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Transient focal ischemia induced in rat brain by occlusion of the middle cerebral artery (MCAo) elicits a generalized induction of the 72 kDa heat-shock protein (hsp72) heralding functional recovery. As this effect implies activation of protein synthesis, and local systems of protein synthesis are present in brain synapses, and may be analyzed in preparations of brain synaptosomes, we evaluated hsp72 expression and protein synthesis in synaptosomal fractions of spontaneously hypertensive rats (SHRs) subjected to permanent MCAo. SHRs were randomly divided in ischemics and sham controls, anaesthesia controls and passive controls. Focal ischemia was induced under chloral hydrate anaesthesia by unilateral permanent MCAo. Protein synthesis was determined by [ 35 S]methionine incorporation into synaptosomal proteins from ischemic and contralateral cortex/striatum, and from cerebellum. Hsp72 expression was measured in the same fractions by immunoblotting. Our data demonstrate that under these conditions synaptic hsp72 markedly increases in the ischemic hemisphere 1 and 2 days after MCAo, progressively declining in the following 2 days, while no significant change occurs in control rats. In addition, in the ischemic hemisphere the rate of synaptic protein synthesis increases more than two-fold between 1 and 4 days after MCAo, without showing signs of an impending decline. The present data provide the first demonstration that synaptic protein synthesis is massively involved in brain plastic events elicited by permanent focal ischemia.

Octopus vulgaris maintained under a 12/12h light/dark cycle exhibit a pronounced nocturnal activity pattern. Animals deprived of rest during the light period show a marked 'rebound' in activity in the following 24h. 'Active' octopuses attack faster than 'quiet' animals and brain activity recorded electrically intensifies during 'quiet' behaviour. Thus, in Octopus as in vertebrates, brain areas involved in memory or 'higher' processes exhibit 'off-line' activity during rest periods.

Dithiocarbamates, a class of compounds widely used in medicine and agriculture, have been reported to impair sleep structure. These effects have been attributed to the decrease in norepinephrine levels induced by these drugs. However, it has also been recently demonstrated that most of the mechanisms by which dithiocarbamates damage cell function involve changes in oxidative environment. To verify the potential relevance of the latter mechanism in the sleep impairment, we examined the sleep response of adult rats to an acute administration of diethyldithiocarbamate (DDTC). At the dose of 0.6 g/kg, DDTC induced fragmentation and a decrease in slow wave sleep (SWS), and a dramatic loss of paradoxical sleep (PS). These changes occurred soon after the treatment (day 0), persisted the following day (day 1), partially recovered on day 3, and regained near basal values on day 6. No sleep anomalies were observed with a lower dose of DDTC (0.06 mg/kg). On the other hand, when the higher dose of DDTC was given in association with either one of two antioxidants, α-tocopherol or melatonin, the amounts of SWS and PS significantly improved even on day 1, suggesting that the DDTC effects on sleep involved an impairment of the brain oxidative balance. Likewise, administration of the lower dose of DDTC 5 days before the higher dose induced a much earlier recovery of normal sleep, presumably due to the development of a tolerance to DDTC. On the whole, the data suggest that the brain oxidative environment may play a role in the mechanisms subserving sleep regulation.

This article focuses on local protein synthesis as a basis for maintaining axoplasmic mass, and expression of plasticity in axons and terminals. Recent evidence of discrete ribosomal domains, subjacent to the axolemma, which are distributed at intermittent intervals along axons, are described. Studies of locally synthesized proteins, and proteins encoded by RNA transcripts in axons indicate that the latter comprise constituents of the so-called slow transport rate groups. A comprehensive review and analysis of published data on synaptosomes and identified presynaptic terminals warrants the conclusion that a cytoribosomal machinery is present, and that protein synthesis could play a role in long-term changes of modifiable synapses. The concept that all axonal proteins are supplied by slow transport after synthesis in the perikaryon is challenged because the underlying assumptions of the model are discordant with known metabolic principles. The flawed slow transport model is supplanted by a metabolic model that is supported by evidence of local synthesis and turnover of proteins in axons. A comparison of the relative strengths of the two models shows that, unlike the local synthesis model, the slow transport model fails as a credible theoretical construct to account for axons and terminals as we know them. Evidence for a dynamic anatomy of axons is presented. It is proposed that a distributed “sprouting program,” which governs local plasticity of axons, is regulated by environmental cues, and ultimately depends on local synthesis. In this respect, nerve regeneration is treated as a special case of the sprouting program. The term merotrophism is proposed to denote a class of phenomena, in which regional phenotype changes are regulated locally without specific involvement of the neuronal nucleus.

High resolution computerized EEG analyses, and behavioral observations were used to identify slow wave sleep (SS), paradoxical sleep (PS) and transition sleep (TS) in adult male Wistar rats exposed to a session of two-way active avoidance training. Of the four sleep sequences that could be identified, two included TS (SS--TS--W and SS--TS--PS), while the other two did not (SS--W and SS--PS). Comparison of post-trial sleep variables between fast learning rats (FL, reaching criterion in the training session), slow learning rats (SL, reaching criterion in the retention session the following day), and non learning rats (NL, failing to reach criterion) indicated that the total amounts of SS, TS and PS of the SS--TS--PS sequence was markedly higher in FL rats than in SL rats. In addition, in comparison with the corresponding baseline period, the average duration and total amount of SS and TS episodes of the SS--TS--PS sequence increased in FL rats, while the number of SS--TS--W sequences decreased. On the other hand, the average duration of SS episodes increased in the SS--TS--W and SS--W sequences of SL rats, and in the SS--W and SS--TS--PS sequences of NL rats. Correlative analyses between number of avoidances and post-trial sleep variables demonstrated that avoidances were directly correlated with the duration of SS episodes of the SS--TS--PS sequence and with the duration of TS episodes of the SS--TS--W sequence, but inversely correlated with the number and amount of SS episodes of the SS--W sequence and with the duration and amount of SS episodes of the SS--PS sequence. On the whole, the data supported the view that TS-containing sleep sequences are involved in long-term storage of novel adaptive behavior, while sleep sequences lacking TS are involved in the maintenance of innate behavioral responses.

By scoring 5-s EEG epochs and calculating spectral power of consecutive EEG segments as short as 1 s, transition sleep (TS) episodes were identified in baseline recordings of adult rats. TS episodes were characterized by the abrupt appearance of theta and alpha waves within an ongoing period of slow-wave sleep (SS). They were followed by paradoxical sleep (PS) or, somewhat more frequently, by a period of wakefulness (W) that often led to an additional SS. Statistical values of the main variables of TS → (W) and TS → (PS) episodes are presented, together with comparable data concerning previous SS and following W or PS episodes. On the whole, TS episodes were more numerous than PS episodes, and less numerous than SS episodes. Their average duration was considerably shorter. As a consequence of the identification of TS and of brief W or PS epochs intervening within SS, the number of SS episodes was estimated to be considerably higher than previously assessed, and their average duration considerably shorter.

The aim of these studies was to map the neural consequences of exposure to a spatial novelty on the expression of immediate gene (IEG) and on unscheduled brain DNA synthesis (UbDS) in two genetic models of altered activity and hippocampal functions, i.e., the Naples High- (NHE) and Low-excitability (NLE) rats. Adult male rats of NLE and NHE lines, and of a random-bred stock (NRB) were tested in a Lat-maze, and corner crossings, rearings, and fecal boll were counted during two 10-min tests 24 h apart. For IEG expression, rats were exposed to a Lat-maze with nonexposed or repeatedly exposed rats used as controls, and were sacrificed at different time intervals thereafter. For UBDS, rats were sacrificed immediately after the first or the second exposure o a Lat-maze. IEG expression was measured by immunocytochemistry for the FOS and JUN proteins. NRB rats exposed for the first time to the maze showed extensive FOS and JUN positive cells in the reticular formation, the granular and pyramidal neurons of hippocampus, the amygdaloid nuclei, all layers of somatosensory cortex, and the granule cells of the cerebellar cortex. The positivity, stronger in rats exposed for the first time, was present between 2 and 6 h and was prevented by the NMDA receptor antagonist CPP (5 mg/kg). The positivity was very low in NHE rats, and it was stronger in NLE compared to NRB rats. UBDS was measured in ex vivo homogenates of brain areas by the incorporation into DNA of 3 H-[methyl]-thymidine given intraventricularly 15 min before test trial 1 or 2 (pulse of 0.5 h). The basal level of UBDS was higher in the hippocampus and neostriata of both NLE/NHE compared to NRB rats. On test trial 1 only, there was a significant decrease in UBDS in the neocortex and hippocampus of the NLE/NHE rats, and in the hypothalamus, midbrain, and cerebellum of NLE rats only. This decrease was lower in NHE than in NLE rats. In conclusion, the differential genotype-dependent profile of FOS and JUN peptides and UBDS in the NLE/NHE rats might cast some light on the neural substrates of spatial and emotional information processing in these neurogenetic models.

Using electroencephalographic methods (EEG), we have analyzed the basal sleep structure and the EEG power spectra of gerbils and rats during periods of wakefulness (W), synchronized sleep (SS) and paradoxical sleep (PS). During the 6hr light period examined, duration of sleep was similar for rats and gerbils, but gerbils showed fewer PS episodes and a longer amount of SS episodes followed by wakefulness. In addition, SS episodes preceding PS were of longer duration in gerbils than in rats. EEG power spectral analysis indicated a higher relative output in the 1–4 Hz range in gerbils in comparison with rats. On the whole, the data indicate the existence of significant differences in the basal sleep structure and EEG power spectra of gerbils and rats. This background information might be useful in the comparison of the effects of a given experimental treatment, such as cerebral ischemia, on the EEG activity of these two animal species.

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.

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.

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.

1. 1. The SE values and the molecular weights of ribosomal RNA extracted from the giant fibre lobe of the squid have been determined by agarose-polyacrylamide gel electrophoresis. The SE value and the molecular weight of the larger rRNA (28.0 and 1.75 × 106 daltons respectively) were similar to those reported for the corresponding mammalian species. On the other hand, the values of the smaller rRNA (21.7 and 1.03 × 106 daltons, respectively) were significantly larger. 2. 2. The data appear to contradict the belief that in the animal kingdom the molecular weight of the smaller ribosomal RNA has remained constant throughout evolution.

Using a translation assay we have shown that the axoplasm of the squid giant axon contains significant amounts of mRNA coding for a heterogeneous group of prot sets of proteins specified by glial and neuronal perikaryal mRNA. Messenger RNA is associated with the “microsomal” fraction of the axoplasm. The possible involvement of axoplasmic mRNA in protein synthesis remains to be ascertained. It is known that axoplasmic proteins are synthesized by the isolated giant axon, presumably by the surrounding glia cells.

1. 1. Using a specific assay method based on the oxidation of tissue extracts with d -amino acid oxidase, we have detected sizable amounts of d -alanine (from 1.5 to about 7 μmol/g) in the muscle and hepatopancreas of several crustacean species. The amino acid has been identified by analysis of the pyruvate formed in the oxidative reaction and, in some species, also by the selective decrease of the alanine peak separated by column chromatography. 2. 2. d -alanine is lacking in the hemolymph of the two crustacean species examined. 3. 3. Using a similar method of assay, but d -aspartate oxidase as the oxidizing enzyme, we have been unable to detect the presence of dicarboxylic amino acids with a d -configuration in the same crustacean species.

Cerebral NAD(P)H:(acceptor) oxidoreductase I (diaphorase I) is inhibited by barbiturates and nervous depressants which compete with the substrate. Determination of K i values for each compound provides a quantitative measure of their inhibitory effect. An investigation has been made on the chemical structures required for inhibition. The importance of the -CO-NR- group and of the size substituent groups is discussed. Other groups of atoms can, however, substitute for -CO-NR-. Cyclic structures are not required for inhibition. A parallel investigation on the structural requirements for inhibition of NADH oxidase of heart mitochondria has revealed a general similarity in the chemical structures required. The possible nature of inhibitory groups involved in binding with the enzyme is discussed.