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P Boksa

Publications and source records attributed to P Boksa.

At least 55 records · Page 3Linked to original sources
PubMed1992

Phorbol ester enhances excitatory amino acid-induced dopamine release from mesencephalic cell cultures.

The hypothesis that protein kinase C activation can modulate excitatory amino acid-induced dopamine release was tested by investigating effects of phorbol esters, direct activators of protein kinase C, on dopamine release stimulated by N-methyl-D-aspartate (NMDA) and non-NMDA sub-types of excitatory amino acid agonists in fetal rat mesencephalic cell cultures. The phorbol ester, 12-O-tetradecanoyl phorbol-13-acetate (TPA), enhanced dopamine release evoked by NMDA, kainate, quisqualate and by K+ depolarization. Release in the presence of NMDA and TPA was completely abolished by the NMDA antagonist, MK-801. TPA enhancement of NMDA-stimulated dopamine release was likely due to protein kinase C activation by the phorbol ester since (1) the NMDA response was enhanced by nanomolar concentrations of TPA, (2) two phorbol esters capable of activating protein kinase C enhanced the NMDA response while an inactive phorbol ester did not, (3) staurosporine, a potent protein kinase C inhibitor, blocked TPA enhancement of the NMDA response. TPA enhancement of NMDA-stimulated dopamine release was not blocked by H8, an inhibitor with high affinity for cyclic nucleotide dependent kinases, while forskolin, a direct activator of adenylate cyclase, had no effect on NMDA-stimulated release, indicating a lack of involvement of cAMP-dependent kinase in the TPA effect. TPA enhanced NMDA-stimulated release both in the presence and absence of Mg2+, indicating that TPA enhancement was not due to reversal of a Mg2+ blockade of the NMDA receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗
PubMed1992

Serotonergic regulation of type II corticosteroid receptor binding in hippocampal cell cultures: evidence for the importance of serotonin-induced changes in cAMP levels.

The development of the hypothalamic-pituitary-adrenal response to stress is profoundly altered by environmental events. One target for environmental regulation within the hypothalamic-pituitary-adrenal axis is the hippocampal type II corticosteroid (or glucocorticoid) receptor system, which mediates the negative-feedback effects of glucocorticoids on hypothalamic-pituitary-adrenal activity. Thus, adult rats handled early in life show increased hippocampal type II corticosteroid receptor density and increased sensitivity to the inhibitory effects of circulating glucocorticoids on post-stress hypothalamic-pituitary-adrenal activity. Both effects persist throughout life. The effects of handling on type II corticosteroid receptor development are, at least in part, mediated by changes in hippocampal 5-hydroxytryptamine turnover. Moreover, 5-hydroxytryptamine can regulate type II corticosteroid receptor density in cultured hippocampal cells, providing a paradigm for examining the neurochemical mechanisms by which environmental stimuli might regulate neural differentiation. In the present studies, we examined the intracellular mechanisms underlying the effects of 5-hydroxytryptamine on type II corticosteroid receptors ([3H]RU 28362 binding) in hippocampal cell cultures. cAMP, but not cGMP, levels in cultured hippocampal cells were significantly increased by the addition of 5-hydroxytryptamine to the medium. The cAMP response to 5-hydroxytryptamine was biphasic: an initial increase in cAMP levels occurred in response to nanomolar 5-hydroxytryptamine concentrations (EC50 = 7.2 nM), while a second increase was apparent at low micromolar concentrations. 5-Hydroxytryptamine also increased [3H]RU 28362 binding (EC50 = 4.5 nM) with a maximal effect at a concentration of 10 nM. There was no further increase in [3H]RU 28362 binding even with higher, micromolar concentrations of 5-hydroxytryptamine.(ABSTRACT TRUNCATED AT 250 WORDS)

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1992

Adrenal phenylethanolamine N-methyltransferase induction in relation to glucocorticoid receptor dynamics: evidence that acute exposure to high cortisol levels is sufficient to induce the enzyme.

Glucocorticoids (GCs) are thought to regulate, in a permissive fashion, the basal activity of adrenal medullary phenylethanolamine N-methyltransferase (PNMT). However, it is unclear whether a large short-term increase in GC release, such as occurs during an acute stress response, may also play a role in PNMT regulation. The present study investigated how the GC influence over PNMT activity varies in relation to dynamic changes in the hormone-receptor signal. Using [3H]dexamethasone (DEX) and [3H]RU 28362 as radioligands, we have confirmed the presence of GC receptors in bovine adrenal medullary cells. A concentration-dependent decline in soluble GC receptor sites and an increase in nuclear uptake of [3H]DEX were found in response to GC levels as low as 5 x 10(-8) M. The loss of soluble sites plateaued between 5 x 10(-8) and 10(-6) M cortisol, with further losses occurring at 10(-5) and at 10(-4) M. The functional consequence of GC receptor binding was confirmed by measuring PNMT activity following 3-day exposure to cortisol. The pattern of PNMT induction was similar to that seen with GC receptor occupancy; at cortisol concentrations between 10(-8) and 10(-5) M, PNMT induction was at a plateau, with a further increase in activity at 10(-4) M. The increase in PNMT activity following 3-day exposure to low (10(-7) M) and high (5 x 10(-5), 10(-5) M) cortisol was blocked by the GC receptor antagonist RU 38486, suggesting a GC receptor-mediated event. Finally, a short (2 h) pulse of GC, which mimics the time course of physiological elevation of GC following acute stress, elevated adrenal medullary PNMT activity measured 3 days later. Therefore, our results provide novel evidence that short-term exposure of adrenal medullary cells to high cortisol levels can elevate PNMT activity.

Adrenal Medulla↗
PubMed1991

Transient postnatal increases in excitatory amino acid binding sites in rat ventral mesencephalon.

The developmental profile of three sub-types of excitatory amino acid (EAA) binding sites was determined in the ventral mesencephalon and the striatum of rats from prenatal day 19 to adult (3 months) using membrane binding assays. In the ventral mesencephalon, there was a transient increase of EAA receptor binding sites beyond adult levels, which peaked at postnatal day 7 (P7) for [3H]glutamate binding to NMDA receptors and at P14 for [3H]AMPA and [3H]kainate binding. In the striatum, [3H]glutamate/NMDA and [3H]kainate binding reached adult levels during the early postnatal period, stabilizing at this level with no transient overexpression beyond adult levels. [3H]AMPA binding also showed an increase above adult levels at P14 in the striatum. These results raise the possibility that the transient overexpression of EAA receptors in the ventral mesencephalon may affect the developmental fate of dopaminergic and other neurons in this region.

Aging↗
PubMed1991

Inhibitory and potentiating influences of glycine on N-methyl-D-aspartate-evoked dopamine release from cultured rat mesencephalic cells.

In the presence of 1.2 mM Mg2+, glycine (30-100 microM) inhibited [3H]dopamine ([3H]DA) release stimulated by N-methyl-D-aspartate (NMDA), in fetal rat mesencephalic cell cultures. Strychnine (1 microM) blocked the inhibitory effect of 100 microM glycine, indicating an action via strychnine-sensitive inhibitory glycine receptors. A higher concentration of strychnine (100 microM), by itself, inhibited NMDA-evoked [3H]DA release in the presence or absence of Mg2+. Spontaneous [3H]DA release and [3H]DA release stimulated by kainate and quisqualate were unaffected by glycine (less than or equal to 100 microM) or strychnine (less than or equal to 100 microM), indicating that glycine and strychnine modulatory effects are only associated with the NMDA receptor subtype. [3H]DA release evoked by K+ (56 mM) was unaffected by glycine (less than or equal to 100 microM) but was attenuated by a high concentration of strychnine (100 microM). In the absence of exogenous Mg2+, glycine (30-100 microM) potentiated NMDA-evoked [3H]DA release by a strychnine-insensitive mechanism. A selective antagonist of the NMDA-associated glycine receptor, 7-chlorokynurenate (10 microM), attenuated NMDA-evoked [3H]DA release in the absence of Mg2+. The effect of 10 microM 7-chlorokynurenate was overcome by 1 microM glycine. Also, when tested in the presence of 1.2 nM Mg2+ and 1 microM strychnine, 100 microM 7-chlorokynurenate inhibited NMDA-evoked [3H]DA release, and this antagonism was overcome by 30 to 100 microM glycine. These results indicate that two distinct glycine receptors modulate NMDA-stimulated [3H]DA release from mesencephalic cells in culture. Manipulation of extracellular Mg2+ permits the differentiation of a strychnine-sensitive glycine response (inhibition of NMDA-evoked [3H]DA release) from a strychnine-insensitive glycine response (potentiation of NMDA-evoked [3H]DA release). It is suggested that voltage-dependent Mg2+ blockade of the NMDA response may allow for the expression of these opposing effects of glycine.

Amino Acids↗
PubMed1990

Chromaffin cell grafts to rat cerebral cortex reverse lesion-induced memory deficits.

Adrenal chromaffin cells were isolated from donor adult rats and transplanted to the cerebral cortex of bilaterally nucleus basalis magnocellularis-lesioned rats. Chromaffin cell grafts to lesioned animals completely reversed the spatial memory deficit seen in lesioned alone animals on a T-maze alternation task. Although chromaffin cell grafts have been used previously to reverse motor abnormalities arising from defective nigro-striatal aminergic transmission, the present report is the first evidence that chromaffin cell transplants can reverse deficits in memory function. Grafts also enhanced cortical acetylcholinesterase staining.

Adrenal Medulla↗
PubMed1990

Subtypes of excitatory amino acid receptors involved in the stimulation of [3H]dopamine release from cell cultures of rat ventral mesencephalon.

N-methyl-D-aspartic acid (NMDA), quisqualic acid (QUIS), and kainic acid (KAIN), respective agonists for three excitatory amino acid (EAA) receptor subtypes, stimulated [3H]dopamine ([3H]DA) release from dissociated cell cultures of fetal rat ventral mesencephalon. Release evoked by all three agonists was Ca2(+)-dependent and inhibited by broad-spectrum antagonists (D,L-cis-2,3-piperidine dicarboxylic acid [PDA] and kynurenic acid [KYN]). However, both of these antagonists were more potent against KAIN than against QUIS and only KAIN-evoked release was blocked by gamma-D-glutamyl-aminomethyl sulfonic acid (GAMS, IC50 700 microM). NMDA-stimulated [3H]DA release was selectively inhibited by competitive (3-[2-carboxypiperazine-4-yl]propyl-1-phosphonic acid [CPP] and D,L-2-amino-5-phosphonovaleric acid [APV]) and non-competitive (phencyclidine and MK-801) NMDA receptor antagonists. In 1.2 mM Mg2+, NMDA-stimulated [3H]DA release was Na(+)-dependent and inhibited by tetrodotoxin (TTX, 2 microM) or by the local anaesthetic, lidocaine (200 microM). However, in 0 Mg 2+, NMDA-evoked release was not inhibited by TTX or lidocaine. Thus, TTX-sensitivity of the NMDA response in 1.2 mM Mg2+ apparently occurs because Na(+)-action potentials are required to alleviate a Mg2+ blockade. Neither QUIS- nor KAIN-evoked release was affected by Mg2+ or TTX. When extracellular NaCl was replaced by sucrose or Na2SO4, the QUIS response was increased. KAIN-evoked release was unaffected by the sucrose substitution and was attenuated in the Na2SO4-containing buffer. It is concluded that NMDA and QUIS/KAIN release [3H]DA via separate receptor subtypes.

Animals↗
PubMed1990

Dopamine release from bovine adrenal medullary cells in culture.

The present study tested whether release of dopamine from isolated bovine adrenal medullary cells in culture could be stimulated or inhibited by secretagogues and modulators known to affect noradrenaline and adrenaline release from adrenal medullary chromaffin cells. K+ depolarization or activation of voltage-sensitive Na+ channels by veratridine both stimulated dopamine release. Ca2+-dependent dopamine release was also stimulated by the mixed nicotinic-muscarinic agonist, carbachol. Carbachol-induced dopamine release was inhibited by a nicotinic but not by a muscarinic antagonist and dopamine release was also stimulated by a selective nicotinic agonist, 1,1-dimethyl-4-phenyl-piperazinium. Carbachol-induced dopamine release was inhibited by substance P and by neuropeptide Y. Histamine also stimulated dopamine release, while angiotensin II and glutamate produced no significant stimulation of dopamine release. Noradrenaline and adrenaline were released in response to the above agents with a profile almost identical to that of dopamine. The results indicate that dopamine can be directly released from adrenal medullary cells in response to stimulation of those cells and suggest that the dopamine release originates from chromaffin cells similar or identical to those storing noradrenaline and adrenaline. A possible role for dopamine, released from adrenal chromaffin cells, in modulating catecholamine release from the chromaffin cells and/or contributing to circulating plasma dopamine is discussed.

Adrenal Medulla↗
PubMed1990

Stimulation of dopamine release from cultured rat mesencephalic cells by naturally occurring excitatory amino acids: involvement of both N-methyl-D-aspartate (NMDA) and non-NMDA receptor subtypes.

In rat mesencephalic cell cultures, L-glutamate at concentrations ranging from 100 microM to 1 mM stimulated release of [3H]dopamine that was attenuated by the non-N-methyl-D-aspartate (non-NMDA) receptor antagonist 6,7-dinitroquinoxalinedione, but not by the selective NMDA receptor antagonists (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (MK-801; 10 microM) and 3-(2-carboxypiperazine-4-yl)propyl-1-phosphonate (300 microM). Even at 1 mM glutamate, this release was Ca2+ dependent. These observations suggest that the release was mediated by a non-NMDA receptor. Only release stimulated by a lower concentration (10 microM) of glutamate was inhibited by MK-801 (10 microM), indicating that glutamate at this concentration activates the NMDA receptor. By contrast, L-aspartate at concentrations of 10 microM to 1 mM evoked [3H]dopamine release that was completely inhibited by MK-801 (10 microM) and was also Ca2+ dependent (tested at 1 and 10 mM aspartate). Thus, effects of aspartate involved activation of the NMDA receptor. Sulfur-containing amino acids (L-homocysteate, L-homocysteine sulfinate, L-cysteate, L-cysteine sulfinate) also evoked [3H]dopamine release. Release evoked by submillimolar concentrations of these amino acids was attenuated by MK-801 (10 microM), indicating involvement of the NMDA receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗
PubMed1990

Phencyclidine and related compounds evoked [3H]dopamine release from rat mesencephalic cell cultures by a mechanism independent of the phencyclidine receptor, sigma binding site, or dopamine uptake site.

At concentrations greater than or equal to 100 microM, phencyclidine (PCP), N-(1-(2-thienyl)-cyclohexyl)piperidine (TCP), and MK-801 induced [3H]dopamine release from dissociated cell cultures of rat mesencephalon. This release was Ca2+ independent and tetrodotoxin insensitive. Tetrodotoxin (2 microM) itself had no effect on spontaneous release of [3H]dopamine. [3H]Dopamine release was induced by 1,3-di(2-tolyl)guanidine, a sigma ligand, and by 4-aminopyridine (1-3 mM), a K+ channel blocker. No stereoselectivity was observed for [3H]dopamine release evoked by the dioxadrol enantiomers, dexoxadrol, and levoxadrol, or by enantiomers of N-allylnormetazocine (SKF 10,047). The selective dopamine uptake inhibitor 1-(2-[bis(4-fluorophenyl)methoxy]ethyl)-4-(3-phenylpropyl)piperazine dihydrochloride (GBR 12909) did not affect spontaneous or TCP-evoked [3H]dopamine release. Together, these data suggest that the dopamine-releasing effects of PCP-like compounds on the mesencephalic cells were not mediated by actions at the PCP receptor or sigma binding site, Ca2+, or Na+ channels, or at the high affinity dopamine uptake site. It remains conceivable that blocking actions of PCP-like compounds at voltage-regulated K+ channels may at least partly explain the response. These results are discussed in comparison with findings in intact brain.

2-Amino-5-phosphonovalerate↗
PubMed1990

Serotonin regulates type II corticosteroid receptor binding in hippocampal cell cultures.

Previous work from our laboratory has shown that early postnatal handling of rat pups permanently increases hippocampal type II, but not type I, corticosteroid receptor binding. Handling also increases hippocampal 5-HT turnover, and the effect of handling on type II corticosteroid receptor binding is blocked by concurrent administration of the 5-HT2 receptor antagonist ketanserin. In view of these findings, the present studies examined the effects of 5-HT on type I ([3H]corticosterone) and type II ([3H]RU 28362) corticosteroid receptor binding in dispersed hippocampal cell cultures derived from animals killed at E19-20 in order to verify that 5-HT can act directly on hippocampal cells to alter corticosteroid receptor binding. Both type I and type II receptors were measurable in cultured hippocampal cells and the apparent affinity (Kd) for [3H]corticosterone (0.4 +/- 0.1 nM) and [3H]RU 28362 (0.8 +/- 0.1 nM) was similar to that from studies with intact animals. 5-HT increased type II, but not type I, corticosteroid receptor binding capacity in a dose-related manner, with the maximal effect (+188%) observed at 10 nM 5-HT and no change in the affinity of the receptor for [3H]RU 28362. The effect of 10 nM 5-HT on [3H]RU 28362 binding required a minimum of 4 d exposure and persisted for at least 7 d following the removal of 5-HT. The effect of 10 nM 5-HT on [3H]RU 28362 binding was completely blocked by the 5-HT2 receptor antagonists ketanserin and mianserin. There were no effects of the 5-HT1a antagonist, BMY 7378, or the 5-HT3 antagonist, MDL 72222.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Pharmacological activity of N-methyl-carbamylcholine, a novel acetylcholine receptor agonist with selectivity for nicotinic receptors.

N-Methyl-carbamylcholine (also called N-methyl-carbachol) is an analogue of the mixed muscarinic-nicotinic acetylcholine receptor agonist, carbachol. Previous studies have provided evidence that radiolabelled N-methyl-carbachol can bind selectively to nicotinic acetylcholine receptors in rat brain. To determine whether N-methyl-carbachol acts as an agonist or an antagonist at nicotine and/or muscarinic receptor sites, the present study examined the pharmacological activity of this compound on some cholinergically innervated tissues. N-Methyl-carbachol, like carbachol, depolarized rat isolated sympathetic ganglia and these effects were inhibited by a nicotinic antagonist, d-tubocurarine, but not by a muscarinic antagonist, atropine. Exposure of rat sympathetic ganglia to N-methyl-carbachol blocked the compound action potential generated in ganglia by stimulation of the pre-ganglionic trunk; this effect of N-methyl-carbachol was likely due to desensitization of the nicotinic response. N-Methyl-carbochol, like carbachol, stimulated the release of [3H]noradrenaline from cultured adrenal medullary cells that had been pre-loaded with [3H]noradrenaline; these effects were largely inhibited by a nicotinic antagonist, mecamylamine, while atropine produced less blockade. N-Methyl-carbachol contracted the frog isolated rectus abdominis muscle and the effect was completely blocked by d-tubocurarine. By contrast, contracture of the rectus abdominis produced by carbachol was partially inhibited by either atropine or d-tubocurarine. N-Methyl-carbachol, like carbachol, contracted the rat isolated ileum and these effects were completely blocked by atropine; however, N-methyl-carbachol was about 42 times less potent than carbachol for this effect. Intravenous injection of N-methyl-carbachol, like nicotine, to the rat produced a transient decrease followed by a more sustained rise in blood pressure while carbachol produced only a sustained decrease in blood pressure. The effects of N-methyl-carbachol and nicotine on blood pressure were blocked by pretreatment of the animal with a nicotinic antagonist, hexamethonium. N-methyl-carbachol, like nicotine, stimulated the release of [3H]dopamine from rat striatal synaptosomes, pre-loaded with [3H]dopamine; release induced by either N-methyl-carbachol or nicotine was inhibited by mecamylamine but not by atropine. In rat cerebral cortical slices pre-loaded with [3H]inositol, carbachol, but not N-methyl-carbachol, stimulated the accumulation of [3H]inositol-1-phosphate, an effect blocked by atropine but not by mecamylamine.(ABSTRACT TRUNCATED AT 400 WORDS)

Adrenal Medulla↗
PubMed1989

Effects of the catecholaminergic neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) on adrenal chromaffin cells in culture.

N-(2-Chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) is an inhibitor of noradrenaline uptake and a neurotoxin known to deplete noradrenaline levels with little effect on dopamine, serotonin or adrenaline in the central nervous system. The present study tested the effects of DSP-4 on catecholamine uptake, release and content in cultures of isolated bovine adrenal chromaffin cells. DSP-4 selectively inhibited the acute uptake of [3H]noradrenaline with little or no effect on [3H]adrenaline or [3H]dopamine uptake. In cultures preloaded with [3H]catecholamines, DSP-4 stimulated the release of [3H]noradrenaline and, to a small extent, also [3H]adrenaline and [3H]dopamine. However, the drug did not stimulate the release of appreciable amounts of endogenous adrenaline, noradrenaline or dopamine. A high concentration of DSP-4 inhibited the carbachol-stimulated release of adrenaline, noradrenaline and dopamine from the cells. Following a 1-hr exposure to the drug, DSP-4 decreased adrenaline, noradrenaline and dopamine levels in the cells with no gross morphologic changes in the cells. Reductions in adrenaline and noradrenaline levels were almost equal in magnitude, while dopamine was depleted to a somewhat greater extent under some conditions. Longer exposure to DSP-4 resulted in morphological changes in the cells, suggesting that the drug is also toxic to chromaffin cells in culture.

Animals↗
PubMed1989

Characterization and quantitative autoradiographic distribution of [3H]acetylcholine muscarinic receptors in mammalian brain. Apparent labelling of an M2-like receptor sub-type.

[3H]Acetylcholine receptor binding characteristics (under muscarinic conditions) have been investigated using membrane binding assays and in vitro receptor autoradiography. In rat, guinea-pig and monkey brain membrane preparations, [3H]acetylcholine binds with high affinity (25-50 nM) to an apparently single class of sites which is differentially distributed across brain regions. The ligand selectivity pattern reveals that the potency of (-)quinuclidinyl benzylate is greater than (greater than) atropine greater than scopolamine greater than oxotremorine greater than carbamylcholine greater than pirenzepine greater than methylcarbamyl-choline = nicotine in competing for [3H]acetylcholine binding sites, indicating that [3H]acetylcholine selectively binds to muscarinic sites under these incubation conditions. Moreover, the low potency of pirenzepine suggests that [3H]acetylcholine does not label a significant proportion of the M1 receptor sub-type but most likely binds to putative M2-like receptor sites. This hypothesis is also supported by the autoradiographic distribution of [3H]acetylcholine binding sites in all species studied here. High densities of [3H]acetylcholine binding sites are seen in various nuclei of the medulla and pons, certain thalamic nuclei, medial septum, laminae III, V and VI of the cortex and just above the pyramidal cell layer of the hippocampus. Such localization is much different from that seen with the non-selective antagonist [3H]quinuclidinyl benzylate and the selective M1 receptor ligand [3H]pirenzepine, although it resembles that of the selective M2 receptor antagonist [3H]AF-DX 116. Thus, [3H]acetylcholine apparently mostly binds with high affinity mainly to non-M1 muscarinic receptor types in mammalian brain tissues. Moreover, the ligand selectivity pattern and in vitro receptor autoradiographic data suggest that at low concentrations (10-20 nM) most of [3H]actylcholine labelled sites are of the M2-like receptor class.

Acetylcholine↗
PubMed1989

Glutamate stimulation of [3H]dopamine release from dissociated cell cultures of rat ventral mesencephalon.

In dissociated cell cultures of fetal rat ventral mesencephalon preloaded with [3H]dopamine, glutamate (10(-5)-10(-3) M) stimulated the release of [3H]dopamine. Glutamate stimulation of [3H]dopamine release was Ca2+ dependent and was blocked by the glutamate antagonist, cis-2,3-piperidine dicarboxylic acid. Glutamate stimulation of [3H]dopamine release was not due to glutamate neurotoxicity because (1) glutamate did not cause release of a cytosolic marker, lactate dehydrogenase, and (2) preincubation of cultures with glutamate did not impair subsequent ability of the cells to take up or release [3H]dopamine. Thus, these dissociated cell cultures appear to provide a good model system to characterize glutamate stimulation of dopamine release. Release of [3H]dopamine from these cultures was stimulated by veratridine, an activator of voltage-sensitive Na+ channels, and this stimulation was blocked by tetrodotoxin. However, glutamate-stimulated [3H]dopamine release was not blocked by tetrodotoxin or Zn2+. Substitution of NaCl in the extracellular medium by sucrose, LiCl, or Na2SO4 had no effect on glutamate stimulation of [3H]dopamine release; however, release was inhibited when NaCl was replaced by choline chloride or N-methyl-D-glucamine HCl. Glutamate-stimulated [3H]-dopamine release was well maintained (60-82% of control) in the presence of Co2+, which blocks Ca2+ action potentials, and was unaffected by the local anesthetic, lidocaine. These results are discussed in terms of the receptor and ionic mechanisms involved in the stimulation of dopamine release by excitatory amino acids.

Animals↗
PubMed1988

Arachidonic acid inhibits choline uptake and depletes acetylcholine content in rat cerebral cortical synaptosomes.

The effects of arachidonic acid on [3H]choline uptake, on [3H]acetylcholine accumulation, and on endogenous acetylcholine content and release in rat cerebral cortical synaptosomes were investigated. Arachidonic acid (10-150 microM) produced a dose-dependent inhibition of high-affinity [3H]choline uptake. Low-affinity [3H]choline uptake was also inhibited by arachidonic acid. Fatty acids inhibited high-affinity [3H]choline uptake with the following order of potency: arachidonic greater than palmitoleic greater than oleic greater than lauric; stearic acid (up to 150 microM) had no effect. Inhibition of [3H]choline uptake by arachidonic acid was reversed by bovine serum albumin. In the presence of arachidonic acid, there was an increased accumulation of choline in the medium, but this did not account for the inhibition of [3H]choline uptake produced by the fatty acid. Arachidonic acid inhibited the synthesis of [3H]acetylcholine from [3H]choline, and this inhibition was equal in magnitude to the inhibition of high-affinity [3H]choline uptake produced by the fatty acid. A K+-stimulated increase in [3H]acetylcholine synthesis was inhibited completely by arachidonic acid. Arachidonic acid also depleted endogenous acetylcholine stores. Concentrations of arachidonic acid and hemicholinium-3 that produced equivalent inhibition of [3H]choline uptake also produced equivalent depletion of acetylcholine content. In the presence of eserine, arachidonic acid had no effect on acetylcholine release. The results suggest that arachidonic acid may deplete acetylcholine content by inhibiting high-affinity choline uptake and subsequent acetylcholine synthesis. This raises the possibility that arachidonic acid may play a role in the impairment of cholinergic transmission seen in cerebral ischemia and other conditions in which large amounts of the free fatty acid are released in brain.

Acetylcholine↗
PubMed1987

[3H]N-methyl-carbamylcholine, a new radioligand specific for nicotinic acetylcholine receptors in brain.

The present study characterized the binding of [3H]N-methyl-carbamylcholine ([3H]methyl-carbachol), a new radioligand, to rat cerebral cortex membranes and demonstrated the autoradiographic distribution of these sites in rat brain. With atropine used to block muscarinic acetylcholine sites and nicotine to define non-specific binding, [3H]methyl-carbachol bound specifically, saturably and with high affinity (Kd = 11.0 nM, Bmax = 118.4 fmol/mg protein and Hill coefficient = 0.92) to a population of presumably nicotinic sites in cerebral cortex membranes. When nicotine was used to block nicotinic acetylcholine sites and atropine to define non-specific binding there was no specific binding of [3H]methyl-carbachol (concentrations up to 45 nM) to possible muscarinic sites in cerebral cortex membranes. The binding parameters under non-selective conditions (without blockade of either muscarinic or nicotinic acetylcholine sites) had very similar values to those obtained under nicotinic conditions (Kd = 8.0 nM, Bmax = 125.0 fmol/mg protein and Hill coefficient = 0.98). [3H]Methyl-carbachol binding was potently inhibited by nicotinic agonists and antagonists but only poorly displaced by muscarinic agents. Autoradiographic studies evidenced high densities of [3H]methyl-carbachol binding sites in the interpeduncular nucleus, various thalamic nuclei, superior colliculus and layers III/IV of the cortex. Such a distribution was very similar to those previously reported for nicotinic acetylcholine sites and other radioligands. These results suggest that [3H]methyl-carbachol is a specific radioligand of the neuronal nicotinic receptor. Its stability and high selectivity constitute distinct advantages over previously used nicotinic radioligands such as acetylcholine and nicotine.

Animals↗