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Biomedical subjects

A Giuditta

Publications and source records attributed to A Giuditta.

At least 55 records · Page 3Linked to original sources
PubMed1993

Neurofilament proteins are synthesized in nerve endings from squid brain.

It is generally believed that the proteins of the nerve endings are synthesized on perikaryal polysomes and are eventually delivered to the presynaptic domain by axoplasmic flow. At variance with this view, we have reported previously that a synaptosomal fraction from squid brain actively synthesizes proteins whose electrophoretic profile differs substantially from that of the proteins made in nerve cell bodies, axons, or glial cells, i.e., by the possible contaminants of the synaptosomal fraction. Using western analyses and immunoabsorption methods, we report now that (a) the translation products of the squid synaptosomal fraction include neurofilament (NF) proteins and (b) the electrophoretic pattern of the synaptosomal newly synthesized NF proteins is drastically different from that of the NF proteins synthesized by nerve cell bodies. The latter results exclude the possibility that NF proteins synthesized by the synaptosomal fraction originate in fragments of nerve cell bodies possibly contaminating the synaptosomal fraction. They rather indicate that in squid brain, nerve terminals synthesize NF proteins.

Animals↗
PubMed1992

The sequential hypothesis of sleep function. III. The structure of postacquisition sleep in learning and nonlearning rats.

EEG methods were used to examine the structure of postacquisition sleep in learning (L) and nonlearning (NL) rats previously exposed to a session of two-way active avoidance training, and in control rats (C) left in their home cages. In agreement with literature data, the number and total amount of paradoxical sleep (PS) episodes were higher in L rats than in NL rats. In addition, significant differences between L and NL rats concerned the episodes of synchronized sleep followed by wakefulness or by PS (SS-W and SS-PS, respectively). The average duration and related parameters of SS-W episodes, and the average duration, number, amount and related parameters of SS-PS episodes increased in NL and L rats in comparison with C rats. Longer SS-W episodes occurred early in NL and L rats, but the effect lasted longer in NL rats. On the other hand, the increments concerning SS-PS episodes occurred earlier, were more pronounced and laster longer in L rats. The results support a role of SS in brain information processing, as envisaged by the sequential hypothesis on the role of sleep. They suggest, furthermore, that memory traces lacking adaptive value may be destabilized and cleared away during SS-W and SS-PS episodes, while the remaining memory traces may be retained and eventually stored again in more integrated form during SS-PS and PS episodes, respectively.

Animals↗
PubMed1992

The sequential hypothesis of sleep function. IV. A correlative analysis of sleep variables in learning and nonlearning rats.

Female adult rats were trained for a two-way active avoidance task (4 h), and allowed free sleep (3 h). Control rats (C) were left in their home cages during the acquisition period. Dural electrodes and an intraventricular cannula, implanted one week in advance, were used for EEG recording during the period of sleep and for the injection of [3H]thymidine at the beginning of the training session, respectively. Rats were killed at the end of the sleep period, and the DNA-specific activity was determined in the main brain regions and in liver. Correlations among sleep, behavioral and biochemical variables were assessed using Spearman's nonparametric method. In learning rats (L), the number of avoidances was negatively correlated with SS-W variables, and positively correlated with SS-PS variables (episodes of synchronized sleep followed by wakefulness or paradoxical sleep, respectively) and with PS variables. An inverse pattern of correlations was shown by the number of escapes or freezings. No correlations occurred in rats unable to achieve the learning criterion (NL). In L rats, the specific activity of brain DNA was negatively correlated with SS-W variables and positively correlated with SS-PS variables, while essentially no correlation concerned PS variables. On the other hand, in NL rats, comparable correlations were positive with SS-W variables and negative with SS-PS and PS variables. Few and weak correlations occurred in C rats. The data support a role of SS in brain information processing, as postulated by the sequential hypothesis on the function of sleep. In addition, they suggest that the elimination of nonadaptive memory traces may require several SS-W episodes and a terminal SS-PS episode. During PS episodes, adaptive memory traces cleared of nonadaptive components may be copied in more suitable brain sites.

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PubMed1991

Distributed changes in rat brain DNA synthesis with long-term habituation and potentiation of the perforant path-granule cell synapse.

The involvement of brain deoxyribonucleic acid (DNA) synthesis in adaptive neural events was studied in the adult rat during long-term habituation (LTH) or potentiation (LTP) of the perforant path-granule cell synapse. Male Long-Evans rats were given 50 muCi [3H]thymidine intraventricularly under urethane anesthesia. Soon thereafter, field excitatory postsynaptic potential (EPSP) slope and population spike were monitored from the right dentate gyrus before and at various times (5, 10, 15, 60 min) following the delivery to the ipsilateral perforant bundle of a low frequency (LFS: 1.0 Hz, 160 s) or a high-frequency train (HFS: 400 Hz, 200 ms), repeated once after 5 min. Unstimulated implanted rats served as controls. DNA synthesis was evaluated by the incorporation of the radioactive precursor into DNA of several brain areas at the end of a 1 h incorporation period. In CA1, LTH and LTP increased DNA synthesis by 30% on the stimulated side. In the entorhinal cortex, LTH but not LTP increased DNA synthesis (by 30%) on the stimulated side. Conversely, in the frontal cortex, LTP but not LTH increased DNA synthesis (by 100%) on both sides. Long-lasting changes in synaptic efficacy covaried non-linearly with DNA synthesis in mono- and polysynaptically stimulated hippocampal regions, and in functionally associated neocortical areas. The co-variations of population spike amplitude were positive for LTH and negative for LTP in the dentate gyrus and frontal cortex of both sides, and in CA3/CA1 of the stimulated side, indicating higher DNA synthesis at lower values of LTH and LTP, and viceversa. Further, regional cross-correlation analyses revealed a high degree of synchronization among brain sites, following low- or high-frequency train pulses, indicating that (i) extra-target sites participate on the stimulated and on the contralateral side, and (ii) small distributed changes take place across the sampled neural networks. A modulatory role of information flow on brain DNA synthesis is inferred to take place in a diffuse, distributed manner.

Animals↗
PubMed1991

Active polysomes in the axoplasm of the squid giant axon.

Axons and axon terminals are widely believed to lack the capacity to synthesize proteins, relying instead on the delivery of proteins made in the perikaryon. In agreement with this view, axoplasmic proteins synthesized by the isolated giant axon of the squid are believed to derive entirely from periaxonal glial cells. However, squid axoplasm is known to contain the requisite components of an extra-mitochondrial protein synthetic system, including protein factors, tRNAs, rRNAs, and a heterogeneous family of mRNAs. Hence, the giant axon could, in principle, maintain an endogenous protein synthetic capacity. Here, we report that the squid giant axon also contains active polysomes and mRNA, which hybridizes to a riboprobe encoding murine neurofilament protein. Taken together, these findings provide direct evidence that proteins (including the putative neuron-specific neurofilament protein) are also synthesized de novo in the axonal compartment.

Animals↗
PubMed1990

Environmental stimuli modulate the circadian rhythm of [3H-methyl]thymidine incorporation into brain DNA of male rats.

A circadian rhythm of DNA synthesis is present in most organs of adult mammals, with peak levels (acrophase) generally in the rest period. We have recently reported that in the rat brain the acrophase of the rhythm of thymidine incorporation into DNA occurs on the contrary during the active period. To determine whether the brain waking activity was exerting a modulatory action we measured the circadian rhythm of incorporation in the brain and kidney of young adult male rats housed in conditions of sensory and social enrichment or impoverishment for four days. Biochemical and autoradiographic data show that the brain rhythm persists in the enriched condition, but is abolished in the impoverished condition. On the other hand, the rhythm of incorporation in the kidney is maintained in both conditions. These results suggest that the permanence of the brain oscillation is selectively dependent on the complexity of the sensory and social stimulation.

Animals↗
PubMed1990

Visualization of differential neurofilament phosphorylation in the pre- and postsynaptic axoplasm of the squid giant synapse: an electron spectroscopic study.

When inelastically scattered electrons with an energy loss specific for interaction with phosphorus atoms were used for visualization of sections of squid axons, bead-like domains of elongated proteins, presumably neurofilaments, exhibited distinct phosphorus signals. A marked asymmetry of these phosphorus signals was detectable between the pre- and the postsynaptic cytoskeleton of the giant synapse. Signals were very numerous and intense in the presynaptic terminal, while rare and weak in the postsynaptic axoplasm. The giant axon revealed a delayed appearance of phosphorus signals in its course from the cell bodies in the giant fibre lobe to its exit from the stellate ganglion. Numerous and intense phosphorylation signals were evident only in the peripheral giant axon. Asymmetry in the distribution of phosphorus signals between pre- and postsynaptic axoplasm paralleled differences in Ca(2+)-buffering mechanisms, as shown in a previous study. In the presynaptic terminal patterns of phosphorus signals correlated with precipitates which had formed after intra-axonal injection of calcium. Our observations suggest a role of phosphorylated neurofilaments in binding of calcium in the squid synapse.

Animals↗
PubMed1989

Visualization of polyribosomes in the postsynaptic area of the squid giant synapse by electron spectroscopic imaging.

Giant axons from squids (Loligo vulgaris and Loligo pealii) were studied by conventional electron microscopy and by electron spectroscopic imaging: electrons with a characteristic energy loss due to interaction with phosphorus atoms were used to image phosphorus-rich elements such as ribosomes. Patches of granular material were seen in the proximal region of giant axons, especially in the postsynaptic area of the giant synapse, but not on the presynaptic side. They were rare or absent in the more distal part of the axon. With electron spectroscopic imaging the patches emitted strong phosphorus signals, indistinguishable from signals of ribosomes in neuronal perikarya, confirming their identification as RNA-rich elements. In the part of the axon near the perikarya in the giant fibre lobe, the patches consisted of sacs of rough endoplasmic reticulum. It is suggested that patches of rough endoplasmic reticulum are transported to the synaptic region where ribosomes detach from the cisterns and aggregate into polyribosomes.

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PubMed1988

Circadian oscillations of DNA synthesis in rat brain.

The possibility that the synthesis of brain DNA undergoes a circadian fluctuation was examined in male adult Wistar rats, kept under natural lighting conditions or born and raised under artificial lighting conditions. Groups of rats were taken every 4 h during the 24 h, injected subcutaneously with [methyl-3H]thymidine and killed 4 h later. By cosinor analysis, the DNA specific activity of cerebral hemispheres and brainstem was found to show a significant 24 h rhythm with the peak at the beginning of the dark period (waking period). By contrast, in kidney, the peak of the circadian rhythm of DNA specific activity occurred during the light period (sleep period), in agreement with literature data. On the other hand, in 4-week-old rats, born and raised in artificial lighting conditions, brain DNA specific activity followed a 12 h rhythm, in agreement with the lack of a significant diurnal oscillation of the sleep--waking structure. It is concluded that brain DNA synthesis undergoes a circadian fluctuation in association with the circadian rhythm of waking.

Aging↗
PubMed1988

Axoplasmic RNA species synthesized in the isolated squid giant axon.

Isolated squid stellate nerves and giant fiber lobes were incubated for 8 hr in Millipore filtered sea water containing [3H]uridine. The electrophoretic patterns of radioactive RNA purified from the axoplasm of the giant axon and from the giant fiber lobe (cell bodies of the giant axon) demonstrated the presence of RNA species with mobilities corresponding to tRNA and rRNA. The presence of labeled rRNAs was confirmed by the behavior of the large rRNA component (31S) which, in the squid, readily dissociates into its two constituent moyeties (17S and 20S). Comparable results were obtained with the axonal sheath and the stellate nerve. In all the electrophoretic patterns, additional species of radioactive RNA migrated between the 4S and the 20S markers, i.e. with mobilities corresponding to presumptive mRNAs. Chromatographic analysis of the purified RNAs on oligo(dT)cellulose indicated the presence of labeled poly(A)+ RNA in all tissue samples. Radioactive poly(A)+ RNA represented approximately 1% of the total labeled RNA in the axoplasm, axonal sheath and stellate nerve, but more than 2% in the giant fiber lobe. The labeled poly(A)+ RNAs of the giant fibre lobe showed a prevalence of larger species in comparison to the axonal sheath and stellate nerve. In conclusion, the axoplasmic RNAs synthesized by the isolated squid giant axon appear to include all the major classes of axoplasmic RNAs, that is rRNA, tRNA and mRNA.

Animals↗
PubMed1988

The sequential hypothesis on sleep function. I. Evidence that the structure of sleep depends on the nature of the previous waking experience.

The sequential hypothesis on sleep function assumes that the information gathered by brain during the waking period is processed during sleep in two main steps occurring during synchronized sleep (SS) and, eventually, during paradoxical sleep (PS). To verify the main consequences of the hypothesis, i.e., (1) that SS is involved in brain information processing; and (2) that the structure of sleep is dependent on the nature of the previous waking experience, an experiment was designed involving rats exposed to a training session (two-way active avoidance) but failing to learn (NL), and rats left in their home cages in the same training room (C). The structure of sleep, determined by EEG techniques in the postacquisition period (3 hr), was different in NL rats in comparison to C rats, chiefly because SS episodes were markedly longer in the former group. A more detailed analysis indicated that, in NL rats, SS episodes not followed by PS increased their duration first, while those followed by PS became longer in the second half of the sleep period. Comparable results were obtained in the comparison of NL and C subgroups deprived of PS at the end of the acquisition period by chlomipramine treatment. The data support the sequential hypothesis and provide evidence for a primary role of SS in brain information processing.

Animals↗
PubMed1988

The sequential hypothesis on sleep function. II. A correlative study between sleep variables and newly synthesized brain DNA.

The information acquired by brain during wakefulness (W) may be processed in two sequential steps occurring during synchronized sleep (SS) and paradoxical sleep (PS), respectively. On the assumption that brain molecules synthesized during the acquisition step undergo a comparable sleep processing, we have designed an experiment aimed at the verification of the sequential hypothesis. Groups of adult female Wistar rats received [3H-methyl] thymidine by intraventricular injection 30 min before being exposed to a 4 hr session of a two-way active avoidance training. Animals failing to achieve the learning criterion were further allowed a period of 3 hr during which they were left free to sleep, or were deprived of PS or of total sleep. Control rats were similarly treated, but were left in their home cages in the same training room during the period of acquisition. The results of correlative study among behavioral, sleep and biochemical variables demonstrate that the specific radioactivity of DNA in the cerebral cortex, cerebellum and brainstem is correlated with several variables of postacquisition sleep, mostly SS parameters. The correlations depend on the previous waking experience of the rats. The data substantiate the two main consequences of the hypothesis, i.e., (1) the involvement of SS in brain information processing; and (2) the dependence of the operations performed by the sleeping brain on the nature of the previous waking experience. The results also provide some insight into the kind of processing which occurs in the sleeping brain.

Animals↗
PubMed1987

Occurrence and sequence complexity of polyadenylated RNA in squid axoplasm.

Axoplasmic RNA from the giant axon of the squid (Loligo pealii) comprises polyadenylated [poly (A)+] RNA, as judged, in part, by hybridization to [3H]polyuridine and by in situ hybridization analyses using the same probe. The polyadenylate content of axoplasm (0.24 ng/microgram of total RNA) suggests that the poly(A)+ RNA population makes up approximately 0.4% of total axoplasmic RNA. Axoplasmic poly(A)+ RNA can serve as a template for the synthesis of cDNA using a reverse transcriptase and oligo(deoxythymidine) as primer. The size of the cDNA synthesized is heterogeneous, with most fragments greater than 450 nucleotides. The hybridization of axoplasmic cDNA to its template RNA reveals two major kinetic classes: a rapidly hybridizing component (abundant sequences) and a slower-reacting component (moderately abundant and rare sequences). The latter component accounts for approximately 56% of the total cDNA mass. The rapidly and slowly hybridizing kinetic components have a sequence complexity of approximately 2.7 kilobases and 3.1 X 10(2) kilobases, respectively. The diversity of the abundant and rare RNA classes is sufficient to code for one to two and 205, respectively, different poly(A)+ RNAs averaging 1,500 nucleotides in length. Overall, the sequence complexity of axoplasmic poly(A)+ RNA represents approximately 0.4% that of poly(A)+ mRNA of the optic lobe, a complex neural tissue used as a standard. Taken together, these findings indicate that the squid giant axon contains a heterogeneous population of poly(A)+ RNAs.

Animals↗
PubMed1986

Synthesis of rat brain DNA during acquisition of an appetitive task.

We have examined the incorporation of [3H-methyl]thymidine into DNA extracted from several brain regions of rats learning a reverse handedness task, of control rats allowed to use their preferred paw, and of control rats left in their home cages. In learning animals, decrements in percent incorporation were observed in the visual cortex, remaining brain, hippocampus and entorhinal cortex. In the latter two regions less marked decreases were present in the active control group. No variation occurred in the sensory-motor cortex. In learning rats the specific radioactivity of neuronal DNA was markedly decreased in the hippocampus and remaining brain. In the former region, a less marked decrease was present in active control rats. In subcellular fractionation studies it was observed that decreases in DNA specific radioactivity prevailed in the mitochondrial fraction isolated from the hippocampus and visual cortex of learning rats. Brain radioactive DNA was widely distributed among fractions differing in their degree of repetitiveness. Its pattern of distribution did not coincide with that of bulk DNA and differed significantly among behavioural groups. The results suggest a non random origin of newly-synthesized brain DNA and its involvement in learning.

Animals↗
PubMed1986

Paradoxical sleep deprivation of the mother enhances DNA synthesis in fetal rat brain: autoradiographic and biochemical evidence.

Pregnant rats were deprived of paradoxical sleep for 3 days starting on the 18th gestational day. The condition of PS-D was imposed by confinement on a small platform surrounded by water or by daily injections of clomipramine. Four hours before the killing rats received a s.c. injection of [3H]-thymidine. The amount of radioactive DNA determined by autoradiography in several regions of fetal brain was found to be markedly increased under both experimental conditions in comparison with the control fetal brain. Considerably more limited effects were observed in kidney. Comparable changes of lower magnitude were obtained by comparing the specific radioactivity of DNA samples purified by chlorophorm extraction and digestion with RNase and proteinase K. The results fully confirm our previous data obtained under similar experimental conditions but based on the analysis of an acid-washed DNA fraction.

Animals↗
PubMed1986

Complexity of nuclear and polysomal RNA from squid optic lobe and gill.

The sequence complexity of nuclear and polysomal RNA from squid optic lobe and gill was measured by RNA-driven hybridization reactions with single-copy [3H]DNA. At saturation, brain nuclear and polysomal RNAs were complementary to 22.8 and 7.9% of the DNA probe, respectively. Assuming asymmetric transcription, the complexity of nuclear and polysomal RNA was equivalent to 2.5 X 10(8) and 8.8 X 10(7) nucleotides, respectively. Approximately 80-85% of the sequence complexity of brain total polysomal RNA was found in the polyadenylated RNA fraction. In contrast to these findings, nuclear and polysomal RNAs from gill hybridized to 9.1 and 2.9%, respectively, of the single-copy DNA, values that were 2.5-fold lower than those obtained in the CNS. Taken together, the results focus attention on the striking diversity of gene expression in the squid CNS and extend to the cephalopod mollusks the observation that nervous tissue expresses significantly more genetic information than other somatic tissues or organs.

Animals↗
PubMed1985

Effect of sleep on cerebral DNA synthesized during shuttle-box avoidance training.

Female Wistar rats weighing 200 g were implanted with cortical electrodes and two intraventricular cannulae. Five days later they were given 3H-thymidine and exposed to shuttle-box training for four hours. They were then left free to sleep in the following three hours during which their EEG activity was recorded. In comparison with control animals (C), learning (L) and non-learning (NL) rats exhibited an increase in SS. In comparison to the EEG recording made the previous day, all animal groups displayed an increase in SS, but only NL rats suffered a decrease in PS(%). The specific radioactivity of DNA measured in several brain regions was tendentially lower in NL rats, but significance was achieved only in the cerebellum in the comparison between NL rats and C rats. No change occurred in liver. More marked and significant decrements in the DNA specific radioactivity of all brain regions were observed in the subgroup of NL rats displaying relatively high values of PS time in comparison to the analogous subgroups of C and L animals. Comparable decrements were present with regard to the subgroup of NL rats endowed with relatively low PS time. Less widespread and more limited changes were observed in the concentration of acid-soluble radioactivity. In addition, several significant correlations were detected by Spearman's analysis among behavioral, biochemical and sleep parameters. The results are consistent with the interpretation that the selective decrease in brain radioactive DNA observed in NL rats reflects a loss of DNA synthesized during the training period. The loss is related to the amount of post-training PS and is associated to a lengthening of the mean duration of PS episodes. It may be concluded that the loss of newly-synthesized brain DNA reflects the elimination of molecules associated with neural information devoid of adaptive value.

Animals↗