Several peptides, including penetratin and Tat, are known to translocate across the plasma membrane. Dynorphin opioid peptides are similar to cell-penetrating peptides in a high content of basic and hydrophobic amino acid residues. We demonstrate that dynorphin A and big dynorphin, consisting of dynorphins A and B, can penetrate into neurons and non-neuronal cells using confocal fluorescence microscopy/immunolabeling. The peptide distribution was characterized by cytoplasmic labeling with minimal signal in the cell nucleus and on the plasma membrane. Translocated peptides were associated with the endoplasmic reticulum but not with the Golgi apparatus or clathrin-coated endocytotic vesicles. Rapid entry of dynorphin A into the cytoplasm of live cells was revealed by fluorescence correlation spectroscopy. The translocation potential of dynorphin A was comparable with that of transportan-10, a prototypical cell-penetrating peptide. A central big dynorphin fragment, which retains all basic amino acids, and dynorphin B did not enter the cells. The latter two peptides interacted with negatively charged phospholipid vesicles similarly to big dynorphin and dynorphin A, suggesting that interactions of these peptides with phospholipids in the plasma membrane are not impaired. Translocation was not mediated via opioid receptors. The potential of dynorphins to penetrate into cells correlates with their ability to induce non-opioid effects in animals. Translocation across the plasma membrane may represent a previously unknown mechanism by which dynorphins can signal information to the cell interior.

Transcription from multiple promoters along with alternative mRNA splicing constitutes the basis for cell-specific gene expression and mRNA and protein diversity. The prodynorphin gene (PDYN) gives rise to prodynorphin (PDYN), precursor to dynorphin opioid peptides that regulate diverse physiological functions and are implicated in various neuropsychiatric disorders. Here, we characterized PDYN transcripts and proteins in the adult human brain and studied PDYN processing and intracellular localization in model cell lines. Seven PDYN mRNAs were identified in the human brain; two of the transcripts, FL1 and FL2, encode the full-length PDYN. The dominant, FL1 transcript shows high expression in limbic-related structures such as the nucleus accumbens and amygdala. The second, FL2 transcript is only expressed in few brain structures such as the claustrum and hypothalamus. FL-PDYN was identified for the first time in the brain as the dominant PDYN protein product. Three novel PDYNs expressed from spliced or truncated PDYN transcripts either lack a central segment but are still processed into dynorphins, or are translated into N-terminally truncated proteins. One truncated PDYN is located in the cell nucleus, suggesting a novel nonopioid function for this protein. The complexity of PDYN expression and diversity of its protein products may be relevant for diverse levels of plasticity in adaptive responses for the dynorphin system.

Studies on parietal cortex slices from guinea pigs showed that a decrease in temperature from 37 degrees C to 24 degrees C significantly decreased the level of spontaneous neuron activity, while activatory reactions to iontophoretic administration of glutamate showed no significant change.

No abstract

Effects of a chronic combined unpredictable stress on activities of two cell death-related proteases, calpain and cathepsin B, were studied along with indices of nitrergic system in rat brain structures. Male Wistar rats were subjected to a 2-week-long combined stress (combination of unpaired flash light and moderate footshock associated with a white noise session). Stress resulted in a significant loss in the body and thymus weight and increased defecation in the open field test, though neither motor and exploratory activity, nor plasma corticosterone differed from the respective control levels. Decreased calpain activity and increased cathepsin B activity were demonstrated in the hippocampus of stressed rats (previously we have shown that caspase-3 activity was significantly suppressed in the brain of rats subjected to same type of stress). A significant reduction in the number of NOS-containing neurons was accompanied by a chronic stressinduced decline in NOS activity in the neocortex. Similar changes were observed in the hippocampus. However, levels of NO metabolites were elevated in both structures. Thus, stress-induced structural modifications in the brain may be mediated by disturbances in the nitrergic system and increased lysosomal proteolysis.