Biomedical subjects

T Bartfai

Publications and source records attributed to T Bartfai.

At least 145 records · Page 8Linked to original sources

PubMed1992

Galanin and galanin antagonists: molecular and biochemical perspectives.

The neuropeptide galanin potently inhibits insulin release, hippocampal acetylcholine release and firing of locus coeruleus cells, and stimulates feeding and release of growth hormone. Galanin regulates K+ channels, adenylyl cyclase and phospholipase C by acting at Gi/Go protein-coupled high-affinity receptors. Galanin receptor agonists such as the N-terminal fragment galanin1-16 act synergistically with morphine in the somatosensory system and have potential analgetic application. Galanin antagonists may be useful therapeutic agents in endocrinology, neurology and psychiatry. The enhancing effect of such agents on hippocampal cholinergic function would be useful in treatment of Alzheimer's disease. Recent synthesis of a series of high-affinity galanin antagonists, reviewed, along with galanin's actions, by Tamas Bartfai and colleagues, opens the possibility of examining the functions of endogenous galanin and test the pharmacological usefulness of antagonism of galanin function in the endocrine, somatosensory and central nervous systems.

Acetylcholine↗

PubMed1992

Interleukin-1 alpha expression is inducible by cholinergic stimulation in the rat adrenal gland.

Interleukin-1-like immunoreactivity has earlier been demonstrated by immunohistochemistry in the noradrenaline-containing chromaffin cells of the rat adrenal gland [Schultzberg et al. (1989) Neuroscience 30, 805-810; Schultzberg et al. (1987) J. Neurosci. Res. 18, 184-189]. In this study, we examine the regulation, upon cholinergic stimulation, of the expression of the cytokine interleukin-1 alpha in the rat adrenal gland. Interleukin-1 alpha and interleukin-1 alpha mRNA levels in the adrenal gland are affected by systemic administration of the cholinergic agonists nicotine (0.5 mg/kg, i.p.) and carbachol (0.5 mg/kg, i.p.). Both drugs cause an increase in interleukin-1 alpha mRNA levels. In contrast to the increased mRNA levels, nicotine and carbachol reduce the interleukin-1 alpha protein level measured in the rat adrenal gland: nicotine by approximately 30%, 60 min after injection, and carbachol by approximately 55%, 30 min after injection. The interleukin-1 alpha protein level returns to control level 90 min after nicotine injection, and 120 min after carbachol injection. We thus found a large, constitutively expressed and inducible pool of interleukin-1 alpha in the rat adrenal gland, which appears to be sensitive to cholinergic stimulation and which may be responsible for some of the local and systemic effects of interleukin-1 alpha. Experiments with Escherichia coli lipopolysaccharide show that this substance, which induces interleukin-1 expression and secretion in macrophages, is also able to induce the expression of interleukin-1 alpha mRNA and interleukin-1 alpha in the adrenal gland when injected at the dose of 2 mg/kg, i.p.

Adrenal Glands↗

PubMed1992

Evidence for a role of the neuropeptide galanin in spatial learning.

The neuropeptide galanin coexists with acetylcholine (ACh) in the basal forebrain cholinergic neurons and modulates cholinergic activity in the forebrain. The cholinergic forebrain neurons appear to play a significant role in learning and memory, as suggested by a severe loss of these neurons in Alzheimer's disease. The involvement of endogenous galanin in learning is demonstrated here by the use of the recently synthesized high-affinity galanin antagonist M35 [galanin(1-13)-bradykinin(2-9) amide] (Kd = 0.1 nM). Intracerebroventricular (i.c.v.) administration of M35 (6 but not 3 nmol) produced a significant (P < 0.025) facilitation of acquisition in a spatial learning test (Morris swim maze) without any increase in swim speed. Thus, M35 (6 nmol) shortened the escape latency, reduced the number of failures to reach the platform, and shortened the path length to reach the hidden platform. M35 (3 and 6 nmol) tended to enhance retention performance seven days after the last training session. Receptor autoradiographic studies on the distribution of [125I]M35 following i.c.v. administration show that it binds preferentially in the periventricular regions including the hippocampus. These results suggest that galanin may modulate spatial learning and memory and that galanin antagonists may provide a new principle in the treatment of Alzheimer's disease.

Amino Acid Sequence↗

PubMed1992

Design of chimeric peptide ligands to galanin receptors and substance P receptors.

Several chimeric peptides were synthesized and found to be high-affinity ligands for both galanin and substance P receptors in membranes from the rat hypothalamus. The peptide galantide, composed of the N-terminal part of galanin and C-terminal part of substance P (SP), galanin-(1-12)-Pro-SP-(5-11) amide, which is the first galanin antagonist to be reported, recognizes two classes of galanin binding sites (KD(1) less than 0.1 nM and KD(2) approximately 6 nM) in the rat hypothalamus, while it appears to bind to a single population of SP receptors (KD approximately 40 nM). The chimeric peptide has higher affinity towards galanin receptors than the endogenous peptide galanin-(1-29) (KD approximately 1 nM) or its N-terminal fragment galanin-(1-13) (KD approximately 1 microM), which constitutes the N-terminus of the chimeric peptide. Galantide has also higher affinity for the SP receptors than the C-terminal SP fragment-(4-11) amide (KD = 0.4 microM), which constitutes its C-terminal portion. Substitution of amino acid residues, which is of importance for recognition of galanin by galanin receptors, such as [Trp2], in the galanin portion of the chimeric peptide or substitution of ([Phe7] or [Met11]-amide) in the SP portion of chimeric peptide both cause significant loss in affinity of the analogs of galantide for both the galanin- and the SP-receptors. These results suggest that the high affinity of the chimeric peptide, galantide, may in part be accounted for by simultaneous recognition/binding to both receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1992

Cytokine regulation of neuronal survival.

Interleukin-1 is a cytokine involved in the immune response to infection and inflammation as well as a growth promotor for several cell types. Interleukin-1-like immunoreactive material has been found in the nervous system. We now show that antisera, which blocked the T-cell proliferative effects of interleukin-1 alpha, decreased neuronal cell counts (to 40% of control) in dissociated spinal cord cultures derived from fetal mice. This neuronal loss was prevented by addition of interleukin-1 alpha, and to a lesser extent by interleukin-1 beta. Exogenous interleukin-1 alpha increased the survival of neurons when added to cultures in which the electrical activity was blocked with tetrodotoxin, whereas no such cytokine-related increase in neuronal survival was observed in electrically active cultures. The antiserum-induced death could also be prevented by cotreatment of the cultures with 0.1 nM vasoactive intestinal peptide, a substance that induces the secretion of neuronal trophic factors from nonneuronal spinal cord cells and thereby increases neuronal survival in electrically inactive cultures. These studies indicate that the cytokine interleukin-1, or an immunologically cross-reactive protein, can increase neuronal survival.

Animals↗

PubMed1992

GEA 857, a putative blocker of potassium conductance, enhances muscarinic agonist-evoked responses: dissociation from an action on 5-HT mechanisms.

GEA 857 [2-(4-chlorophenyl)-1,1-dimethylethyl 2-amino-3-methylbutanoate], a structural analogue of the 5-HT uptake blocker alaproclate, was tested for its ability to modify tremor and salivation induced by muscarinic agonists (oxotremorine, arecoline) and acetylcholinesterase inhibitors (physostigmine, THA) in the male rat. These agents were employed at submaximal doses. GEA 857, similarly to alaproclate (Ogren et al. 1985a & b), produced a dose-dependent, statistically significant (in the 5-20 mg/kg dose range) enhancement of the tremor response induced by all four cholinergic stimulants. However, unlike alaproclate, GEA 857 failed to enhance salivation in a consistent manner. GEA 857 itself did not produce tremor in the absence of the muscarinic agonists or the acetylcholinesterase inhibitors. The potentiation of oxotremorine tremor by GEA 857 could be fully blocked by atropine (1 mg/kg intraperitoneally). Unlike alaproclate, GEA 857 failed to affect 5-HT uptake or 5-HT metabolism in the 10-20 mg/kg dose range. However, similarly to the action of alaproclate, the potentiating effect of GEA 857 on muscarinic responses could be explained neither by actions on serotonergic mechanisms nor by actions on muscarinic receptor mechanisms in the striatum. Evidence is presented suggesting that the ability of GEA 857 to enhance responses evoked by muscarinic agonists involves inhibitory properties of GEA 857 at certain membranal Ca(2+)-dependent K+ channels, the blockade of which can potentiate or prolong muscarinic cholinergic actions.

Alanine↗

PubMed1992

Galanin receptors and their second messengers in the lumbar dorsal spinal cord.

The ligand binding properties of galanin receptors were examined in crude synaptosomal fraction preparations of lumbar dorsal spinal cord, using chloramin-T mono-iodinated porcine Tyr26 galanin as ligand. The equilibrium binding of [125I]galanin showed the presence of a single population of high-affinity binding sites with KD = 0.6 +/- 0.2 nM in a concentration of 55 +/- 15 fmol mg-1 protein (Bmax). The N-terminal fragments galanin (1-16) and galanin (1-12) fully displaced specific [125I]galanin binding from membranes with IC50 values 6 nM and 4 microM, respectively. The C-terminal fragment galanin (17-29) did not displace [125I]galanin when applied in the concentration range 10(-11)-10(-4) M. GTP inhibited the specific binding of [125I] galanin in a concentration dependent manner, with 54% inhibition at 1 mM, suggesting that the galanin receptor found in lumbar dorsal spinal cord is G-protein coupled. Second messenger systems, through which the galanin receptor in lumbar dorsal spinal cord may exert its effect, were also studied. A galanin (10 microM) produced inhibition (58%) of the depolarization induced cGMP increase was found, whereas galanin (10 microM) did not inhibit the noradrenalin (100 microM) activated cAMP synthesis or phosphoinositide turnover in tissue slices of the spinal cord. Bilateral transection of the sciatic nerve at midthigh level 14 days prior to the binding experiment was performed, a treatment which is known to cause a dramatic increase of galanin-like immunoreactivity in the superficial layers of the dorsal spinal cord, dorsal root ganglion and in galanin mRNA levels, but no significant effect on Bmax or KD of the galanin receptor was found.

Animals↗

PubMed1992

Galanin in sensory neurons in the spinal cord.

The distribution and physiological effects of the neuropeptide galanin (GAL) have been examined in the somatosensory system. GAL is normally present in a few sensory neurons that terminate in the dorsal horn of the spinal cord and it is colocalized with substance P and calcitonin gene-related peptide. After peripheral nerve section, but not dorsal root section, the amount of GAL produced and present in sensory fibers proximal to the section is dramatically upregulated. In parallel functional studies, we could demonstrate that exogenous GAL has a complex effect on the spinal cord reflex excitability, facilitatory at low doses and inhibitory at high doses. Furthermore, GAL inhibits the effect of excitatory neuropeptides physiologically released at the peripheral and central terminals of small diameter afferents that subserve a nociceptive function. After axotomy, the inhibitory effect of GAL is increased. We conclude that GAL may have an important role in the control of nervous impulses that underlie pain states that can occur after peripheral nerve injury.

Animals↗

PubMed1991

Galanin inhibits the potassium-evoked release of acetylcholine and the muscarinic receptor-mediated stimulation of phosphoinositide turnover in slices of monkey hippocampus.

In the ventral hippocampus of Cynomologus monkey, galanin, a 29 amino acid long neuropeptide, reduced the potassium-evoked release of [3H]acetylcholine from slices preloaded with [3H]choline and diminished the carbachol-stimulated accumulation of [3H]inositol polyphosphates in hippocampal microprisms preincubated with myo-[2-3H]inositol. Using receptor autoradiography a strong, specific binding of iodinated galanin was observed in the molecular layer of the dentate gyrus. These may thus be the sites where galanin exerts its inhibitory effects on acetylcholine (ACh) release and phosphoinositide breakdown. These data provide evidence that galanin is a modulator of cholinergic function in septo-hippocampal neurons of primates.

Acetylcholine↗

PubMed1991

M-15: high-affinity chimeric peptide that blocks the neuronal actions of galanin in the hippocampus, locus coeruleus, and spinal cord.

The 20-amino acid peptide M-15 binds with high affinity (IC50 approximately 0.1 nM) to 125I-labeled galanin (125I-GAL) binding sites in membranes from the ventral hippocampus, midbrain, and rat spinal cord. Receptor autoradiographic studies show that M-15 can displace 125I-GAL from all labeled sites. M-15 acts as a reversible high-affinity antagonist in blocking the inhibitory effects of GAL on the evoked release of acetylcholine in vivo in the hippocampus and on the GAL-induced hyperpolarization of locus coeruleus neurons in slices. M-15 also blocks the facilitatory effects of GAL on the spinal flexor reflex. Thus, the chimeric peptide M-15 [GAL-(1-13)-substance P-(5-11)amide] represents the first antagonist to the neuronal actions of GAL.

Acetylcholine↗

PubMed1991

Interleukin 1 expression is inducible by nerve growth factor in PC12 pheochromocytoma cells.

Expression of the cytokine interleukin 1 alpha (IL-1 alpha) was demonstrated in the rat PC12 pheochromocytoma cell line by (i) immunohistochemistry using rabbit polyclonal antisera raised against the recombinant murine IL-1 alpha, (ii) an ELISA, and (iii) a specific cell conversion bioassay based on the use of LBRM33-1A5 cells. IL-1 alpha mRNA was demonstrated in the PC12 cells, by PCR amplification. Constitutive expression of IL-1 alpha in PC12 cells was demonstrated in all experiments, although the cellular levels of IL-1 alpha-like immunoreactivity varied. The expression of IL-1 alpha, as studied at the mRNA level, was inducible by mouse nerve growth factor (7S NGF), and the gene product level was inducible in a dose- and time-dependent fashion by 7S NGF. The maximum induction corresponds to a 600% increase in IL-1 alpha-like immunoreactivity above the expression level found in noninduced cells and occurred after a 3-day incubation of the cells with NGF at 0.75 micrograms/ml of culture medium. The significance of the ability of NGF to induce IL-1 expression lies in the fact that IL-1 itself also acts as a growth factor that promotes glial proliferation and, even more importantly, IL-1 itself induces the expression of NGF at peripheral nerve injury [Lindholm, D., Heumann, R., Meyer, M. & Thoenen, H. (1987) Nature (London) 330, 658-659].

Animals↗

PubMed1991

Galanin-induced inhibition of insulin secretion from rat islets: effects of rat and pig galanin and galanin fragments and analogues.

The 29-amino acid neuropeptide galanin occurs in intrapancreatic nerves and inhibits insulin secretion. To study the structure-activity relations of galanin, we examined the effects of pig and rat galanin, three galanin fragments (galanin-(1-11), galanin-(1-16) and rat galanin-(17-29) and four galanin analogues ([Ala2]pig galanin, [Ala2]rat galanin, [D-Trp2]rat galanin and [D-Trp2]galanin-(1-16] on glucose-stimulated insulin secretion from isolated rat islets. Pig and rat galanin and galanin-(1-11) equipotently inhibited glucose-stimulated (8.3 mM) insulin secretion at and above 10(-7) M (P less than 0.05), whereas galanin-(1-16), inhibited insulin secretion at 10(-6) M (P less than 0.01). In contrast, the C-terminal rat galanin-(17-29) and the galanin analogues did not influence insulin secretion. Thus, rat and pig galanin are equipotent in inhibiting glucose-stimulated insulin secretion from rat islets. The active site resides in the N-terminal part of the molecule. Furthermore, the binding of galanin to its receptor depends on structural characteristics governed by the N-terminal position and in particular by the Trp2 residue.

Amino Acid Sequence↗

PubMed1991

Hypothalamic degradation of galanin(1-29) and galanin(1-16): identification and characterization of the peptidolytic products.

The degradation of the neuropeptide galanin(1-29) and its fully active synthetic N-terminal fragment galanin(1-16) in hypothalamic tissue, where these peptides potently affect feeding behaviour, is studied. Galanin(1-29) had a half-life of 100 min while galanin(1-16) had a half-life of 28 min when incubated with a hypothalamic membrane preparation. The putative sites of peptidolytic cleavage of the active N-terminal fragment galanin(1-16) were determined as being between amino acids Leu4 and Asn5, between Asn5 and Ser6, and between His14 and Ala15, respectively. The synthetic analogs of galanin(1-16) where Leu4, Asn5 or Ser6 was substituted by Ala were all more stable to peptidolysis; [Ala4]galanin(1-16) had a half-life of 55 min. Cleavage of the galanin(1-16) between His14-Ala15 yields a ligand-galanin(1-14) which binds to the receptor with high affinity (KD approximately 10(7) M), while cleavage at amino acid residues Leu4, Asn5 and Ser6 results in inactive peptide fragments with affinities for the galanin receptor below 10(-4) M. The enzyme(s) responsible for degradation of galanin were identified as endopeptidase(s), which were partially inhibited by bacitracin (1 mg/ml) by up to 50%, but not significantly by EDTA (1 mM), phosphoramidon (1 microM), phenylmethylsulfonyl fluoride, (100 microM) or aprotinin (10 micrograms/ml).

Amino Acid Sequence↗

PubMed1991

Characterization of a neuronal interleukin-1 receptor and the corresponding mRNA in the mouse anterior pituitary cell line AtT-20.

Interleukin-1 (IL-1) mediates numerous responses on the mouse anterior pituitary cell line AtT-20. We have studied the ligand binding properties of the IL-1 receptors (IL-1R) of the AtT-20 cells and found that they possess 1.5 x 10(3) receptors/cell with a Kd of 0.15 nM for [125I]IL-1 alpha. Using oligonucleotide primers, which define a 236 bp region of the cDNA coding for the cytosolic part of the mouse T-cell IL-1R, and cDNA prepared from AtT-20 cells, we obtained by polymerase chain reaction (PCR) a DNA fragment which was shown to possess an identical sequence to that of the corresponding region of the mouse T-cell IL-1R. Thus the AtT-20 pituitary cell line IL-1 receptor appears to be similar to those found on mouse T-cells and fibroblasts.

Adrenocorticotropic Hormone↗

PubMed1991

Pertussis toxin-sensitive G-protein mediates galanin's inhibition of scopolamine-evoked acetylcholine release in vivo and carbachol-stimulated phosphoinositide turnover in rat ventral hippocampus.

The effects of intracerebroventricular (i.c.v.) injections of pertussis toxin were investigated on the inhibitory action of galanin on acetylcholine release and phosphoinositide breakdown stimulated by muscarinic agents in rat ventral hippocampus. Pertussis toxin (0.6 micrograms, i.c.v., 96 h) counteracted the in vitro inhibitory effect of galanin (3.1 nmol) on phosphoinositide breakdown stimulated by carbachol without altering the stimulatory action of the cholinergic agonist on signal transduction, in miniprisms from rat ventral hippocampus. Pertussis toxin also abolished the in vivo effect of galanin on scopolamine-stimulated acetylcholine release in vivo but did not affect basal acetylcholine release. The results indicate that pertussis toxin-sensitive G-protein(s) mediates the galanin receptor regulation of pre- and postsynaptic cholinergic functions in the ventral hippocampus.

Acetylcholine↗

PubMed1991

A sensitive method for measuring neutralizing antibodies to Bordetella pertussis toxin: optimized ADP-ribosylation of transducin.

Pertussis toxin (PT), preactivated with 20 mm dithiothreitol (DTT), was incubated with different serum dilutions (1/10-1/200) before addition to the reaction mixture. Final concentrations of the reagents were: PT (50 ng/ml), dithiothreitol (DTT)(less than or equal to 0.3 mM), bovine transducin (2 micrograms/ml), ATP (1 mM). GTP (1 mM), lysophosphatidylcholine (LTC) (0.1 mg/ml), sodium acetate (NaAc) (0.1 M), Tris-HCl, pH 7.1 (0.06 M) and 32P-NAD+ (10 microCi 28 Ci/mM). The reaction was stopped by precipitation with 10% TCA (w/v), the pellet was collected and the samples were submitted to SDS-PAGE followed by autoradiography. ADP-ribosylation was detected as the radiolabelling of a protein band (m.w. approximately 40 kD) corresponding to the alpha-subunit of transducin. 32P-ADP incorporation was a linear function of PT concentration. By this assay quantitative differences in PT neutralizing antibodies were found between sera which were not revealed by measuring PT neutralizing antibodies by a Chinese hamster ovary (CHO) cell culture test or by an enzyme-linked immunosorbent assay (ELISA). The conditions of the ADP-ribosylation of bovine transducin have been optimized to permit detection of the enzymic activity of as low amounts of PT as 0.5 ng. This opens the possibility of the study of the presence and avidity of neutralizing antibodies to PT in post-vaccination sera without preceding purification and concentration of the antibodies and thus may provide a useful tool for evaluation of whooping cough vaccines.

Adenosine Diphosphate Ribose↗

PubMed1991

Linear and cyclic N-terminal galanin fragments and analogs as ligands at the hypothalamic galanin receptor.

The neuropeptide galanin (1-29) binds with high affinity to hypothalamic receptors (KD approximately 0.9 nM) and regulates feeding behavior. The N-terminal fragments (1-16), (1-16)NH2 are high affinity (KD approximately 6 nM) full agonists in vivo and in vitro. L-Ala substitutions show that amino acid residues Gly1, Trp2, Asn5, Tyr9, and Gly12 are important for the high affinity binding of galanin (1-16). Shortening the fragment (1-16) to galanin (1-7) causes a gradual drop of affinity: galanin (1-15), (1-14), and (1-13) have submicromolar KD values and galanin (1-12) has KD approximately 3 microM. Cyclic analogs of galanin (1-12) of different ring size were synthesized by condensing Gly1 and Gly12 without or with spacer groups. These analogs, independent of ring size, had a lower affinity than the linear galanin (1-12). Derivatization of the N-terminus of galanin (1-29), (1-16), and (1-12) all resulted in a large drop of affinity for the receptors, suggesting again the importance of the free N-terminal Gly.

Alanine↗

PubMed1991

Galanin reduces carbachol stimulation of phosphoinositide turnover in rat ventral hippocampus by lowering Ca2+ influx through voltage-sensitive Ca2+ channels.

The 29-amino-acid peptide galanin (GAL) caused concentration-dependent inhibition of the accumulation of 3H-inositol phosphates (3H-InsPs) induced by the muscarinic agonist carbachol (CARB; 10(-3)-10(-5) M) in the presence of 5 mM lithium, specifically in tissue miniprisms from rat ventral hippocampus. The inhibitory effect of GAL involved the mono-, bis-, tris-, and tetrakisphosphates formed during activation for 2 min of phospholipase C by CARB (1 mM) in the absence of lithium. GAL (1 microM) did not affect alpha-adrenergic or serotonergic type 2 receptor-mediated phosphoinositide (PI) breakdown in the same tissue. GAL by itself neither acted on basal levels of 3H-InsPs nor affected muscarinic receptors in binding studies. Blockade of the T-, N-, and L-types of voltage-sensitive calcium channel (VSCC) with 200 microM Cd2+ reduced muscarinic receptor-mediated PI breakdown by 50% and abolished the inhibitory effect of GAL (1 microM). Reduction of the extracellular Ca2+ concentration from 1.3 mM to 0.49 microM abolished the GAL inhibition of CARB-stimulated PI hydrolysis. Ca2+ influx promoted by 18 mM K+ depolarization or by 1 microM Bay K 8644, a selective agonist of the L-type VSCC, prevented the inhibitory effect of GAL. Blockade of the L-type VSCC with nifedipine (1 microM) potentiated the inhibitory effects of GAL without affecting muscarinic stimulation of PI breakdown.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗