonist rauwolscine (maximum inhibition by 25% and 50% in the absence and presence of rauwolscine, respectively). At a supramaximal concentration (10 microM), the inhibitory action of DAGO, a selective micro-opioid receptor agonist, was less pronounced than that of nociceptin. The inhibitory effect of nociceptin was counteracted by naloxone benzoylhydrazone (3 microM) which by itself did not change the stimulation-evoked noradrenaline overflow. Naloxone (10 microM), a non-selective opioid receptor antagonist, did not affect the inhibitory effect of nociceptin whereas it abolished that of DAGO. In conclusion, these results suggest that nociceptin modulates noradrenergic neurotransmission by acting on prejunctional ORL1 receptors located on nerve terminals innervating the rat tail artery. They also demonstrate that prejunctional ORL1 receptors interact with prejunctional alpha2-adrenoceptors. The physiological significance of this phenomenon remains to be determined.

Inhibition by interleukin-1 beta of noradrenaline release in rat spleen: involvement of lymphocytes, NO and opioid receptors.

Effects of indomethacin, N omega-nitro-L-arginine (NNA) and naloxone, and of pretreatment with cyclophosphamide (CY), on the interleukin (IL)-1 beta induced inhibition of exocytotic noradrenaline release were investigated in the isolated, vascularly perfused spleen of the rat. Neurotransmitter release was evoked by perivascular electrical stimulation (4 Hz) and the overflow of endogenous noradrenaline was determined by HPLC with electrochemical detection. Perfusion of the spleen with Tyrode's solution containing IL-1 beta (100 pg/ml) for 90 min caused an inhibition of the stimulation-evoked noradrenaline overflow which persisted for at least 20 min after washout of the IL. The evoked overflow was reduced in the presence of NNA 30 mumol/l, but remained unaffected by indomethacin 3 mumol/l, naloxone 0.1 mumol/l or treatment of the rats with CY (250 mg/kg). The opioid agonist etorphine 10 mumol/l inhibited the evoked overflow of noradrenaline and this effect was prevented by naloxone 0.1 mumol/l. The inhibition of evoked overflow by IL-1 beta was not affected by indomethacin but was reduced or even prevented in the presence of NNA or naloxone, or after lymphocyte depletion of spleens by CY. The results are compatible with the idea that in the rat spleen exocytotic noradrenaline release is accompanied by a concomitant secretion of a nitric oxide (NO)-like compound which, in turn, reinforces noradrenaline release, and that the release can be inhibited via prejunctional opioid receptors. The IL-1 beta induced inhibition of evoked release appears to be a complex process which involves as one of many steps a decrease of the facilitatory NO-like compound and the release of endogenous opioids probably from spleen lymphocytes.

1-Aminocyclopropanecarboxylic acid is a partial agonist at rat hippocampal presynaptic NMDA receptors

Pre- and postjunctional effects of the α1-selectiv adrenoceptor antagonist prazosin and the α1- and α2-selective adrenoceptor agonists methoxamine and 6-fluoronoradrenaline, respectively, were studied in skeletal muscle in situ. Prazosin reduced the vasoconstriction and enhanced the overflow of endogenous noradrenaline elicited by sympathetic nerve stimulation (1–4 Hz, 2 min); the threshold concentration was 10–100 times lower for postjunctional than for prejunctional α-adrenoceptor blockade. The enhancement of noradrenaline overflow by prazosin was not inversely frequency-dependent, as shown elsewhere for α2-adrenoceptor antagonists. Thus, different mechanisms may be involved. Inhibition of prostaglandin synthesis by diclofenac did not alter the stimulation-evoked noradrenaline overflow, indicating a minor importance of prostaglandin-mediated transjunctional mechanisms in the modulation of noradrenaline overflow. Methoxamine and 6-fluoronoradrenaline elevated the basal vascular tone and, at higher concentrations, reduced the stimulation-evoked noradrenaline overflow. Methoxamine was 20 times more selective than 6-fluoronoradrenaline for postjunctional receptors. Our results are compatible with a pre- and postjunctional localization of α2-adrenoceptors and a predominantly, but not exclusively, postjunctional localization of α1-adrenoceptors. The postjunctional selectivity for prazosin was less marked than previously reported from in vitro studies. Hence, care should be taken when extrapolating in vitro findings to the more complex in vivo situation.

Role of angiotensin and sodium intake in cardiac noradrenaline release

The effects of exogenous and of endogenous angiotensin on noradrenaline overflow were investigated in saline perfused rat hearts with intact sympathetic nerves. The overflow of endogenous noradrenaline induced by electrical stimulation of the left stellate ganglion was determined in the coronary venous effluent by HPLC. The activity of the renin-angiotensin system was modulated by varying of the nutritional sodium load prior to the experiments. Endogenous angiotensin formation was blocked by angiotensin-converting enzyme inhibitors. Following high sodium intake, both angiotensin II (100 nmol/l) and angiotensin I (1 mumol/l) caused a marked increase of the electrically evoked noradrenaline overflow. After inhibition of the angiotensin-converting enzyme using captopril (350 nmol/l) or ramiprilat (50 nmol/l), angiotensin I (1 mumol/l) did not enhance noradrenaline overflow. This indicates an active cardiac angiotensin conversion, since the sole administration of captopril and ramiprilat did not affect noradrenaline overflow in rats with high sodium intake. Following low sodium intake, neither angiotensin II (100 nmol/l) nor angiotensin I (1 mumol/l) significantly affected noradrenaline overflow. Both captopril and ramiprilat, however, significantly reduced noradrenaline overflow induced by electrical stimulation, suggesting a facilitory action of endogenous angiotensin under these conditions. This concept was substantiated when evaluating the noradrenaline overflow during control stimulations. Following low sodium intake, stimulation evoked noradrenaline overflow was higher as compared to that after nutritional sodium load. The results are in keeping with a sodium-dependent intracardiac formation of angiotensin II which facilitates noradrenaline release from sympathetic nerve terminals. Following low sodium intake, cardiac angiotensin II formation is active, as indicated by the suppression of noradrenaline release by angiotensin-converting enzyme inhibitors and the ineffectiveness of exogenous application of angiotensin II.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of the renin-angiotensin system on sympathetic neurotransmission in canine skeletal muscle in vivo.

The physiological importance of interactions between angiotensin II and sympathetic neurotransmission was studied in an in vivo model with constant flow blood perfused gracilis muscle in situ in dogs pretreated with desipramine and atropine. Sympathetic nerve stimulation-(2 and 8 Hz, 480 pulses) evoked over-flow of endogenous noradrenaline and vasoconstriction, and vasoconstrictor responses to exogenous noradrenaline (0.5 nmol, locally i.a.) were evaluated. Angiotensin converting enzyme inhibition by benazeprilat (10 mg i.v.; n = 8) reduced arterial angiotensin II levels from 26 +/- 8 to 2 +/- 1 pM and reduced mean arterial and basal muscle perfusion pressures. Subsequent resubstitution of angiotensin II (3, 30 and 90 ng kg-1 min-1 i.v.) elevated arterial angiotensin II dose-dependently (to 67 +/- 14, 622 +/- 63 and 1940 +/- 251 pM, respectively), as well as mean arterial and muscle perfusion pressures. Nerve stimulation-evoked noradrenaline overflow was unchanged following benazeprilat (-4 +/- 4 and +1 +/- 8% at 2 and 8 Hz, respectively) and during subsequent infusions of angiotensin II. Vasoconstrictor responses to nerve stimulation and exogenous noradrenaline were also uninfluenced by these treatments. Thus, angiotensin II did not enhance sympathetic neurotransmission at the postjunctional level. Another group of animals was pretreated with noncompetitive alpha-adrenoceptor blockade locally by phenoxybenzamine and benextramine (0.5 mg kg-1 i.a. of each; n = 7), which abolished vasoconstrictor responses to nerve stimulation. The effects of benazeprilat and subsequent angiotensin II infusions (3 and 30 ng kg-1 min-1 i.v.) on circulating angiotensin II levels, mean arterial and muscle perfusion pressures were similar in this group.(ABSTRACT TRUNCATED AT 250 WORDS)

Involvement of neuropeptide Y in sympathetic vascular control of skeletal muscle in vivo

The possibility that neuropeptide Y, a vasoconstrictor peptide co-stored with noradrenaline in sympathetic nerves, participates in neurogenic vascular control was investigated in canine gracilis muscle in situ. Sympathetic nerve stimulation with recordings of the normal irregular sympathetic discharge to human skeletal muscle elicited frequency-dependent overflows of neuropeptide Y-like immunoreactivity and noradrenaline, and vasoconstriction. The overflow of neuropeptide Y-like immunoreactivity was more markedly enhanced with increasing frequency. Exogenous neuropeptide Y reduced nerve stimulation-evoked noradrenaline overflow, possibly through a prejunctional mechanism, and caused dose-dependent vasoconstriction. Nerve stimulation elicited significant vasoconstrictor responses following alpha- and beta-adrenoceptor blockade with phenoxybenzamine and propranolol, which abolished the vasoconstriction to exogenous noradrenaline. Nerve stimulation-evoked overflows of neuropeptide Y-like immunoreactivity and noradrenaline were enhanced, consistent with prejunctional alpha-adrenoceptor-mediated inhibition of the release. Thus, neuropeptide Y is likely to be involved in the non-adrenergic component of vasoconstriction, and may participate in the physiological control of vascular tone at moderate to high impulse frequencies.

ACTH increases noradrenaline release in the rabbit heart

The effects of ACTH on the release of noradrenaline and the increase of heart rate produced by sympathetic nerve stimulation (1 Hz) were studied in isolated perfused rabbit hearts. ACTH-(1-24) 0.1-100 nmol/l increased the stimulation-evoked overflow of noradrenaline concentration-dependently, reversibly and up to two-fold. The basal outflow of noradrenaline, the basal heart rate and the stimulation-evoked increase in heart rate were not changed. Human ACTH-(1-39) also increased the evoked overflow of noradrenaline. The effect of ACTH-(1-24) 0.3 nmol/l persisted after blockade of beta-adrenoceptors with propranolol and blockade of neuronal catecholamine uptake by cocaine. ACTH-(1-24) 3 nmol/l did not change the removal of noradrenaline from the perfusion fluid, when hearts were perfused with medium containing 59 nmol/l noradrenaline. The results show that ACTH increases the action potential-evoked release of noradrenaline from cardiac postganglionic sympathetic neurones, probably by activating specific presynaptic ACTH receptors. The high potency of ACTH suggests that these presynaptic receptors may be activated in vivo by circulating ACTH under certain pathophysiological conditions.

Adenosine inhibits exocytotic release of endogenous noradrenaline in rat heart: a protective mechanism in early myocardial ischemia.

The effects of exogenous and endogenous adenosine on exocytotic noradrenaline release were studied in rat hearts perfused in situ. Exocytotic release of endogenous noradrenaline (determined by high pressure liquid chromatography) was induced by electrical stimulation of the left cervicothoracic ganglion. Exogenous adenosine significantly reduced noradrenaline overflow from the heart. This suppression of noradrenaline overflow was not influenced by desipramine, indicating a mechanism independent from noradrenaline reuptake. The A1 subtype specific agonists cyclohexyladenosine and R-phenylisopropyladenosine had inhibitory effects at lower concentrations than adenosine and S-phenylisopropyladenosine, suggesting the relevance of presynaptic inhibitory adenosine receptors of the A1 subtype. Short ischemic periods of 3 minutes resulted in a marked coronary venous overflow of adenosine during reperfusion. This was accompanied by an inhibition of noradrenaline release evoked by nerve stimulation during ischemia. The adenosine antagonists theophylline and 8-phenyltheophylline prevented this suppression of noradrenaline release. Blockade of oxidative phosphorylation by cyanide in combination with glucose-free perfusion induced an increased formation of endogenous adenosine and suppression of stimulation-evoked noradrenaline overflow. Again, in the presence of the adenosine antagonists theophylline or 8-phenyltheophylline, this suppression was abolished. These results indicate that adenosine is a potent inhibitor of exocytotic noradrenaline release in the heart with relevance during conditions of increased endogenous adenosine formation such as myocardial ischemia.

Pre- and postjunctional alpha-adrenoceptor-mediated effects of prazosin, methoxamine and 6-fluoronoradrenaline in blood-perfused canine skeletal muscle in situ.

Pre- and postjunctional effects of the alpha 1-selective adrenoceptor antagonist prazosin and the alpha 1- and alpha 2-selective adrenoceptor agonists methoxamine and 6-fluoronoradrenaline, respectively, were studied in skeletal muscle in situ. Prazosin reduced the vasoconstriction and enhanced the overflow of endogenous noradrenaline elicited by sympathetic nerve stimulation (1-4 Hz, 2 min); the threshold concentration was 10-100 times lower for postjunctional than for prejunctional alpha-adrenoceptor blockade. The enhancement of noradrenaline overflow by prazosin was not inversely frequency-dependent, as shown elsewhere for alpha 2-adrenoceptor antagonists. Thus, different mechanisms may be involved. Inhibition of prostaglandin synthesis by diclofenac did not alter the stimulation-evoked noradrenaline overflow, indicating a minor importance of prostaglandin-mediated transjunctional mechanisms in the modulation of noradrenaline overflow. Methoxamine and 6-fluoronoradrenaline elevated the basal vascular tone and, at higher concentrations, reduced the stimulation-evoked noradrenaline overflow. Methoxamine was 20 times more selective than 6-fluoronoradrenaline for postjunctional receptors. Our results are compatible with a pre- and postjunctional localization of alpha 2-adrenoceptors and a predominantly, but not exclusively, postjunctional localization of alpha 1-adrenoceptors. The postjunctional selectivity for prazosin was less marked than previously reported from in vitro studies. Hence, care should be taken when extrapolating in vitro findings to the more complex in vivo situation.

Kinetic analysis of stimulation-evoked overflow of noradrenaline and dopamine β-hydroxylase from the isolated rabbit heart. The effect of dopa decar☐ylase inibition

Rabbit isolated hearts were perfused according to a modified Langendorff procedure. Noradrenaline was measured in the venous effluents whereas dopamine β-hydroxylase was determined in the transmyocardial fluid drained from the interstitium. Calcium-dependent release of noradrenaline and dopamine β-hydroxylase was evoked by electrical stimulation of the heart for 6 periods of 1 min duration at 30 min intervals. Tetraethylammonium was added to enhance release. As a result of repeated stimulation, the dopamine β-hydroxylase overflow from the heart declined monoexponentially, whereas the decline of noradrenaline was biexponential. The two phases of decline were unaltered by cocaine, an inhibitor of noradrenaline uptake. However, pretreatment of the rabbits with monofluoromethyldopa, an irreversible inhibitor of DOPA decar☐ylase, transformed the biexponential decline of noradrenaline overflow into a monoexponential one with the same rate constant as that of dopamine β-hydroxylase overflow. The sum of noradrenaline released plus that remaining in the heart was perfectly matched by the area under the curve of noradrenaline overflow values. Using the dopamine β-hydroxylase overflow curve, the pool size of the soluble enzyme was calculated which, conversely, agreed with the enzyme contained in the 100,000 g supernatant of strongly homogenized unstimulated hearts. Before stimulation, 18% of the total dopamine β-hydroxylase activity was in the soluble form. Stimulation depleted the hearts of noradrenaline and soluble dopamine β-hydroxylase in a stoichiometric manner, provided that noradrenaline synthesis was blocked. The present work utilizes a novel method of measuring soluble dopamine β-hydroxylase that is available for exocytotic release. Under the conditions of repeated stimulation and prolonged action potentials, the only factor interfering with the stoichiometry of noradrenaline and dopamine β-hydroxylase release is the re-synthesis of noradrenaline.