Purification and properties of phosphotransacetylase from Lactobacillus fermenti.
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Acyltransferases↗
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A Kv3-like persistent, outwardly rectifying, Cs+-permeable, K+ current in rat subthalamic nucleus neurones.
A persistent outward K+ current (IPO), activated by depolarization from resting potential, has been identified and characterized in rat subthalamic nucleus (SThN) neurones using whole-cell voltage-clamp recording in brain slices. IPO both rapidly activated (tau = 8 ms at +5 mV) and deactivated (tau = 2 ms at -68 mV), while showing little inactivation. Tail current reversal potentials varied with extracellular K+ concentration in a Nernstian manner. Intracellular Cs+ did not alter either IPO amplitude or the voltage dependence of activation, but blocked transient (A-like) outward currents activated by depolarization. When extracellular K+ was replaced with Cs+, IPO tail current reversal potentials were dependent upon the extracellular Cs+ concentration, indicating an ability to conduct Cs+, as well as K+. IPO was blocked by Ba2+ (1 mM), 4-aminopyridine (1 mM) and tetraethylammonium (TEA; 20 mM), with an IC50 for TEA of 0.39 mM. The IPO conductance appeared maximal (38 nS) at around +27 mV, half-maximal at -13 mV, with the threshold for activation at around -38 mV. TEA (1 mM) blocked the action potential after-hyperpolarization and permitted accommodation of action potential firing at frequencies greater than around 200 Hz. We conclude that IPO, which shares many characteristics of currents attributable to Kv3.1 K+ channels, enables high-frequency spike trains in SThN neurones.
4-Aminopyridine↗
Hormone-releasing silastic intrauterine devices: effect or provera-releasing intrauterine devices on two species of primates.
Silastic intrauterine contraceptive devices (IUDs) 13 mm long and 1.07 mm in diameter could be inserted easily into patas monkey uteri which, like human uteri, expelled them. Addition of 10% Provera to these devices did not reduce the expulsion rate significantly in our study. Control devices had no effect on cycle length in rhesus monkeys. After insertion, the active IUDs frequently caused a delay in onset of menstruation; however, cycles did occur with the device in situ, and normal-length cycles were resumed following removal of the device. A short period of rapid release (almost 35% of the total amount) of Provera from the device was followed by a longer period of sustained release of low levels of the hormone. Even 9 mug/day were sufficient to maintain a decidual reaction in the endometrium of the rhesus monkey. The drug could not be detected in the blood stream at 3,6, or 12 hours in patas monkeys or at 1 or 2 months in rhesus monkeys and so may never have reached the systemic circulation. Devices currently under study in baboons catain Provera or one of three other steroids to determine whether these compounds improve retention rates as well as meet the other two criteria set for the ideal IUD incorporation, for unless we meet this first criterion we can never achieve, let alone test, the others.
Animals↗
An ATP-dependent inwardly rectifying potassium channel, KAB-2 (Kir4. 1), in cochlear stria vascularis of inner ear: its specific subcellular localization and correlation with the formation of endocochlear potential.
Cochlear endolymph has a highly positive potential of approximately +80 mV. This so-called endocochlear potential (EP) is essential for hearing. Although pivotal roles of K+ channels in the formation of EP have been suggested, the types and distribution of K+ channels in cochlea have not been characterized. Because EP was depressed by vascular perfusion of Ba2+, an inhibitor of inwardly rectifying K+ (Kir) channels, but not by either 4-aminopyridine or tetraethylammonium, we examined the expression of Kir channel subunits in cochlear stria vascularis, the tissue that is supposed to play the central role in the generation of positive EP. Of 11 members of the Kir channel family examined with reverse transcription-PCR, we could detect only expression of KAB-2 (Kir4.1) mRNA in stria vascularis. KAB-2 immunoreactivity was specifically localized at the basolateral membrane of marginal cells but not in either basal or intermediate cells. Developmental expression of KAB-2 in marginal cells paralleled formation of EP. Furthermore, deaf mutant mice (viable dominant spotting; WV/WV) expressed no KAB-2 in their marginal cells. These results suggest that KAB-2 in marginal cells may be critically involved in the generation of positive EP.
4-Aminopyridine↗
Nitration of benzo[a]pyrene adsorbed on coal fly ash particles by nitrogen dioxide: role of thermal activation.
Nitration of benzo[a]pyrene (BaP) by nitrogen dioxide (NO2) adsorbed on the surface of thermally activated coal fly ash and model aluminosilicate particles led to the formation of nitrobenzo[a]pyrenes as verified by extraction and gas chromatography/mass spectrometry (GC/MS). In situ diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS) was utilized to follow the nitration reaction on the surface of zeolite Y. Nitrobenzo[a]pyrene formation was observed along with the formation of nitrous acid and nitrate species. The formation of the BaP radical cation was also observed on thermally activated aluminosilicate particles by electron spin resonance (ESR) spectroscopy. On the basis of GC/MS, DRIFTS, and ESR spectroscopy results, a mechanism of nitration involving intermediate BaP radical cations generated on thermally activated aluminosilicate particles is proposed. These observations have led to the hypothesis that nitration of adsorbed polyaromatic hydrocarbons on coal fly ash by reaction with nitrogen oxides can occur in the smokestack, but with the aging of the fly ash particles, the extent of the nitration reaction will be diminished.
Adsorption↗
Cerebellar GABAB receptors modulate function of GABAA receptors.
Interactions between GABAA and GABAB receptors were studied using muscimol-stimulated uptake of 36Cl- by membrane vesicles from mouse cerebellum. Baclofen inhibited muscimol-stimulated 36Cl- uptake and this action was more pronounced with longer flux times (30 vs. 3 s) and after predesensitization of GABAA receptors. Baclofen also inhibited 36Cl- flux by cortical membranes but was more effective with cerebellar preparations. The action of baclofen was stereoselective, calcium-dependent, and blocked by the GABAB receptor antagonist 2-OH-saclofen. It was mimicked by GTP-gamma-S but not by GDP-beta-S, which suggests that baclofen may be acting via a G protein. The action of baclofen was inhibited by U73122, an inhibitor of phospholipase C. However, the potassium channel blockers tetraethylammonium or Ba2+ did not affect the action of baclofen. The results show that activation of GABAB receptors can inhibit the function of GABAA receptors and suggest that this action involves either a nondesensitizing subtype of GABAA receptor or the rate or recycling of desensitized to nondesensitized receptors. We speculate that this action of baclofen results from activation of phospholipase C and phosphorylation of a subtype of GABAA receptor by protein kinase C.
Animals↗
Transient and sustained depolarization of retinal ganglion cells by Ih.
1. Using whole cell patch-clamp methods, we have identified an inward cationic current activated by hyperpolarization (Ih) in somata of goldfish retinal ganglion cells. 2. Ih activated at test potentials between -70 and -105 mV, and did not appear to inactivate during prolonged hyperpolarizations under voltage clamp. During step hyperpolarizations from holding potentials between -70 and -40 mV, apparent activation was faster at more negative test potentials. On repolarization from -105 mV to holding potentials between -75 and -55 mV, Ih deactivated exponentially at rates showing no marked voltage dependence (tau = approximately 100 ms). 3. Ih tail currents reversed at membrane potentials consistent with a relative permeability to Na+ and K+ of roughly 0.5, when pipette and bath solutions both contained Na+ and K+. 4. Ih was readily blocked by extracellular Cs+ (3 mM), but was resistant to block by tetraethylammonium (30 mM), Ba2+ (1 mM), or Co2+ (2.4 mM). 5. Time-dependent voltage rectification developed during injection of hyperpolarizing current under current clamp. After current injection ceased, membrane potential depolarized beyond resting potential, often leading to anode-break-like spikes. Both voltage rectification and voltage overshoot were suppressed by extracellular Cs+. 6. Voltage-clamp measurements in the presence and absence of Cs+ were used to model membrane potential changes produced by exogenous current injections, by hyperpolarizing synaptic inputs, and by termination of both. Modeled responses resembled membrane potential changes measured under current clamp when terms for activation and deactivation of Ih were included. 7. The voltage rectification and anode-break-like spikes observed in isolated cells resemble those recorded during and after light-evoked hyperpolarizations of retinal ganglion cells in situ. Ih may transiently augment retinal ganglion cell excitability after termination of hyperpolarizing light stimuli, and thus promote encoding of stimulus timing.
Animals↗
k-Opioid receptor activation of a dendrotoxin-sensitive potassium channel mediates presynaptic inhibition of mossy fiber neurotransmitter release.
Activation of kappa-opioid receptors on mossy fiber terminals in the hippocampus inhibits excitatory amino acid release. The mechanism of presynaptic inhibition at the mossy fiber synapse was investigated through whole-cell voltage-clamp of CA3 pyramidal cells. The application of a kappa-opioid agonist, U69593, reduced the amplitude of the excitatory postsynaptic current response, and this effect was reversed with a k receptor antagonist. Presynaptic potassium channels were blocked by bath application of channel toxins, and the effect of kappa receptor activation was tested. The inhibition caused by U69593 was blocked by low doses of 4-aminopyridine (30 microM) and the selective peptide toxins dendrotoxin and mast cell degranulating peptide. The inhibition was not blocked by low doses of tetraethylammonium chloride (1 mM), barium, or glibenclamide. Thus, we conclude that presynaptic kappa-opioid receptors are coupled to a Shaker-type voltage-dependent potassium channel that is sensitive to dendrotoxin and mast cell degranulating peptide. An increase in presynaptic potassium conductance would enhance the rate of repolarization after action potential invasion, thereby limiting calcium influx and neurotransmitter release. This is the first physiological demonstration of the involvement of a dendrotoxin-sensitive potassium current in presynaptic inhibition mediated by a G protein-coupled receptor.
4-Aminopyridine↗
Assay of phenolic compounds in red wine by on-line combination of capillary isotachophoresis with capillary zone electrophoresis.
The on-line combination of capillary zone electrophoresis (CZE) with capillary isotachophoresis (ITP) increases significantly the separation capability and sensitivity of capillary electrophoresis. This technique was used for separation and quantification of fourteen selected natural constituents in red wine belonging to flavonoids and phenolic acids. The leading electrolyte (LE) in the ITP pre-separation step was 10 mM HCl of pH* 7.2 with Tris as counterion, the terminating electrolyte (TE) was 50 mM boric acid of pH* 8.2 (adjusted with barium hydroxide). The background electrolyte in the electrophoretic step contained 25 mM beta-hydroxy-4-morpholinopropanesulfonic acid (MOPSO), 50 mM Tris, 15 mM boric acid and 5 mM beta-cyclodextrin of pH* 8.5. The content of methanol in all electrolytes was 20% (v/v). For exact timing of the transfer of isotachophoretically stacked analyte zones into the CZE column and for the control of the residual amount of leading and terminating ITP electrolytes picric acid was used as coloured marker. The R.S.D. values (n = 6) ranged between approximately 0.1% (for 0.25 microg ml(-1) rutin) and approximately 11% (for 0.25 microg ml(-1) of quercitrin). Detection limits were 30 ng mi(-1) for phenolic acids, quercitrin and rutin, 100 ng ml(-1) for quercetin, kaempferol and epicatechin and 250 ng ml(-1) for catechin. A single analysis took 45 min.
Electrophoresis↗
"Calcium antagonists": a class of drugs with a bright future. Part II. Determination of basic pharmacological properties.
This minireview discusses some simple pharmacological tests useful in detecting biological activity (screening), characterizing mechanisms of action and predicting possible therapeutic applications for calcium antagonists in general and calcium slow channel blockers in particular. In smooth muscle preparations these agents inhibit mechanical effects evoked by K+-depolarization which selectively opens voltage-operated calcium channels (VOC) to allow extracellular Ca++ into the cytosol. In contrast, any inhibition of receptor-mediated responses by calcium antagonists appears to depend on the transduction system and the specific cellular mechanism (e.g. VOC opening consequent to partial depolarization) activated by the receptor and, evidently, on ancillary pharmacological properties of the studied compound. For instance, whereas calcium slow channel blockers antagonize contractions produced by norepinephrine and K+-depolarization in the rat isolated portal vein, they inhibit effectively only the latter response in the rabbit aorta. This apparent discrepancy may be accounted for by the different pool of Ca++ mobilized in the two tissues by norepinephrine. Agents (e.g. diphenylalkylamines, calmodulin blockers) that impair the interaction of Ca++ with intracellular proteins produce effects which are less specific than those of slow channel blockers. Currently, the pharmacological profile of calcium antagonists can be appropriately defined by studying their effects on radioligand (dihydropyridine) binding, radioactive calcium movements through biological membranes, electrophysiological parameters in cardiac and vascular smooth muscle and on various in vivo cardiovascular preparations. Together, these approaches allow a functional classification of new calcium antagonists in relation to already known compounds and some hypotheses on their potential clinical applications. Finally, desirable pharmacokinetics and pharmacological properties for novel calcium antagonists are mentioned. This point will be further explored in the forthcoming minireview which will deal with the clinical applications of calcium antagonists.
Animals↗
Naked clusters of 56 tin atoms in the solid state.
Isolated, gigantic tin clusters of 56 atoms are discovered in the ternary compound Ba(16)Na(204)Sn(310) (cubic, F(-)43m, Z = 1, a = 25.2041(8) A) made by direct fusion of the elements at 800 degrees C. The cluster, made of four face-fused pentagonal dodecahedra, has 36 pentagonal faces and 90 edges, and resembles a concave fullerene "dented" at four places. It is made of three- and four-bonded tin atoms and is "stuffed" with four barium cations, [Ba(4)@Sn(56)](36-). This is the largest main group naked cluster in the solid state besides the fullerenes. Also occurring in the structure are two other isolated clusters of tin, Sn(16-n) (n = 0, 1, 2, 3, or 4) and Sn(8).
Journal Article↗
99m-technetium-HmPAO-labelled leucocytes in the diagnosis of acute colonic diverticulitis.
Eighteen patients with abdominal symptoms clinically resembling acute colonic diverticulitis were studied with 99mTc-labelled leucocytes on average 5 days after the onset of symptoms. The diagnosis of diverticular disease was confirmed in all cases by means of barium enema or sigmoidoscopy. The leucocyte scan was positive in 13 patients, nonspecific in four and unclear in one patient. The diagnostic possibilities in acute colonic diverticulitis are discussed.
Acute Disease↗
[Effects of bis(dimethyl amino)-diphenyl methane (BDDM) on action potentials in guinea pig papillary muscle].
The effects of BDDM on action potentials and slow response action potentials in guinea pig papillary muscle were investigated by microelectrode technique. BDDM prolonged action potentials duration at 30, 50, 90, and 100% repolarization (APD30, APD50, APD90, APD100) and prolonged the effective refractory period (ERP), so ERP/APD value became greater. BDDM (51.6 mumol/L) decreased the APA, Vmax and prolonged APD50, APD90. In barium-induced ventricular autorhythmicity, BDDM suppressed the maximal diastolic potential (MDP), APA and reduced the rate of spontaneous rhythm. The results suggest that BDDM may unspecifically inhibit the currents of Ca2+, K+, and Na+.
Action Potentials↗
[Dissolution and radio-opacity of apatite cement].
Dissolution and remineralization behaviors of self-setting apatite cement is organic acids and synthetic saliva and the effect of barium apatite (BaAp)on physicochemical and mechanical properties of the apatite cement were investigated. One-week solubility of several types of apatite cement was considerably smaller than that of commercially available dental cements in 1 mM of acetic, lactic and citric acids (pH4.0 at 37 degrees C). For example, the one-week solubility in citric acids was less than 4.5%. To investigate the dissolution behavior of the apatite cement in detail, two types of 45Ca labeled apatite cement were prepared. In one, the seed apatite was labeled with 45Ca(45Ca-HAp cement) and in the other the matrix apatite was labeled with 45Ca through use of 45Ca-DCPD (45Ca-DCPD cement). Solubility, estimated from the concentration of 45Ca released in 1mM of the organic acid was approximately zero for 45Ca-HAp cement, whereas the solubility of 45Ca-DCPD cement was similar to that of unlabeled cements. This suggests cat dissolution of the matrix apatite governs dissolution of the set cement. In synthetic saliva, the solution phosphate decreased with time once the set cement was introduced to the solution, whereas the solution calcium increased. The former finding suggests that some phosphate compounds precipitated in the synthetic saliva and the latter finding indicates that some portion of the set cement dissolved. The thermodynamic analysis of the solution compositions strongly suggests that remineralization is a major process and dissolution a minor one. In fact, in systhetic saliva labeled with 45Ca having a degree of supersaturation with respect to apatite comparable to rest saliva, 45Ca concentration in solution decreased once the cement pellet was introduced. This finding clearly suggests that the set apatite cement has the ability to remineralize but not to dissolve in synthetic saliva even if the degree of supersaturation with respect to apatite is relatively low. Addition of barium hydroxyapatite (BaAp) successfully bestowed clinically acceptable radio-opacity to the apatite cement. To accelerate the setting reaction which was retarded by Ba2+ released from BaAp at a lower pH during the first stage of spatulation, calcium hydroxyapatite (CaAp) was added to the cement mixture. At 20wt% of BaAp and 20wt% CaAp, the setting reaction proceeded at a neutral or weak alkaline pH, which is one of the most promising aspects of the apatite cement. The cement spaturated at L/P = 0.4 set within 10 minutes and its radio-opacity was comparable to or more than that of tooth enamel.(ABSTRACT TRUNCATED AT 400 WORDS)
Acetates↗
Characterisation of Ca(2+)-dependent inwardly rectifying K+ currents in HeLa cells.
The whole-cell configuration of the patch-clamp technique was used to examine K+ currents in HeLa cells. Under quasi-physiological ionic gradients, using an intracellular solution containing 10(-7) mol/l free Ca2+, mainly outward currents were observed. Large inwardly rectifying currents were elicited in symmetrical 145 mmol/l KCl. Replacement of all extracellular K+ by isomolar Na+, greatly decreased inward currents and shifted the reversal potential as expected for K+ selectivity. The inwardly rectifying K+ currents exhibited little or no apparent voltage dependence within the range of from -120 mV to 120 mV. A square-root relationship between chord conductance and [K+] at negative potentials could be established. The inwardly rectifying nature of the currents was unaltered after removal of intracellular Mg2+ and chelation with ATP and ethylenediaminetetraacetic acid (EDTA). Permeability ratios for other monovalent cations relative to K+ were: K+ (1.0) > Rb+ (0.86) > Cs+ (0.12) > Li (0.08) > Na+ (0.03). Slope conductance ratios measured at -100 mV were: Rb+ (1.66) > K+ (1.0) > Na+ (0.09) > Li (0.08) > Cs+ (0.06). K+ conductance was highly sensitive to intracellular free Ca2+ concentration. The relationship between conductance at 0 mV and Ca2+ concentration was well described by a Hill expression with a dissociation constant, KD, of 70 nmol/l and a Hill coefficient, n, of 1.81. Extracellular Ba2+ blocked the currents in a concentration- and voltage-dependent manner. The dependence of the KD for the blockade was analysed using a Woodhull-type treatment, locating the ion interaction site at 19% of the distance across the electrical field of the membrane and a KD (0 mV) of 7 mmol/l. Tetraethylammonium and 4-aminopyridine were without effect whilst quinine and quinidine blocked the currents with concentrations for half-maximum effects equal to 7 mumol/l and 3.5 mumol/l, respectively. The unfractionated venom of the scorpion Leiurus quinquestriatus (LQV) blocked the K+ currents of HeLa cells. The toxins apamin and scyllatoxin had no detectable effect whilst charybdotoxin, a component of LQV, blocked in a voltage-dependent manner with half-maximal concentrations of 40 nmol/l at -120 mV and 189 nmol/l at 60 mV; blockade by charybdotoxin accounts for the effect of LQV. Application of ionomycin (5-10 mumol/l), histamine (1 mmol/l) or bradykinin (1-10 mumol/l) to cells dialysed with low-buffered intracellular solutions induced K+ currents showing inward rectification and a lack of voltage dependence.
Adenosine Triphosphate↗
Latent adenosine deaminase in mouse brain. II. Purification and properties of mitochondrial and supernatant adenosine deaminases.
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Adenosine Triphosphate↗
Tegaserod (HTF 919) stimulates gut motility in normal horses.
REASONS FOR PERFORMING STUDY: It has been shown that the selective 5-HT4 receptor agonist tegaserod induces an increase in frequency and amplitude of contractions in isolated muscle preparations of equine ileum and pelvic flexure. OBJECTIVES: To investigate the effects of tegaserod on gut motility and transit of spheres in normal horses. METHODS: Six mature Freiberger horses were kept under standardised conditions. Effects of tegaserod (0.02 mg/kg bwt i.v. b.i.d. for 2 days) or vehicle on intestinal transit of barium-filled spheres, defaecation and gut sounds were studied in a cross-over design. Spheres were given via stomach tube prior to the first dosing of tegaserod or vehicle. Faeces were collected every 3 h and spheres eliminated were identified radiologically in the faeces. RESULTS: Tegaserod significantly accelerated the gastrointestinal (GI) transit time of spheres and increased the frequency of defaecation and scores of gut sounds compared to vehicle. The compound was well tolerated; no effects on behaviour, body temperature, heart rate, respiratory rate and clinical laboratory data were observed. CONCLUSIONS: Tegaserod efficaciously stimulated motility and accelerate GI transit in healthy horses. POTENTIAL RELEVANCE: Tegaserod may offer therapeutic potential in horses suffering from impaction or paralytic ileus.
Animals↗
Characterization of K+ channels in the basolateral membrane of rat tracheal epithelia.
To study K+ channels in the basolateral membrane of chloride-secreting epithelia, rat tracheal epithelial monolayers were cultured on permeable filters and mounted into an Ussing chamber system. The mucosal membrane was permeabilized with nystatin (180 microg/ml) in the symmetrical high K+ (145 mm) Ringer solution. During measurement of the macroscopic K+ conductance properties of the basolateral membrane under a transepithelial voltage clamp, we detected at least two types of K+ currents: one is an inwardly rectifying K+ current and the other is a slowly activating outwardly rectifying K+ current. The inwardly rectifying K+ current is inhibited by Ba2+. The slowly activating K+ current was potentiated by cAMP and inhibited by clofilium, phorbol 12-myristate 13-acetate (PMA) and lowering temperature. This is consistent with the biophysical characteristics of ISK channel. RT-PCR analysis revealed the presence of ISK cDNA in the rat trachea epithelia. Although 0.1 mM Ba2+ only had minimal affect on short-circuit current (Isc) induced by cAMP in intact epithelia, 0.1 mM clofilium strongly inhibited it. These results indicate that ISK might be important for maintaining cAMP-induced chloride secretion in the rat trachea epithelia.
Animals↗