Infant Rat Brain Research Articles

The activity of deoxyribonucleic acid polymerase and deoxyribonucleic acid synthesis in nuclei from brain fractionated by zonal centrifugation.

1. The DNA polymerase (EC 2.7.7.7) activity in purified intact brain nuclei from infant rats was investigated. The effects of pH, Mg(2+), glycerol, sonication and storage of the nuclei under different conditions were examined and a suitable assay system was established. 2. The nuclei from infant brain cells were fractionated by zonal centrifugation in a discontinuous sucrose gradient into five zones: zone (I) contained neuronal nuclei (59%) and astrocytic nuclei (41%); zone (II) contained astrocytic nuclei (81%) and neuronal nuclei (19%); zone (III) contained astrocytic nuclei (82%) and oligodendrocytic nuclei (18%); zone (IV) contained oligodendrocytic nuclei (92%) and zone (V) contained oligodendrocytic nuclei (100%). 3. The content of DNA, RNA and protein for each fraction was measured. 4. The distribution of DNA polymerase activity in the fractionated infant and adult rat brain nuclei was determined. The highest activity was found in the neuronal nuclei from zone (I) and the following zones exhibited a progressive decline. In contrast with the nuclei from infant rats those from adults had a much higher activity and expressed a preference for native DNA as template. 5. The deoxyribonuclease activity in all classes of nuclei was measured with [(3)H]DNA as substrate. A general correspondence in the pattern of the relative activities in the nuclear fractions with the distribution of DNA polymerase was found. 6. The incorporation of [(3)H]thymidine into nuclear DNA in infant and adult rat brain was investigated. The specific radioactivity of the DNA in the 10-day-old rats was highest in zone (V) whereas in the nuclei of adult rats, which exhibited a comparatively low incorporation, the highest specific radioactivity was associated with zones (I) and (V).

The effect of cannabinoids on the number of nuclear membrane-attached ribosomes in infant rat brain

The pharmacologically active cannabinoids Δ 9-THC, Δ 9-THC and 11-OH-Δ 9, -THC each caused a highly significant reduction in nuclear membrane-attached ribosomes of infant rat brain cells. The pharmacologically inactive agents cannabinol and cannabidiol had only a slight effect while cannabigerol had a moderate effect. The activity was measured 3 hr after a 10 mg/kg s.c. dose. The duration of effect was tested with Δ 9-THC. It was significant after 1 hr, maximally developed after 3 hr, and almost completely diminished after 6 hr.

DNA synthesis in isolated nuclei from the brains of rats at different post-partal stages and the infant rat brain cytosol factor stimulating the DNA synthesis in infant rat brain nuclei

[ 3H]TTP incorporation into DNA of isolated nuclei from the brains of rats at different post-partal stages, as well as the effects of the adult and the infant rat brain cytosols upon the DNA synthesis in the isolated nuclear system, have been investigated. The nuclear system requires the four deoxyribonucleoside triphosphates, ATP and Mg 2+ for a maximum activity and DNA synthesis proceeds linearly within 30 min of incubation. A large portion of [ 3H]TTP incorporated into DNA is sensitive to actinomycin D inhibition and seems to be DNA polymerase dependent, while the remaining incorporation (approx. 20 % in the infant nuclear system and approx. 40 % in the adult nuclear system) is less sensitive to actinomycin D inhibition, proceeds without dATP, dCTP and dGTP and seems to be due to the terminal DNA nucleotidyltransferase activity. The nuclear activity of [ 3H]TTP incorporation into DNA increases after birth, reaches a maximum on the fifth day and then declines rapidly, reaching the adult level ( i. e. one half the maximum level) on the 22nd day after birth. [ 3H]TTP incorporation into DNA of isolated infant rat brain nuclei is stimulated specifically by the infant rat brain cytosol. The active principle in the infant rat brain cytosol seems to be thermostable and low molecular, and differs from the stimulatory factor present in the serum.

Studies on gangliosides of goldfish brain.

Abstract— Purified gangliosides from goldfish brain have been separated on thin layer chromatography. Incorporation of radioactivity into gangliosides from intracranially‐ injected [3H]galactose was not affected by concentrations of puromycin (170 μg intra‐ cranially) known to exert behavioural effects. Puromycin (500 μg intracranially) inhibited the incorporation of radiolabelled galactose into infant rat brain gangliosides, a result which confirms a previous report in the literature.

Reduction in number of nuclear membrane-attached ribosomes in infant rat brain following acute Δ 9-tetrahydrocannabinol administration

Reduction in number of nuclear membrane-attached ribosomes in infant rat brain following acute Δ 9-tetrahydrocannabinol administration

Inhibition of rat brain protein and nucleic acid synthesis by cannabinoids in vitro.

The effect of Δ9-tetrahydrocannabinol (THC), cannabidiol, and cannabigerol on some metabolic processes in infant and adult rat brain cortex slices was studied in vitro. With L-leucine-U-14C as the tracer substrate, the incorporation of radioactivity into the protein and nucleic acid fractions was significantly inhibited by THC. The oxygen consumption of the slices, the uptake of L-leucine into the slices, and the evolution of 14CO2 were, however, unaffected by THC. Cannabidiol was comparable in activity to THC but cannabigerol was less active. The pattern of inhibition by THC was also observed when the rat brain cortex slices were stimulated by 100 mM K+ or 10 μM protoveratrine. THC also brought about a significant decrease in the incorporation of uridine-2-14C into the nucleic acid fraction of infant and adult rat brain cortex slices. There was a decreased formation of uridine nucleotides in the presence of THC and an increase in uridine and uracil in the low molecular weight fraction. Experiments with THC-2,4-14C established that there was rapid uptake and a concentration of radioactivity in the incubated brain tissue.

The activity of D- -hydroxybutyrate dehydrogenase in fetal, infant and adult rat brain and the influence of starvation.

The activity of D- -hydroxybutyrate dehydrogenase in fetal, infant and adult rat brain and the influence of starvation.

Undernutrition and cerebellar development

Undernutrition has previously been shown to affect the cerebellum to a greater extent than the remainder of the brain. This study showed that the vermis is the primary area of the cerebellum affected. In contrast to the deficits in the cerebellar hemispheres, the reductions in the vermis wet weight, DNA, and protein content in brains of infant rats were persistent in rehabilitated adult animals. In 20-day-old malnourished rats, microscopic studies of the vermis showed increased density of Purkinje cells and internal granule cells and a disproportionately greater reduction in the size of the molecular layer compared to the internal granule cell layer. In rehabilitated adult rats, the increased density of Purkinje cells remained, but the size of the molecular layer compared to the internal granule cell layer was not different.

Effect of Xylocaine on infant rat brain mitochondria

Local anesthetics are widely used in obstetric practice to facilitate painless delivery. Their usage has been associated with neonatal respiratory depression and, in the case of fetal injection, with neonatal seizures. Because of these observations the effect of Xylocaine on infant rat brain mitochondrial enzymes was investigated. The neurochemlcal development of 5- to 10-day-old infant rats approximates that of the human newborn. Xylocaine was administered intraperitoneally to infant rats at this age in doses calculated to produce blood levels comparable to those in human newborns with toxic signs and symptoms. Two hours after injection there was a 65% decrease in brain D(—)-beta-hydroxybutyrate dehydrogenase (HBDH) activity and a 40% decrease in succinic dehydrogenase (SDH) activity. Brain cytochrome oxidase (CyO) activity was not significantly decreased. Temporal examination of these enzymes after injection suggested that HBDH was the first enzyme inhibited. These observations are in contrast to the lack of inhibition of liver HBDH, SDH, and CyO. The inhibition of oxidative enzymes in infant rat brain mitochondria persists up to 5 hours after injection. This enzyme inhibition is also demonstrable in measurements of respiration of intact brain mitochondria. Oxygen consumption of normal infant rat brain mitochondria is best supported by D(—)-beta-hydroxybutyrate (BOH), but succinate was also effective. BOH dependent respiration of infant rat brain mitochondria was inhibited by Xylocain in vivo and in vitro. The data suggest that further studies of Xylocaine usage and effects in the perinatal period are indicated.

DNA nucleotidyltransferase activity of the developing rat brain

DNA polymerase (deoxynucleoside triphosphate: DNA deoxynucleotidyltransferase, EC 2.7.7.7) activity in soluble extracts from developing rat brain has been studied. The DNA polymerase from infant rat brain required the presence of template DNA, Mg 2+ and all four deoxynucleoside triphosphates. The activity was enhanced by the addition of dithiothreitol. In developing rat brain the DNA polymerase activity of the cerebellum was, depending upon age, 20–50 times more than that of the cortex. The activity in the cerebellum peaked at around 6 days of age and then dropped during maturation. The activity in the cortex was highest immediately after birth and went down gradually. The activity was higher with heat-denatured DNA as a template than with native DNA except at the older ages where the activity seemed to be higher with native DNA. Spermidine at a concentration of 1 mM stimulated the activity of DNA polymerase but at a concentration of 3 mM or higher it inhibited the activity.

Acetoacetate metabolism in infant and adult rat brain in vitro.

1. Acetoacetate or dl-beta-hydroxybutyrate increases the rate of oxygen consumption to a smaller extent than that brought about by glucose or pyruvate in adult rat brain-cortex slices but to the same extent as that in infant rat brain-cortex slices. 2. The rate of (14)CO(2) evolution from [1-(14)C]glucose considerably exceeds that from [6-(14)C]glucose in respiring infant rat brain-cortex slices, in contrast with adult brain-cortex slices, suggesting that the hexose monophosphate shunt operates at a greater rate in the infant rat brain than in the adult rat brain. 3. The rate of (14)CO(2) evolution from [3-(14)C]acetoacetate or dl-beta-hydroxy[3-(14)C]butyrate, in the absence of glucose, is the same in infant rat brain slices as in adult rat brain slices. It exceeds that from [2-(14)C]glucose in infant rat brain but is less than that from [2-(14)C]glucose in adult rat brain. 4. Acetoacetate is oxidized in the brain through the operation of the citric acid cycle, as shown by the accelerating effect of glucose on acetoacetate oxidation in adult brain slices, by the inhibitory effects of malonate in both infant and adult brain slices and by its conversion into glutamate and related amino acids in both tissues. 5. Acetoacetate does not affect glucose utilization in adult or infant brain slices. It inhibits the rate of (14)CO(2) formation from [2-(14)C]glucose or [U-(14)C]-glucose the effect not being wholly due to isotopic dilution. 6. Acetoacetate inhibits non-competitively the oxidation of [1-(14)C]pyruvate, the effect being attributed to competition between acetyl-CoA and CoA for the pyruvate-oxidation system. 7. Acetoacetate increases the rate of aerobic formation of lactate from glucose with both adult and infant rat brain slices. 8. The presence of 0.1mm-2,4-dinitrophenol diminishes but does not abolish the rate of (14)CO(2) formation from [3-(14)C]acetoacetate in rat brain slices. This points to the participation of ATP in the process of oxidation of acetoacetate in infant or adult rat brain. 9. The presence of 5mm-d-glutamate inhibits the rate of (14)CO(2) formation from [3-(14)C]acetoacetate, in the presence or absence of glucose. 10. Labelled amino acids are formed from [3-(14)C]acetoacetate in both adult and infant rat brain-cortex slices, but the amounts are smaller than those found with [2-(14)C]glucose in adult rat brain and greater than those found with [2-(14)C]glucose in infant rat brain. 11. Acetoacetate is not as effective as glucose as a precursor of acetylcholine in adult rat brain but is as effective as glucose in infant rat brain slices. 12. Acetoacetate or beta-hydroxybutyrate is a more potent source of acetyl-CoA than is glucose in infant rat brain slices but is less so in adult rat brain slices.

Effect of excess phenylalanine and of other amino acids on brain development in the infant rat.

Extract: The effects of excess phenylalanine and of a number of other amino acids on brain weight, total brain lipid, and formation of the myelin lipid cerebroside sulfate (sulfatide) have been studied in the newborn rat. Brain weight, amount of brain lipids, and the formation in vivo of sulfatide were reduced as a result of injections of phenylalanine or of certain other amino acids. Total brain cholisterol, cerebroside, and sulfatide were all significantly reduced in phenylalanine-injected animals, although sulfatide was the only lipid that was reduced per gram wet weight of brain. The in vitro activity of the brain enzyme, galactolipid sulfotransferase, which catalyzes the formation of sulfatide from cerebroside and adenosine 3′-phosphate 5′-phosphosulfate (PAPS), was significantly reduced only when exogenous PAPS was not added to the assay medium. This finding suggests that in the brains of animals injected with phenylalanine there was impairement in the formation of PAPS from ATP and sulfate. In both cerebrums and cerebellums of animals injected with phenylalanine for the first 18 days of life, the amount of brain DNA and protein was reduced. Speculation: Increased ambient tissue levels of a number of amino acids, including phenylalanine, may interfere with normal development of the infant rat brain.

Ribonucleic acid biosynthesis in adult and infant rat brain in vitro.

The rate of biosynthesis of ribonucleic acid (as judged by the rate of incorporation of uridine into ribonucleic acid) in infant and adullt rat brain cortex slices, incubated (aerobically in tile presenice of various substrates, is directly proportionial to the adenosine triphosphate concentration. This suggests that the adenosine triphosphate concentration is one of the factors involved in the control of ribonucleic acid biosynthesis in infant and adult rat brain. Acetoacetate or beta-hydroxybutyrate is about 70 percent as effective as glucose, with both infant and adult brain, for the promotion of ribonucleic acid biosynthesis. but they are considerably mnore effective than succinate in infant brain than in adult brain.

DNA synthesis in the developing rat brain.

The in vitro synthesis of DNA as measured by the incorporation of thymidine-2-14C into DNA has been studied for various regions of the infant rat brain. Both intact cerebellum and cell-free extracts of cerebellum from newborn and infant rat brain showed a very rapid rate of DNA synthesis which was highest around 6 days after birth and decreased rapidly thereafter up to 18 days. This DNA synthesis in developing rat brain was strongly inhibited by hydroxyurea but was much less sensitive than was RNA synthesis to inhibition by actinomycin D.

Differential radiosensitivity of stationary and migratory primitive cells in the brains of infant rats

The heads of infant rats were irradiated with 200 r X-ray. In one group the animals were injected with thymidine- 3H immediately after irradiation, in the other, 24 hours before irradiation. All were killed at 3 days of age, 6 hours after irradiation and 6 or 30 hours after injection. In autoradiograms from the brains of unirradiated animals killed 6 hours after injection, the proportion of labeled cells, reflecting regional rate of cell poliferation, was determined at several brain sites. The relative reduction in the rate of cell proliferation in irradiated animals was also established. In matched stained sections the proportion of pyknotic cells, reflecting rate of cell death, was determined in the same brain regions. Finally, pyknosis index/labeling index ratios were calculated to relate radiation-induced cell death to rate of cell proliferation at different loci. It was established that migrating cells and prospective migratory cells, whether mitotic or postmitotic, are extremely radiosensitive, with a large proportion of them killed by exposure to 200 r. Relatively few of the stationary proliferating cells were killed with this dose. Stationary differentiating cells, and mature cells were not visibly affected. These results suggest that damage to the cellular mechanisms that enable primitive cells to locomote, rather than damage to DNA, is the principal cause of cell death in the nervous system exposed to low-level radiation. Moreover, the selective effect of this procedure could provide a technique for the identification of migratory cells at different brain sites during different stages of development.

Deoxyribonucleases from rat brain.

Abstract—Two distinctly different DNases were isolated from rat brain and could be separated easily by ammonium sulphate fractionation. One of the DNases acts optimally at pH 5.0 hydrolysing preferentially native DNA and requiring an optimal Mg2+ concentration of about 0.03 m. The other DNase has its optimal pH between 7.4 and 8.9, acts preferentially on heat‐denatured DNA and requires a lower Mg2+ concentration, the optimum being 1 × 10−4m.Cerebellum from adult rat brain has a lower acid DNase activity and higher alkaline DNase activity, and therefore has a higher ratio of alkaline DNase to acid DNase than the other areas of brain studied. This unique activity ratio in cerebellum of adult rat brain was not observed in infant rat brain.

Catecholamine accumulation in the brains of infant and adult rats after monoamine oxidase inhibition

Measurement of brain catecholamines after monoamine oxidase inhibition shows a faster and greater increase in the adult rat than the in the infant.

RECENT PROGRESS IN RESEARCH ON THE NEUROPHYSIOLOGIC BASIS OF MORPHINE ADDICTION

RECENT PROGRESS IN RESEARCH ON THE NEUROPHYSIOLOGIC BASIS OF MORPHINE ADDICTION