Biomedical subjects

S Boehm

Publications and source records attributed to S Boehm.

At least 37 records · Page 2Linked to original sources

PubMed1998

Presynaptic inhibition by concanavalin A: are alpha-latrotoxin receptors involved in action potential-dependent transmitter release?

Effects of concanavalin A on transmitter release were investigated in primary cultures of chick sympathetic neurons. The lectin reduced electrically evoked [3H]noradrenaline release by up to 30% with half-maximal inhibition at 0.16 microM. Concanavalin A also reduced the release triggered by extracellular Ca2+ in neurons depolarized by 25 mM K or rendered Ca2+-permeable by the ionophore A23187. The inhibitory action of concanavalin A on electrically evoked release was additive to that of the alpha2-adrenergic agonist UK 14,304. Inactivation of Gs and Gi/Go type G proteins by either cholera or pertussis toxin did not alter the inhibitory effect of the lectin. Concanavalin A failed to affect the resting membrane potential, action potential waveforms, or voltage-dependent K+ and Ca2+ currents. In contrast, the lectin efficiently blocked both the Ca2+-dependent and -independent alpha-latrotoxin-induced transmitter release, but only when applied before the toxin. The reduction of electrically evoked, as well as alpha-latrotoxin-evoked, release by concanavalin A was attenuated in the presence of glucose and abolished by methyl alpha-D-mannopyranoside. The dimeric derivative, succinyl-concanavalin A, was significantly less active than tetrameric concanavalin A. In bovine adrenal chromaffin cells, which displayed only weak secretory responses to alpha-latrotoxin, concanavalin A failed to alter K+-evoked catecholamine secretion. These results show that concanavalin A causes presynaptic inhibition in sympathetic neurons and indicate that cross-linking of alpha-latrotoxin receptors may reduce action potential-dependent transmitter release.

Action Potentials↗

PubMed1997

Glycine receptors in cultured chick sympathetic neurons are excitatory and trigger neurotransmitter release.

1. Total RNA isolated from embryonic chick paravertebral sympathetic ganglia was used in a reverse transcription-polymerase chain reaction (RT-PCR) assay with a pair of degenerate oligonucleotide primers deduced from conserved regions of mammalian glycine receptor alpha-subunits. Three classes of cDNA were identified which encode portions of the chicken homologues of the mammalian glycine receptor alpha 1, alpha 2 and alpha 3 subunits. 2. The presence of functional glycine receptors was investigated in the whole-cell configuration of the patch-clamp technique in neurons dissociated from the ganglia and kept in culture for 7-8 days. In cells voltage clamped to -70 mV, glycine consistently induced inward currents in a concentration-dependent manner and elicited half-maximal peak current amplitudes at 43 microM. 3. The steady-state current-voltage relation for glycine-induced currents was linear between +80 and -60 mV, but showed outward rectification at more hyperpolarized potentials. Reversal potentials of these currents shifted with changes in intracellular chloride concentrations and matched the calculated Nernst potentials for chloride. 4. beta-Alanine and taurine were significantly less potent than glycine in triggering inward currents, with half-maximal responses at 79 and 86 microM, respectively. At maximally active concentrations, beta-alanine-evoked currents were identical in amplitude to those induced by glycine. Taurine-evoked currents, in contrast, never reached the same amplitude as glycine-induced currents. 5. The classical glycine receptor antagonist strychnine reversibly reduced glycine-induced currents, with half-maximal inhibition occurring at 62 nM. Two more recently characterized glycine receptor antagonists, isonipecotic acid (half-maximal inhibition at 2 mM) and 7-trifluoromethyl-4-hydroxyquinoline-3-carboxylic acid (half-maximal inhibition at 67 microM), also blocked glycine-evoked currents in a reversible manner. The chloride channel blocker picrotoxin reduced glycine-evoked currents, with half-maximal effects at 348 microM. Inhibition by the glycine receptor channel blocker cyanotriphenylborate was half-maximal at 4 microM. 6. Apart from evoking inward currents, glycine occasionally triggered short (< 100 ms) spike-like currents which were abolished by hexamethonium and thus reflected synaptic release of endogenous acetylcholine. In addition, glycine caused Ca(2+)-dependent and tetrodotoxin-sensitive tritium overflow from neurons previously labelled with [3H]noradrenaline. This stimulatory action of glycine was reduced in the presence of strychnine and after treatment with the chloride uptake inhibitor furosemide (frusemide). 7. In 65% of neurons loaded with the Ca2+ indicator fura-2 acetoxymethyl ester, glycine increased the ratio of the fluorescence signal obtained with excitation wavelengths of 340 and 380 nm, respectively, which indicates a rise in intracellular Ca2+ concentration. 8. The results show that sympathetic neurons contain transcripts for different glycine receptor alpha-subunits and carry functional heteromeric glycine receptors which depolarize the majority of neurons to trigger transmitter release.

Amino Acid Sequence↗

PubMed1997

Somatostatin inhibits excitatory transmission at rat hippocampal synapses via presynaptic receptors.

Somatostatin is one of the major peptides in interneurons of the hippocampus. It is believed to play a role in memory formation and to reduce the susceptibility of the hippocampus to seizure-like activity. However, at the cellular level, the actions of somatostatin on hippocampal neurons are still controversial, ranging from inhibition to excitation. In the present study, we measured autaptic currents of hippocampal neurons isolated in single-neuron microcultures. Somatostatin and the analogous peptides seglitide and octreotide reduced glutamatergic, but not GABAergic, autaptic currents via pertussis toxin-sensitive G-proteins. This effect was observed whether autaptic currents were mediated by NMDA or non-NMDA glutamate receptors. Furthermore, somatostatin did not affect currents evoked by the direct application of glutamate, but reduced the frequency of spontaneously occurring excitatory autaptic currents. These results show that presynaptic somatostatin receptors of the SRIF1 family inhibit glutamate release at hippocampal synapses. Somatostatin, seglitide, and octreotide also reduced the frequency of miniature excitatory postsynaptic currents in mass cultures without affecting their amplitudes. In addition, all three agonists inhibited voltage-activated Ca2+ currents at neuronal somata, but failed to alter K+ currents, effects that were also abolished by pertussis toxin. Thus, presynaptic somatostatin receptors in the hippocampus selectively inhibit excitatory transmission via G-proteins of the Gi/Go family and through at least two separate mechanisms, the modulation of Ca2+ channels and an effect downstream of Ca2+ entry. This presynaptic inhibition by somatostatin may provide a basis for its reportedly anticonvulsive action.

6-Cyano-7-nitroquinoxaline-2,3-dione↗

PubMed1997

Receptors controlling transmitter release from sympathetic neurons in vitro.

Primary cultures of postganglionic sympathetic neurons were established more than 30 years ago. More recently, these cultures have been used to characterize various neurotransmitter receptors that govern sympathetic transmitter release. These receptors may be categorized into at least three groups: (1) receptors which evoke transmitter release: (2) receptors which facilitate; (3) receptors which inhibit, depolarization-evoked release. Group (1) comprises nicotinic and muscarinic acetylcholine receptors, P2X purinoceptors and pyrimidinoceptors. Group (2) currently harbours beta-adrenoceptors, P2 purinoceptors, receptors for PACAP and VIP, as well as prostanoid EP1 receptors. In group (3), muscarinic cholinoceptors, alpha 2- and beta-adrenoceptors, P2 purinoceptors, and receptors for the neuropeptides NPY, somatostatin (SRIF1) and LHRH, as well as opioid (delta and kappa) receptors can be found. Receptors which regulate transmitter release from neurons in cell culture may be located either at the somatodendritic region or at the sites of exocytosis, i.e. the presynaptic specializations of axons. Most of the receptors that evoke release are located at the soma. There ionotropic receptors cause depolarizations to generate action potentials which then trigger Ca(2+)-dependent exocytosis at axon terminals. The signalling mechanisms of metabotropic receptors which evoke release still remain to be identified. Receptors which facilitate depolarization-evoked release appear to be located preferentially at presynaptic sites and presumably act via an increase in cyclic AMP. Receptors which inhibit stimulation evoked release are also presynaptic origin and most commonly rely on a G protein-mediated blockade of voltage-gated Ca2+ channels. Results obtained with primary cell cultures of postganglionic sympathetic neurons have now supplemented previous data about neurotransmitter receptors involved in the regulation of ganglionic as well as sympatho-effector transmission. In the future, this technique may prove useful to identify yet unrecognized receptors which control the output of the sympathetic nervous system and to elucidate underlying signalling mechanisms.

Animals↗

PubMed1997

Noradrenaline release from rat sympathetic neurones triggered by activation of B2 bradykinin receptors.

1. The role of bradykinin receptors in the regulation of sympathetic transmitter release was investigated in primary cultures of neurones dissociated from superior cervical ganglia of neonatal rats. These cultures were loaded with [3H]-noradrenaline and the outflow of radioactivity was determined under continuous superfusion. 2. Bradykinin (100 nmol l[-1] applied for 10 min) caused a transient increase in tritium outflow that reached a peak within four minutes after the beginning of the application and then declined towards the baseline, despite the continuing presence of the peptide. ATP (100 micromol l[-1]) and nicotine (10 micromol l[-1]) caused elevations in 3H outflow with similar kinetics, whereas outflow remained elevated during a 10 min period of electrical field stimulation (0.5 ms, 50 mA, 50 V cm[-1], 1.0 Hz). 3. When bradykinin was applied for periods of 2 min, the evoked 3H overflow was half-maximal at 12 nmol l(-1) and reached a maximum of 2.3% of cellular radioactivity. The preferential B1 receptor agonist des-Arg9-bradykinin failed to alter 3H outflow. The B2 receptor antagonists, [D-Phe7]-bradykinin (1 micromol l[-1]) and Hoe 140 (10 nmol l[-1]), per se did not alter 3H outflow, but shifted the concentration-response curve for bradykinin-evoked 3H overflow to the right by a factor of 7.9 and 4.3, respectively. 4. Bradykinin-induced overflow was abolished in the absence of extracellular Ca2+ and in the presence of either 1 micromol l(-1) tetrodotoxin or 300 micromol l(-1) Cd2+, as was electrically-induced overflow. Activation of alpha2-adrenoceptors by 1 micromol l(-1) UK 14,304 reduced both bradykinin- and electrically-triggered overflow. The Ca2+-ATPase inhibitor thapsigargin (0.3 micromol l[-1]) failed to alter either type of stimulated overflow. Caffeine (10 mmol l[-1]) enhanced bradykinin-induced overflow, but reduced overflow triggered by electrical field stimulation. 5. Inclusion of Ba2+ (0.1 to 1 mmol l[-1]) in the superfusion medium enhanced electrically induced overflow by approximately 100% and potentiated bradykinin-triggered overflow by almost 400%. Application of 1 mmol l(-1) Ba2+ for periods of 2 min triggered 3H overflow, and this overflow was abolished by 1 micromol l(-1) tetrodotoxin and enhanced by 10 mmol l(-1) caffeine. In contrast, inclusion of tetraethylammonium (0.1 to 1 mmol l[-1]) in the superfusion buffer caused similar increases of bradykinin- and electrically evoked 3H overflow (by about 100%), and tetraethylammonium, when applied for 2 min, failed to alter 3H outflow. 6. Treatment of cultures with 100 ng ml(-1) pertussis toxin caused a significant increase in bradykinin-, but not in electrically-, evoked tritium overflow. Treatment with 100 ng ml(-1) cholera toxin reduced both types of stimulated 3H overflow. 7. These data reveal bradykinin as a potent stimulant of action potential-mediated and Ca2+-dependent transmitter release from rat sympathetic neurones in primary cell culture. This neurosecretory effect of bradykinin involves activation of B2-receptors, presumably linked to pertussis- and cholera toxin-insensitive G proteins, most likely members of the Gq family. Results obtained with inhibitors of muscarinic K+ (KM) channels, like caffeine and Ba2+, indicate that the secretagogue action of bradykinin probably involves inhibition of these K+ channels.

Adrenergic alpha-Agonists↗

PubMed1996

Pharmacological characterization of P2 purinoceptor types in rat locus coeruleus neurons.

The frequency of spontaneous action potentials of locus coeruleus neurons was recorded extracellularly in pontine slices of the rat brain. The adenosine 5'-triphosphate (ATP) analogues alpha,beta-methylene ATP (alpha,beta-meATP) and 2-methylthio ATP increased the firing rate with a similar potency, while uridine 5'-triphosphate (UTP) was inactive. Diadenosine 5'-pentaphosphate (Ap5A), diadenosine 5'-tetraphosphate (Ap4A) and diadenosine 5'-triphosphate (Ap3A) all facilitated the firing. When equimolar concentrations were compared, Ap5A had the largest effect followed by Ap4A and Ap3A. Suramin markedly inhibited responses to alpha,beta-meATP and 2-methylthio ATP; the effect of Ap4A was only slightly depressed by suramin. Pyridoxalphosphate-6-azophenyl-2,4-disulfonic acid (PPADS) strongly antagonized alpha, beta-meATP, but failed to alter the effects of 2-methylthio ATP and Ap4A. Reactive blue 2 weakly antagonized alpha,beta-meATP and did not interfere with 2-methylthio ATP and Ap4A. Moreover, suramin depressed responses to (S)-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-D-aspartic acid (NMDA), but not to substance P. PPADS failed to affect the AMPA- and NMDA-induced increases in firing. Hence, locus coeruleus neurons may possess receptors for adenosine nucleotides (P2X and P2Y purinoceptors) and dinucleotides (P2D purinoceptors); receptors for uridine nucleotides (P2U purinoceptors or pyrimidinoceptors) are probably absent.

Adenosine↗

PubMed1996

Involvement of a phorbol ester-insensitive protein kinase C in the alpha2-adrenergic inhibition of voltage-gated calcium current in chick sympathetic neurons.

alpha2-Adrenoceptors regulate the efficacy at the sympathoeffector junction by means of a feedback inhibition of transmitter release. In chick sympathetic neurons, the mechanism involves an inhibition of N-type calcium channels, and we now present evidence that this effect involves an atypical, phorbol ester-insensitive protein kinase C (PKC). The inhibition of voltage-gated Ca2+ currents by the specific alpha2-adrenergic agonist UK 14,304 was significantly attenuated when the PKC inhibitors PKC(19-36), staurosporine, or calphostin C were included in the internal solution used to fill the patch pipettes, or if staurosporine or calphostin C were applied extracellularly; however, phorbol esters as classical activators of PKC or oleoylacetylglycerol did not mimic the effect of UK 14,304, and chronic exposure to 4-beta-phorbol dibutyrate (PDBu) did not attenuate it, ever though PKCalpha and -epsilon isozymes were translocated to plasma membranes by PDBu. The atypical isozyme PKCzeta was translocated by 100 micrometer AA and this effect was attenuated when PKC(19-36) was added to the patch pipette solution. Our observations indicate that classical, new, and atypical PKC isozymes are present in chick sympathetic neurons and that an atypical, phorbol ester-insensitive PKC is involved in the inhibition of voltage-activated calcium currents by alpha2-adrenoceptor activation.

Animals↗

PubMed1996

Synthesis and calcium antagonistic activity of 8-[N-[2-(3,4-dimethoxy- phenyl)ethyl]-beta-alanyl]-5,6,7,8-tetrahydrothieno[3,2-b][1,4]thiazepi ne fumarate.

KT-362 is an antiarrhythmic and antihypertensive agent with vasodilating activity. Since it carries a homoveratryl group in the side chain, an obvious relation exists to the verapamil-type calcium antagonists. Replacement of the fused aromatic moiety in KT-362 with thiophene provided 8-[N-[2-(3,4-dimethoxyphenyl) ethyl]-beta-alanyl]-5,6,7,8-tetrahydrothieno[3,2-b][1,4] thiazepine (1). Compound 1 shows a negative chronotropic activity in spontaneously beating right atria (IC50 = 23 microM, n = 7), and a negative inotropic effect in papillary muscles (IC50 = 2.7 microM, n = 7) and left atria (IC50 = 4 microM, n = 6) of the guinea-pig heart. The decrease of contractility in papillary muscles could be antagonized by increasing the extracellular calcium concentration. Compound 1 was found to affect high (IC50: 70 +/- 5 microM) and low (IC50: 129 +/- 34 microM) voltage-activated calcium channel currents as well as voltage-activated sodium channel currents (IC50: 80 +/- 13 microM) in chick dorsal root ganglion neurons. In addition nicotine-induced currents were potently inhibited (IC50: 6 +/- 0.7 microM) in bovine chromaffin cells.

Animals↗

PubMed1996

A somatostatin receptor inhibits noradrenaline release from chick sympathetic neurons through pertussis toxin-sensitive mechanisms: comparison with the action of alpha 2-adrenoceptors.

The effects of somatostatin and analogues were investigated in cultures of chick sympathetic neurons. Electrically evoked tritium overflow from cultures labelled with [3H]noradrenaline was reduced by somatostatin-14 in a concentration-dependent manner, with half maximal effects at 0.3 nM and a maximum of 45% inhibition. Somatostatin-28 was equipotent to somatostatin-14 (half maximal concentration at 0.5 nM), and seglitide was less potent, the effects being half maximal at 4.2 nM. The inhibitory action of somatostatin-14 on stimulation-evoked overflow desensitized within minutes at 100 nM, but not at 10 nM, and was abolished by a pretreatment of neurons with pertussis toxin. All somatostatin analogues reduced voltage-activated Ca2+ currents recorded in the whole-cell configuration of the patch-clamp technique, with somatostatin-14 being equipotent to somatostatin-28, but more potent than seglitide. However, the inhibition of Ca2+ currents occurred at concentrations more than ten-fold higher than those required for the reduction of stimulation evoked 3H overflow. The action of somatostatin upon Ca2+ currents was also abolished by pertussis toxin and desensitized within minutes. In preceding experiments, alpha 2-adrenoceptor activation had been found to reduce transmitter release and Ca2+ currents of chick sympathetic neurons through a pertussis toxin-sensitive mechanism. In the present study, the alpha 2-adrenergic agonist UK 14,304 completely occluded the inhibition of Ca2+ currents and of electrically evoked overflow by somatostatin-14. Neither UK 14,304 nor somatostatin affected the resting membrane potential or voltage-dependent K+ currents. These results demonstrate that chick sympathetic neurons possess SRIF1 type somatostatin receptors which control transmitter release. This effect is mediated by pertussis toxin-sensitive GTP binding proteins and apparently involves an inhibition of voltage-activated Ca2+ channels, but not a modulation of K+ channels. Since alpha 2-adrenergic agonists share all of these actions and occlude the effects of somatostatin, alpha 2-adrenoceptors and SRIF1 receptors seem to regulate sympathetic transmitter release via common signalling mechanisms.

Adrenergic alpha-Agonists↗

PubMed1996

Cholera toxin induces cyclic AMP-independent down-regulation of Gs alpha and sensitization of alpha 2-autoreceptors in chick sympathetic neurons.

The role of the stimulatory GTP-binding protein (Gs) in the alpha 2-autoinhibitory modulation of noradrenaline release was investigated in cultured chick sympathetic neurons. The alpha 2-adrenoceptor agonist UK 14,304 caused a concentration-dependent reduction of electrically evoked [3H] noradrenaline release with half-maximal effects at 14.0 +/- 5.5 nM. In neurons treated with 100 ng/ml cholera toxin for 24 h, the half-maximal concentration was lowered to 3.2 +/- 1.4 nM without changes in the maximal effect of UK 14,304. The pretreatment with cholera toxin also increased the inhibitory action of 10 nM UK 14,304 when compared with the inhibition of noradrenaline release in untreated cultures derived from the same cell population. In cultures treated with either 10 microM forskolin or 100 microM 8-bromo-cyclic AMP, neither the half-maximal concentration nor the maximal effect of UK 14,304 was altered. Cholera toxin, forskolin, and 8-bromo-cyclic AMP all induced an increase in spontaneous outflow and a reduction in electrically evoked overflow, effects not observed after a pretreatment with dideoxyforskolin. Exposure of neurons to cholera toxin, but not to forskolin or 8-bromo-cyclic AMP, induced a translocation of alpha-subunits of Gs (Gs alpha) from particulate to soluble fractions and led ultimately to a complete loss of Gs alpha from the neurons. In contrast, no effect was seen on the distribution of either alpha-subunits of Gi- or Ga-type G proteins or of beta-subunits. These results indicate that cholera toxin causes a selective, cyclic AMP-independent down-regulation of Gs alpha. This down-regulation of Gs alpha is associated with the sensitization of alpha 2-autoreceptors.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1996

Inhibition of N-type calcium channels: the only mechanism by which presynaptic alpha 2-autoreceptors control sympathetic transmitter release.

Alpha 2-Adrenoceptors are known to inhibit voltage-dependent Ca2+ channels located at neuronal cell bodies; the present study investigated whether this or alternative mechanisms, possibly downstream of Ca2+ entry, underlie the presynaptic alpha 2-adrenergic modulation of transmitter release from chick sympathetic neurons. Using chick sympathetic neurons, overflow of previously incorporated [3H]noradrenaline was elicited in the presence of extracellular Ca2+ by electrical pulses, 25 mM K+ or 10 microM nicotine, or by adding Ca2+ to otherwise Ca(2+)-free medium when cells had been made permeable by the calcium ionophore A23187 or by alpha-latrotoxin. Pretreatment of neurons with the N-type Ca2+ channel blocker omega-conotoxin GVIA and application of the alpha 2-adrenergic agonist UK 14304 reduced the overflow elicited by electrical pulses, K+ or nicotine, but not the overflow caused by Ca2+ after permeabilization with alpha-latrotoxin or A23187. In contrast, the L-type Ca2+ channel blocker nitrendipine reduced the overflow due to K+ and nicotine, but not the overflow following electrical stimulation or alpha-latrotoxin- and A23187-permeabilization. The inhibition of electrically evoked overflow by UK 14304 persisted in the presence of nitrendipine and the L-type Ca2+ channel agonist BayK 8644, which per se enhanced overflow. In omega-conotoxin GVIA-treated cultures, electrically evoked overflow was also enhanced by BayK 8644 and almost reached the value obtained in untreated neurons. However, UK 14304 lost its effect under these conditions. Whole-cell recordings of voltage-activated Ca2+ currents corroborated these results: UK 14304 inhibited Ca2+ currents by 33%, nitrendipine caused a 7% reduction, and BayK 8644 increased the currents by 30%. Moreover, the dihydropyridines failed to abolish the inhibition by UK 14304, but pretreatment with omega-conotoxin GVIA, which reduced mean amplitude from 0.95 to 0.23 nA, entirely prevented alpha 2-adrenergic effects. Our results indicate that the alpha 2-autoreceptor-mediated modulation of noradrenaline release from chick sympathetic neurons relies exclusively on the inhibition of omega-conotoxin GVIA-sensitive N-type Ca2+ channels. Mechanisms downstream of these channels and voltage-sensitive Ca2+ channels other than N-type appear not to be important.

Animals↗

PubMed1996

Suramin analogues as subtype-selective G protein inhibitors.

G protein alpha subunits expose specific binding sites that allow for the sequential, conformation-dependent binding of protein reaction partners, e.g., G protein beta gamma dimers, receptors, and effectors. These domains represent potential sites for binding of low-molecular-weight inhibitors. We tested the following suramin analogues as G protein antagonists: 8-(3-nitrobenzamido)-1,3,5-naphtalenetrisulfonic acid (NF007), 8-(3-(3-nitrobenzamido)benzamido)-1,3,5-naphtalenetrisulfonic++ + acid NF018), 8,8'-(carbonylbis(imino-3,1-phenylene))bis-(1,3,5-naphtalenetri sulfonic acid) (NF023), 8,8'-(carbonylbis(imino-3,1-phenylene)carbonylimino-(3,1-phe nylene))bis-(1,3, 5-naphtalenetrisulfonic acid) (NF037), and suramin. The compounds suppressed [35S]GTPgammaS binding to purified, recombinant G protein alpha subunits, an effect that is due to inhibition of GDP release. Suramin is selective for recombinant Gsalpha-s (EC50 values o f approximately 240 nM; rank order of potency, suramin > NF037 > NF023 > NF018 > NF007), whereas NF023 is selective for recombinant Gi alpha-1 and recombinant Go alpha (EC50 value of approximately 300 nM; rank order of potency, NF023 > / = NF037 > suramin >0 NF018 > NF007). Selectivity was also demonstrated on a cellular level. In rat sympathetic neurons, alpha-2-adrenergic and muscarinic receptor-dependent inhibition of the voltage-sensitive calcium current is mediated by Gi/Go, whereas inhibition by vasoactive intestinal peptide (VIP) is mediated by Gs. Calcium current inhibition by alpha2-adrenergic and muscarinic receptors was greatly reduced when 100 microM NF023 was applied intracellularly, whereas the response to VIP was unaffected; in contrast, the response to VIP was blunted only with 100 microM suramin in the recording pipette. The suramin analogues do not interfere with the interaction between alpha subunits and G protein beta gamma dimer but compete with binding of the effector. The addition of purified adenylyl cyclase reverses the inhibitory effect of suramin on the rate of [35S]GTPgammaS binding to recombinant Gsalpha-s, indicating direct competition for a common site; similarly, immunoprecipitation by an antibody directed against an epitope of the effector binding site is inhibited by suramin. Our results show that it is possible to design G protein inhibitors that target the effector binding site on the alpha subunits.

Animals↗

PubMed1995

alpha 2-Adrenoreceptor-mediated inhibition of acetylcholine-induced noradrenaline release from rat sympathetic neurons: an action at voltage-gated Ca2+ channels.

[3H]Noradrenaline release was studied in cultured sympathetic neurons derived from superior cervical ganglia of neonatal rats. Acetylcholine elicited a concentration- and time-dependent increase in 3H outflow which was half-maximal at about 300 microM and within 5 s. The overflow induced by 10 s exposure to 300 micro A acetylcholine was reduced by the nicotinic antagonist hexamethonium, but increased by the muscarinic antagonist atropine. Cd2+ (300 microM) prevented the overflow evoked by electrical field stimulation, but reduced acetylcholine-induced overflow by less than 50%. Removal of extracellular Ca2+ abolished stimulation-evoked tritium overflow irrespective of the stimulus. The selective alpha2-adrenoceptor agonist UK 14,304 inhibited acetylcholine-evoked overflow to a significantly smaller extent (approximately 25% maximal inhibition) than electrically induced overflow ( > or = 45% maximal inhibition). These inhibitory effects were antagonized by the alpha2-adrenoceptor antagonist yohimbine. Noradrenaline (0.1 microM) reduced acetylcholine-evoked overflow to the same extent as did UK 14,304 (0.1 microM). UK 14,304 had no effect when 3H overflow was evoked by acetylcholine in the presence of 300 microM Cd2+. Currents through nicotinic acetylcholine receptors and voltage-activated Ca2+ currents were studied with the whole-cell variant of teh patch-clamp technique. UK 14,304 reduced nicotinic acetylcholine receptor currents and voltage-activated Ca2+ currents with similar potency and efficacy. Yohimbine, however, antagonized only the inhibition of voltage-activated Ca2+ currents, but not the effects of UK 14,304 on nicotinic receptor currents. Furthermore, yohimbine per se reduced currents through nicotinic receptors. Noradrenaline (10 microM) inhibited voltage-dependent Ca2+ currents just as did UK 14,304 (10 microM), but failed to reduce currents through nicotinic acetylcholine receptor channels. Cd2+ (300 microM) abolished voltage-activated Ca2+ currents and reduced nicotinic acetylcholine receptor currents by 65%. These results indicate that acetylcholine evokes noradrenaline release from rat sympathetic neurons by activation of nicotinic receptors and restricts this release via muscarinic receptors. The acetylcholine-induced transmitter release is based on two mechanisms, one involving and the other one bypassing voltage-dependent Ca2+ channels. alpha2-Adrenoceptor activation reduces voltage-activated Ca2+ currents and effects exclusively the component of acetylcholine-induced release which involves voltage-dependent Ca2+ channels. These results support the hypothesis that voltage-activated Ca2+ channels are the sole site of autoinhibitory alpha2-adrenergic effects on transmitter release from rat sympathetic neurons. The inhibitory effects of alpha2-adrenoceptor agonists and antagonists on currents through nicotinic acetylcholine receptors are not mediated by an alpha2-adrenoceptor.

Acetylcholine↗

PubMed1995

Behavioral analysis and nursing interventions for reducing disruptive behaviors of patients with dementia.

In this study, two cases were used to examine the potential of behavioral analysis as an intervention to decrease disruptive behaviors of institutionalized individuals with dementia. Behavioral modeling was used to teach the principles of behavioral analysis. The nurses observed a behavior change plan implemented by the investigator, revised the plan based on behavioral analysis techniques, and subsequently implemented the revised behavioral strategies. The patients' disruptive behaviors decreased markedly when the behavioral intervention was implemented. Patients resumed disruptive behaviors when care was provided without the behavioral plan. The findings suggest the need to address interventions in nursing research, education, and practice that use behavioral analysis to reduce disruptive behaviors in individuals with dementia.

Aged↗

PubMed1995

Rapid, agonist-induced desensitization of alpha 2-autoreceptors modulating transmitter release.

1. The release of previously incorporated [3H]-noradrenaline was investigated in cultures of dissociated chick or rat sympathetic neurones and in cerebrocortical slices from neonatal or adult rats. Noradrenaline, in the presence of 10 mumol l-1 of the uptake inhibitor, cocaine, or the selective alpha 2-adrenoceptor agonist, 5-bromo-N-(4,5-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine (UK 4,304), was applied for different periods of time in order to detect a possible time-dependence of the alpha 2-adrenoceptor-mediated inhibition of electrically evoked tritium outflow. 2. In chick sympathetic neurones, stimulation-evoked overflow was reduced to 30%, 42%, or 56% of control when noradrenaline (1 mumol l-1) was present for 2, 8, or 16 min, respectively. Likewise, UK 14,304 (1 mumol l-1) present for these periods of time reduced 3H overflow to 35%, 51%, and 53% of control, respectively. Addition of 1 nmol l-1 to 10 mumol l-1 UK 14,304 for either 2 or 16 min did not produce significantly different IC50 values, but the inhibitory effects were smaller with 16 min as compared to 2 min exposure at concentrations > or = 10 nmol l-1. 3. In rat sympathetic neurones, noradrenaline (100 nmol l-1) reduced stimulation-evoked overflow to 33%, 56%, or 57% of control, when present for 2, 8, or 16 min, respectively. Addition of UK 14,304 (1 mumol l-1) for these periods of time caused inhibition to 11%, 41%, and 46% of control. Applying UK14,304 for either 2 or 16 min did not result in significantly different IC5o values, but the inhibition induced by 16 min as compared to 2 min exposure was smaller at concentrations > 10 nmol 1-1.4. In cerebrocortical slices from either neonatal or adult rats, exposure to 0.1 to 1.0 micromol 1-1 UK14,304 for 16 min never caused a smaller inhibition than a corresponding 3 min exposure, although various experimental conditions were investigated.5 The results demonstrate that alpha 2-adrenoceptors which regulate noradrenaline release from sympathetic neurones undergo agonist-induced desensitization within minutes. Such rapid desensitization of alpha 2-autoreceptors was not detected in brain slice preparations.

Adrenergic alpha-Agonists↗

PubMed1995

UTP- and ATP-triggered transmitter release from rat sympathetic neurones via separate receptors.

In rat cultured sympathetic neurones, UDP, UTP and ATP at micromolar concentrations triggered Ca(2+)-dependent and tetrodotoxin-sensitive [3H]-noradrenaline release. The overflow evoked by UTP or ATP was similar at 100 mumol l-1, the concentration used in all subsequent experiments. Pre-exposure of the neurones to 100 mumol l-1 UTP significantly reduced ensuing secretory effects of UTP but not of ATP. Conversely, pre-exposure to ATP diminished the overflow due to ATP but not that due to UTP. In the presence of 10 mumol l-1 pyridoxal-5'-phosphate or 30 mumol l-1 suramin, the secretory response to ATP was reduced, but the effect of UTP was unaltered. Zn2+ (10 mumol l-1) reduced the overflow triggered by UTP, but increased the overflow due to ATP. These results indicate the presence of separate receptors for pyrimidine nucleotides and for purine nucleotides which both trigger transmitter release.

Adenosine Triphosphate↗

PubMed1995

Prostate cancer in African American men: increasing knowledge and self-efficacy.

An educational and screening program for prostate cancer designed to increase knowledge and self-efficacy in African American men was provided in African American churches in a major midwestern city. Modeling was provided by trained lay educators who were African American men previously diagnosed and treated for prostate cancer and who could serve as role models of the desired behaviors. Pretests and posttests developed for this study and provided to the participants were the Prostate Cancer Screening Knowledge Inventory and the Prostate Cancer Screening Self-Efficacy Scale. Paired t tests demonstrated that after completing the church-based intervention, participants had significantly improved knowledge and self-efficacy scores related to prostate cancer screening.

Adult↗

PubMed1995

Synovial fibroblast-like cells strongly express jun-B and C-fos proto-oncogenes in rheumatoid- and osteoarthritis.

To identify recently activated cells in the synovial membrane (SM) of patients with rheumatoid arthritis (RA), the in situ expression of the proto-oncogenes jun-B, c-jun, jun-D, and c-fos was assessed by means of immunohistochemistry and in situ hybridization techniques. SM from patients with osteoarthritis (OA) or joint trauma (JT), as well as from normals (No) were used as controls. Numerous cells expressing high levels of jun-B and c-fos were found within lining layer and diffuse infiltrates in the vicinity of inflammatory cells, but only a few in lymphoid follicles and endothelia. The positive cells were spindle-shaped, CD14- and CD3-negative and, in addition, expressed mRNA for collagen alpha 2 (I) and alpha 1 (III), indicating that they were fibroblasts. In control OA, JT, and even No SM, individual fibroblast-like cells stained as strongly as in RA; however, the density of positive cells was substantially lower. In RA SM, fibroblasts, but not lymphocytes or macrophages, appear to undergo in situ activation. Quantitative differences among RA, OA, and JT may be related to different degrees of inflammatory infiltration.

Arthritis, Rheumatoid↗