Effect of Exogenous Gangliosides on Amino Acid Uptake and Na+,K+-ATPase Activity in Superior Cervical and Nodose Ganglia of Rats

The effects of some gangliosides on active uptake of nonmetabolizable alpha-aminoisobutyric acid (AIB) and Na+, K+-ATPase and Ca2+, Mg2+-ATPase activities in superior cervical ganglia (SCG) and nodose ganglia (NG) excised from adult rats were examined during aerobic incubation at 37 degrees C for 2 h. In NG, amino acid uptake was greatly accelerated with the addition of galactosyl-N-acetylgalactosaminyl-[N-acetylneuraminyl]-galactosylgluc osyl ceramide (GM1) (85%) and also with N-acetylgalactosaminyl-[N-acetylneuraminyl]-galactosylglucosyl ceramide (GM2) or [N-acetylneuraminyl]-galactosyl-N-acetylgalactosaminyl-[N-acetyl- neuraminyl]-galactosylglucosyl ceramide (GD1a) (43% each) compared with a nonaddition control at a 5 nM concentration. Under identical conditions, Na+, K+-ATPase activity was strongly stimulated with GM1 (180%) and GD1a (93%), whereas Ca2+, Mg2+-ATPase activity showed no change. In SCG, on the other hand, AIB uptake was apparently inhibited (-27%) by addition of GM1, with a slight decrease in Na+, K+-ATPase but no change in Ca2+, Mg2+-ATPase activity in the tissue. Both asialo-GM1, in which N-acetylneuraminic acid is deficient, and Forssman glycolipid, which is not present in nervous tissue, failed to produce any significant increase in both SCG and NG not only in amino acid uptake, but also in Na+, K+-ATPase activity. A kinetic study of active AIB uptake showed that GM1 ganglioside produced an increase in Km with no change in Vmax in SCG, whereas it caused a decrease in Km with a slight increase in Vmax in NG. Treatment of NG and SCG with neuraminidase from Vibrio cholerae, an enzyme that split off sialic acid from polysialoganglioside, leaving GM1 intact, caused little inhibition of the amino acid uptake.(ABSTRACT TRUNCATED AT 250 WORDS)     

[3H]Dipyridamole Binding to Guinea Pig Brain Membranes: Possible Heterogeneity of Central Adenosine Uptake Sites

The binding of [3H]dipyridamole ([3H]DPR) to guinea pig brain membranes is described and compared to that of [3H]nitrobenzylthioinosine ([3H]NBI). The binding of [3H]DPR is saturable, reversible, and specific with pharmacologic evidence indicating that this ligand is binding to the adenosine uptake site. Compared to [3H]NBI the binding of [3H]DPR is of higher capacity (Bmax = 208 +/- 16 fmol/mg protein for [3H]NBI and 530 +/- 40 fmol/mg protein for [3H]DPR) and lower affinity (KD = 0.35 +/- 0.02 nM for [3H]NBI and 7.6 +/- 0.7 nM for [3H]DPR). The adenosine uptake inhibitors are the most potent inhibitors of binding (Ki of 10(-8)-10(-7) M) whereas adenosine receptor ligands such as cyclohexyladenosine, 2-chloroadenosine, and various methylxanthines are several orders of magnitude less potent (Ki 10(-5)-10(-2). The inhibition of [3H]DPR binding by NBI is biphasic, with only 40% of binding being susceptible to inhibition of NBI concentrations less than 10(-5) M. The tissue distribution of [3H]DPR binding parallels that of [3H]NBI although in most cases significantly more sites are observed with [3H]DPR. Calcium channel blocking agents such as nifedipine, nimodipine, and verapamil are also inhibitors of [3H]DPR binding with potencies in the micromolar range. The data are consistent with [3H]DPR being a useful additional ligand for the adenosine uptake site and provide evidence that multiple uptake binding sites exist of which only about 40% are NBI-sensitive.     

Effect of Quinolinic Acid in the Nucleus Basalis Magnocellularis on Cortical High-Affinity Choline Uptake

A transient 45% increase in cortical high-affinity choline uptake (HACU) was observed after an injection of quinolinic acid (QUIN) into the nucleus basalis magnocellularis (nbM) of the rat. This was followed by a steady decline in choline uptake, which resulted in a 46% decrease by day 7. Specific [3H]hemicholinium-3 binding to coronal brain sections showed a similar pattern following injections of QUIN into the nbM. The increase in cortical HACU elicited by QUIN appeared to be dose dependent.     

Low-Level Microwave Irradiations Affect Central Cholinergic Activity in the Rat

Sodium-dependent high-affinity choline uptake was measured in various regions of the brains of rats irradiated for 45 min with either pulsed or continuous-wave low-level microwaves (2,450 MHz; power density, 1 mW/cm2; average whole-body specific absorption rate, 0.6 W/kg). Pulsed microwave irradiation (2-microseconds pulses, 500 pulses/s) decreased choline uptake in the hippocampus and frontal cortex but had no significant effect on the hypothalamus, striatum, and inferior colliculus. Pretreatment with a narcotic antagonist (naloxone or naltrexone; 1 mg/kg i.p.) blocked the effect of pulsed microwaves on hippocampal choline uptake but did not significantly alter the effect on the frontal cortex. Irradiation with continuous-wave microwaves did not significantly affect choline uptake in the hippocampus, striatum, and hypothalamus but decreased the uptake in the frontal cortex. The effect on the frontal cortex was not altered by pretreatment with narcotic antagonist. These data suggest that exposure to low-level pulsed or continuous-wave microwaves leads to changes in cholinergic functions in the brain.     

N1-Methyl-2-125I-Lysergic Acid Diethylamide, a Preferred Ligand for In Vitro and In Vivo Characterization of Serotonin Receptors

Methylation of 2-125I-lysergic acid diethylamide (125I-LSD) at the N1 position produces a new derivative, N1-methyl-2-125I-lysergic acid diethylamide (125I-MIL), with improved selectivity and higher affinity for serotonin 5-HT2 receptors. In rat frontal cortex homogenates, specific binding of 125I-MIL represents 80-90% of total binding, and the apparent dissociation constant (KD) for serotonin 5-HT2 receptors is 0.14 nM (using 2 mg of tissue/ml). 125I-MIL also displays a high affinity for serotonin 5-HT1C receptors, with an apparent dissociation constant of 0.41 nM at this site. 125I-MIL exhibits at least 60-fold higher affinity for serotonin 5-HT2 receptors than for other classes of neurotransmitter receptors, with the dopamine D2 receptor as its most potent secondary binding site. Studies of the association and dissociation kinetics of 125I-MIL reveal a strong temperature dependence, with very slow association and dissociation rates at 0 degree C. Autoradiographic experiments confirm the improved specificity of 125I-MIL. Selective labeling of serotonin receptors was observed in all brain areas examined. In vivo binding studies in mice indicate that 125I-MIL is the best serotonin receptor label yet described, with the highest frontal cortex to cerebellum ratio of any serotonergic radioligand. 125I-MIL is a promising ligand for both in vitro and in vivo labeling of serotonin receptors in the mammalian brain.     

Host immunoglobulin G titre and antibody activity in haemolymph of the tick, Ornithodoros moubata

Immunoglobulin G (IgG) in tick haemolymph was analysed immunochemically and biochemically for its antigenicity, antibody activity and relative concentration in a soft tick, Ornithodoros moubata (Murray) sensu Walton 1962 (Acari: Argasidae). Ouchterlony immunodiffusion tests showed that haemolymph from a tick engorged on rabbit IgG (or human IgG) through an artificial membrane, reacted with anti-rabbit IgG (anti-human IgG) but not with anti-human IgG (anti-rabbit IgG). This indicates that haemolymph of the fed tick contains IgG with a similar antigen specificity to host blood IgG. IgG from tick haemolymph was demonstrated by enzyme immunoassay to have the same antibody activity as ingested IgG. The IgG concentration in tick haemolymph was measured by a quantitative single immunodiffusion test. Changes of IgG titre after a bloodmeal were correlated with IgG activity, which was low for 5 days after a bloodmeal and then suddenly increased. The IgG titre reached a maximum 7 days post-engorgement, and remained high for over 4 months during and after oviposition. 125I-labelled IgG was injected into the tick haemocoel to determine the persistence of IgG in the haemolymph. Recovery of labelled IgG was low at 1 and 3 days, and high at 5, 8 and 16 days after engorgement. The data suggest that IgG in haemolymph disappears quickly soon after engorgement possibly by degradation and/or absorption (adhesion to tissues).     

Atrial granules in the dog heart after cardiac denervation

Summary Because the increase in sodium excretion during left atrial distension in conscious dogs is abolished after chronic cardiac denervation, we have investigated whether this is a result of the disappearance of specific atrial granules. Electron microscopy and light-microscopical and ultrastructural immunohistochemistry of canine atria show that atrial granules displaying immunoreactivity for cardiac hormones of the cardiodilatin/atrial natriuretic polypeptide (CDD/ANP) family are still present in denervated left and right atria, although reduced in quantity. It is concluded that the atrial-induced natriuresis is not only related to the existence of specific atrial granules. The functional link between atrial-induced natriuresis provoked by atrial distension and the release of atrial polypeptide hormones remains uncertain because the denervated heart can secrete CDD although the diuretic-natriuretic effect is altered.     

Detection and characterization of acidic compartments (vacuoles) in Chlorella vulgaris 11h cells by 31P-in vivo NMR spectroscopy and cytochemical techniques

Acidic inorganic phosphate (Pi) pool (pH around 6) was detected besides the cytoplasmic pool in intact cells of Chlorella vulgaris 11h by ³¹P-in vivo nuclear magnetic resonance (NMR) spectroscopy. It was characterized as acidic compartments (vacuoles) in combination with the cytochemical technique; staining the cells with neutral red and chloroquine which are known as basic reagents specifically accumulated in acidic compartments. Under various conditions, the results obtained with the cytochemical methods were well correlated with those obtained from in vivo NMR spectra; the vacuoles were well developed in the cells at the stationary growth phase where the acidic Pi signal was detected. In contrast, cells at the logarithmic phase in which no acidic Pi signal was detected contained only smaller vesicles that accumulated these basic reagents. No acidic compartment was detected by both cytochemical technique and ³¹P-NMR spectroscopy when the cells were treated with NH₄OH. The vacuolar pH was lowered by the anaerobic treatment of the cells in the presence of glucose, while it was not affected by the external pH during the preincubation ranging from 3 to 10. Possible vacuolar functions in unicellular algae especially with respect to intracellular pH regulation are discussed.Non-standard abbreviations EDTA ethylenediaminetetraacetic acid - HEPES N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid - MDP methylene diphosphonic acid - NMR nuelear magnetic resonance - PCA perchloric acid - PCV packed cell volume - Pi inorganic phosphate - Pic sytoplasmic inorganic phosphate - Piv vacuolar inorganic phosphate - ppm parts per million - SP sugar phosphates - TCA trichloroacetic acid     

Stimulation of phospholipase D activity and indication of acetylcholine synthesis by oleate in rat brain synaptosomal preparations

It had been previously demonstrated that the oleate activation of synaptosomal membrane phospholipase D liberated choline which was available for acetylcholine formation. The present investigations were undertaken to determine if oleate might have an effect on choline uptake by synaptosomes. It was observed that oleate interfered with choline uptake when incubations were carried out at 37°C but uptake was stimulated at 3°C. Oleate was the most effective fatty acid of several tested. Preliminary observations suggest the presence of a membranous form of choline acetyltransferase.     

Mutagenicity,sister chromatid exchange inducibility andin vitro cell transforming ability of particulates from Athens air

Airborne particulates were collected over a period of twelve months by the use of Hi-Vol samplers in the basin of Athens, Greece. N-Hexane extracts were tested in a battery ofin vitro tests for their ability to induce mutation in bacteria as well as mutation, sister chromatid exchange and morphological transformation in cultured mammalian cells. Positive results were found for mutagenicity withSalmonella strain TA98 in the Ames assay, for sister chromatid exchange induction in CHO cells and for transformation in BALB/c 3T3 cells in culture. They also showed weak non-doserelated induction of ouabain resistance in BALB/c 3T3 cells. The contribution of oxidizing and nitrating agents found in the Athens atmosphere, together with sunlight UV irradiation in the formation of direct acting mutagens and potential carcinogens from ambient polycyclic aromatic hydrocarbons, is suggested.Abbreviations FCS fetal calf serum - FPG fluorescent-plus-Giemsa technique - oua^R ouabain resistant - PAH polycyclic aromatic hydrocarbon - SCE sister chromatid exchange - TSP total suspended particulate