Posttranslational deamidation of crystalline lens proteins during animal aging

The amide content of total proteins and protein fractions (alpha-, beta-, gamma-cristallins and albuminoid) from cortex and nuclear lens zones of cattle has been investigated. The amide content in proteins of cortex and nuclear lens of young animals (1,5-2 years old) is the same. The decrease of the amide content in the proteins of nuclear lenz zone of old animals (6-12 years old) is due to fraction of readily hydrolysed amides. The data obtained prove the posttranslational desamidation of lens proteins at ageing. beta-Cristallines--the proteins with the highest amides content is exposed to posttranslational desamidation most of all.     

Prostaglandins and functional specificity of central neurons of Helix pomatia

The effect of extra- and intracellularly injected prostaglandins (PG) E₂ and F₂ on electrical activity and responses to acetylcholine and serotonin were studied in experiments on identified neurons ofHelix pomatia. As a rule prostaglandins modified the typical electrical activity of the identified neurons: PG E₂ enhanced and PG F₂ depressed it. These substances mainly weakened responses of the nerve cells to mediators: PG E₂ caused a greater change in the response to serotonin and PG F₂ in the response to acetylcholine. Effects of the prostaglandins when injected extracellularly and intracellularly differed. The possible molecular-cellular mechanisms of the central action of prostaglandins are discussed in the light of their functional connections with other universal regulators of cellular metabolism and with proteins specific for nerve tissues.P. K. Anokhin Research Institute of Normal Physiology, Academy of Medical Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 13, No. 6, pp. 580–588, November–December, 1981.     

Heterochronic effects of neurotrophic factors in neurochemical organization of learning and memory in the adult organism

Following elaboration of long-term habituation to a startle-response, antibodies to neurotrophic factor protein S100b exerted selective and dose-dependent influence on different learning processes and memory of learned behavioural patterns in adult rats. S100b increased at all stages of behavioural skill development in hippocampus, hypothalamus, frontal cortex, cerebellar hemispheres and vermis, basal ganglia.     

Biochemical markers of apoptosis in different brain regions after training

Activity of caspase-3 and content of DNA fragments with sizes 0.2-0.6 kbp and more than 4.0 kbp in the worm of the cerebellum, hippocampus and prefrontal cortex of the adult rat brain were estimated in 4 and 24 hours after the procedure of acoustic startle habituation and fear conditioning. Heterochronic changes in apoptotic markers in the examined brain structures were observed after training. Caspase-3 activity was decreased in the worm of the cerebellum and hippocampus, and DNA fragmentation was suppressed in the hippocampus and brain cortex. At the same time, both caspase activity and DNA fragmentation were increased in the hypothalamus. These results provide evidence for the involvement of apoptosis in the mechanisms of learning and memory in adult brain.     

Inhibition of protein synthesis during associative memory reactivation produces reversible or irreversible amnesia in the snail Helix lucorum

Effects of protein synthesis inhibitors on reactivation processes of food aversion conditioning were inverstigated in snail Helix lucorum. Protein synthesis inhibitor (PSI, anisomycin, 0.4 mg, or cycloheximede, 0.6 mg) was injected into snail body cavity 24 hours after 3-day training; then conditioned stimulus (banana) was presented and memory was tested. It was found that 2.5-3 hours after first reminding, associative food conditioning was suppressed, recovering of the conditioning was observed 4.5-5.5 hours after first reminding. In other group of snails, PSI injections were single (1.8 mg) or triple (0.6 mg with 2-hour interval). Reminding stimulus was presented after each injection. In this case, suppression of food aversion conditioning was also observed 2.5-3 hours after first reminding, while amnesia in this case lasted over 30 days. Repeated training of the group of snails recovered the food aversion conditioning only partially. In control snails (saline instead of PSI or 3 injections of PSI without reminding), foot aversion conditioning was detected 30 days after first training. Thus we found that PSI effects during reminding of food aversion conditioning produced two phases amnesia: (1) the easily suppressed by PSI transient phase lasted 2-3 hours, and (2) irreversible phase, its suppression by high doses of PSI-initiated amnesia lasting over 1 month. Second phase of amnesia was not recovered after repeated training. It was suggested that reminding induced reconsolidation of initial memory. Its suppression by protein synthesis inhibitors results in erasing of memory trace and disturbs repeated consolidation.     

Neuroprotective effect of the hexapeptide HLDF-6 on neurons of rat hippocampus on the model of Alzheimer's disease in vivo and in vitro

The neuroprotective effect of Thr-Gly-Glu-Asn-His-Arg hexapeptide (HLDF-6), a biologically active fragment of the differentiation factor of human leukemia cells (HLDF), was demonstrated on models of Alzheimer's disease in vivo and in vitro. The syndromes of this pathology were induced in male rats by administration of the peptide corresponding to the 25-35 sequence of beta-amyloid peptide (25-35) and ibotenic acid into the hippocampus. HLDF-6 prevented loss of long-term memory and decrease in the orientation-investigation activity of these animals and significantly decreased the number of pyknotic neurons in the CA1 area of the hippocampus. This peptide also exerts a protective effect in vitro on the primary cultures of neurons of the hippocampus and cerebellum of rats under conditions of the beta-amyloid toxicity. An increase in the dihydrotestosterone (DHT) content was demonstrated in the blood plasma of rats with the syndrome of Alzheimer's disease and in the medium of the culture of hippocampus neurons in the presence of the Abeta(25-35) peptide. HLDF-6 inhibited this increase in both cases. A probable mechanism of the neuroprotective effect of HLDF-6 was suggested as being connected to its possible effect on both the biosynthesis and the metabolism of sex steroid hormones.     

How many biological replicates are needed in an RNA-seq experiment and which differential expression tool should you use?

RNA-seq is now the technology of choice for genome-wide differential gene expression experiments, but it is not clear how many biological replicates are needed to ensure valid biological interpretation of the results or which statistical tools are best for analyzing the data. An RNA-seq experiment with 48 biological replicates in each of two conditions was performed to answer these questions and provide guidelines for experimental design. With three biological replicates, nine of the 11 tools evaluated found only 20%–40% of the significantly differentially expressed (SDE) genes identified with the full set of 42 clean replicates. This rises to >85% for the subset of SDE genes changing in expression by more than fourfold. To achieve >85% for all SDE genes regardless of fold change requires more than 20 biological replicates. The same nine tools successfully control their false discovery rate at ≲5% for all numbers of replicates, while the remaining two tools fail to control their FDR adequately, particularly for low numbers of replicates. For future RNA-seq experiments, these results suggest that at least six biological replicates should be used, rising to at least 12 when it is important to identify SDE genes for all fold changes. If fewer than 12 replicates are used, a superior combination of true positive and false positive performances makes edgeR and DESeq2 the leading tools. For higher replicate numbers, minimizing false positives is more important and DESeq marginally outperforms the other tools.