Arun R. Wakade

Wayne State University | WSU · Department of Pharmacology

Inhibitory effect of adenosine on the isolated heart muscle and vascular system were first described in 1929. Since then, numerous reviews have been published on the diverse actions of this nucleoside on a wide variety of cell types. Essentially all effects of adenosine in neurons and non-neuronal cells are mediated by activation of nucleoside memb...

The novel chromogranin A fragment catestatin (bovine chromogranin A(344-364); RSMRLSFRARGYGFRGPGLQL) is a potent inhibitor of catecholamine release (IC50, approximately 0.2-0.3 microM) by acting as a nicotinic cholinergic antagonist. To define the minimal active region within catestatin, we tested the potencies of synthetic serial three-residue del...

Previous work has shown that nucleosides produce apoptosis in sympathetic ganglion (SG) cells in vitro. The present study examined the effects of nucleosides on the development of the chick embryo in vivo with special attention to the SG and the optic tectum of the central nervous system. In the presence of an adenosine deaminase inhibitor, adenosi...

Removal of nerve growth factor (NGF) from sympathetic neurons initiates a neuronal death program and apoptosis. We show that pituitary adenylyl cyclase-activating polypeptide (PACAP) prevents apoptosis in NGF-deprived sympathetic neurons. PACAP (100 nM) added to culture medium at the time of plating failed to support neuronal survival. However, in...

Our past work on nucleoside toxicity in sympathetic neurons has clearly revealed that adenosine and 2'-deoxyadenosine (dAdo) have different mechanisms of action in inducing apoptotic death. For example, adenosine is toxic to neurons only during early phase of growth whereas dAdo kills even mature neurons. In this study, we hypothesize that dAdo-ind...

Our previous work has established that adenosine is toxic to chick embryonic sympathetic neurons and kills freshly plated neurons by a process of apoptosis. Although the exact mechanism remains unknown, we found that phosphorylation of adenosine was essential to the toxicity. Using markers for RNA ([3H]uridine) and protein ([35S]methionine) synthes...

Recently, we have demonstrated that adenosine and 2'-deoxyadenosine are toxic to embryonic sympathetic neurons and proposed that purine and pyrimidine metabolism may play a critical role in the growth and development of sympathetic neurons. To extend this hypothesis further, we examined the effects of these nucleosides on two other neuronal populat...

An increase in the release of norepinephrine (NE) when an identical number of stimulating pulses are delivered at increasing frequencies was first observed in perfused spleen of a cat. A classic agent used to increase neuronal action potential duration is tetraethylammonium (TEA), which selectively inhibits repolarizing K+ currents. The use of cell...

Chronic activation of protein kinase C (PKC) has been implicated in regulation of Ca2+ entry responsible for normal development of transmitter properties in cultured sympathetic neurons. The idea that PKC alters the expression of Ca2+ channels was tested using phorbol 12,13-dibutyrate (PDB) which activates PKC and also supports survival of chick sy...

We show here that 2'-deoxyadenosine (2'-dAdo) but not adenosine was toxic to chromaffin cells of 3-4-week-old rat adrenal glands. More than 75% of the cells plated in culture gradually died over a 3-day period in the presence of 100 microM 2'-dAdo plus 3 microM deoxycoformycin (DCF). Morphological observations together with bisbenzimide staining an...

Increasing evidence suggests that adenosine and ATP not only modulate cell growth and differentiation, but may also act as inducers of cell death. In the session “Purines and cell death,” held during the Purines '96 Symposium and chaired by Claudio Franceschi (Modena, Italy) and Geoffrey Burnstock (London, UK) presentation and discussion of new stu...

We demonstrate that 1-methyl-4-phenylpyridinium (MPP+) is toxic to chick peripheral sympathetic neurons maintained in culture in the presence of nerve growth factor (NGF). When MPP+ was added to the culture medium at the time the neurons were plated, cell loss after 3 days in culture was evident at concentrations as low as 3 nM, and near maximal at...

These experiments characterize the nucleoside transport and quantify the neurotoxicity of adenosine and 2'-deoxyadenosine (dAdo) in chick sympathetic neurons. We show that [3H]adenosine transport was sensitive to low temperature, specific inhibitors of nucleoside transport, and an excess concentration of adenosine. However, many of these treatments...

1. The biochemical basis for differences in noradrenaline (NA) transporter function between chromaffin cells in the adrenal medulla and those maintained in primary culture was investigated. 2. Intact adrenal medullae of neonatal rats accumulated small amounts of [3H]NA. In contrast, dissociated chromaffin cells placed in culture for 2-6 days accumu...

We used cultured rat chromaffin cells to test the hypothesis that Ca2+ entry but not release from internal stores is utilized for exocytosis. Two protocols were used to identify internal versus external Ca2+ sources: (a) Ca2+ surrounding single cells was transiently displaced by applying agonist with or without Ca2+ from an ejection pipette. (b) In...

The hypothesis that multiple trophic inputs are essential for normal development of transmitter release properties in sympathetic neurons was tested using two supportive agents (excess KCl and phorbol 12,13-dibutyrate which produce marked activation of protein kinase C and also support survival of chick sympathetic neurons in culture) in addition t...

Pituitary adenylate cyclase-activating polypeptide (PACAP) is the most potent non-cholinergic neurotransmitter to stimulate catecholamine secretion from rat chromaffin cells; however, the mechanism of action is not clear. We used amperometric detection of exocytosis and indo-1 monitoring of [Ca] to identify PACAP actions in cultured chromaffin cell...

Rat adrenal chromaffin cells were invested by a dense network of nerve fibers immunoreactive to pituitary adenylate cyclase activating polypeptide-38 (PACAP-IR). Immunohistochemical studies demonstrated the presence of PACAP-IR in nodose and dorsal root ganglion cells, but not in neurons of the intermediolateral cell column and other autonomic nucl...

1. The contribution of target cells in controlling the functional properties of sympathetic neurons was investigated using pure neuronal cultures and co-cultures of neurons with their physiological target cells. 2. Chick embryo sympathetic neurons cultured alone exhibited maximal elevation of cytosolic free Ca2+ ([Ca2+]i) and release of tritiated n...

1. A newly found action of adenosine in neurons, which may have an important physiological function in the growth and development of the sympathetic nervous system, is described. Adenosine (1-100 microM) inhibited neurite outgrowth within the first 24 h and killed about 80% of sympathetic neurons supported by nerve growth factor over the next 2 day...

The stimulated elevation of [Ca2+]i can either promote neuronal survival or lead to Ca(2+)-mediated neurotoxicity. Similarly, growth cone mobility and neurite outgrowth may be promoted or arrested by elevated [Ca2+]i. We examined survival, development and transmitter release properties of chick sympathetic neurons maintained in culture medium conta...

Postmitotic sympathetic neurons are known to undergo a programmed cell death (apoptosis) when they are deprived of nerve growth factor (NGF) or treated with arabinofuranosyl nucleoside antimetabolites. Here we report the existence of a biochemical mechanism for the induction of neuronal death by an endogenous nucleoside in the presence of NGF. In s...

1. Rat adrenal medulla is stimulated by cholinergic and peptidergic transmitters released from splanchnic nerves. The peptidergic transmitter has been identified as vasoactive intestinal polypeptide (VIP) and its contribution in comparison to that of acetylcholine (ACh) is more prominent at low neuronal activity. The purpose of this study is to det...

Secretion of catecholamines from chromaffin cells is mediated by cholinergic and peptidergic neurotransmitters. The cholinergic transmitter acetylcholine activates both nicotinic and muscarinic receptors to trigger catecholamine secretion in rat adrenal medulla. Vasoactive intestinal polypeptide (VIP) has been identified as the peptidergic transmit...

A single stimulus applied to sympathetic neurons co-cultured with cardiac cells produced a very small increase in the release of tritiated norepinephrine and intracellular free Ca2+ concentration ([Ca2+]i), but evoked a typical neuronal action potential. Treatment with 10 mM tetraethylammonium caused a dramatic increase in the responses to a single...

Entry of Ca2+ through voltage-dependent Ca2+ channels is known to be linked to the exocytotic release of transmitter from sympathetic neurons. In this paper we provide evidence that transmitter release can also be stimulated by Ca2+ influx via the Na-Ca exchanger. Furthermore, the release linked to Na-Ca exchange is regulated by cardiac target cell...

Fluorescence imaging of indo-1 loaded cells was used to monitor influx and distribution of Ca²⁺ in cell bodies, neurites and growth cones of sympathetic neurons cultured from embryonic chick. Similar experiments on release of tritiated noradrenaline were performed to assess the relationship between intracellular Ca²⁺ concentration ([Ca²⁺]) and tran...

The concentration of cytosolic free Ca2+ ([Ca2+]i) and the release of tritiated norepinephrine ([3H]NE) were monitored during Ba2+ stimulation of sympathetic neurons cultured from chick embryos. Ba2+ (2.5 mM in Ca(2+)-free medium) caused a rise in [Ca2+]i in all regions (cell bodies, neurites, and growth cones) of sympathetic neurons and evoked [3H...

A study of the effects of dihydropyridine Ca2+ channel modulators on the release of catecholamines from perfused rat adrenal glands, evoked by electrical stimulation of their splanchnic nerves, is presented. Electrically mediated secretory responses were compared to chemically mediated responses (exogenous acetylcholine, nicotine, or high K+). Inte...

We studied the effects of lanthanum (La3+) on the release of 3H-norepinephrine(3H-NE), intracellular Ca2+ concentration, and voltage clamped Ca2+ and K+ currents in cultured sympathetic neurons. La3+ (0.1 to 10 μm) produced concentration-dependent inhibition of depolarization induced Ca2+ influx and 3H-NE release. La3+ was more potent and more effi...

Tryptic digestion of tyrosine hydroxylase (TH) isolated from rat adrenal glands labeled with 32Pi produced five phosphopeptides. Based on the correspondence of these phosphopeptides with those identified in TH from rat pheochromocytoma cells, four phosphorylation sites (Ser8, Ser19, Ser31, and Ser40) were inferred. Field stimulation of the splanchn...

1. The perfused adrenal gland of the rat was used to establish the identity of a non-cholinergic substance involved in splanchnic nerve-mediated secretion of catecholamines. 2. The perfused adrenal medulla was rich in vasoactive intestinal polypeptide (VIP) content (28 pmol g-1 of wet tissue). VIP-immunoreactive nerve fibres were present in the adr...

Purification of amyloid plaque core proteins (APCP) from Alzheimer's disease brains to complete homogeneity and in high yield permitted its chemical fractionation and characterization of its components. APCP is mainly made of beta-amyloid (beta A) and an assortment of glycoproteins (accounting for 20%) rich in carbohydrates compatible with N- and O...

1. We studied the relation between cyclic AMP, intracellular Ca2+ concentration and release of [3H]noradrenaline ([3H]NA) in sympathetic neurons cultured from chick embryos. 2. Forskolin (10 microM) and vasoactive intestinal polypeptide (VIP, 3 microM) increased cellular levels of cyclic AMP 8- and 3-fold, respectively, either in the absence or pre...

Several types of extracellular signals affect the function of peripheral neurons. Depolarizing stimuli cause sudden increases in permeability to various ions leading to propagation of nerve impulses and release of transmitter substances. Neurons also receive external signals via neurotransmitter receptors located on the membrane. Different types of...

Effects of the inorganic calcium channel blockers zinc, manganese, cadmium, and nickel on secretion of catecholamines from the perfused adrenal gland of the rat were investigated. Secretion of catecholamines evoked by splanchnic nerve stimulation (1 and 10 Hz) was not affected by nickel (100 microM), partially blocked (50%) by cadmium (100 microM),...

Calcium is an important regulator of a variety of neuronal activities including gene expression. However, it is not clear how Ca2+ influx affects intracellular Ca2+ concentration [( Ca2+]i) in the nucleus. We have taken advantage of laser photometry, the Ca2(+)-sensitive dye Indo-1 that allows ratio imaging, and confocal microscopy to eliminate the...

The significance of the amyloid plaque core proteins (APCP) in Alzheimer's disease (AD) and its consequences for neuronal survival have been controversial. To address this problem we purified the APCP and beta A obtained from brains with AD, and assessed their biological effects in tissue culture. APCP and beta A caused severe toxicity to chick and...

The effects of various neurotransmitters on phosphoinositide hydrolysis, mobilization of Ca2+ and release of [3H]-norepinephrine ([3H]-NE) were studied in cultures of sympathetic neurons of chick embryos. [3H]-inositol-1,4,5-triphosphate ([3H]-IP3) was increased in sympathetic neurons by acetylcholine (ACh), muscarine and serotonin (5-HT). Dopamine...

The effects of electrical stimulation, muscarinic and serotonergic agonists, and caffeine on [3H]inositol 1,4,5-trisphosphate ([3H]Ins(1,4,5)P3) content, intracellular free Ca2+ concentration ([Ca2+]i), and release of [3H]norepinephrine ([3H]NE) were studied in cultured sympathetic neurons. Neuronal cell body [Ca2+]i was unaffected by muscarinic or...

1. Phorbol esters are known to inhibit phospholipase C-mediated hydrolysis of membrane phosphoinositide. This inhibition is attributed to participation of protein kinase C (PKC) in a negative-feedback control of phosphoinositide metabolism. We have tested this hypothesis by using different types of activators and inhibitors of PKC. 2. Phorbol-12,13...

The effects of neurotransmitters and peptides on phosphoinositide hydrolysis were studied by measuring [3H]inositol monophosphate ([3H]IP) and protein kinase C (PKC) activity in the sympathetic and sensory neuronal cultures of the chick embryo. [3H]IP was increased in sympathetic neurons by acetylcholine (ACh), muscarine, serotonin (5-HT), and vaso...

Forskolin has become an invaluable tool for exploring the involvement of cyclic AMP in a variety of cellular functions. The diterpine directly activates the catalytic subunit of adenylate cyclase, causing a marked increase in cyclic AMP content. Because of this well-characterized action, practically all the observed effects of forskolin are commonl...

Sensory neurons of the chick embryo are supported in culture by several neurotrophic factors, including the phorbol esters. Because phorbol esters are known to activate one of the second messengers, namely, protein kinase C, it was of interest to see if the neurotrophic action of phorbol 12,13-dibutyrate (PDB) was related to the activation of prote...

Veratrine (VT), an alkaloid known to act on the sodium channels and cause depolarization of a cell membrane, was found to support the survival of cultured sympathetic neurons. At 30 microM it was as effective as nerve growth factor (NGF), as determined by the cell counts and [3H]norepinephrine ([3H]NE) uptake. Protein kinase C (PKC) activity of the...

The purpose of the present study was to determine the molecular mechanism of stimulatory actions of ACh and vasoactive intestinal polypeptide (VIP) by determining the role of various second messengers in the neurohumoral secretion. Toward such a goal, we measured cAMP, cGMP, protein kinase (PKC) activity, 3H-inositol triphosphate (3H-IP3), and 45Ca...

Evidence is presented for morphological proof of exocytosis in the rat adrenal medulla in situ. Techniques were modified to allow perfusion of the intact adrenal gland with secretagogues (or electrical stimulation) followed by tannic acid. Unstimulated specimens demonstrated exocytotic (omega-shaped) profiles filled with flocculent material. This f...

Isolated perfused rat adrenal glands were prelabeled with 3H-norepinephrine and catecholamine secretion was evoked by nerve stimulation (10 Hz, supramaximal voltage for 30 seconds) or acetylcholine (ACh)(5.4 micrograms) injection. Nerve stimulation evoked significant increases in tritium (16371 +/- 2109 cpm) and catecholamine (11.5 +/- 1.0 ng norep...

The functional behavior of embryonic chick sympathetic neurons was determined by inducing release of [3H]norepinephrine by electrical stimulation of sympathetic neurons growing in the chick heart and in culture, with and without heart cells. A very close correspondence between the functional behavior of neurons developing with the heart cells, eith...

Experiments were carried out on cultured sympathetic neurons of the chick embryo; first, to demonstrate the presence of adrenergic and dopaminergic receptors, and then to see if these receptors are involved in regulation of transmitter release. We show that alpha 2-agonists, norepinephrine, epinephrine and clonidine, had no effect on neuronal cycli...

The effects of phorbol esters [phorbol 12,13-dibutyrate (PDB), 12-O-tetradecanoylphorbol 13-acetate (TPA), and phorbol 13-acetate] were investigated on the release of [3H]norepinephrine, 45Ca2+ accumulation, and protein kinase C activity in cultured sympathetic neurons of the chick embryo. Sympathetic neurons derived from 10-day-old chick embryo we...

Experiments were carried out to determine the cholinergic properties of sensory neurons of the chick embryo by measuring the choline acetyltransferase activity (ChAT) and [3H]choline uptake. The choline acetyltransferase activity in the dorsal root ganglia of an 8-day-old chick embryo was 24.2 +/- 2.52, which increased to 45.4 +/- 9.69 pmol ACh/mg...

The effects of phorbol esters were investigated on the survival of chick sympathetic neurons in a serum-free culture medium. The protein kinase C activator phorbol 12,13-dibutyrate (PDB) supported about 40% of the plated sympathetic neurons. This number was comparable to that supported by nerve growth factor (NGF). A combination of phorbol ester an...

Noradrenergic properties of peripheral sympathetic neurons obtained from 10-12-day-old chick embryos were examined under various culture conditions. Sympathetic neurons supported by nerve growth factor in serum-free or serum-containing medium took up significant and almost equivalent amounts of [3H]norepinephrine. The uptake was markedly enhanced w...

The functional integrity of adrenal chromaffin storage vesicles was studied in the perfused rat adrenal gland subjected to intense exocytosis. Continuous perfusion with 55 mM K+-Krebs solution produced a large and uninterrupted secretion of catecholamines. Total amounts secreted within 45 min were 4.66 μg and represented almost 30% of the total tis...

The effects of continuous exposure of chromaffin cells to nicotine, muscarine and excess potassium on the secretion of catecholamines and uptake of radiolabeled calcium were investigated in the isolated perfused adrenal gland of the rat. Perfusion with 20 microM nicotine-Krebs solution increased catecholamine secretion from a basal value of 5 ng to...

The effect of continuous stimulation of splanchnic nerves at 1, 3, and 10 Hz on the secretion of catecholamines from the isolated rat adrenal gland was examined. Secretion evoked at 10 Hz declined over 60% in 1 h, and by the end of 4 h the secretion was only 10% of the initial value. The secretion evoked at 3 Hz was unchanged in the first hour, but...

We have recently shown that vasoactive intestinal polypeptide (VIP) is as potent as acetylcholine in inducing the secretion of catecholamines from the rat adrenal medulla. In the present study we have investigated the molecular mechanism involved in the exocytotic secretion of catecholamines by VIP and the effects of VIP on Ca45 uptake and phosphoi...

The ability of nerve growth factor to cause rapid activation of the Na+K+ pump of its responsive cells was examined by measuring the uptake of 86Rb+. A significant increase in 86Rb+ uptake in E8 chick dorsal root ganglion sensory neurons after NGF treatment was seen only if the cells had been damaged during the preparation procedure. Such damaged c...

We have recently shown that vasoactive intestinal polypeptide (VIP) is as potent as acetylcholine in inducing the secretion of catecholamines from the rat adrenal medulla. In the present study we have investigated the molecular mechanism involved in the exocytotic secretion of catecholamines by VIP and the effects of VIP on Ca45 uptake and phosphoi...

1. Our previous studies have indicated that splanchnic nerves release a substance(s), other than acetylcholine, that induces the secretion of catecholamines from the rat adrenal medulla. To identify the nature of the non-cholinergic substance, the effects of met-enkephalin and vasoactive intestinal polypeptide (VIP) were investigated in the perfuse...

It had previously been thought that muscarinic cholinergic receptors utilize an influx of extracellular calcium for activation of adrenomedullary catecholamine secretion. However, it has recently been demonstrated that muscarinic receptors on isolated adrenal chromaffin cells can elevate cytosolic free calcium levels in a manner independent of extr...

1. Effects of nicotinic (mecamylamine) and muscarinic (atropine) receptor antagonists were investigated on the secretion of catecholamines evoked by stimulation of splanchnic nerve terminals and acetylcholine in the isolated perfused adrenal gland of the rat to determine whether non-cholinergic substances released from nerve terminals participate i...

Pirenzepine, McN-A-343 and oxotremorine were used to determine the subtypes of muscarinic receptors involved in the secretion of catecholamines from the isolated perfused adrenal gland of the rat. In the presence of 0.1 microM pirenzepine, the concentration-secretion curve for muscarine was shifted in parallel to the right by almost one log unit. W...

Secretion of catecholamines (CA) from the isolated rat adrenal gland during and after perfusion with hypertonic Krebs bicarbonate solution was studied. Perfusion with hypertonic solution made by adding either 156 mM sodium chloride, choline chloride, arginine hydrochloride, sodium thiocyanate, or 312 mM sucrose had no effect on the spontaneous secr...

The uptake and release of catecholamines was investigated in the isolated perfused adrenal gland of the rat after preloading the preparation with [³H]norepinephrine, and the effects of various agents were examined on the stimulation-evoked secretion of catecholamines and total tritium. Large quantities of tritium were found in the adrenal medulla a...

Secretion of catecholamines (CA) from the isolated rat adrenal gland during and after perfusion with hypertonic Krebs bicarbonate solution was studied. Perfusion with hypertonic solution made by adding either 156 mM sodium chloride, choline chloride, arginine hydrochloride, sodium thiocyanate, or 312 mM sucrose had no effect on the spontaneous secr...

Several investigators have shown that tumour promoter phorbol esters mimic the effects of endogenous diacylglycerol to activate a second messenger, protein kinase C. These phorbol esters have proved to be valuable tools for exploring the role of protein kinase C in many cellular functions. We demonstrate here that secretion of catecholamines evoked...

The question of local regulation of catecholamine secretion by alpha-adrenoceptors of either presynaptic splanchnic nerve terminals or postsynaptic chromaffin cells was investigated in the isolated perfused adrenal gland of the rat. The most selective agonists and antagonists of alpha-adrenoceptors such as clonidine, phenylephrine, yohimbine, etc....

The purpose of the present study was twofold: one, to establish the metabolic pathway responsible for the synthesis of ATP of chick embryonic sympathetic neurons maintained in culture and supplemented with nerve growth factor; two, to establish the relationship between ATP content and the uptake and storage of [3H]norepinephrine. We show that ATP c...

The effect of gallamine on spontaneous and stimulation-evoked overflow of tritium was studied in the submandibular gland of the rat. The gland was perfused retrogradely and labeled with3H-noradrenaline. The stimulation-evoked (1 Hz for 60 s) overflow of tritium was facilitated by increasing concentrations of gallamine (0.3–20 mM). None of the conce...

A culture medium circulated through the rat heart and supplemented with insulin, transferrin and nerve growth factor leads to a massive proliferation of neurite outgrowth from neurons of peripheral sympathetic ganglia of the chick embryo. Addition of 1 mM cyanide or 50 microM dinitrophenol to such medium for 2 days had no adverse effect on the neur...

The effect of phorbol ester, phorbol 12,13-dibutyrate, was investigated on the overflow of tritium from 3H-noradrenaline-loaded sympathetic neurons of the isolated perfused salivary gland of the rat. Stimulation (1 Hz for 60 s)-evoked overflow of tritium was enhanced by phorbol ester. A significant enhancement was seen at 1 nmol/l, which increased...

The left adrenal gland of the rat was perfused in vitro via the adrenal vein with Krebs-bicarbonate solution to study accumulation of exogenous catecholamines and secretion of endogenous norepinephrine and epinephrine by exposing the gland to various sympathomimetic agents. Perfusions with high concentrations of norepinephrine for 30 min did not re...

The effects of various drugs, known to be inhibitors of calmodulin, were tested on the secretion of catecholamines (CA) from the adrenal medulla and sympathetic nerves of the salivary gland of the rat. Secretion of CA from the perfused adrenal gland was evoked by injection of acetylcholine (ACh, 50 micrograms), excess K (700 micrograms), or transmu...

The results of the present experiments demonstrate that embryonic sympathetic neurons initially selected in culture by nerve growth factor could be subsequently supported by raising potassium concentration of the medium to 35 mM. Alternatively, neurons initially selected by 35 mM potassium could be supported in a normal culture medium but supplemen...

The purpose of the present study was to obtain evidence for the functional role of presynaptic alpha adrenergic receptors in the intact animal. This aim was approached by investigating the effects of alpha adrenergic blocking agents and other hypotensive agents on the norepinephrine content of the heart, spleen and the salivary gland of the normal...

The aim of the present study was (1) to evaluate the functional contribution of nicotinic and muscarinic receptors of the rat adrenal medullary cells in the secretion of catecholamines evoked by exogenous and endogenous acetylcholine, (2) to know whether the ratio of epinephrine to norepinephrine in the perfusate of the adrenal gland stimulated wit...

The mechanism of alpha-adrenergic receptor-mediated feedback regulation of [3H]norepinephrine release was studied in the guinea-pig heart. The overflow of [3H]norepinephrine evoked by stimulation (1 Hz for 60 s) was inhibited up to 90% in a dose-dependent manner by norepinephrine (58.5 and 177 nM) and epinephrine (54 and 164 nM). These inhibitory e...

Dissociated neurons from paravertebral sympathetic chains of 11- to 13-day-old chick embryos were maintained in culture medium supplemented with 10% (v/v) heat-inactivated horse serum and nerve growth factor and used to study changes in resting membrane potential, calcium influx and release of [3H]norepinephrine ([3H]NE) on various K concentrations...

Recent reports indicate that the presynaptic negative feedback mechanism involved in regulating sympathetic neurotransmission is only functional within a narrow range of neuronal activity. The specific aim of this report is to examine the release of sympathetic transmitter at varying frequencies of stimulation, and the effects of alpha-adrenoceptor...

1. The present investigation was specifically aimed to study the question of re‐use of sympathetic storage vesicles of the isolated guinea‐pig heart. 2. Tetraethylammonium (TEA, 20 mM) caused a massive enhancement of noradrenaline (NA) overflow upon transmural stimulation of the heart. The enhancement was inversely related to the frequency of stimu...

The aim of the present investigation was to examine the question of reutilization of norepinephrine storage vesicles in the cardiac sympathetic nerves. More specifically, the question was whether exocytotically depleted stores of norepinephrine could be restored by epinephrine and the restoration of epinephrine could be demonstrated by chemical and...

Transmural stimulation of the isolated adrenal gland of the rat and guinea pig results in secretion of catecholamine. The secretion is due to activation of cholinergic receptors of the adrenal medulla by acetylcholine released from splanchnic nerve terminals after transmural stimulation. Our aim was to see whether the same experimental technique co...

Isolated heart of the guinea-pig was stimulated transmurally (1 Hz), or by excess [K+] to evoke release of [3H]noradrenaline. The interactions of tetraethylammonium, acetylcholine and adenosine were investigated on [3H]noradrenaline overflow at different [Ca2+] concentrations. One mM tetraethylammonium doubled [3H]noradrenaline overflow, and at 30...

The effect of ammonium (0.3 to 3.6 mM) was studied on the overflow of 3H-norepinephrine in the vas deferens and the brain of the rat. Ammonium enhanced the overflow of 3H-norepinephrine in both organs and this response was greatly facilitated by alkaline pH (7.8) and was blocked by pH 7.0. Calcium was not needed for ammonium-induced overflow. In fa...

1. The influence of varying concentrations of Ca and tetraethylammonium (TEA) was investigated on spontaneous and stimulation-evoked overflow of 2H-noradrenaline in the isolated heart of the guinea pig. 2. Perfusion of the heart with Krebs-bicarbonate solution containing 0-2.5 mM Ca does not modify the spontaneous overflow of 3H-noradrenaline. 3. E...

1. A method of studying the secretion of catecholamines (CA) in the isolated perfused rat adrenal gland by transmural stimulation or by application of acetylcholine (ACh) has been described. 2. Secretion of CA was practically linear in response to ACh administration, starting from 4.42 microM to 1.32 mM. Transmural stimulation enhanced secretion fr...

1. The effect of K deprivation on catecholamine (CA) secretion induced by transmural stimulation of the isolated perfused adrenal glands of the rat was studied. 2. In the absence of external K, secretion of CA evoked by transmural stimulation was greatly (2- to 7-fold) facilitated. The facilitation was inversely related to the frequency of stimulat...

1. The primary aim of this work was two-fold. To introduce a simpe method for studying sympathetic neurosecretion in the mammalian heart in response to nerve stimulation. The other aim was to determine whether or not presynaptic alpha-adrenoceptors and high affinity uptake mechanism of cardiac sympathetic nerves play some role in the regulation of...

In the normal and phenoxybenzamine‐treated vasa deferentia of the rat the overflow of tritium after preloading with [ ³ H]‐noradrenaline increased with the increase in frequency of transmural stimulation. Phenozybenzamine enhanced overflow by 5 to 7 fold. After tetraethylammonium (TEA, 10 m m ), fractional tritium overflow at 5 Hz increased from a...

The noradrenaline (NA) content of various peripheral sympathetic neuroeffector organs and brain was measured at various times after different doses of reserpine administered to the rat. About a 25% reduction in the NA content of the heart was observed 24 h after 0.005 mg/kg reserpine. Two to ten times more reserpine was needed to obtain an approxim...