Noradrenergic Nerve Terminals Research Articles

INHIBITION OF SYMPATHETIC NORADRENERGIC TRANSMISSION BY GUANABENZ AND GUANETHIDINE IN RAT ISOLATED MESENTERIC ARTERY: INVOLVEMENT OF NEURONAL POTASSIUM CHANNELS

The present study investigated the effects of the α 2-adrenoceptor agonist guanabenz and the adrenergic neurone blocking drug guanethidine on the resting and stimulation-induced (S-I) effluxes of radioactivity from rat isolated mesenteric artery preparations in which the noradrenergic transmitter stores had been radiolabelled with [ 3H]-noradrenaline. The efflux of radioactivity evoked by electrical field stimulation of periarterial sympathetic nerves (60s trains of 1ms pulses, 2Hz, 12V) was taken as an index of transmitter noradrenaline release. Guanabenz (0.1–10μ M) decreased, in a concentration-dependent manner, both the resting and S-I effluxes of radioactivity. Guanethidine (0.1 and 1μ M) also decreased S-I efflux but increased resting efflux, both effects being concentration dependent. The inhibitory effects of guanabenz on both resting and S-I effluxes were reduced by blockade of the neuronal amine carrier with desipramine (1μ M). The inhibitory effect of guanabenz on resting efflux was prevented by inhibition of monoamine oxidase with pargyline (100μ M). The inhibitory effect of guanabenz on S-I efflux was not due to activation of prejunctional α 2-adrenoceptors since the inhibition was not blocked by the selective α 2-adrenoceptor antagonist idazoxan (0.1μ M). However, the inhibitory effect of guanabenz and guanethidine on S-I efflux was reduced by the inhibitor of Ca 2+-activated potassium channels apamin (0.1μ M). The findings suggest that guanabenz, like guanethidine, enters noradrenergic nerve terminals by the neuronal amine carrier. The inhibition of resting efflux produced by guanabenz may be due to inhibition of neuronal monoamine oxidase. The enhancement of resting efflux produced by guanethidine is attributable to its indirect sympathomimetic action. Finally, guanabenz and guanethidine may inhibit transmitter noradrenaline release by activating potassium channels on sympathetic noradrenergic nerve terminals. These findings may be relevant to the mechanism of adrenergic neurone blockade.

Presynaptic NMDA Receptors Display Physiological Characteristics of Homomeric Complexes of NR1 Subunits that Contain the Exon 5 Insert in the N‐Terminal Domain

The subunit composition of the N-methyl-D-aspartate (NMDA) glutamate receptor affects both its channel activity and its sensitivity to modulation by a wide variety of substances. Expression studies in oocytes and physiological studies in neurons indicate that endogenous postsynaptic NMDA receptors are heterooligomeric complexes of NR1 and NR2 subunits. To deduce the subunit composition of the presynaptic NMDA receptor on noradrenergic nerve terminals, we examined the modulation of NMDA-evoked norepinephrine (NE) release from hippocampal synaptosomes. At high glycine concentrations, the NMDA-evoked release was not potentiated by reducing reagents, low micromolar Zn2+ or Ni2+, polyamines, or 100 microM histamine. It was also not inhibited by oxidizing agents or physiological concentrations of protons but was inhibited by high micromolar Co2+, Zn2+, and Ni2+, but not Fe3+, by high micromolar ifenprodil, and by 1 mM histamine. At low glycine concentrations, it was potentiated by spermine. These characteristics are similar to those displayed by homooligomeric complexes of NR1 subunits that contain in the N-terminal domain the 21-amino-acid insert encoded by exon 5. These data provide physiological evidence that some endogenous NMDA receptor complexes may contain only the NR1 (+ exon 5) subunits.

Oxidation chemistry of (-)-norepinephrine in the presence of L-cysteine.

The noradrenergic neurotransmitter (-)-norepinephrine (1) is very easily oxidized at physiological pH to an o-quinone (2) that normally cyclizes and subsequently oxidatively polymerizes to black melanin. In this investigation it is demonstrated that L-cysteine (CySH) can divert the melanin pathway by efficiently scavenging o-quinone 2 to give, initially, 5-S-cysteinylnorepinephrine (6) and 2-S-cysteinylnorepinephrine (7). These cysteinyl conjugates are appreciably more easily oxidized than 1 to o-quinones that, in part, are further attacked by CySH to give 2,5-bi-S-cysteinylnorepinephrine (8), an even more easily oxidized compound. The o-quinone intermediates formed upon oxidation of 6-8 can also undergo facile intramolecular cyclizations to bicyclic o-quinone imines that oxidize the cysteinyl conjugates from which they are derived in a reaction sequence that leads initially to a number of dihydrobenzothiazines. At least two of these compounds, 7-(1-hydroxy-2-aminoethyl)-3,4-dihydro-5- hydroxy-2H-1,4-benzothiazine-3-carboxylic acid (9) and 8-(1-hydroxy-2-aminoethyl)-3,4-dihydro-5-hydroxy-2H-1, 4-benzothiazine-3-carboxylic acid (10) are lethal when administered into the brains of mice. The in vitro chemical pathways elucidated in this investigation might be of relevance to the depigmentation and degeneration of neuromelanin-pigmented noradrenergic cell bodies in the locus ceruleus in Parkinson's Disease and to the degeneration of noradrenergic nerve terminals in Alzheimer's Disease and following transient cerebral ischemia (stroke).

3H]Nisoxetine—A radioligand for noradrenaline reuptake sites: Correlation with inhibition of [ 3H]noradrenaline uptake and effect of DSP-4 lesioning and antidepressant treatments

Nisoxetine is a potent and selective inhibitor of noradrenaline uptake into noradrenergic neurones. [ 3H]Nisoxetine binding to rat frontal cortical membranes was of high affinity. The binding data of both competition and saturation studies fitted a single site binding model. [ 3H]Nisoxetine binding was potently inhibited by the selective noradrenaline uptake inhibitors desipramine and protriptyline. In addition, a very good correlation was obtained between the ability of 25 monoamine reuptake inhibitors and related compounds both to inhibit [ 3H]nisoxetine binding and to inhibit [ 3H]noradrenaline uptake in rat frontal cortex. DSP-4 (10–100 mg/kg, i.p.) dose-dependently depleted cortical noradrenaline concentrations (51–100%), with no significant effects on 5-HT and dopamine. These depletions, which were used as a marker of loss of noradrenergic nerve terminals, were associated with a dose-dependent decrease in the number of [ 3H]nisoxetine binding sites (20–97%) with no change in binding affinity. Furthermore, a good correlation was obtained between cortical noradrenaline concentrations and the number of [ 3H]nisoxetine binding sites. These data support the view that [ 3H]nisoxetine binds to a single population of homogeneous sites associated with the noradrenaline transporter complex. Using this ligand, the effects of repeated administration of both antidepressant drugs with a range of pharmacological actions and of electroconvulsive shock on noradrenaline reuptake sites were examined. The number and affinity of [ 3H]nisoxetine binding sites were unaltered by all treatments. It is unlikely, therefore, that antidepressant therapy would produce adaptive changes in noradrenaline uptake sites.

Decrease of anesthetics activity by electroacupuncture on Jen-Chung point in rabbits

The effect of acupuncture at life-saving point on the central nervous depressive action of anesthetics was investigated in rabbits. Stimulation with electroacupuncture (EA) inserted in Jen-Chung point, which is located at the mid-point on the upper lip, decreased the sleeping time induced by pentobarbital or propofol. However, this action of acupuncture was not modified by naloxone at the doses sufficient to block opiate receptors. Plasma β-endorphin detected by radioimmunoassay was also not markedly changed in rabbits which received similar electrostimulation. Moreover, pretreatment with para-chlorophenylalanine at a dose sufficient to deplete endogenous 5-hydroxytryptamine (5-HT) failed to influence the action of EA. Mediation of endogenous opioids and/or 5-HT in this action of EA was then ruled out. Prazosin reversed the sleeping time decreasing action of acupuncture in a dose-dependent manner. Also, the action of acupuncture was eliminated in rabbits which received intracerebroventricular injection of guanethidine at a dose which could block noradrenergic nerve terminals. It is suggested that stimulation of Jen-Chung point through EA can activate noradrenergic neurotransmission in the brain, which in turn reduces the central nervous depressive activity of anesthetics.

Subcellular localization of synaptophysin in noradrenergic nerve terminals: a biochemical and morphological study.

The subcellular localization of synaptophysin was investigated in noradrenergic nerve terminals of bovine vas deferens and dog spleen and compared with membrane-bound and soluble markers of noradrenergic storage vesicles. At the light microscopical level chromogranin A- and cytochrome b561-immunoreactivity revealed an identical and very dense innervation of the entire vas deferens. In the case of synaptophysin, most immunoreactivity was found only in the outmost varicosities closest to the lumen, which were also positive for chromogranin A. Small dense-core vesicles of dog spleen were purified using a combination of velocity gradient centrifugation and size exclusion chromatography. Small dense-core vesicles were enriched 64 times as measured by the noradrenaline content. Enrichments for dopamine-beta-hydroxylase were in a similar range. Synaptophysin-containing vesicles were smaller in size and they did not contain the typical noradrenergic markers dopamine-beta-hydroxylase, cytochrome b561, and noradrenaline. Instead, they might store adenosine triphosphate (ATP). A greater part of synaptophysin immunoreactivity was consistently found at high sucrose densities at the position of large dense-core vesicles. We conclude that in the noradrenergic nerve terminal: (1) small dense-core vesicles have a membrane composition similar to large dense-core vesicles, indicating that the former are derived from the latter, and (2) synaptophysin seems not to be present on small dense-core vesicles. We suggest the possibility that synaptophysin-containing vesicles form a residual population whose role in neurotransmission has been taken over by large and small dense-core vesicles following noradrenergic differentiation.

4-Aminopyridine-induced phasic contractions in rat caudal epididymis are mediated through release of noradrenaline

4-Aminopyridine, a K + channel blocker, evoked phasic contractions in the caudal duct of the rat epididymis. The 4-aminopyridine-induced contractile response was either inhibited or prevented by the α 1-adrenoceptor antagonists, prazosin (IC 50 = 2.7 nM) and benoxathian (IC 50 = 14.6 nM). Blockers (1 μM) of α 2-adrenoceptors and purinoceptors but not of β-adrenoceptors or muscarinic receptors caused a small but statistically significant reduction of the 4-aminopyridine-induced response. 4-Aminopyridine lost its ability to induce contractions after noradrenergic nerves had been destroyed by 6-hydroxydopamine. In addition, protriptyline and xylamine, blockers of noradrenaline uptake, also inhibited the 4-aminopyridine-induced contractile response. However, other putative K + channel blockers (tetraethylammonium ion, quinine, quinidine and glibenclamide) did not cause the muscle to contract. These findings demonstrate that the 4-aminopyridine-induced release of noradrenaline and adenosine 5′-triphosphate as co-transmitters results from membrane depolarization due to 4-aminopyridine blockade of K + channels in noradrenergic nerve terminals. The 4-aminopyridine-sensitive K + channels might thus play a physiological role in regulating the nerve membrane potential and neurotransmission in the rat caudal epididymis.

Histamine, a neuromodulator of noradrenergic transmission in uterine horns from mice in diestrus

The effect of histamine on [ 3H]norepinephrine ([ 3H]NE) release in uterine horns from mice in estrous and diestrous states was studied under two different experimental conditions: resting NE release and stimulus-evoked NE release. It was found that [ 3H]NE release was higher for the diestrous state under both resting and stimulus-evoked (100 mM K + and electrical stimulus) conditions. Histamine only potentiated the stimulus-evoked [ 3H]NE outflow in uterine horns from mice in the diestrous state and from ovariectomized mice treated with progesterone. This effect was dose dependent and was antagonized by H 1 but not by H 2 or H 3 antagonists. R- α-Methylhistamine, a H 3 agonist, had no effect on stimulus-evoked [ 3H]NE release. According to these results, it could be concluded that: (a) histamine regulates the NE release from noradrenergic nerve terminals in uterine tissues; (b) this heterologous regulation depends on progesterone predominance and on terminal depolarization; and (c) presynaptic H 1 receptors located on noradrenergic terminals could be responsible for such an effect.

Participation of noradrenergic neurotransmission in the enhancement of baroreceptor reflex response by substance P at the nucleus tractus solitarii of the rat: a reverse microdialysis study.

We applied reverse microdialysis and HPLC analysis to evaluate the participation of noradrenergic neurotransmission in modulation of the baroreceptor reflex response by substance P at the nucleus tractus solitarii in Sprague-Dawley rats anesthetized with pentobarbital sodium (50 mg/kg, i.p., with 20 mg/kg/h.i.v. supplement). Continuous infusion of substance P (600 microM) at 1 microliter/min into the nucleus tractus solitarii through a stereotaxically positioned microdialysis probe (active exchange length, 180-200 microns; diameter, 220 microns) for 1 h elicited an enhancement of the baroreceptor reflex response. This facilitatory effect correlated positively, during the 60-min infusion period, with the time course of increase in the extracellular concentration of substance P and noradrenaline in the nucleus tractus solitarii. Experimentally elevating the concentration of noradrenaline at this medullary nucleus also augmented the baroreceptor reflex sensitivity. On the other hand, depletion of the noradrenergic fibers and nerve terminals at the nucleus tractus solitarii with DSP4 diminished the enhancement of baroreceptor reflex response and the corresponding elevation in extracellular concentration of noradrenaline by substance P. Microinfusion of noradrenaline into the nucleus tractus solitarii in DSP4-treated animals, however, potentiated the baroreceptor reflex response. These results suggested that the enhancement of baroreceptor reflex response by substance P may involve an increase in the concentration of noradrenaline at the nucleus tractus solitarii via a presynaptic mechanism.

Sigma receptor regulation of norepinephrine release from rat hippocampal slices

Multiple sigma receptor subtypes have been identified in the hippocampus, yet their physiological role remains largely undefined. In the current study, we examined the role of sigma receptors in the regulation of N-methyl- d-aspartate (NMDA)-stimulated [ 3H]norepinephrine ([ 3H]NE) release from rat hippocampal slices. Both sigma agonists (+)pentazocine and BD737 inhibited stimulated norepinephrine release in a concentration-dependent manner. The sigma 1 antagonist DuP 734 completely antagonized the inhibition of release by all concentrations of BD737 tested. However, DuP 734 only partially reversed inhibition of release by (+)pentazocine concentrations above 100 nM. 1,3 Di- o-tolylguanidine (DTG), but not haloperidol, antagonized BD737-mediated inhibition of release. DTG also completely antagonized inhibition of release by 100 nM (+)pentazocine yet haloperidol produced only a partial reversal. A combination of DuP 734 and haloperidol produced complete reversal of (+)pentazocine-mediated inhibition, suggesting potential involvement of multiple sigma receptor subtypes in the regulation of norepinephrine release. Both (+)pentazocine and BD737 failed to inhibit stimulated release in the presence of tetrodotoxin, suggesting that sigma receptors regulating NE release are not located on noradrenergic nerve terminals. These results suggest that sigma receptors may be a therapeutic target for disorders resulting from noradrenergic imbalance in hippocampus.

Hypothalamic changes in norepinephrine release in rats with estradiol valerate-induced polycystic ovaries.

Chronic anovulation and polycystic ovaries (PCO) can be induced by a single i.m. injection of estradiol valerate (EV, 2 mg in oil) in the rat. Constant exposure to high plasma levels of estradiol provokes a neurotoxic effect on the hypothalamic neurons, including those from the arcuate nucleus. Because of the important participation of hypothalamic norepinephrine (NE) in the regulation of GnRH release and the possible noxious effect of prolonged exposure of these neurons to estradiol, our interest was to study the activity of the noradrenergic neurons innervating the hypothalamus. We analyzed the biosynthesis, content, and release of NE from the noradrenergic nerve terminals of the hypothalamus during the PCO condition. We found a decrease in tyrosine hydroxylase (TH) activity and in the content of dopamine (DA) in the anterior hypothalamus after 2 mo of EV injection, whereas dopamine-beta-hydroxylase (D beta H) was increased without changes in NE content. No variations in TH activity or in DA and NE contents in the medial hypothalamus were observed, but a decrease in D beta H activity was evident. After 2 mo of EV administration, an increase in the electrically induced release of NE from anterior hypothalamic blocks incubated in vitro was detected; this effect was not evidenced in the medial hypothalamus. After 5 mo of EV administration, release of NE increased in anterior hypothalamic blocks but decreased in medial hypothalamic tissue. The inhibitory effect of morphine on NE release found in control animals was increased in the hypothalamus from PCO rats, suggesting an increased number of mu-opioid binding sites in noradrenergic neurons.(ABSTRACT TRUNCATED AT 250 WORDS)

Y2‐receptor‐mediated selective inhibition of slow, inhibitory postsynaptic potential in submucous neurones of guinea‐pig caecum

1. The subtype of neuropeptide Y receptor mediating the selective inhibition of the slow inhibitory postsynaptic potential (i.p.s.p.) of submucous neurones in guinea-pig caecum was investigated by use of conventional intracellular electrophysiological recording techniques. 2. Neuropeptide Y (NPY) (1-300 nM) was found to depress or abolish reversibly the slow i.p.s.p. evoked by focal stimulation of internodal fibre tracts. At low concentrations (1-30 nM), a reduction in the duration of the slow i.p.s.p. was often apparent before any inhibition of the amplitude of this synaptic potential. 3. These inhibitory effects of NPY were mimicked by peptide YY (PYY; 0.3-100 nM), NPY13-36 (1-300 nM) and NPY22-36 (10-100 nM); [Leu31,Pro34]NPY ([Pro34]NPY) and bovine pancreatic polypeptide (bPP) were without pre- or postsynaptic effects at concentrations of up to 300 nM. The IC50 +/- s.e. mean values for PYY, NPY, and NPY13-36 were 2.7 +/- 0.3, 7.8 +/- 2.1 and 30 +/- 4.8 nM, respectively, and were significantly different from each other. Thus, the apparent rank order of potency was PYY > NPY > NPY13-36 >> [Pro34]NPY and bPP. 4. In concentrations of up to 300 nM, NPY and its analogues had no depressant effects on the active and passive properties of the impaled neurone and did not affect the amplitude or duration of either cholinergic fast synaptic potentials or non-cholinergic, slow excitatory postsynaptic potentials (e.p.s.ps). Furthermore, none of these peptides altered the amplitude or time-course of changes in membrane potential induced by focal application of acetylcholine or noradrenaline. 5. It is, therefore, concluded that the selective inhibition of the slow i.p.s.p. is mediated by Y2-receptors,located presynaptically on noradrenergic nerve terminals.

Punicalagin‐induced release of norepinephrine reverses orthostatic hypotension in rats

AbstractPunicalagain, an active principle isolated from the leaves of Lumnitzera racemosa (Combretaceae), reversed the fall in arterial blood pressure in conscious Wistar rats with orthostatic hypotension induced by injection of hexamethonium into animals subjected to 90° head‐up tilts for 60 s. Punicalagin was found to be less effective on prazosin‐induced orthostatic hypotension. Plasma norepinephrine (NE) was elevated by intravenous injection of punicalagin in a dose‐dependent manner from 0.5 to 15 mg/kg, both in normotensive rats and the orthostatic hypotensive animals. An increase of mean blood pressure was also found in rats that received the injection of punicalagin; this increase in blood pressure was totally abolished by prazosin. Failure of hexamethonium or pentolinium, antagonists of ganglionic nicotinic receptors, to influence the NE releasing action of punicalagin ruled out the participation of ganglionic stimulation. This NE‐releasing action of punicalagin was, however, totally blocked by the inhibitors of noradrenergic nerve terminals, guanethidine or bretylium. Also, the activity of punicalagin was not modified by adrenalectomy. Thus, a direct release of NE from the noradrenergic nerve terminals by punicalagin seems responsible for the reversal of orthostatic hypotension.

Influence of acetonylgeraniin, a hydrolyzable tannin from Euphoria longana, on orthostatic hypotension in a rat model.

Acetonylgeraniin, an active principle isolated from the seeds of Euphoria longana Lam. (Sapindaceae), reversed the fall in arterial blood pressure in conscious hypertensive rats (SHRs) with orthostatic hypotension induced by injection of hexamethonium into animals subjected to 90 degrees head-up tilts for 60 seconds. However, acetonylgeraniin failed to affect prazosin-induced orthostatic hypotension. Plasma noradrenaline (NA) and mean blood pressure were elevated dose-dependently by an intravenous injection of acetonyl-geraniin into the rats; this increase in blood pressure was totally abolished by prazosin. Failure of hexamethonium or pentolinium, the blockers of ganglionic nicotinic receptors, to influence the NA releasing action of acetonylgeraniin ruled out the participation of ganglionic stimulation. This NA-releasing action of acetonylgeraniin was, however, totally abolished by the inhibitors of noradrenergic nerve terminals, guanethidine or bretylium. Also, the activity of this tannin was not modified by adrenalectomy. Thus, a direct release of NA from the noradrenergic nerve terminals by acetonylgeraniin seems responsible for the reversing of orthostatic hypotension.

Trypanosoma cruzi infection in rats induced early lesion of the heart noradrenergic nerve terminals by a complement-independent mechanism.

The acute phase of the experimental Chagas' disease in rats induces extensive lesion of the heart sympathetic nerve terminals. Because of evidence indicating the involvement of immune reactions in neuron destruction provoked by Chagas' disease, we tested the effects of depleting the complement system by cobra venom factor upon the sympathetic denervation. The serum hemolytic activity against sensitized sheep erythrocytes ensured the efficacy of the anticomplementary treatment. Glyoxylic acid-induced histofluorescence and electron-microscopic methods allowed the study of the heart noradrenergic nerves. T. cruzi infection caused marked rarefaction of fluorescent nerve terminals at day 10 of infection and the ultrastructural study guaranteed that this rarefaction involved lesion of noradrenergic terminals. The complement depletion failed to prevent this early heart noradrenergic denervation, excluding the participation of complement-mediated lysis as a main mechanism.

Lack of catecholamine involvement in the increased luteinizing hormone release due to blockade of κ-opioid receptors in the medial basal hypothalamus during midpregnancy in the rat

Blockade of κ-opioid receptors in the medial basal hypothalamus (MBH) with nor-binaltorphimine (nor-BNI) stimulates luteinizing hormone (LH) release during midpregnancy in the rat [48]. The objective of this study was to determine whether norepinephrine (NE) or dopamine (DA) mediates the LH response to blockade of MBH κ-opioid receptors on days 13–17 of pregnancy in the rat. Two experiments were conducted. In the first, push-pull perfusion in conjunction with HPLC was used to monitor in vivo NE release in the MBH occuring in response to (a) artificial CSF followed by CSF containing nor-BNI (40 μg/h), (b) desipramine (DMI, a NE reuptake blocker, 10 μM) in CSF followed by DMI, and (c) DMI followed by DMI + nor-BNI. Blood samples were taken at 12 min intervals concurrent with push-pull perfusate samples. Plasma LH levels were determined by RIA. Nor-BNI significantly increased LH release compared to CSF alone, but perfusate NE was undetectable in either perfusion period. However, perfusion with CSF containing 100 mM K + in these rats markedly increased perfusate NE levels, indicating noradrenergic nerve terminals were present at the perfusion sites in the MBH. Addition of DMI to the CSF significantly increased perfusate NE levels, but produced no changhe in LH release. Nor-BNI + DMI perfusion increased LH secretion similar to nor-BNI alone, but produced no additional increase in MBH perfusate NE levels compared to perfusion with DMI alone. In the second experiment, push-pull perfusion in the MBH with nor-BNI was done in rats pretreated either with the NE synthesis inhibitor FLA-63 (25 mg/kg, s.c.), the α-adrenergic receptor blocker phentolamine (5 mg/kg, i.v.), the DA receptor antagonist d-butaclamol (1 mg/kg, s.c.), or vehicle. None of these drug treatments affected the increased LH release occuring during MBH perfusion with nor-BNI. The present results demonstrate that neither NE nor DA mediates the increased LH release occuring in response to blockade of κ-opioid receptors in the MBH during midpregnancy in the rat.

Mu-Opioid receptors modulate noradrenaline release from the rat hippocampus as measured by brain microdialysis

The modulation of noradrenaline (NA) release via presynaptic opioid receptors in the hippocampus of freely moving rats was studied by the use of brain microdialysis. Extracellular levels of NA were estimated by assaying its concentrations in the perfusion fluid using high-performance liquid chromatography (HPLC) with electrochemical detection (ECD). Spontaneous NA levels were reduced by tetrodotoxin (1 μM) co-perfusion and were increased by peripheral administration of desipramine (5 and 10 mg/kg, i.p.). Addition of potassium (K +, 60 and 120 mM) to the perfusion fluid evoked a concentration-dependent release of NA. K + (120 mM)-evoked NA release was markedly reduced by removal of calcium (Ca 2+) from the perfusion fluid. These results indicate that both the spontaneous and the K +-evoked NA release measured by the use of brain microdialysis coupled with HPLC-ECD can be used as indices of neuronal release from the noradrenergic nerve terminals. A μ-opioid receptor agonist, morphine (0.01–10 μM), when co-perfused with K + (120 mM), produced a reduction of K +-evoked NA release in a concentration-dependent manner. Neither co-perfusion with a high concentration of [ d-Pen 2, d-Pen 5]-enkephalin (DPDPE) (10 μM), an agonist selective for δ-opioid receptors, nor with U-69593 (10 μM), an agonist selective for κ-opioid receptors, modified the K + (120 mM)-evoked release of NA. Morphine-induced (1 μM) inhibition of NA release was blocked by a μ-opioid receptor antagonist, naltrexone (3 and 9 mg/kg, i.p). Naltrexone by itself did not alter the spontaneous NA levels or the K +-evoked NA release. The morphine-induced (1 μM) inhibition of NA release was abolished by pretreatment with pertussis toxin (PTX) that was perfused into the hippocampus by push-pull cannulation. These findings suggest that NA release is functionally modulated via the μ-opioid receptors located on noradrenergic nerve terminals in the rat hippocampus. It indicates, furthermore, the possibility that the inhibition of NA release via the μ-opioid receptors is linked to G-proteins which are substrates of PTX.

Close microtopographical relationships between sympathetic nerve terminals and bulbous process endings of pinealocytes in the pineal gland of the Mongolian gerbil.

Previous studies have shown that pinealocytes of the gerbil pineal gland exhibit processes that form terminal swellings filled with abundant electron-lucent microvesicles. The membrane of these presumptive secretory microvesicles is known to contain synaptophysin, a major integral glycoprotein of neuronal synaptic vesicles. The present study was conducted to evaluate the microtopographical relationships between the vesicle-rich process swellings and intra-pineal nerve terminals. For this purpose, both nerve terminals and pinealocyte process endings were visualized immunohistochemically in the same semi-thin sections of plastic-embedded gerbil pineals, using antibodies directed against synaptophysin. This approach consistently revealed close spatial associations of punctate immunopositive nerve endings with intensity stained bulbous process terminals of pinealocytes in or near the perivascular spaces. The light-microscopic observations of intimate neuronal-pinealocytic relationships were corroborated at the electron-microscopic level. Perivascular varicosities with ultrastructural features characteristic of sympathetic nerve terminals were frequently juxtaposed to vesicle-filled process endings of pinealocytes. Analysis of serial thin sections showed that multiple point-to-point contacts are encountered between noradrenergic nerve terminals and pinealocytic process swellings. Our morphological findings imply that bulbous process terminals, at least in the gerbil pineal gland, are major targets for the neuronal control of the secretory activity of pinealocytes.

Methamphetamine antagonistic property of (+)- and (−)-4-phenyltetrahydroisoquinoline in rat anococcygeus muscle

The antagonistic effects of (+)- and (−)-4-phenyl-1,2,3,4-tetrahydroisoquinoline (4PTIQ) on methamphetamine in the rat anococcygeus muscle were compared with those of cocaine and nomifensine. Methamphetamine contracted the anococcygeus muscle through the release of norepinephrine from noradrenergic nerve terminals. (+)-4PTIQ inhibited the methamphetamine-induced contractions more strongly than cocaine and nomifensine. (+)-4PTIQ had no potentiating effects on exogenous norepinephrine-induced contraction, which was considered to be an index of amine neuronal uptake blockade. On the other hand, (−)-4PTIQ, cocaine and nomifensine produced a significant leftward shift of the norepinephrine concentration-response curve, i.e. they showed a strong blocking effect on amine neuronal uptake. These results suggest that the inhibitory effects of (+)-4PTIQ on the action of methamphetamine are mediated by a mechanism other than inhibition of amine neuronal uptake.

Cardiac noradrenergic nerve terminal abnormalities in dogs with experimental congestive heart failure.

We have shown previously that norepinephrine (NE) uptake activity is reduced in the failing right ventricle of animals with right heart failure (RHF) produced by tricuspid avulsion and progressive pulmonary constriction. However, it is unknown whether this defect in neuronal NE uptake is related to reduction of noradrenergic nerve terminals or whether these changes also occur in animals with left heart failure (LHF). It is also unknown whether increased NE release in heart failure contributes to the noradrenergic nerve abnormalities. We measured myocardial NE content. NE uptake function, and noradrenergic nerve profiles in dogs with either RHF or LHF induced by rapid ventricular pacing. NE uptake activity was measured using [3H]NE, and noradrenergic nerve profiles were visualized by glyoxylic acid (SPG)-induced histofluorescence and tyrosine hydroxylase immunocytochemical staining. To study the effects of excess NE, we exposed normal dogs to 8 weeks of chronic NE infusion using subcutaneous osmotic minipumps. RHF and LHF animals exhibited reduced myocardial contractile function and congestive heart failure, as evidence by reduced cardiac output and elevated right atrial pressure. However, unlike that in LHF, left atrial pressure was not increased in RHF. The animals also showed an increase in plasma NE and a decrease in cardiac NE. In addition, SPG-induced histofluorescence correlated significantly with NE uptake activity (r = .712, P < .001) and tyrosine hydroxylase immunoreactive profiles (r = .569, P < .001) in the right ventricles of RHF dogs and in both ventricles of LHF dogs. The numbers of catecholaminergic profiles and tyrosine hydroxylase profiles significantly correlated with cardiac filling pressures. Chronic infusion of NE decreased heart rate in normal dogs but had no effect on either mean aortic pressure or left atrial pressure; like heart failure, it resulted in significant decreases in myocardial NE uptake activity and numbers of SPG-induced catecholaminergic histofluorescence and immunoreactive tyrosine hydroxylase profiles. Myocardial NE uptake activity was reduced only in the failing ventricles with elevated filling pressure in RHF and LHF. These changes probably were caused by loss of noradrenergic nerve terminals in the failing ventricles, as evidenced by the reductions of catecholaminergic histofluorescence and tyrosine hydroxylase immunostained profiles. Furthermore, since similar reductions of myocardial NE uptake and noradrenergic nerve profiles could be produced by chronic NE infusion in normal dogs, elevated NE levels may play a role in the development of cardiac noradrenergic nerve abnormalities in congestive heart failure.