Biomedical subjects

Ulo Langel

Publications and source records attributed to Ulo Langel.

At least 37 records · Page 2Linked to original sources

PubMed2005

Translocation of dynorphin neuropeptides across the plasma membrane. A putative mechanism of signal transmission.

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.

Animals↗

PubMed2005

Galanin and perseveration.

Galanin is a 29/30 amino acid neuropeptide that has been shown to impair learning and memory task performance and also have roles in somatosensation, stress responses, sexual behavior, and feeding regulation. However, little is known about galanin involvement in higher cognitive processes, especially executive processes. Perseveration is a classic sign of frontal cortex damage and failure of executive control. Galanin has been shown to disrupt the performance of maze delayed alternation tasks and the operant, spatial delayed nonmatch-to-position (DNMTP) working memory task, tests especially sensitive to perseverative responding. To better understand this potential involvement of galanin in executive control, the present study tested the hypothesis that galanin induces perseveration. The first experiment examined the effects of galanin (10, 20 microg i.c.v.) on the performance of a simple operant response alternation task in which stimuli were assigned to one of two spatially distinct locations to produce extended sequences of presentations to one location, separated by a 10-s intertrial interval. The second experiment looked at the effects of galanin (5, 20 microg i.c.v.) on the performance of non-delayed match-to-position and nonmatch-to-position conditional discrimination operant tasks in which a minimal 1.0 s time interval separated responses. Finally, the effects of galanin (10, 20 microg i.c.v.) on delayed match-to-position (DMTP) performance were examined to determine whether response alternation (i.e., nonmatching) was critical to observing a galanin-induced impairment in this task. Galanin reduced the rate of trial completion in all the tasks, but did not alter simple or conditional discrimination accuracy. Galanin (10 microg) impaired DMTP performance in a delay-independent manner. Together, these data suggest that galanin does not produce perseveration, but are consistent with a galanin-induced decrease in reinforcer strength.

Animals↗

PubMed2005

Effects of galnon, a non-peptide galanin-receptor agonist, on insulin release from rat pancreatic islets.

Galanin is a neurotransmitter peptide that suppresses insulin secretion. The present study aimed at investigating how a non-peptide galanin receptor agonist, galnon, affects insulin secretion from isolated pancreatic islets of healthy Wistar and diabetic Goto-Kakizaki (GK) rats. Galnon stimulated insulin release potently in isolated Wistar rat islets; 100 microM of the compound increased the release 8.5 times (p<0.001) at 3.3 mM and 3.7 times (p<0.001) at 16.7 mM glucose. Also in islet perifusions, galnon augmented several-fold both acute and late phases of insulin response to glucose. Furthermore, galnon stimulated insulin release in GK rat islets. These effects were not inhibited by the presence of galanin or the galanin receptor antagonist M35. The stimulatory effects of galnon were partly inhibited by the PKA and PKC inhibitors, H-89 and calphostin C, respectively, at 16.7 but not 3.3 mM glucose. In both Wistar and GK rat islets, insulin release was stimulated by depolarization of 30 mM KCl, and 100 microM galnon further enhanced insulin release 1.5-2 times (p<0.05). Cytosolic calcium levels, determined by fura-2, were increased in parallel with insulin release, and the L-type Ca2+-channel blocker nimodipine suppressed insulin response to glucose and galnon. In conclusion, galnon stimulates insulin release in islets of healthy rats and diabetic GK rats. The mechanism of this stimulatory effect does not involve galanin receptors. Galnon-induced insulin release is not glucose-dependent and appears to involve opening of L-type Ca2+-channels, but the main effect of galnon seems to be exerted at a step distal to these channels, i.e., at B-cell exocytosis.

Animals↗

PubMed2005

Important pharmacophores for binding to galanin receptor 2.

Galanin(2-11) has been introduced as a receptor subtype selective ligand for the GalR2 subtype of the galanin receptors, and has gained use in pharmacological studies of galaninergic signaling in the past two years. By introducing l-Ala substitutions in the galanin(2-11) sequence, we have examined the amino acid residues which are of importance for binding to the GalR2 receptor. Our study shows that Trp2, Asn5, Gly8 and Tyr9 are of great importance for high affinity binding. When placed in an alpha-helical conformation, the side chains of these residues are, with the exception of Tyr9, displayed on the same "side" of the peptide. This information is useful in the rational design of non-peptide type GalR2 receptor ligands.

Amino Acid Substitution↗

PubMed2005

Galnon--a low-molecular weight ligand of the galanin receptors.

Galnon is a low-molecular weight galanin receptor ligand, with affinity towards the three galanin receptors in the micromolar range. Galnon is of interest as a drug candidate due to its stability and ability to pass the blood-brain barrier. Like galanin, galnon has also been shown to affect various physiological functions; however, occasionally galanin and galnon act in opposing ways. Since its introduction in 2002, galnon has been characterized to inhibit seizures, decrease feeding behaviour, diminish physical signs of opiate withdrawal and to alleviate heat-hyperalgesic response to partial sciatic nerve injury. In this review, we will summarize what is known about galnon to date.

Animals↗

PubMed2005

Assessment of new functional roles for galanin in the CNS.

There is considerable evidence that galanin plays an important role in food intake, nociception, stress reactivity, sexual behavior, and learning. However, its involvement in other important behavioral processes, especially higher cognition, has not been systematically investigated. This review summarizes our recent efforts to explore the role of galanin in executive processes as well as in fundamental motivational and motor productive processes that are essential to the behavioral assessment of cognitive constructs in rodents. Our work points to a role in reward processes and physiological water balance regulation, and argues against a role for galanin in fine motor production and maintenance, simple visual discrimination, and perseveration.

Animals↗

PubMed2005

Galanin receptor ligands.

The three galanin receptor subtypes (GalR1-3) belong to the G protein-coupled receptor superfamily. The widespread distribution of galanin and its receptors in the CNS and PNS and the numerous physiological and pharmacological effects of galanin (for review, cf. Vrontakis, 2002) render the three galanin receptors attractive drug targets. The industrial efforts, however, have not yet resulted in a wealth of receptor subtype specific agonists or antagonists with high affinity and selectivity. The present paper summarizes the properties of the galanin ligands used at the end of 2004 in the ca. 2000 publications and complements their pharmacological characterization with new data.

Animals↗

PubMed2005

Internalisation of cell-penetrating peptides into tobacco protoplasts.

Cells are protected from the surrounding environment by plasma membrane which is impenetrable for most hydrophilic molecules. In the last 10 years cell-penetrating peptides (CPPs) have been discovered and developed. CPPs enter mammalian cells and carry cargo molecules over the plasma membrane with a molecular weight several times their own. Known transformation methods for plant cells have relatively low efficiency and require improvement. The possibility to use CPPs as potential delivery vectors for internalisation in plant cells has been studied in the present work. We analyse and compare the uptake of the fluorescein-labeled CPPs, transportan, TP10, penetratin and pVEC in Bowes human melanoma cells and Nicotiana tabacum cultivar (cv.) SR-1 protoplasts (plant cells without cell wall). We study the internalisation efficiency of CPPs with fluorescence microscopy, spectrofluorometry and fluorescence-activated cell sorter (FACS). All methods indicate, for the first time, that these CPPs can internalise into N. tabacum cv. SR-1 protoplasts. Transportan has the highest uptake efficacy among the studied peptides, both in mammalian cells and plant protoplast. The internalisation of CPPs by plant protoplasts may open up a new effective method for transfection in plants.

Carrier Proteins↗

PubMed2005

Evaluation of transportan 10 in PEI mediated plasmid delivery assay.

Cell-penetrating peptides (CPPs) are novel high-capacity delivery vectors for different bioactive cargoes. We have evaluated the CPP transportan 10 (TP10) as a delivery vector in different in vitro plasmid delivery assays. Tested methods include: TP10 crosslinked to a plasmid via a peptide nucleic acid (PNA) oligomer, TP10 conjugation with polyethyleneimine (PEI), and addition of unconjugated TP10 to standard PEI transfection assay. We found that without additional DNA condensing agents, TP10 has poor transfection abilities. However, the presence of TP10 increases the transfection efficiency several folds compared to PEI alone. At as low concentrations as 0.6 nM, TP10-PNA constructs were found to enhance plasmid delivery up to 3.7-fold in Neuro-2a cells. Interestingly, the transfection efficiency was most significant at low PEI concentrations, allowing reduced PEI concentration without loss of gene delivery. No increase in cytotoxicity due to TP10 was observed and the uptake mechanism was determined to be endocytosis, as previously reported for PEI mediated transfection. In conclusion, TP10 can enhance PEI mediated transfection at relatively low concentrations and may help to develop future gene delivery systems with reduced toxicity.

Animals↗

PubMed2005

Cell-penetrating peptides: mechanism and kinetics of cargo delivery.

Cell-penetrating peptides (CPPs) are short peptides of less than 30 amino acids that are able to penetrate cell membranes and translocate different cargoes into cells. The only common feature of these peptides appears to be that they are amphipathic and net positively charged. The mechanism of cell translocation is not known but it is apparently receptor and energy independent although, in certain cases, translocation can be partially mediated by endocytosis. Cargoes that are successfully internalized by CPPs range from small molecules to proteins and supramolecular particles. Most CPPs are inert or have very limited side effects. Their penetration into cells is rapid and initially first-order, with half-times from 5 to 20 min. The size of smaller cargoes does not affect the rate of internalization, but with larger cargoes, the rate is substantially decreased. CPPs are novel vehicles for the translocation of cargo into cells, whose properties make them potential drug delivery agents, of interest for future use.

Animals↗

PubMed2005

Intracerebroventricular administration of galanin decreases free water intake and operant water reinforcer efficacy in water-restricted rats.

The 29/30 amino acid neuropeptide galanin coexists with vasopressin in the hypothalamus and has been shown to inhibit the actions of vasopressin and aldosterone, suggesting an inhibitory role for galanin in physiological water retention mechanisms and water seeking and water consumption behavior. Little work, however, has examined a role for galanin in water intake regulation. Furthermore, many experiments that have reported galanin-induced impairments in the performance of tasks thought to measure learning and memory have used water restriction routines and water reinforcers to maintain responding. Therefore, the present study examined the effects of intracerebroventricularly administered galanin (5.0-20.0 microg/5 microl) on free water consumption during a 10 min test session and a follow up open field exploration, an operant progressive ratio (PR) schedule, a test used to assess reinforcer strength, and an operant fixed time schedule (FT 20) in 23.5h water restricted rats. Finally, in an additional experiment that was designed to simulate the effects of a galanin-induced decrease in water reinforcer efficacy, the rats were allowed access to water prior to testing in an operant delayed non-matching to position (DNMTP) task. A galanin-induced decrease in water consumption was observed in both the free access test and the FT 20 at the 20 microg dose, but no significant galanin-induced alterations in open field behavior. A decrease in responses emitted and rewards received was observed on the PR schedule at the 5, 10, and 20 microg doses. Pre-session access to water significantly reduced the number of trials per session in the DNMTP but did not reduce accuracy. This study is the first to observe a galanin-induced reduction in water intake and reinforced operant behavior, and suggests that galanin may play a role in regulating water intake and reinforcement. However, the present data also suggest that DNMTP choice accuracy deficits observed previously cannot be attributed to a galanin-induced change in reinforcer efficacy.

Animals↗

PubMed2005

Synthesis of cell-penetrating peptides for cargo delivery.

Cell-penetrating peptides (CPPs) have served as vehicles for the delivery of different molecules and particles into cells. The efficiency and noninvasive nature of peptide-mediated cellular transduction provides a promising tool for biomedical research. Studies of cell-penetrating constructs that deliver oligonucleotides, peptides, and proteins have elucidated several important cellular signaling mechanisms. Here we briefly describe the major classes of cell-penetrating peptides. We also present various strategies used to couple different cargoes to CPPs.

Amino Acid Sequence↗

PubMed2005

Cellular delivery of peptide nucleic acid by cell-penetrating peptides.

Peptide nucleic acid (PNA) is a promising tool in biomedical research. PNA oligomers can be synthesized according to peptide synthesis protocols, but they hybridize to complementary RNA and DNA strands like oligonucleotides. The main hindrance to the use of PNA oligomers has been their poor uptake by cells. Fortunately, this limitation can be overcome by the application of peptide vectors. Different strategies are used to conjugate peptide vectors and PNA oligomers. In this chapter we discuss different types of delivery vectors (receptor ligands and cell-penetrating peptides) and strategies for the design and synthesis of PNA-vector conjugates. A protocol for the delivery of antisense PNA oligomer-cell-penetrating peptide conjugates is also described.

Cells↗

PubMed2005

Manual solid-phase synthesis of glutathione analogs: a laboratory-based short course.

This chapter provides a manual for a laboratory-based short course to introduce the common techniques of solid-phase peptide synthesis (SPPS). The course provides students the opportunity to design and manually synthesize analogs of glutathione using relatively simple equipment available in any unsophisticated laboratory. The manual provides compact protocols for both the different steps of SPPS and the final cleavage of peptides from resin supports. We also introduce a simple method for the synthesis of combinatorial libraries of glutathione analogs that is suitable for those relatively unfamiliar with the field of peptide chemistry.

Amino Acids↗

PubMed2005

Galanin and its receptors in neurological disorders.

Galanin is a highly inducible neuropeptide, showing distinct up-regulation after pathological disturbance within the nervous system. Significant increase in galanin expression is observed after peripheral nerve injury, in the basal forebrain in Alzheimer's disease (AD), during neuronal development, and after stimulation with estrogen, while seizure activity deplete galanin in the hippocampus. A wide distribution of galanin and its receptors is seen in the nervous system, often in co-localization with classical neurotransmitters and other neuromodulators. Galanin acts predominantly as an inhibitory, hyperpolarizing neuromodulator on neurotransmitter and glucose-induced insulin release and stimulates growth hormone and prolactin secretion. Galanin has been implicated in several higher order physiological functions including cognition, feeding, nociception, mood regulation, and neuroendocrine modulation. The effects of galanin are mediated via three G protein-coupled receptors with different functional coupling. Moderate to low pharmacological effects are seen by galanin under physiological conditions, in contrast to its dramatic effects on the nervous system after neuronal disturbance. This pathophysiological heavy function of the galaninergic system renders it an interest for disorders such as AD, depression, and epilepsy in terms of side effects. Some properties of the galaninergic system are of particular importance in the context of neurodegeneration. Galanin is highly inducible, 10- to 100-fold, upon nerve injury, whereas most neuropeptides are induced 1.5- to 2-fold. Galanin is strongly neurotrophic during development as well as subsequent to injury. Whereas other neurotrophic neuropeptides like VIP and PACAP activate cAMP synthesis, galanin suppresses its synthesis, yet it is a strong neurotrophic as well as neuroprotective agent. As we delineate which galanin receptor subtype mediates neuroprotective and neurotrophic effects and which mediates synaptic inhibition, pharmacological use of receptor- selective galaninergic ligands for treatment in neurodegenerative diseases are coming closer.

Animals↗

PubMed2005

Cell-penetrating peptides: mechanisms and applications.

A major obstacle in the development of new therapeutic agents is the low bioavailability of hydrophilic substances. Drugs that bind to intracellular targets must penetrate the lipid bilayer surrounding the cell in order to exert their effect. A relatively new research area that addresses this problem by introducing novel transport peptides that facilitate the cellular penetration of potential drugs has emerged. These peptides predominantly have a positive net charge and/or an amphipathic nature, but can otherwise have very different characteristics. This group of peptides, although sometimes called protein transduction domains (PTDs), is here referred to as cell-penetrating peptides (CPPs). For many years it was believed that these peptides were internalized into cells via a non-endocytotic, receptor-independent pathway. However, recent publications have suggested that an endocytotic pathway cannot be ruled out, and that earlier results might be based on artifacts associated with fixation of cells and membrane association of peptides. Although the mechanism of cellular uptake remains unclear, there is an increasing amount of reports on biological effects of CPPs and their cargos. Thus, CPPs are an attractive pharmaceutical and biochemical tool that needs more attention. This review will discuss some recent results in this research field with focus on the cell-penetrating peptide transportan.

Animals↗

PubMed2004

Intracerebroventricularly administered galanin does not alter operant reaction time or differentially reinforced high rate schedule operant responding in rats.

Galanin (Gal) is a 29/30 amino acid neuroendocrine peptide that impairs learning and memory processes, stimulates feeding, and modulates somatosensory, sex, and stress responses. Anatomical markers for Gal are found throughout the brain, including in the caudate-putamen and substantia nigra motor regions. Many of the behavioral tests that have been used to study the involvement of Gal in complex behavioral processes are motorically demanding, but no research has specifically investigated the involvement of Gal in response initiation or the maintenance of fine motor action. Therefore, the present study examined the effects of intraventricularly administered Gal on two highly sensitive operant tasks designed to detect alteration of these response properties. Response initiation was studied using a light-dark discrimination reaction time task that required a correct response within 2.5s of a spatially and temporally uncertain stimulus onset. The ability to perform high local rates of responding was studied using an operant differential reinforcement of high rate (DRH) of responding task. Gal (10-20 microg, i.c.v.) did not alter reaction time or inter-response time distributions in either task, though did substantially reduce the total number of responses and reinforcers obtained on the DRH schedule. These results are consistent with a Gal-induced reduction of reinforcer efficacy rather than Gal-disruption of response initiation or response patterning.

Analysis of Variance↗

PubMed2004

The use of cell-penetrating peptides as a tool for gene regulation.

A novel carrier system that originates from membrane shuttling proteins such as the Drosophila homeobox protein Antennapedia, the HIV-1 transcriptional factor TAT and VP22 from HSV-1 has advantages for targeted delivery compared with standard translocation techniques. This transport system is mediated by so-called cell-penetrating peptides, which consist of short peptide sequences that rapidly translocate large molecules into the cell interior in a seemingly energy- and receptor-independent manner. Cell-penetrating peptides have low toxicity and a high yield of delivery and in the future might become a widely used tool in the field of gene regulation.

Amino Acid Sequence↗