Comparison of the effects of central and peripheral aluminum administration on regional 2-deoxy-D-glucose incorporation in the rat brain

Intracerebroventricular (ICV) Injection of aluminum tartrate (ALT 205.7 mcg) in the rat induces a progressive encephalopathy characterized by neurobehavioral derangements, by the slowing of the background rhythm of the quantitative electroencephalogram and by learning and memory deficits. The condition, lethal within about 35 days, is associated with a reduced ability of cerebral synaptosomes to incorporate radiolabeled 2-Deoxy-D-glucose (2DG) in vitro. The present study surveyed and compared the in vivo regional cerebral glucose uptake (rCGlu) capacity of rats injected with ALT 7 or 14 days previously either by the ICV or intraperitoneal (120 mg/Kg) routes. ICV injection produces transient rCGlu depression in caudate-putamen, geniculate bodies and periaquaeductal gray, resolving by day 14. Thalamic nuclei exhibit depressed rCGlu by the 7th day undergoing further depression by day 14. The rCGlu of occipitoparietal cortices, normal at day 7, was increased by day 14. In contrast, peripheral aluminum administration produced transient rCGlu depression in olfactory bulbs, frontal and occipitoparietal cortices, nucleus accumbens and cerebellum, and transiently increased rCGlu in the geniculate nuclei. These effects, present by day 7, had resolved by day 14 when rCGlu had increased in the previously normal pontine nuclei and decreased in the previously normal hippocampus. Neither treatment changed rCGlu in the septal nuclei, globus pallidus, amygdala, olfactory cortex, substantia nigra, superior or inferior colliculi or the medullary nuclei. The pattern of anomalies in cerebral 2DG incorporation most probably indexes the deranged glucoregulatory and metabolic demands of these brain areas in the aluminum intoxicated state.     

Synergistic effect of salt mixture on the gelation temperature and morphology of methylcellulose hydrogel

Gelation temperature of methylcellulose (MC) can be altered by adding different additives. Pure MC showed sol-gel transition at 60°C. Sodium citrate and sodium tartrate were used alone and in combination to see the effect of individual salt and combination of salts on the gelation temperature of MC. The gelation temperature of all the binary and ternary combinations of MC and salts were measured with different methods such as test tube tilting method (TTM), UV-vis spectroscopy, viscometry, and by rheometer and also the morphology of gels were characterized with the help of environmental scanning electron microscopy (ESEM). It was observed that when 0.1M sodium citrate (NaC) and 0.1M sodium tartrate (NaT) were used separately, the gelation temperature of MC was reduced up to 44°C and 47°C respectively but when mixture of NaC and NaT (0.1(M) NaC and 0.1(M) (NaT)) were used the gelation temperature was further reduced to 36°C. It was clear from ESEM images that when NaC and NaT were used separately the formation of network was not distinguishable. But, well-connected network structure was observed when a mixture 0.1M NaC and 0.1M NaT was used.     

Ontogenetic development of electrocorticogram in the rat

Spontaneous electrocorticogram (ECoG) was recorded in frontal (sensorimotor) temporal (auditory) and occipital (visual) cortical regions of 86 male rats (immobilized with d-tubocurarine) aged from 3 days to adulthood. Activity which could be classified as ECoG was for the first time recorded in 5-day-old rats; it was formed by groups of slow waves with unstable frequency intermingled with periods of isoelectric line. Discontinuous ECoG activity was regularly registered even in 10-day-old rats, exceptionally in 12-day-old rats. During further maturation of the continuous ECoG an increase in frequency and an establishment of a basic rhythmic activity synchronous over both hemispheres took place, so that 25- and 30-day-old rats did not differ from the adult ones. Autocorrelagrams and power frequency spectra demonstrated a broad frequency range of the basic rhythm as well as delay in the development of occipital cortical areas in comparison to frontal areas.     

Reductions in calcium uptake induced in rat brain synaptosomes by ionizing radiation

Gamma irradiation (60Co) reduced KCl-stimulated voltage-dependent 45Ca2+ uptake in whole-brain, cortical, and striatal synaptosomes. The time course (3, 10, 30, and 60 s) of calcium uptake by irradiated (3 Gy) and nonirradiated synaptosomes, as well as the effect of KCl (15-65 mM), was measured in whole-brain synaptosomes. The fastest and highest rate of depolarization-dependent calcium uptake occurred at 3 s with 65 mM KCl. Irradiation reduced calcium uptake at all incubation times and KCl concentrations. Bay K 8644 enhancement of KCl-stimulated calcium influx was also reduced by radiation exposure. Nimodipine binding to dihydropyridine (DHP) L-type calcium channel receptors was not altered following radiation exposure. These results demonstrate an inhibitory effect of ionizing radiation on the voltage-sensitive calcium channels in rat brain synaptosomes that are not mediated by DHP receptors.     

An inhibitor of dopamine uptake,LR5182, CIS-3-(3,4-dichlorophenyl)-2-N,N-dimethylaminomethyl-bicyclo-[2,2,2]-octane,hydrochloride

LR5182 inhibited the uptake of dopamine in rat striatal synaptosomes and the uptake of norepinephrine in cortical synaptosomes with inhibitor constants, Ki values, of 3nM and 58nM, respectively. It was only a week inhibitor of serotonin uptake in cortical synaptosomes with a Ki value of 1.7μM. The uptake of dopamine and norepinephrine were significantly lowered within an hour after an intraperitoneal injection of LR5182. Among known inhibitors of dopamine uptake in synaptosomes of rat brain, LR5182 is most effective and selective. The rigid structure of LR5182 (Figure 1) suggested a gauche conformation of dopamine to be favored by the striatal uptake of dopamine.     

Cortical modulation of cholinergic neurons in the striatum

Previous reports suggest the existence of a cortico-striatal pathway which might use glutamate as the transmitter. In the present study, the possible influence of this pathway on striatal cholinergic neurons was investigated. Two weeks following surgical destruction of the cerebral cortex, the high affinity uptake of glutamate and choline into striatal synaptosomes was significantly reduced whereas GABA uptake was unaffected. In acute experiments (1 hour following decortication), only choline uptake was significantly reduced while the uptake of glutamate and GABA were not altered. Acute injection (2 minutes) of kainic acid into the striatum, 1 hour after decortication, reversed the effect of the decortication on choline uptake, perhaps by simulating an excitatory input to the striatum which was presumably removed by the cortical ablation. These observations are consistent with the existence of a cortical input (perhaps glutamatergic) to the striatum and suggest that striatal cholinergic neurons can be influenced by this cortico-striatal pathway.     

The effect of cyclo (Leu-Gly) on chemical denervation supersensitivity of dopamine receptors induced by intracerebroventricular injection of 6-hydroxydopamine in mice

Chemical denervation supersensitivity of postsynaptic dopamine receptors was induced in mice by intracerebroventricular injection of 6-hydroxydopamine. Fourteen days after the 6-hydroxydopamine injection, mice exhibited greater spontaneous locomotor activity and hypothermic response when challenged intraperitoneally with apomorphine. Whole brain levels of dopamine were reduced by 80%. Daily subcutaneous administration of cyclo (Leu-Gly) (50 μg/mouse/day) for 14 days inhibited the development of dopamine receptor supersensitivity induced by 6-hydroxydopamine as evidenced by the blockade of an apomorphine induced locomotor and hypothermic effect. Cyclo (Leu-Gly) did not alter the depletion of brain dopamine induced by 6-hydroxydopamine. These data suggest that cyclo (Leu-Gly) can block the development of dopamine receptor supersensitivity induced by 6-hydroxydopamine without protecting the neurons from dopamine depletion.     

3H]adrenaline release from hypothalamic synaptosomes and its modulation by clonidine: effects of chronic antidepressant drug regimens

[3H]Adrenaline ([3H]ADR, 40 nM) was accumulated by rat hypothalamic synaptosomes (P2) more rapidly and in significantly greater amounts than by similar preparations from cerebral cortex. There was no significant difference between these two tissues in the rate or amount of [3H]noradrenaline ([3H]NA, 40 nM) accumulation. Talusupram (10 microM), maximally inhibited the uptake of [3H]ADR into hypothalamic synaptosomes by 60%. Nomifensine further inhibited uptake by 14%. From these observations it was concluded that some [3H]ADR was accumulated into non adrenergic neuronal terminals. The effects of desipramine (DMI, 10 mg/kg/day and clorgyline (1 mg/kg/day) administration for 28 days on K+-evoked release of [3H]ADR was investigated using superfused hypothalamic synaptosomes. After both chronic antidepressant drug regimens, total [3H]ADR release (spontaneous + evoked) was significantly reduced. Evoked release of [3H]ADR (by KCl, 16 mM) was significantly reduced after the DMI but not the clorgyline regimens. Presynaptic alpha 2-adrenoceptor function in the hypothalamus was assessed during superfusion by measuring the reduction in K+-evoked release of [3H]ADR caused by clonidine (1 microM). The attenuating effects of clonidine on [3H]ADR release (42% in untreated controls and 36% after chronic clorgyline) was diminished (to 4%) after chronic DMI administration. Alpha 2 adrenoceptor numbers in the rat hypothalamus were not significantly changed after clorgyline or DMI administration, suggesting that the functional subsensitivity seen in synaptosomes after DMI, may not be related to alpha 2 adrenoceptor down regulation.     

Effects of pinealectomy and melatonin treatments on serotonin uptake and release from synaptosomes of rat hypothalamic regions

This study examined the effects induced by long-term pinealectomy, daily melatonin treatment to pinealectomized and intact rats, and a single melatonin injection on [¹⁴C]-serotonin (5-HT) uptake and release from synaptosomes obtained of hypothalamic regions. Pinealectomy inhibited the accumulation of labeled 5-HT by synaptosomes of the preoptic area-anterior hypothalamus (POA-AH), but it failed to alter the [K⁺]-evoked 5-HT release. Melatonin treatment for 10 consecutive days to pinealectomized rats restored 5-HT uptake in POA-AH, and also increased 5-HT release in medial and posterior hypothalamus. These results suggest that pineal melatonin plays a stimulatory role on the serotoninergic terminals of the hypothalamus. Moreover, when daily melatonin treatment was administered to intact rats a significant increase in 5-HT uptake activity by synaptosomes of all the hypothalamic regions was observed, but 5-HT release was unaffected. In contrast, a single melatonin injection induced a significant decrease in 5-HT release from synaptosomes of the POA-AH was observed. The results suggest the existence of a differential sensitivity in the mechanisms mediating melatonin actions on 5-HT uptake/release, which depends on the presence of the pineal gland in the animals and on the frequency of the treatments with the pineal hormone.     

CHOLINE: SELECTIVE ACCUMULATION BY CENTRAL CHOLINERGIC NEURONS

Abstract— Most of the cholinergic input to the hippocampus was destroyed by placement of lesions in the medial septal area. In animals with such lesions we found that hippocampal ChAc activity was reduced by 85–90% and endogenous acetylcholine levels were reduced by more than 80 %. When hippocampal synaptosomes from animals with lesions were incubated with [³H]choline at concentrations of 7.5 nm, 1 μm and 10 μm there was approximately a 60 % reduction in the uptake of [³H]choline, suggesting that cholinergic nerve endings were mainly responsible for [³H]choline uptake. At 0.1 mm concentrations of [³H]choline, there was only a 25 % reduction of choline uptake, suggesting that at higher concentrations of choline there was more nonspecific uptake. The uptake of radiolabelled tryptophan, glutamate and GABA were only slightly or not at all affected by the lesions. There was a significant reduction of uptake of radiolabelled serotonin and norepinephrine, since known monoaminergic tracts were disrupted. Choline uptake was reduced only in brain regions in which cholinergic input was interrupted (i.e. the cerebral cortex and hippocampus) and remained unchanged in other regions (i.e. the cerebellum and striatum). The time course of the reduction in choline uptake was similar to that of the reductions in ChAc activity and endogenous ACh levels; there was no decrease at 1 day, a significant decrease at 2 days, and the maximal decrease at 4 days postlesion. There was a close correlation among choline uptake, ChAc activity and ACh levels in the four brain regions examined (i.e. the striatum, cerebral cortex, hippocampus and cerebellum). Our results suggest that when hippocampal synaptosomes (and perhaps synaptosomes from other brain areas as well) are incubated in the presence of choline, at concentrations of 10 μm m or lower, then cholinergic nerve endings are responsible for the bulk of the choline accumulated by the tissue.     

Irreversible Binding and Recovery of the Norepinephrine Uptake System Using an Alkylating Derivative of Norepinephrine

The effects of bromoacetylaminomenthylnorepinephrine (BAAN) on the sodium-dependent, high-affinity norepinephrine (NE) uptake system in rat brain synaptosomes and CNS neuronal cultures were investigated. BAAN inhibited [3H]NE uptake into synaptosomes in a dose- and time-dependent manner (IC50, 6.5 microM). Pretreatment of cortical synaptosomes or neuronal cells with BAAN alone, followed by washing to remove free drug, reduced the Vmax but did not alter the Km value for [3H]NE uptake. The BAAN-induced reduction in Vmax was attenuated by concurrent pretreatment with desipramine and blocked by the reaction of BAAN with dithiothreitol or cysteine. In contrast, BAAN was 19-fold less potent at inhibiting [3H]dopamine uptake in striatal synaptosomes, and no change in the Vmax or Km value for [3H]dopamine uptake was observed after a pretreatment with BAAN followed by washing. Furthermore, the irreversible beta-antagonist, bromoacetylalprenololmentane, was equipotent to BAAN for inhibiting [3H]NE uptake into cortical synaptosomes, but did not alter the Vmax or Km for [3H]NE after pretreatment. In neuronal cultures, BAAN inhibited sodium-dependent uptake of [3H]NE (IC50, 5.6 microM) with no effect on sodium-independent uptake. After pretreatment of cultures with 30 microM BAAN followed by washing, there was a 74% decrease in the Vmax for [3H]NE uptake. Following a 24-h lag period, uptake recovered to the control level within 48 h; however, recovery was completely blocked by cycloheximide. The data indicate that BAAN irreversibly binds to the [3H]NE uptake system in both CNS synaptosomes and neuronal cultures and may be a useful probe for studying the turnover of the [3H]NE uptake system.     

Inhibition of serotonin uptake in synaptosomes and glia cells by some pharmacological substances

The uptake of serotonin -14 C by glial cells and synaptosomes of the rabbit brain cortex was studied. The Km value of the uptake of serotonin -14 C proved to be equal (0.83 + 0.02 microM) both for synaptosomes and glial cells. Synaptosomes of the rabbit brain cortex take up serotonin -14 C twice as fast as glial cells (uptake rates were compared from protein). Among psychotropic drugs studied the tricyclic antidepressant imipramine and psychostimulant cocaine turned out the most active inhibitors of both synaptosomal and glial uptake of serotonin -14 C. The drugs in 50 microM concentration inhibit the uptake of serotonin -14 C in synaptosomes and glial cells by 90 and 75-80%, respectively.     

REGIONAL TRANSPORT OF TRYPTOPHAN IN RAT BRAIN

Abstract— Tryptophan uptake was studied in brain slices and synaptosomes prepared from regions known to vary in the numbers of serotoninergic cell bodies and nerve endings that they contain. The rate of tryptophan uptake was highest in hypothalamus for both types of preparation. Differences among the regions were much more pronounced in isolated nerve endings (synaptosomes). Loading with tryptophan did not affect the uptake into tissue slices. Tryptophan accumulation in hypothalamus synaptosomes was reduced after intraventricular injection of 5,7–dihydroxytryptamine whereas no change was observed in synaptosomes prepared from cerebellum under the same conditions; accumulation by synaptosomes prepared from the hypothalamic and hippocampal regions was reduced after raphe lesions.     

Studying brain functions with mesoscopic measurements: Advances in electrocorticography for non-human primates

Our brain is organized in a modular structure. Information in different modalities is processed within distinct cortical areas. However, individual cortical areas cannot enable complex cognitive functions without interacting with other cortical areas. Electrocorticography (ECoG) has recently become an important tool for studying global network activity across cortical areas in animal models. With stable recordings of electrical field potentials from multiple cortical areas, ECoG provides an opportunity to systematically study large-scale cortical activity at a mesoscopic spatiotemporal resolution under various experimental conditions. Recent developments in thin, flexible ECoG electrodes permit recording field potentials from not only gyral but intrasulcal cortical surfaces. Our review here focuses on the recent advances of ECoG applications to non-human primates.     

Reductions of Γ-Aminobutyric Acid and Glutamate Uptake and (Na++ K+)-ATPase Activity in Brain Slices and Synaptosomes by Arachidonic Acid

Arachidonic acid, a major polyunsaturated fatty acid of membrane phospholipids in the CNS, reduced the high-affinity uptake of glutamate and gamma-aminobutyric acid (GABA) in both rat brain cortical slices and synaptosomes. alpha-Aminoisobutyric acid uptake was not affected. Intrasynaptosomal sodium was increased concomitant with decreased (Na+ + K+)-ATPase activity in synaptosomal membranes. The reduction of GABA uptake in synaptosomes could be partially reversed by alpha-tocopherol. The inhibition of membrane-bound (Na+ + K+)-ATPase by arachidonic acid was not due to a simple detergent-like action on membranes, since sodium dodecyl sulfate stimulated the sodium pump activity in synaptosomes. These data indicate that arachidonic acid selectively modifies membrane stability and integrity associated with reductions of GABA and glutamate uptake and of (Na+ + K+)-ATPase activity.     

GABA release and uptake measured in crude synaptosomes from Genetic Absence Epilepsy Rats from Strasbourg (GAERS).

GABA release and uptake were examined in Genetic Absence Epilepsy Rats from Strasbourg and in non-epileptic control animals, using crude synaptosomes prepared from the cerebral cortex and thalamus. Uptake of [3H]GABA over time was reduced in thalamic synaptosomes from epileptic rats, compared to controls. The affinity of the uptake process in thalamic synaptosomes was lower in epileptic animals. NNC-711, a ligand for the GAT-1 uptake protein, reduced synaptosomal uptake by more than 95%; beta-alanine, an inhibitor selective for the uptake proteins GAT-2 and -3, did not significantly reduce synaptosomal uptake. Autoradiography studies using [3H]tiagabine, a ligand selective for GAT-1, revealed no differences between the strains in either affinity or levels of binding. Ethanolamine O-sulphate (100 microM), a selective inhibitor of GABA-transaminase, did not affect uptake levels. Aminooxyacetic acid (10-100 microM), an inhibitor of GABA-transaminase and, to a lesser extent, glutamate decarboxylase, caused an increase in measured uptake in both thalamic and cortical synaptosomes, in both strains. We found no difference in in vitro basal or KCl-stimulated endogenous GABA release between epileptic and control rats. These results indicate that GABA uptake in the thalamus of Genetic Absence Epilepsy Rats from Strasbourg was reduced, compared to control animals. The lower uptake affinity in the epileptic animals probably contributed to the reduction in uptake over time. Uptake appeared to be mediated primarily by the 'neuronal' transporter GAT-1. Autoradiography studies revealed no differences in the number or affinity of this uptake protein. It is therefore possible that altered functional modulation of GAT-1 caused the decrease in uptake shown in the epileptic animals. Inhibition of GABA-transaminase activity had no effect on measured GABA uptake, whereas a reduction in glutamate decarboxylase activity may have affected measured uptake levels.     

Uptake of L-glutamine into synaptosomes. Is the γ-glutamyl cycle involved?

Transport of L-glutamine into rat cortical synaptosomes has been investigated by (14C)L-glutamine uptake experiments. This amino acid enters synaptosomes both by an active carrier mediated system, which may be the result of gamma-glutamyl cycle activity and by a Na+-dependent transport system. This view is supported by the following observations: a) as demonstrated previously (10), glutamine inside synaptosomes reaches concentrations higher than those of the incubation medium, and initial rates of uptake approach saturation kinetics; b) the uptake of glutamine is inhibited by uncouplers; c) the uptake is inhibited by methionine sulfoximine, a suicide-inhibitor of an enzyme of the gamma-glutamyl cycle; d) the initial rate of uptake is lowered by decreasing the Na+-level of the incubation medium or by adding ouabain. The validity of this hypothesis is discussed.     

Na+, K+-ATPase activity and serotonin transport in the rat brain synaptosomes fractions after acute 1,2-dichloroethane intoxication and nicotinamide administration

Alterations of Na+,K+-ATPase activity and serotoninergic system functioning were investigated in brain synaptosomes fractions of rats under experimental acute 1,2-dichloroethane (DChE) intoxication. It was shown that Na+,K+-ATPase activity was markedly increased (by 41,8%) in a period of 24 h after DChE intoxication and decreased (by 27%) after 48 h intoxication. The level of [2-14C]-serotonin uptake by synaptosomes was progressively diminished after 24 and 48 h after DChE injection whereas the activity of monoamine uptake proved to be unchanged. Nicotinamide (200 mg/kg of body weight) was administered to rats subjected to DChE 1, 24 and 36 h after poisoning. The treatment of rats with nicotinamide resulted in some normalization of brain synaptosomal Na+, K+-ATPase activity and serotonin uptake controlled at 48 h after DChE intoxication.     

Studies on the Role of α2-Adrenoceptors in the Control of Synaptosomal [3H]5-Hydroxytryptamine Release: Effects of Antidepressant Drugs

The sensitivity of alpha 2-adrenoceptors on 5-hydroxytryptamine (5-HT) nerve endings obtained from rat cerebral cortex was investigated following treatment with the antidepressant drugs desipramine (10 mg/kg/day for 21-28 days) or clorgyline (1 mg/kg/day for 21-28 days). [3H]5-HT (100 nM) was used to load cortical synaptosomes (P2) after experiments with uptake inhibitors confirmed that this concentration of amine ensured exclusive uptake into 5-HT nerve terminals. The sensitivity of K+-stimulated release of [3H]5-HT to alpha 2-adrenoceptor occupancy was assessed in a superfusion system by means of the dose-dependent inhibition of [3H]5-HT release by clonidine. This is blocked by yohimbine (1 microM), which, when administered alone, enhances release, suggesting that endogenous catecholamines released from other synaptosomes act on these alpha 2-heteroreceptors. The effect of addition of citalopram (1 microM) to superfusates suggests that some reuptake of [3H]5-HT occurs during superfusion. Of the tritium released into superfusates during "background" and K+-stimulated release, 17 and 90%, respectively is [3H]5-HT. The attenuation of K+-stimulated release by clonidine is apparently diminished by the chronic clorgyline regimen but not by desipramine. However, clorgyline elevates catecholamine levels, and this might increase endogenous noradrenaline (NA) efflux, which by competition with clonidine could appear to alter alpha 2-adrenoceptor sensitivity. This possibility was investigated by depleting NA with the neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP4). These studies showed that the apparent effect of chronic clorgyline on alpha 2-adrenoceptor sensitivity to clonidine was due to competition with increased levels of endogenous NA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Deficits in dopamine clearance and locomotion in hypoinsulinemic rats unmask novel modulation of dopamine transporters by amphetamine

Insulin affects brain reward pathways and there is converging evidence that this occurs through insulin regulation of the dopamine (DA) transporter (DAT). In rats made hypoinsulinemic by fasting, synaptosomal DA uptake is reduced. Interestingly, [3H]DA uptake is increased in hypoinsulinemic rats with a history of amphetamine self-administration. The possibility that amphetamine and insulin act in concert to regulate DAT activity prompted this study. Here we show that [3H]DA uptake, measured in vitro and clearance of exogenously applied DA in vivo, is significantly reduced in rats made hypoinsulinemic by a single injection of streptozotocin. Strikingly, amphetamine (1.78 mg/kg, given every other day for 8 days) restored DA clearance in streptozotocin-treated rats but was without effect on DA clearance in saline-treated rats. Basal locomotor activity of streptozotocin-treated rats was lower compared to control rats; however, in streptozotocin-treated rats, hyperlocomotion induced by amphetamine increased over successive amphetamine injections. In saline-treated rats the locomotor stimulant effect of amphetamine remained stable across the four amphetamine injections. These results provide exciting new evidence that actions of amphetamine on DA neurotransmission are insulin-dependent and further suggest that exposure to amphetamine may cause long-lasting changes in DAT function.