Stimulation-induced Noradrenaline Release Research Articles

Effects of yohimbine on norepinephrine release by locus coeruleus in stressed rats

Objective To study the cellular and molecular modulation mechanisms of norepinephrine (NE) release in hypothalamus induced by electrical stimulation in locus coeruleus(LC) in the rat model of depression-like behavior . Methods The depression-like behavior model was established by combining separation and chronic unpredictable stress stimulations.After the model establishment, behavior tests were used to verify the success of modeling. NE release in the hypothalamus induced by electrical stimulation in LC was studied in real time and NE signal was recorded with carbon fiber electrode. The peak value, the time to peak and half-life period of NE signal in both group rats were measured and analysed. Results The bodyweight, score of open-field test, percentage of sucrose preference of model group rats((263.9±16.4)g, (19.4±7.9), (44.3±10.8)%) were significantly lower than those((314.3±24.3)g, ( 53.3±19.0), ( 60.6±13.3)%) of control group(P<0.01). The peak value of NE signal in the hypothalamus induced by electrical stimulation in locus coeruleus in depression-like behavior model rats((176.9±50.4) pA)was less than that((361.6±88.6) pA )in control group(P<0.01), and the time to peak of NE signal was also shortened in depression-like behavior model group rats(P<0.05). Intraperitoneal injection of yohimbine (3 mg/kg) potentiated electrical stimulation induced NE release in depression-like behavior model rats but not in control group. Conclusion The chronic unpredictable stresses attenuate the secretion of NE in the hypothalamus induced by electrical stimulation in LC in rats. Yohimbine, a presynaptic α2 receptor antagonist, increased NE release in hypothalamus in depression-like behavior model rats. These findings suggest that the LC projects functionally to the hypothalamus and the projection may contribute to the pathogenesis of depression. Key words: Chronic unpredictable stress; Rats; Locus coeruleus; Hypothalamus; Yohimbine

Inhibitory effects of angiotensin-(1–7) on the nerve stimulation-induced release of norepinephrine and neuropeptide Y from the mesenteric arterial bed

Neuropeptide Y (NPY) is a cotransmitter with norepinephrine (NE) and ATP in sympathetic nerves. There is evidence for increased activity of the sympathetic nervous system and the renin-angiotensin system (RAS), as well as a role for NPY in the development of hypertension in experimental animal models and in humans. Angiotensin II (ANG II) is known to facilitate sympathetic neurotransmission, an effect greater in spontaneously hypertensive rats (SHR) than normotensive Wistar-Kyoto (WKY) rats. A newly discovered product of the RAS is angiotensin-(1-7) [ANG-(1-7)]. There is evidence suggesting that ANG-(1-7) opposes the actions of ANG II, resulting in hypotensive effects. The objective of this study was to investigate the role of ANG-(1-7) on the nerve-stimulated overflow of NE and NPY from the mesenteric arterial bed of SHR and the mechanisms involved in mediating any effects produced. ANG-(1-7) (0.001, 0.01, 0.1 microM) decreased nerve-stimulated NE and NPY overflow, as well as perfusion pressure in preparations obtained from SHR. This effect was greater in preparations of SHR than WKY controls. In addition, ANG-(1-7) decreased NE overflow to a greater extent than NPY overflow. Administration of the Mas receptor antagonist, D-Ala(7) ANG-(1-7), attenuated the decrease in both NE and NPY overflow due to ANG-(1-7) administration. However, the angiotensin type 2 receptor antagonist, PD-123391, attenuated the effect of ANG-(1-7) on NE overflow without affecting the decrease in NPY overflow. Moreover, in the presence of N(G)-nitro-L-arginine methyl ester, ANG-(1-7) decreased NPY overflow, but not NE overflow. ANG-(1-7) decreases the nerve-stimulated overflow of NE and NPY in preparations of SHR, whereas ANG II enhances it. Therefore, ANG-(1-7) may counteract the effects of ANG II by acting on ANG type 2 and Mas receptors.

Abstract 5255: Differential Effect Of A Novel Adenosine-Agonist On Stimulation-Induced Norepinephrine Release From Rat Hearts

Presynaptic modulation of norepinephrine (NE) release plays a major role in the regulation of cardiac sympathetic activity. We tested the effect of the novel, highly selective adenosine A1 subtype agonist BAY 68–4986 on stimulation-induced NE release. Isolated rat hearts (Wistar or SHR) were perfused with a modified Krebs-Henseleit solution (Langendorff technique). Exocytotic NE release was induced by electrical field stimulation with 5 V, 6 Hz for 1 minute. NE was detected by HPLC. Two stimulations (S1, S2) were performed to compare intraindividually NE release under control conditions (S1) and with BAY 68–4986 30ug/ml, 300 ug/ml or CCPA 10–6M (S2). Neuronal uptake was inhibited by desipramine (100 nM) in order to prevent amine elimination and nonexocytotic release of NE. Stimulation of adenosine A1 receptors with BAY 68–4986 led to a concentration dependent decrease of the stimulation-induced NE release only in SHR rats, whereas no effect could be seen in Wistar rats (SHR: S2/S1 = 0.90 +/− 0.08 with 30 ug/l, S2/S1 = 0.54 +/− 0.02 with 300 ug/l, Wistar: S2/S1 = 1.05 +/− 0.12 with 30 ug/l, S2/S1 = 1.03 +/− 0.09 with 300 ug/l, p=0.003). The effect of BAY 68–4986 on stimulation-induced NE release in SHR was similar to the effect of 10–6 M CCPA (S2/S1 = 0.59 ± 0.05). This differential effect in genetically different rats was also observed in vivo, with a blunted increase of the heart rate during stress reaction only in SHR rats, when treated with BAY 68–4986. We could demonstrate that the novel, highly selective A1-adenosine agonist BAY 68–4986 has a differential effect on stimulation-induced NE release and stress responde in SHR rats compared to Wistar rats.

ATP MODULATES THE RELEASE OF NORADRENALINE THROUGH TWO DIFFERENT PREJUNCTIONAL RECEPTORS ON THE ADRENERGIC NERVES OF RAT PROSTATE

1. The effects of adenosine and ATP receptor agonists on the release of endogenous noradrenaline from electrically stimulated (2 Hz, 0.1 msec) rat prostate were examined in order to clarify the pharmacological properties of prejunctional receptors for adenosine and ATP on the adrenergic nerve varicosities in the prostate. Noradrenaline was quantified by HPLC coupled with electrochemical detection techniques. 2. Both adenosine and ATP receptor agonists (1 micromol/L) inhibited noradrenaline release and the relative order of inhibitory effect was N(6)-cyclopentyl-adenosine (CPA) > 5'-N-ethylcarboxamidoadenosine > 2-chloroadenosine > adenosine > 2-methylthio-ATP (2mSATP) > AMP > ATP. 3. The adenosine receptor agonist CPA (1 nmol/L-1 micromol/L) and the ATP receptor agonist 2mSATP (100 nmol/L-100 micromol/L) inhibited the stimulation-induced release of noradrenaline in a concentration-dependent manner. The concentrations of CPA and 2mSATP that produced 50% inhibition of noradrenaline release were 9.6 nmol/L and 1.4 micromol/L, respectively. 4. 1,3-Dipropyl-8-cyclopentylxanthine, an adenosine A(1) receptor antagonist, significantly reduced the inhibitory effects of not only CPA, but also 2mSATP. 5. Suramin, an ATP receptor antagonist, significantly reduced the inhibitory effects of 2mSATP, but not those of CPA. 6. Pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid, another ATP receptor antagonist, had no effect on the inhibitory action of either agonist. 7. These results suggest that, in the sympathetic nerve terminals of rat prostate, adenosine and ATP induce inhibition of noradrenaline release via the activation of adenosine A(1) and/or xanthine-sensitive ATP receptors, which play an inhibitory regulatory role in adrenergic neurotransmission in the prostate.

Hypothermia reduces ischemia- and stimulation-induced myocardial interstitial norepinephrine and acetylcholine releases

Although hypothermia is one of the most powerful modulators that can reduce ischemic injury, the effects of hypothermia on the function of the cardiac autonomic nerves in vivo are not well understood. We examined the effects of hypothermia on the myocardial interstitial norepinephrine (NE) and ACh releases in response to acute myocardial ischemia and to efferent sympathetic or vagal nerve stimulation in anesthetized cats. We induced acute myocardial ischemia by coronary artery occlusion. Compared with normothermia (n = 8), hypothermia at 33 degrees C (n = 6) suppressed the ischemia-induced NE release [63 nM (SD 39) vs. 18 nM (SD 25), P < 0.01] and ACh release [11.6 nM (SD 7.6) vs. 2.4 nM (SD 1.3), P < 0.01] in the ischemic region. Under hypothermia, the coronary occlusion increased the ACh level from 0.67 nM (SD 0.44) to 6.0 nM (SD 6.0) (P < 0.05) and decreased the NE level from 0.63 nM (SD 0.19) to 0.40 nM (SD 0.25) (P < 0.05) in the nonischemic region. Hypothermia attenuated the nerve stimulation-induced NE release from 1.05 nM (SD 0.85) to 0.73 nM (SD 0.73) (P < 0.05, n = 6) and ACh release from 10.2 nM (SD 5.1) to 7.1 nM (SD 3.4) (P < 0.05, n = 5). In conclusion, hypothermia attenuated the ischemia-induced NE and ACh releases in the ischemic region. Moreover, hypothermia also attenuated the nerve stimulation-induced NE and ACh releases. The Bezold-Jarisch reflex evoked by the left anterior descending coronary artery occlusion, however, did not appear to be affected under hypothermia.

Calcitonin gene-related peptide does not interact with sympathetic activity in myocardial ischemia

The aim of the present study was to assess whether calcitonin gene-related peptide (CGRP) modulates exocytotic norepinephrine release in ischemic myocardium. In isolated rat hearts subjected to 30 min of low flow ischemia, CGRP release increased 2.8-fold whereas stimulation-induced norepinephrine release decreased 4.1-fold. Pretreatment of rats with capsaicin almost completely depleted cardiac CGRP stores; however, suppression of norepinephrine release by 30 min of low flow ischemia was not affected. At normal flow, exogenous CGRP (5 μmol l −1) had no effect on norepinephrine release. These findings suggest that CGRP release from sensory neurons does not interact with the cardiac sympathetic system during myocardial ischemia.

AT1 and AT2-receptor antagonists inhibit Ang II-mediated facilitation of noradrenaline release in human atria.

It is generally accepted that regulation of blood pressure and sympathetic neurotransmission by angiotensin (Ang) II is brought about through activation of AT1-receptors. Since recent studies demonstrated a high proportion of AT2-receptors in the human heart, the aim of our study was to investigate whether Ang II modulates noradrenaline release also through activation of AT2-receptors in this tissue. Human atrial appendages were prelabeled with [3H]-noradrenaline and electrically field-stimulated. Stimulation-induced outflow of radioactivity was taken as an index of endogenous noradrenaline release. Ang I and II enhanced noradrenaline release in a dose-dependent manner up to 55 and 72%, respectively. These effects were blocked by the selective AT1-receptor antagonists EXP3174 and irbesartan (10 nmol/L). Moreover, the selective AT2-receptor antagonists PD123319 and CGP42112A (0.1 and 1 micromol/L) also inhibited Ang II-induced facilitation of noradrenaline release. Captopril (5 micromol/L) shifted the dose response curve for Ang I less potent to the right than EXP3174 (10 nmol/L). Ang I and II enhanced the stimulation-induced noradrenaline release significantly more potent in tissues of patients pretreated with ACE inhibitors than without. In conclusion, both AT1- and AT2-receptors seem to play a role in Ang II-mediated facilitation of noradrenaline release in the human heart. Chronic treatment with ACE inhibitors appears to affect cardiac sympathetic neurotransmission possibly by upregulation of presynaptic Ang II receptors.

Age and monosodium glutamate treatment cause changes in the stimulation-induced [ 3H]-norepinephrine release from rat nucleus tractus solitarii-dorsal vagal nucleus slices

In nucleus tractus solitarii-dorsal vagal nucleus slices prepared from young adult rats (180–260 g) 10 −3 M L-glutamate and 10 −5 M baclofen caused a 2-3-fold increase of field stimulation-induced [ 3H]-norepinephrine release without affecting the resting release. In slices prepared from rats treated neonatally with monosodium glutamate neither L-glutamate nor baclofen had any effect on stimulation-induced norepinephrine release, tested between postnatal days 74–99 (350–530 g). In untreated littermates used in the same period (460–580 g) L-glutamate was fully effective whereas baclofen was ineffective. The tritium content in tissue extracts did not differ significantly in the three experimental groups. It is concluded that i) the loss of GABA B receptor-mediated disinhibitory stimulation of norepinephrine release is an age-related phenomenon and ii) neonatal monosodium glutamate treatment causes a damage in the local neural circuitry characterized by the loss of glutamate receptor-mediated mechanism that stimulates the release of norepinephrine.

Prostanoid-induced modulation of neuropeptide Y and noradrenaline release from the rat mesenteric bed.

1. A variety of prostanoids were examined for their ability to alter the periarterial nerve stimulation-induced release of noradrenaline (NA) and neuropeptide Y immunoreactive compounds (NPY-ir) from the perfused mesenteric arterial bed of the rat. 2. Periarterial nerve stimulation (16 Hz) increased the overflow of NA, NPY-ir and perfusion pressure. 3. The prostacyclin (PGI2) analogues, carbaPGI2 and cicaprost both produced a concentration-dependent attenuation of the nerve stimulation-induced increase in NA, NPY-ir overflow and perfusion pressure. 4. The prostaglandin (PG) analogue PGE2 attenuated the evoked increase in NPY-ir overflow as well as a modest decrease in NA. 5. PGE1, sulprostone and iloprost attenuated the nerve stimulation-induced increase in NA overflow but not NPY-ir. 6. Neither PGF2alpha nor the thromboxane A2 analogue U46619 altered the evoked increase in NA or NPY-ir overflow. 7. The results support the view that sympathetic co-transmitter release can be differentially modulated by paracrine/autocrine mediators at sympathetic neuroeffector junctions.

Norepinephrine release is reduced in cardiac tissue of Type 2 diabetic patients.

The aim of this study was to assess whether cardiac catecholamine release is affected in patients with Type 2 diabetes mellitus. A trial tissue was obtained from 19 diabetic (Type 2) and 43 non-diabetic patients undergoing coronary surgery. Endogenous norepinephrine release was examined under baseline conditions as well as during electrical field stimulation (effective voltage 5 V, stimulation frequency 4 Hz, pulse width 2 msec) by high performance liquid chromatography and electrochemical detection. Cardiac function and arterial blood pressure was assessed from coronary angiography. In atrial tissue from diabetic patients, stimulation-induced norepinephrine release was reduced by 25% compared with non-diabetic patients, while baseline norepinephrine release did not differ between both groups. Preoperative plasma glucose and haemoglobin A(1C) concentrations were increased in patients with diabetes, however, no relation was found to catecholamine release. Diabetic and non-diabetic patients did not differ regarding left ventricular ejection fraction and arterial blood pressure. Cardiac norepinephrine release is suppressed in patients with Type 2 diabetes which could contribute to sympathetic neuropathy. The difference of norepinephrine release in diabetic and non-diabetic patients was independent of cardiac function and arterial blood pressure.

Catecholaminergic microcircuitry controlling the output of airway-related vagal preganglionic neurons.

In this study, we have investigated the ultrastructure and function of the catecholaminergic circuitry modulating the output of airway-related vagal preganglionic neurons (AVPNs) in ferrets. Immunoelectron microscopy was employed to characterize the nature of catecholaminergic innervation of AVPN at the ultrastructural level. In addition, immunofluorescence was used to examine the expression of the alpha(2A)-adrenergic receptor (alpha(2A)-AR) on AVPNs, and norepinephrine release within the rostral nucleus ambiguous (rNA) was measured by using microdialysis. Physiological experiments were performed to determine the effects of stimulation of the noradrenergic locus coeruleus (LC) cell group on airway smooth muscle tone. The results showed that 1) catecholaminergic nerve endings terminate in the vicinity of identified AVPNs but very rarely form axosomatic or axodendritic synapses with the AVPNs that innervate the extrathoracic trachea; 2) AVPNs express the alpha(2A)-AR; 3) LC stimulation-induced norepinephrine release within the rNA region was associated with airway smooth muscle relaxation; and 4) blockade of alpha(2A)-AR on AVPNs diminished the inhibitory effects of LC stimulation on airway smooth muscle tone. It is concluded that a noradrenergic circuit originating within the LC is involved in the regulation of AVPN activity within the rNA, and stimulation of the LC dilates the airways by the release of norepinephrine and activation of alpha(2A)-AR expressed by AVPNs, mainly via volume transmission.

Endothelin-induced modulation of neuropeptide Y and norepinephrine release from the rat mesenteric bed.

The effect of three endothelin (ET) agonists [ET-1, ET-3, and sarafotoxin (STX6C)] on the nerve stimulation-induced release of norepinephrine (NE) and neuropeptide Y-immunoreactive compounds (NPY-ir) from the perfused mesenteric arterial bed of the rat as well as the effect on perfusion pressure were examined. ET-1, ET-3, and STX6C all produced a significant, concentration-dependent decrease in the evoked release of NPY-ir but had no effect on the release of NE. In contrast, all three ETs potentiated the nerve stimulation-induced increase in perfusion pressure. The inhibition of nerve stimulation-induced NPY-ir release by ET-1 was significantly blocked by the ET(A)/ET(B) antagonist PD-142893 and the ET(B) antagonist RES-701-1 but not by the ET(A) antagonist BQ-123. The potentiation of the nerve stimulation-induced increase in perfusion pressure by ET-1 was significantly blocked by PD-142893 and BQ-123 and attenuated by RES-701-1. Prior exposure of the preparation to indomethacin or meclofenamate failed to alter the attenuation of the evoked release of NPY-ir or the potentiation of the increase in perfusion pressure produced by ET-1 or ET-3. These results are consistent with the idea that sympathetic cotransmitters can be preferentially modulated by paracrine mediators at the vascular neuroeffector junction.

Regarding the unitary theory of agonist and antagonist action at presynaptic adrenoceptors.

1. The linkage between potentiation of field stimulation-induced noradrenaline release and blockade of the presynaptic inhibitory effect of exogenous noradrenaline by a presynaptic antagonist was examined in superfused rabbit aorta preparations. 2. Rauwolscine clearly potentiated the release of noradrenaline in response to 100 pulses at 2 Hz but reduced the capacity of noradrenaline to inhibit transmitter release to a questionable extent, and then only when comparisons were made with untreated, rather then to rauwolscine-treated, controls. 3. Aortic preparations exposed for 60 min to rauwolscine followed by superfusion with antagonist-free Krebs for 60 min retained the potentiation of stimulation-induced transmitter release but no antagonism of the noradrenaline-induced inhibition could be detected at either of two noradrenaline concentrations when comparisons were made with rauwolscine treated controls. 4. Comparisons of the inhibitory effect of exogenous noradrenaline (1.8 x 10-6 M) on transmitter efflux in the presence and absence of rauwolscine pretreatment revealed that the antagonist enhanced rather than antagonized the presynaptic inhibition by noradrenaline. 5 It is concluded that the unitary hypothesis that asserts that antagonist enhancement of transmitter release and its blockade of noradrenaline induced inhibition are manifestations of a unitary event are not supportable.

Effects of maturation on adrenergic neurotransmission in ovine cerebral arteries.

The present studies examine the hypothesis that multiple adrenergic neuroeffector mechanisms are not fully developed in fetal, compared with adult, ovine middle cerebral arteries. In arteries denuded of endothelium and pretreated with 1 microM atropine to block involvement of muscarinic receptors, 10 microM capsaicin to deplete sensory peptidergic neurons, and 10 microM nitro-L-arginine methyl ester (L-NAME) to block possible influences from nitric oxidergic innervation, transmural stimulation at 16 Hz increased contractile tensions to 9.5 +/- 3.7% (n = 6) of the potassium maximum in adult arteries. Corresponding values in fetal arteries, however, were significantly less and averaged only 1.1 +/- 0.6% (n =10). However, postsynaptic sensitivity to norepinephrine (NE) was similar in the two age groups; NE pD(2) values (-log EC(50)) averaged 6.11 +/- 0.12 (n = 6) and 6.33 +/- 0.09 M (n = 9) in fetal and adult arteries, respectively. Similarly, NE content measured via HPLC was also similar in the two age groups and averaged 32.4 +/- 5.0 (n = 17) and 32.5 +/- 3.9 ng/ng wet wt (n = 13) in fetal and adult middle cerebral arteries, respectively. In contrast, stimulation-induced NE release was greater in fetal than in adult arteries, whether calculated as total mass released [883 +/- 184 (n = 17) vs. 416 +/- 106 pg NE/mg wet wt (n = 13)] or as fractional release [51.1 +/- 5.3 (n = 17) vs. 22.8 +/- 3.8 pg/pg NE content per pulse x 10(-6)]. Measured as an index of synaptic density, neuronal cocaine-sensitive NE uptake was similar in fetal and adult arteries [1.55 +/- 0.40 (n = 10) and 1.84 +/- 0.51 pmol/mg wet wt (n = 7), respectively]. Overall, age-related differences in postsynaptic sensitivity to NE, NE release, and NE uptake capacity cannot explain the corresponding age-related differences in response to stimulation. The data thus suggest that total synaptic volume and cleft width, in particular, are probably greater and/or that adrenergic corelease of vasoactive substances other than NE is altered in fetal compared with adult middle cerebral arteries.

The possible effect of nitric oxide on relaxation and noradrenaline release in the isolated rabbit urethra

We evaluated the effects of N ω-nitro- l-arginine ( l-NNA, a nitric oxide (NO) synthase inhibitor) and carboxy-2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide (carboxy-PTIO, a NO scavenger) on NO-mediated relaxation and noradrenaline release from adrenergic nerve endings induced by electrical field stimulation in the rabbit urethra. Electrical field stimulation caused frequency-dependent relaxation of rabbit urethral smooth muscles precontracted with phenylephrine. The relaxation responses were significantly inhibited by treatment with l-NNA or carboxy-PTIO. The inhibitory effect of carboxy-PTIO was significantly weaker than that of l-NNA. Electrical field stimulation caused significant noradrenaline release from adrenergic nerve endings in the rabbit urethra. Treatment with carboxy PTIO enhanced electrical field stimulation-induced noradrenaline release, and simultaneous application of l-NNA and carboxy-PTIO did not further enhance noradrenaline release in the rabbit urethra. As carboxy-PTIO reacts only with the free radical NO, the present results suggest that free radical NO and NO-containing compounds are involved in the l-NNA-sensitive nitrergic nerve-mediated relaxation in the rabbit urethra. At the same time free radical NO has a prejunctional action by which it may inhibit noradrenaline release from adrenergic nerves.

PACAP enhances stimulation-induced norepinephrine release in canine pancreas in vivo

The present study was to investigate whether pituitary adenylate cyclase activating polypeptide (PACAP) can modify norepinephrine (NE) release in response to pancreatic nerve stimulation in anesthetized dogs. Plasma catecholamine concentrations in aortic and superior pancreaticoduodenal (SPD) venous blood were determined by a high performance liquid chromatography method. SPD venous blood flow was measured with an electromagnetic flowmeter. Pancreatic nerves were directly stimulated for 1 min (2 ms, 12 V) at various frequencies at the level of the SPD artery. Various doses of PACAP1-27 (PACAP27) were locally infused into the pancreas through the SPD artery. Nerve stimulation significantly increased both SPD venous NE concentration and its output from the pancreas in a frequency-dependent manner. With PACAP27 alone, neither SPD venous NE concentration nor its output changed significantly following the local administration of PACAP27 at any dose tested. In the presence of PACAP27, however, the net increases in NE concentration and its output in response to nerve stimulation at 2 Hz were significantly enhanced in a dose-dependent manner. The enhanced NE responses to nerve stimulation by PACAP27 were thus significantly greater than those obtained from the group receiving either PACAP27 or stimulation alone. Increases in NE concentration and its output induced by local administration of tyramine were virtually abolished by desipramine, a neural amine uptake inhibitor. However, the NE response to tyramine was not diminished by PACAP27. The results indicate that PACAP27 enhances the stimulation-induced NE release in the pancreas, and that this facilitatory effect of PACAP27 does not result from an inhibition of the neural amine uptake mechanism. The study suggests that PACAP receptor-mediated mechanisms may be involved either directly or indirectly in the local modulation of neural NE release in the canine pancreas in vivo.Key words: presynaptic, sympathetic nerve, neuropeptide, tyramine, reuptake, anesthetized dog.

PACAP enhances stimulation-induced norepinephrine release in canine pancreas in vivo.

The present study was to investigate whether pituitary adenylate cyclase activating polypeptide (PACAP) can modify norepinephrine (NE) release in response to pancreatic nerve stimulation in anesthetized dogs. Plasma catecholamine concentrations in aortic and superior pancreaticoduodenal (SPD) venous blood were determined by a high performance liquid chromatography method. SPD venous blood flow was measured with an electromagnetic flowmeter. Pancreatic nerves were directly stimulated for 1 min (2 ms, 12 V) at various frequencies at the level of the SPD artery. Various doses of PACAP1-27 (PACAP27) were locally infused into the pancreas through the SPD artery. Nerve stimulation significantly increased both SPD venous NE concentration and its output from the pancreas in a frequency-dependent manner. With PACAP27 alone, neither SPD venous NE concentration nor its output changed significantly following the local administration of PACAP27 at any dose tested. In the presence of PACAP27, however, the net increases in NE concentration and its output in response to nerve stimulation at 2 Hz were significantly enhanced in a dose-dependent manner. The enhanced NE responses to nerve stimulation by PACAP27 were thus significantly greater than those obtained from the group receiving either PACAP27 or stimulation alone. Increases in NE concentration and its output induced by local administration of tyramine were virtually abolished by desipramine, a neural amine uptake inhibitor. However, the NE response to tyramine was not diminished by PACAP27. The results indicate that PACAP27 enhances the stimulation-induced NE release in the pancreas, and that this facilitatory effect of PACAP27 does not result from an inhibition of the neural amine uptake mechanism. The study suggests that PACAP receptor-mediated mechanisms may be involved either directly or indirectly in the local modulation of neural NE release in the canine pancreas in vivo.

Bradykinin B2-receptor-mediated Stimulation of Exocytotic Noradrenaline Release from Cardiac Sympathetic Neurons

Myocardial ischemia, as well as angiotensin-converting-enzyme-inhibitors, increase cardiac concentrations of the non-apeptide bradykinin. Cardiac effects of bradykinin are potentially mediated by modulation of sympathoadrenergic neurotransmission. Accordingly, the present study was designed to examine the influence of bradykinin on exocytotic noradrenaline release from rat isolated perfused heart. Exocytotic noradrenaline release was induced by electrical field stimulation (1 min, 5 V, 6 Hz) twice to compare the effect of intervention (S2) with respective control stimulation (S1). The overflow of endogenous noradrenaline was determined by high pressure liquid chromatography and electrochemical detection. The results are expressed as the mean S2/S1ratio±s.e.m.Bradykinin (1μmol/l) evoked a significant increase in noradrenaline release (S2/S1: 1.60±0.12;P<0.01), which was even more pronounced after inhibition of neuronal reuptake of noradrenaline by desipramine (0.1μmol/l: S2/S1: 1.83±0.15;P<0.01) excluding interference of bradykinin with the noradrenaline uptake1carrier. The concentration–response curve for bradykinin (0.1 nmol/l to 10μmol/l) revealed a maximum effect at 1μmol/l and an EC50-value of 7.5 nmol/l. The effect of bradykinin was unaltered by the B1-receptor antagonist des-Arg9(Leu8)-bradykinin (1μmol/l; S2/S1: 1.69±0.17), whereas it was reduced significantly by the B2-receptor antagonist Hoe 140 (1μmol/l; S2/S1: 1.14±0.11;P<0.05). Des-Arg9-bradykinin (1μmol/l), a specific B1-agonist, had no effect on stimulation-induced noradrenaline release (S2/S1: 0.94±0.08). Utilizing pharmacological interventions, we attempted to characterize the intraneuronal signal transduction pathway mediating the effect of bradykinin on exocytosis. Neither inhibition of cyclooxygenase nor blockade of nitric oxide synthesis affected bradykinin-induced stimulation of noradrenaline release. Likewise, inhibition of protein kinase C by bisindolylmaleimide (1μmol/l) or tyrosine kinase by genistein (10μmol/l) had no effect on the promoting action of bradykinin. In contrast, inhibition of cytosolic phospholipase A2activity by the specific inhibitor AACOCF3(1μmol/l) prevented bradykinin-induced increase in noradrenaline release (S2/S1: 1.09±0.15;P<0.01). In conclusion, bradykinin increases exocytotic release of endogenous noradrenaline from cardiac sympathetic neurons via activation of presynaptic B2-receptors. Intraneuronal coupling of B2-receptors to phospholipase A2appears to mediate the facilitatory effect of bradykinin on noradrenaline release in rat heart.

Prejunctional neuropeptide Y receptors in human kidney and atrium.

The aim of our study was to characterize functionally prejunctional neuropeptide Y (NPY) receptors in human and rabbit renal cortex, as well as in human right atrium. Segments of human atrial appendages and of human and rabbit renal cortex were preincubated with [3H]noradrenaline, superfused with Krebs-Henseleit solution and stimulated electrically in superfusion chambers. The stimulation-induced outflow of radioactivity was taken as an index of endogenous noradrenaline release. The effects of subtype-selective NPY analogs on the stimulation-induced noradrenaline release were studied. NPY, its endogenous analog, peptide YY, and its C-terminal fragment, NPY13-36, but not its analog, [Leu31,Pro34]NPY, concentration dependently (1-100 nM) inhibited [3H]noradrenaline release in all tissues studied. NPY-induced inhibition of [3H]noradrenaline release in human and rabbit kidney was abolished by pretreatment with pertussis toxin. We conclude that prejunctional inhibition of noradrenaline release in human heart and human and rabbit kidney occurs through NPY receptors of the Y2 subtype, which appear to couple to a pertussis toxin-sensitive G protein.

Second messenger pathways in the modulation of neurotransmitter release.

Activation of receptors on postganglionic sympathetic nerve endings can alter the amount of noradrenaline release during a train of nerve impulses. These changes may be produced by the enzyme-linked synthesis of second messenger molecules within the nerve terminal. Cyclic AMP analogues enhance noradrenaline release and two hormones adrenaline and ACTH appear to enhance noradrenaline release through activation of adenylate cyclase. Activation of the phospholipase C/protein kinase C pathway also elevates stimulation-induced noradrenaline release and angiotensin enhancement of noradrenaline release appears to act through this pathway. On the other hand, receptors which inhibit noradrenaline release (alpha 2-adrenoceptors, muscarinic M2 receptors and neuropeptide Y receptors) do not act through either of these signal transduction pathways. Since these inhibitory systems are neurotransmitter activated and relay information on a nerve pulse to nerve pulse time scale back to the nerve ending a fast activation and deactivation rate of modulation is required. This may be better served by direct modulation of ion channels without a slow intervening enzyme step. Activation of protein kinase C by phorbol esters produces relatively large increases (two-threefold) in stimulation-induced noradrenaline release and this enzyme may also have a physiological role. Protein kinase C may be an appropriate target for drugs to manipulate transmitter release and development of selective activators and inhibitors of the many protein kinase C isoenzymes may prove clinically useful in diseases with inappropriate transmitter release profiles.