Showing papers in "Journal of Neuroscience Research in 1980"

A sensitive radioenzymatic assay for catechol drugs.

Abstract: This assay measures picogram quantities of catechol drugs and endogenous catecholamines in body tissues and fluids. The catechols are converted to their 3H-O-methyl metabolites during incubation with 3H-S-adenosylmethionine then separated by solvent extraction and thin-layer chromatography. Most drugs containing the catechol structure can be radiolabeled and separated from norepinephrine and epinephrine by this technique to provide simultaneous measurement of endogenous and exogenously administered catechols. The disposition of isoproterenol in tissues and fluids of man and experimental animals is measured to illustrate the utility of this assay. The reactivity of several commonly administered catechol drugs with COMT is described and the possible implications discussed.

Starvation-induced changes in transport of ketone bodies across the blood-brain barrier.

Abstract: The permeability of the blood brain barrier (BBB) to beta-hydroxybutyrate (beta-HB) was computed in fed and starved (five days) rats by the simultaneous measurement of cerebral blood flow (diffusible indicator method-123I-iodoantipyrine) and brain uptake of 14C-beta-HB (relative to a 3H2O reference). The results from the present study demonstrate that the movement of beta-HB across the BBB in rat is by a carrier-mediated process. During starvation, total movement (carrier-mediated and diffusionary) of this ketone body into brain was observed to be enhanced because of an increase in the diffusionary loss across the cerebral capillary. The calculated transport kinetics also suggest that the beta-HB molecule has a greater affinity for the transport (mediator) protein during starvation, although the maximal rate of uptake by brain due to a carrier processes mediated Vmax is decreased either because there is a smaller quantity of the mediating molecule or because there is trans inhibition by a high cellular concentration of beta-HB or some analog.

Induction of pinocytosis in cerebral vessels by acute hypertension and by hyperosmolar solutions.

Abstract: A new approach is described to study the mechanism of protein leakage through the endothelial cells in acute hypertension and after intracarotid infusion of hyperosmolar solutions. The luminarl surface of the cerebrovascular endothelial cells was labeled with choleragenoid-peroxidase before the blood-brain barrier (BBB) dysfunction was induced. Numerous pinocytotic vesicles and some transendothelial channels were formed and showed labeling of parts of their membranes. Our results indicate that the mechanism behind the protein leakage induced by hypertension and by hyperosmolar solutions is the same--ie, transendothelial pinocytosis.

The interrelation between brain PO2 and NADH oxidation-reduction state in the gerbil.

Abstract: The present investigation was aimed at studying the interrelation between hemodynamic, metabolic and oxygen tension in the brain of the gerbil exposed to various physiological and pathological conditions in the awake and anesthetized states. The hemodynamic and metabolic activities were evaluated by the DC fluorometer/reflectometer and were correlated to the pO2 values obtained by a surface electrode. When the awake brain was exposed to spreading depression (SD), a typical oxidation cycle of the NADH was recorded concomitant with a decrease cycle of the pO2. Under anesthetic effect the same dip in pO2 response was found but the NADH showed a "reduction cycle." The pO2 values were in a very good correlation with the changes in the reflectance trace, namely, the pO2 was a good indicator of the vasoconstriction-vasodilatation responses under various conditions. The same model was used under hypoxic and ischemic conditions, as well as under the effects of anesthetics.

Respiration in rat cerebral astrocytes from primary culture.

Abstract: Respiration was measured polarographically in astrocytes from dissociated neonatal rat cerebra grown in primary culture. Cells grown in a modified Eagle's minimal essential medium containing 20% fetal calf serum for 14 to 21 days respired at a rate of 34.23 n at. equiv 0 mg protein−1 min−1. Cells which were grown in this medium for 14 days and then grown in serum-free medium containing 0.25 mM dibutyrylcyclic-AMP (dbcAMP) for three to six days, had respiratory rates 20% higher than in cells from untreated control cultures (P = 0.005). Maximal inhibition by oligomycin required 200 p mole per mg protein for dbcAMP-treated cells and 80 pmole per mg protein for untreated cells. Oligomycin inhibited the respiration of both dbcAMP-treated and untreated cells by about 70%. At a concentration of 175 nmole per mg protein, 2,4-dinitrophenol (DNP) produced a maximal stimulation of respiration, which was 191% of the resting rate in dbc AMP-treated cells and 218% of the resting rate in untreated cells. The maximal DNP-stimulated respiratory rates were the same in dbcAMP-treated and untreated cells.

Chemical requirements of vasopressins for barrel rotation convulsions and reversal by oxytocin.

Abstract: When vasopressin is administered into the lateral ventricles of rats it produces severe convulsive activity characterized by a rapid barrel rotation. Electrical recordings from the dorsal hippocampus indicate marked elevations in the amplitude and frequency at doses of 5 microliter of 2 x 10(-5) M vasopressin. No significant behavioral effects were noted with oxytocin, somatostatin, beta-melanophore-stimulating hormone, adrenocorticotropin, or leu-enkephalin. Pretreatment of the rats with intraventricularly administered oxytocin, beta-MSH, or systemically administered Dilantin prevented the vasopressin-induced seizures. With the use of chemical and enzymic modification procedures, the essential fragment and amino acids of vasopressin needed for the activity were determined. It was concluded that although the peptide could be acting by vasoconstricting blood arterioles and capillaries in the brain, it may also be exerting a direct excitatory action on neurons.

Simultaneous fluorescent retrograde axonal tracing and immunofluorescent characterization of neurons.

Abstract: A simple method is described for simultaneously double labeling single neurons with fluorescent retrograde axonal tracers and immunofluorescence using an antibody to serotonin. Caudata-putamen injection of orange fluorescent propidium iodide, in particular, proved effective in retrogradely labeling dorsal raphe cell bodies. These same perikarya could be stained green with the indirect immunofluorescent method, using green fluorescent fluorescein isothiocynate (FITC) as the second antibody.

Ultrastructural differentiation and synaptogenesis in aggregating rotation cultures of rat cerebral cells

Abstract: Cerebral cortices from fetal rats were dissociated into single cells by either trypsinization or mechanical sieving. Then the cells were allowed to form aggregates in rotation cultures. At 7, 14, 21, 28, and 35 days, aggregates were processed for electron microscopic study of morphological differentiation with special emphasis on synaptogenesis. Whether the tissue was initially dissociated by trypsin treatment or by mechanical sieving, the aggregating cultures did not exhibit any apparent differences in ultrastructural differentiation and synaptogenesis. At day 7, most neurons were immature and the extracellular space was large. Cell processes had not yet branched extensively but did contain numerous microtubules. A few immature synapses were observed. Astrocytes and oligodendrocytes displayed many of their typical cytological features. At day 14, dendritic spines had developed, some of which formed axodendritic spinous synapses. Multilayered myelin sheaths were tightly wrapped around axons. At day 21, synapses appeared mature and their number per unit area was maximal. The extracellular space had greatly decreased. At this age, asymmetrical synapses had increased approximately sixfold, whereas symmetrical synapses increased only fourfold when compared with the 7-day-old aggregates. Multifocal degeneration became apparent at 28 days and was accompanied by a significant decline in the number of synapses.

Choline acetyltransferase: purification and characterization.

Abstract: Choline acetyltransferase catalyzes the synthesis of the neurotransmitter acetylcholine and is the specific enzyme marker for cholinergic neurons. This review summarizes and discusses different purification procedures, as well as the characteristics of this important enzyme. Explanations for conflicting or controversial results concerning this enzyme are also discussed.

Immunohistochemical localization of the myelin basic protein and of the glial fibrillary acidic protein: comparative study in normal, quaking and jimpy mice.

Abstract: The topographical distribution of myelin basic protein (MBP) and of glial fibrillary acidic protein (GFA) have been compared in the CNS of 18-day-old normal, quaking, and jimpy mice using indirect immunofluorescence. Comparison was made in brain and spinal cord sections. MBP was strongly decreased in quaking and jimpy mice in all parts of the CNS. A pronounced gliosis was observed in jimpy mice with occurrence of numerous hypertrophied astrocytes. To a lesser extent, an astrocytic reaction was also observed in quaking CNS.

GABA uptake in astrocytes in primary cultures: coupling with two sodium ions.

Abstract: The influence of sodium ions on GABA uptake into astrocytes in primary cultures has been investigated performing kinetic analysis of GABA uptake at different sodium concentrations in the range 16 to 151 mM. These investigations reveal that sodium affects both the Km and the Vmax of the saturable component of the astroglial GABA uptake. Uptake rates as a function of the sodium concentration at high GABA concentrations (greater than or equal to 50 microM) were clearly sigmoid whereas at lower GABA concentrations this sigmoid shape was not obvious. Accordingly, Hill plots of the sodium dependency at high GABA concentrations exhibited straight lines with slopes of 2.0 to 2.5, suggesting that the coupling ratio between sodium and GABA is at least two. Corresponding Hill plots at lower GABA concentrations exhibited slopes of 1.6 to 1.8. Moreover, plots of 1/v versus 1/Na2 gave better fits to straight lines than plots of 1/v versus 1/Na which were curvilinear upward. Again, this curvilinearity was more pronounced at high GABA concentrations that at low GABA concentrations. From these results it is concluded that GABA uptake into astrocytes in primary cultures requires the binding of at least two sodium ions per GABA molecule transported.

Accumulation of GFA, the monomeric precursor of the gliofilaments, during development in normal mice and dysmyelinating mutants.

Abstract: Astrocytic reactivity during the myelination period in the mouse was studied with immunochemical method, ie, quantitative determination of the soluble pool of the GFA protein. There is normally a maximum content at the time of early myelinogenesis in any structure; then the GFA level decreases and finally keeps constant for a long period during the adult life. This evolutive pattern is also observed in the dysmyelinating mutant quaking, with a permanent shift toward higher values especially in areas of earlier maturation. In the jimpy mutant, practically devoid of myelin, the increase of GFA occurs but does not stop until death, at 25 days postnatal. This study points out (1) the capacity of astrocytes to synthesize surprisingly high amounts of soluble GFA at periods of intense metabolic activity, and (2) the reactivity of astrocytes in relation to the degree of deficiency of the myelinating oligodendrocytes.

Spatial distribution of catecholaminergic cells in the fish retina.

Abstract: The cell density, distribution pattern, and morphology of catecholaminergic (CA) cells have been studied by fluorescence microscopy of retinal flatmounted preparations from various species of fresh water, estuary, and marine fishes, and the toad. In all fish retinas examined the CA-cell density (cells/mm2) is lowest in the central region surrounding the optic disc, slightly higher in the intermediate region, and highest in the periphery. The size of CA-cells is smaller the higher their density. Following administration of L-Dopa, dompamine, or noradrenaline, the density of CA-cells approximately doubled, due to the appearance of small fluorescent cells. CA-cells are arranged in rows along radial lines which fan out from the optic disc. In large cells of the central and intermediate regions three to five processes arise from the soma and extend and ramify irregularly in the inner plexiform layer, while in small cells from the intermediate and peripheral regions two processes arise from opposite poles and extend regularly in a direction perpendicular to the rows of cells and parallel to the retinal margin. In the retina of the marine fish Holocentrus sp CA-cells are fewer in number compared to other fish studied and their processes extend without any regular pattern. In the toad their size and density are homogenous throughout the retina, and their processes show a regular arrangement.

Effects of morphine on sensory‐evoked responses recorded from central gray, reticular formation, thalamus, hypothalamus, limbic system, basal ganglia, dorsal raphe, locus ceruleus, and pineal body

Abstract: Field potential recordings of acoustic and photic-evoked responses were obtained from 15 brain sites of freely behaving unanesthetized rats previously implanted stereotaxically with permanent electrodes. Several dosages of morphine (1, 5, 10, 30, and 50 mg/kg) were examined. The activities recorded from all the structures in this study, except the cochlear nucleus (CoN), were affected by morphine. Different sensitivities to morphine threshold were observed between structures, and several structures exhibited dose-related patterns (ventromedial hypothalamus (VMH), caudate nucleus (CN), central gray (CG), hippocampus (Hipp), and lateral septum (Spt)). Several brain sites, after the initial dose of morphine, did not recruit more responses to subsequent doses of the drug, ie, exhibited all-or-none responses (pineal body (PB), medial thalamus (MTh), anterior hypothalamus (AH), mesencephalic reticular formation (MRF), and the dorsal raphe (DR)). In some structures, morphine induced increases in the response amplitudes, while in other sites decreases in response amplitudes were elicited. Biphasic responses, ie, increases in response amplitude after low doses of the drug and decreases in response amplitude after higher dosages, were also observed (VMH, CN, DR, CG, and MRF). The acoustic-evoked responses were affected by morphine more than the photic responses. The present observations indicated that (1) morphine exerts effects in many parts of the central nervous system (CNS); (2) some structures are more sensitive to morphine than others; (3) only a few structures exhibit dose-related patterns and, thus, may represent sites of direct morphine action; (4) some structures exhibit all-or-none responses; and (5) morphine depressed activity in some structures and increased activity in others, ie, morphine elicited different effects in different structures.

Glycerol oxidation in rat brain: subcellular localization and kinetic characteristics.

Abstract: The oxidation of [1,3-14C] glycerol to 14CO was measured in slices, whole homogenates, and subcellular fractions of rat brain. In all of these tissue preparations, the Lineweaver-Burk plots of glycerol oxidation were biphasic, yielding two apparent Km and V values. Similar kinetic characteristics were obtained with brain homogenates from guinea pig, mouse, rabbit, monkey, and pig. In other tissues of the rat, including heart, kidney, liver, and skeletal muscle, the Lineweaver-Burk plots for glycerol oxidation were not biphasic but were linear. Heating the brain homogenates for five minutes at 5 degrees C caused a 50% decrease in the rate of oxidation of glycerol without a change in the biphasic double reciprocal plot. The addition of purified glycerol kinase (EC 2.7.1.30) to the homogenate caused an increase in the rate of oxidation and resulted in linear Lineweaver-Burke plot. Brain mitochondria were prepared by two different methods, both of which yielded an enrichment of glycerol oxidation. In contrast, the rate of glucose oxidation was higher in homogenates than in mitochondria, and glucose competed with glycerol as substrate only extramitochondrially. The effects of various metabolic inhibitors suggested the participation of intact, coupled mitochondria, of glycolytic enzymes, and of electron transport in the oxidation of glycerol. The data support the primary localization of glycerol oxidation in nonsynaptic mitochondria in brain and the presence in that organelle of enzymes of the Embden-Meyerhoff pathway or an as yet unidentified system for oxidizing this compound.

Neuron-specific enolase levels in primary cultures of neurons.

Abstract: Primary neuronal cell cultures, derived by centrifugal elutriation of cells dissociated from embryonic rat cerebra, have been analyzed for their content of neuron-specific enolase (NSE) and non-neuronal enolase (NNE). Both immunocytochemical staining and radioimmunoassay establish that NSE is present in these cells. Furthermore, the observed increase in NSE and decrease in the NNE/NSE ratio with time in culture parallels that observed in vivo. It has recently been shown that during in vivo neurogenesis and maturation a "switch over" occurs from NNE to NSE which correlates with neuronal differentiation. The presence of a neuron-specific protein in these primary neuronal cultures and the changes observed during growth further support their use as a model system for the study of nervous tissue differentiation.

Isolation of ribosome containing synaptosome subpopulation with active in vitro protein synthesis.

Abstract: Subpopulations of synaptosomes harvested from neonatal rat brain cortices revealed a differential ability to synthesize protein in vitro. Incubation of synaptosomes with radiolbeled leucine, followed by continuous sucrose gradient centrifugation produced an asymmetric shift in the radioactivity toward the higher density sucrose fractions. The bulk of the mitochondrial cytochrome c-oxidase activity was also found in these fractions, however, subfractionation studies of osmotically-lysed synaptosomes suggested that the newly-synthesized proteins reside in an osmotically sensitive, non-mitochondrial compartment. The ability of each subpopulation of synaptosomes to synthesize protein in vitro was assessed after their isolation from linear continuous sucrose gradients. There was an enrichment of highly active protein synthesizing particles in the “heavy” subpopulations of neonatal synaptosomes. The inhibitory effects of chloramphenicol and cycloheximide on the protein synthesis in these particles were similar to those of the original synaptosome fraction. Electron microscopic analysis revealed an increase in the numbers of ribosome-containing structures resembling dendritic dendritic and axonal growth cones.

Effect of methionine, glycine and serine on serine hydroxymethyltransferase activity in rat glioma and human neuroblastoma cells.

Abstract: Human neuroblastoma SK-N-SH-SY5Y (5Y) and rat glioma (C6) cells were cultured with supplemental methionine, glycine, or serine for three to six days. Serine hydroxmethyltransferase (SHMT: L-serine: tetrahydrofolate 5, 10-hydroxymethyltransferase, EC 2.12.1) was assayed radiometrically in whole cell homogenates, crude supernatant fractions and crude particulate fractions. No significant changes in specific activity or cellular morphology were noted at methionine, glycine, or serine concentrations up to 16 mM. Serine concentrations of 20 and 40 mM led to significantly lower gliomal enzyme specific activities. This activity was unevenly distributed between soluble and particulate fractions, with 190 and 398 nmoles of HCHO formed per mg of protein per hour, respectively. Growth stage and time of incubation were major determinants of enzyme specific activity. C6 cells' specific activity rose slowly with increasing time in culture until cellular confluence. At this time there was a pronounced elevation in specific activity, occurring more rapidly in cells grown in 1.2 mM methionine. Intracellular amino acid analysis of C6 cells demonstrated a significant rise in methionine after four days in media containing 0.2 mM methionine. No appreciable diminution in the intracellular levels of glycine or serine occurred following incubation in excess methionine. It is concluded that SHMT-specific activity in C6 and 5Y cells is not regulated by glycine, serine, or methionine levels and that high concentrations of these amino acids (> 30 mM) are not detrimental to these cells derived from the CNS.

Intraventricular ACTH and vasopressin cause regionally specific changes in cerebral deoxyglucose uptake.

Abstract: The cerebral uptake of subcutaneously injected [3H] 2-deoxyglucose (2DG) was used to determine regional changes of cerebral glucose uptake associated with peptide-induced behaviors in mice. Evidence is presented that the use of [3H] 2DG (s.c.) gives results qualitatively similar to those obtained using intravenous [14C] 2DG. Lysine vasopressin, 1 microgram intracerebroventricularly (i.c.v.) induced the characteristic hyperactivity previously described, and significantly decreased [3H] 2DG uptake in frontal cortex. ACTH1-24 (1 microgram i.c.v.) induced excessive grooming and concomitant decreases in [3H] 2DG uptake in the olfactory bulb, pyriform cortex, and amygdala, and an increase in cerebellum. These results are only partly consistent with previous results on the cerebral sites of action of ACTH and vasopressin. These patterns of 2DG uptake are distinct from those observed following footshock, indicating that the peptide hormones mediate the effects of footshock on [3H] 2DG uptake only partially if at all.

Sodium dependence of GABA transport in rat hypothalamic synaptosomes

Abstract: The sodium dependence of gamma-aminobutyric acid (GABA) uptake has been studied in rat hypothalamic synaptosomes, and the results compared to previous studies in cortical synaptosomes. Initial velocity of GABA uptake was measured as a function of both GABA and sodium concentration, and these data were fitted to the rate equation for each of several plausible models. The minimal best-fit model was found to be identical to that for cortical synaptosomes. However, the constants which quantitate the model were found to differ for hypothalamus and cortex. As a result, uptake at any combination of [Na] and [G] in hypothalamic synaptosomes is approximately double that in cortical synaptosomes. The rate equation for the minimal best-fit model was utilized to define and compute certain parameters which are useful in comparing transport mechanisms (Vmax, Va, Kt, Jm, and k Na). In all cases, differences were found between hypothalamus and cortex.

Some mechanisms underlying actions of ketamine on electromechanical coupling in skeletal muscle

Abstract: The effects on excitation contraction coupling (ECC) of ketamine (a dissociative general anesthetic) were investigated using the sartorius muscle of the frog. Extracellular studies revealed that ketamine depressed action potential production in a concentration-dependent manner. Ketamine decreased both the conduction velocity and the compound action potential while concomitantly increasing the threshold current. Intracellular studies showed that ketamine caused a slight non-significant decrease in the membrane potential and also decreased the threshold potential (mechanical threshold). Ketamine (1.5 X 10(-4) M and 3.0 X 10(-4) M) initially potentiated and then blocked the twitch response elicited by direct muscle stimulation. Both of these effects were statistically different from control values. These findings suggest that ketamine alters action potential production in frog skeletal muscle. This property of ketamine contributes in part to the disruption of ECC observed with this drug. The results suggest the ketamine probably interferes with calcium binding, its release and/or its fluxes which may contribute to the intial potentiation and subsequent depression of twitch tension.

Neurophysiological evidence for tolerance and dependence on opiates: simultaneous multiunit recordings from septum, thalamus, and caudate nucleus.

Abstract: Multiunit activity was recorded simultaneously from the septum (Spt), medial thalamus (PF-CM complex), and caudate nucleus (CN) in freely behaving rats previously implanted with permanent nichrone semimicroelectrodes (62 mu in diameter) A challenge dose of morphine (10 mg/kg) and its antagonist (naloxone 10 mg/kg) was examined in the same animals while they were morphine naive and after they had become morphine physically dependent Electrophysiological observations indicated that it would be possible to identify three physiological phenomena: 1) activity related to morphine dependency; 2) activity related to tolerance; and 3) activity related to withdrawal Experimental observations yielded three different electrophysiological patterns of activity related to tolerance Each of the three central sites (Spt, PF-CM, and CN) exhibited a different response pattern to morphine in morphine physically dependent animals

Ionic mechanism of inhibition of long duration in Aplysia synapse.

Abstract: In the abdominal ganglion of Aplysia kurodai, three neurons were identified that responded with inhibition of long duration (ILD) to a single stimulation of the siphon nerve and to bath and iontophoretic application of dopamine. The ILD is a long-lasting (15-60 sec) hyperpolarizing potential, typically associated with a conductance increase but with a reversal potential, typically negative than the potassium equilibrium potential. Exchange of half the external chloride with acetate induced no change in the ILD. The ILD was reduced in amplitude in the presence of 0.1 normal K+ in the external medium and enhanced in two times normal K+ solution. The ILD amplitude, but not conductance, was progressively impaired by cooling and by the metabolic inhibitors ouabain and strophanthidin. These results suggest that the ILD is composed of an increase in K++ conductance and activity of the electrogenic Na-pump. theophylline potentiated the ILD, especially its duration. Furthermore, bath application of dibutyryl cAMP augmented the amplitude of the synaptically elicited ILD and the dopamine responses by 30% over controls without any effect on the change in membrane conductance. Intracellularly injected cAMP also increased the ILD, but 5'-AMP showed no such effect. These data suggest that dopamine acts on the ILD neurons by increasing K+ conductance and the activity of the electrogenic Na-pump and that cAMP might selectively stimulate the pump component of the ILD.

The role of the medial septal nucleus in mediating adrenocortical responses to somatosensory stimulation.

Abstract: In view of the demonstrated involvement of the hippocampus in the mediation of adrenocortical responses following sciatic nerve stimulation, the role of the septum and the preoptic area in the transmission of this response was investigated. Changes in plasma corticosterone following ether stress and photic, acoustic, or sciatic nerve stimulation were studied in intact rats and in animals with lesions in the medial septal nucleus and the preoptic area. The response to ether stress and to photic and acoustic stimulation was normal in these animals. However, the adrenocortical response to sciatic nerve stimulation was partially reduced in the rats with lesions in the superior, but not in the inferior preoptic area, and it was completely blocked in those with medial septal lesions. Our previous and present experiments indicate that the ventral hippocampus, the fimbria, the septum, and anterior hypothalamic afferents mediate the adrenocortical response to somatosensory stimulation.

In vitro evidence of photoreception in the chick pineal gland and its interaction with the circadian clock controlling N-acetyltransferase (NAT).

Abstract: The response of the circadian rhythm in N-acetyltransferase (NAT) activity to phase-shifted light cycles was examined in vitro in explant cultures of chick (Gallus domesticus) pineal glands. Bisected portions of glands, obtained from birds housed in a light-dark cycle (LD 12:12), were explanted into culture and maintained under one of three light cycles (LD 12:12), the phase of which was either 1) similar to that of the birds' previous cycle, 2) seven hours phase-delayed, or 3) six hours phase-advanced. Following two to three days of exposure to the respective light cycles, cultures were placed into continuous darkness (DD). Sampling from cultures during exposure to DD revealed a circadian rhythm of NAT activity. In each case, the phase of the subsequent rhythm of enzyme activity in DD reflected that of the preceding in vitro light cycle. A distinct phase difference of approximately 180 degrees was observed between cultures exposed to opposite lighting regimes. These results indicate entrainment of the circadian rhythm of NAT activity to the prevailing in vitro light cycle and suggest a direct interaction between pineal photoreception and the circadian "clock" controlling NAT.

The uptake of 3-deoxy-3-fluoro-D-glucose by synaptosomes from rat brain cortex.

Abstract: D-[3-3H]-3-deoxy-3-fluoroglucose was synthesized chemically and shown to be transported into rat brain synaptosomes by a saturable process with a Km 6.2 x 10(-4) M and a Vmax 2.8 nmole x mg protein-1. After an initial, rapid period of transport, further uptake of the fluorosugar is restricted by the rate of its phosphorylation. Both D-glucose and cytochalasin B are competitive inhibitors of 3-deoxy-3-fluoro-D-glucose transport with Ki values of 93 micron and 6.0 x 10(-7) M, respectively. Phloretin, N-ethylmaleimide and p-chloromercuribenzoate also inhibit the fluorosugar uptake, whereas ouabain and changes in K+, Na+, Mg2+ and Ca2+ ions have only a small effect. The recorded 3-deoxy-3-fluoro-D-glucose influx is slightly reduced by potassium cyanide, antimycin A, 2,4-dinitrophenol, and rotenone. The uptake reduction caused by these four reagents is relieved by the addition of exogenous ATP. The possible influence of hexokinse activity on the uptake process is discussed.

Drug‐induced changes in catecholaminergic cells of the fish retina

Abstract: The changes in fluorescence intensity and number of visible catecholaminergic cells (CA-cells), as revealed by means of a histofluorescence technique, were used as indicators of the effects of various pharmacological agents upon CA-cells in the retina of fishes (Cyprinus carpio and Eugerres plumieri). The study includes in vivo and in vitro experiments. In the in vivo experiments, intravitreal injection, two or three hours before eye enucleation, of 10 microgram L-DOPA, dopamine, or noradrenaline accentuated CA-cell fluorescence and increased the number of visible cells, whereas 10 microgram of tyramine, octopamine, synephrine, or adrenaline reduced the endogenous fluorescence. Intramuscular injection of reserpine (3 mg/kg) abolished CA-cell fluorescence. In the in vitro experiments, pieces of isolated retinas were incubated for three or 30 minutes in media containing different drugs. Only minor changes in fluorescence were detected after three minutes of incubation, but after 30 minutes, dopamine (20 microM) markedly enhanced CA-cell fluorescence. Carbachol (20 mM), acetylcholine (10 mM) plus BW-anticholinesterase (1 mM) or substance P(1.6 x 10(-2) mM), all reduced CA-cell fluorescence. Kainic acid (20 mM) abolished fluorescence from CA-cell somata, while fluorescent fiber networks remain unchanged. L-aspartate (5 mM) and GABA (10 mM) in the incubation medium did not influence fluorescence intensity. The results are relevant to, and consistent with, electrophysiological observations of dopamine-mediated spatial effects on horizontal cell potentials.

Studies on AMP deaminase and 5'-nucleotidase in rat brain under different experimental conditions.

Abstract: Adenosine monophosphate (AMP) deaminase and 5'-nucleotidase, the two enzymes involved in the disposal of AMP, have been detected in different regions of normal rat brain and in animals subjected to heightened neuronal activity (leptazol-induced convulsions) and to depression of the central nervous system (CNS) by the administration of barbiturates. They have also been estimated in the CNS of animals subjected to anoxia or treated with lithium and ammonium salts. The AMP deaminase activity was found to be highest in cerebellum and lowest in cerebral cortex, while the 5'-nucleotidase activity was found to be highest in brain stem and lowest in cerebellum. The AMP deaminase activity was elevated in all the regions of brain during the preconvulsive and convulsive periods. The activity returned to normal during recovery. The activity of 5'-nucleotidase was found to be depressed in the preconvulsive and post-convulsive periods. The enzyme was also found to be depressed in all the three regions after the administration of barbiturates. Administration of lithium or ammonium salts of induction of anoxic states resulted in an increase in the activity of AMP deaminase in all the three regions of brain. These results are discussed in relation to the probable production of cyclic AMP and cyclic guanosine monophosphate (GMP) which may have depressive and excitatory roles, respectively, in brain. It appears that increased AMP deaminase activity is associated with increased neuronal activity while depression of 5'-nucleotidase activity is associated with conditions of decreased CNS excitability.

On some derivatives of ferrocene, novel acetylcholinesterase inhibitors.

Abstract: Eight novel methylcarbamates, derivates of ferrocenylketoximes and aldoximes, and ferrocenyl-p-hydroxyphenylaldimine were prepared. They are the first carbamates in the field of ferrocene derivatives. Their anticholinesterase activity was tested and found to fall in the range of 10(-4) through 10(-6) M (I50). The correlation between the activity of the compounds and the kind of substituents grafted on the methylene carbon atom is discussed.

Adrenal and hypothalamic dopamine-beta-hydroxylase activity in the hamster: evidence for species-distinctive day-night and post-pinealectomy differences in activity.

Abstract: Dopamine-beta-hydroxylase (DBH) activities were measured in the adrenal gland and hypothalamus of the golden hamster (Mesocricetus auratus) by means of a sensitive radioenzymatic method. Groups of unoperated control (C), sham pinealectomized (SPX), and pinealectomized (PX) adult males were sacrificed shortly before (09:00 h) and after (11:00 h) the onset of darkness of the daily photoperiod of LD 12:12 (lights on 22:00 h to 10:00 h). In neither adrenal gland nor the hypothalamus were there any day-night changes in the C groups. Pinealectomy led to no significant differences in hypothalamic DBH activity either in light or in dark. However, intracranial surgery resulted in increased DBH activity in the adrenal gland, evident in both light and dark. Although this increase was greatest in the PX groups, intracranial surgery in SPX groups also showed a trend of increase in adrenal DBH, evident even one month after surgery. Such an increase, however, was not found in hypothalamic DBH in the same SPX groups. In both the adrenal gland and the hypothalamus, copper-sensitive endogenous inhibitors of DBH did not show any notable change in concentration, either in relation to the daily light-to-dark change or as a consequence of intracranial surgery. These and related results suggest that important interspecies variations exist in DBH activities in relation to diurnal changes, and also in control or modulation by pineal and other factors.