Annals of the New York Academy of SciencesVolume 805, Issue 1 p. 159-169 Development of VIP Agonists and Antagonists with Tissue and Receptor Specificity: Effects on Behavioral Maturation, Sexual Function, and the Biologic Clocka I. GOZES, Corresponding Author I. GOZES Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, Israel Tel: 972-3-640-7240; fax: 972-3-640-8541; e-mail: igozes@post.tau.ac.il.Search for more papers by this authorG. LILLING, G. LILLING Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. DAVIDSON, A. DAVIDSON Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. BARDEA, A. BARDEA Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. RESHEF, A. RESHEF Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorR. GLAZER, R. GLAZER Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorR. ZAMOSTIANO, R. ZAMOSTIANO Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorO. ASHUR-FABIAN, O. ASHUR-FABIAN Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. TICHER, A. TICHER Department of Human Genetics Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorI. E. ASHKENAZI, I. E. ASHKENAZI Department of Human Genetics Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorT. W. MOODY, T. W. MOODY Biomarkers and Prevention Research Branch NCI National Institutes of Health Rockville, Maryland 20850Search for more papers by this authorS. RUBINRAUT, S. RUBINRAUT Department of Organic Chemistry Weizmann Institute of Science Rehovot 76100, IsraelSearch for more papers by this authorM. FRIDKIN, M. FRIDKIN Department of Organic Chemistry Weizmann Institute of Science Rehovot 76100, IsraelSearch for more papers by this authorD. E. BRENNEMAN, D. E. BRENNEMAN Section on Developmental and Molecular Pharmacology Laboratory of Developmental Neurobiology National Institute of Child Health and Human Development National Institutes of Health Bethesda, Maryland 20842Search for more papers by this author I. GOZES, Corresponding Author I. GOZES Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, Israel Tel: 972-3-640-7240; fax: 972-3-640-8541; e-mail: igozes@post.tau.ac.il.Search for more papers by this authorG. LILLING, G. LILLING Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. DAVIDSON, A. DAVIDSON Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. BARDEA, A. BARDEA Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. RESHEF, A. RESHEF Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorR. GLAZER, R. GLAZER Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorR. ZAMOSTIANO, R. ZAMOSTIANO Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorO. ASHUR-FABIAN, O. ASHUR-FABIAN Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. TICHER, A. TICHER Department of Human Genetics Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorI. E. ASHKENAZI, I. E. ASHKENAZI Department of Human Genetics Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorT. W. MOODY, T. W. MOODY Biomarkers and Prevention Research Branch NCI National Institutes of Health Rockville, Maryland 20850Search for more papers by this authorS. RUBINRAUT, S. RUBINRAUT Department of Organic Chemistry Weizmann Institute of Science Rehovot 76100, IsraelSearch for more papers by this authorM. FRIDKIN, M. FRIDKIN Department of Organic Chemistry Weizmann Institute of Science Rehovot 76100, IsraelSearch for more papers by this authorD. E. BRENNEMAN, D. E. BRENNEMAN Section on Developmental and Molecular Pharmacology Laboratory of Developmental Neurobiology National Institute of Child Health and Human Development National Institutes of Health Bethesda, Maryland 20842Search for more papers by this author First published: December 1996 https://doi.org/10.1111/j.1749-6632.1996.tb17481.xCitations: 4 a This work was supported in part by Fujimoto Pharmaceutical Corporation, and in part by the Israel-U.S. Binational Science Foundation. AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Citing Literature Volume805, Issue1VIP, PACAP, Glucagon, and Related PeptidesDecember 1996Pages 159-169 RelatedInformation

A bstract : The complete coding sequence of a novel protein (828 amino acids, pI 5.99), a potential new mediator of vasoactive intestinal peptide (VIP) activity was recently revealed. The expression of this molecule, activity‐dependent neuroprotective protein (ADNP), was augmented in the presence of VIP, in cerebral cortical astrocytes. The mRNA transcripts encoding ADNP were enriched in the mouse hippocampus and cerebellum. The protein deduced sequence contained the following: (1) a unique peptide, NAPVSIPQ, sharing structural and immunological homologies with the previously reported, activity‐dependent neurotrophic factor (ADNF) and exhibiting neuroprotection in vitro and in vivo ; (2) a glutaredoxin active site; and (3) a classical zinc binding domain. Comparative studies suggested that the peptide, NAPVSIPQ (NAP), was more efficacious than peptides derived from ADNF. ADNP, a potential mediator of VIP‐associated neuronal survival, and the new peptide, a potential lead compound for drug design, are discussed below.

Abstract: Vasoactive intestinal peptide has neurotrophic and neuroprotective properties that influence the survival of activity‐dependent neurons in the central nervous system. Investigations of the mechanism of this neurotrophic peptide indicated that these actions are contingent on interactions with astroglia. The complex mixture of neurotrophic mediators released from astroglia include cytokines, a protease inhibitor, and activity‐dependent neurotrophic factor, a protein with apparent structural similarities to hsp60. Investigations of ADNF resulted in the discovery of active peptides of extraordinary potency and broad neuroprotective properties. These studies indicate that a nine‐amino acid core peptide of ADNF had significantly greater neuroprotective properties in comparison to the parent growth factor and these advantages identify ADNF‐9 as an attractive lead compound for drug development.

Vasoactive intestinal peptide (VIP) expression is restricted to interneurons in the hippocampus of normal adult rats. However, 3-6 hours after a 60-minute walk in an activity wheel, VIP was transiently expressed in most pyramidal and granular neurons of the hippocampus. Locomotion was also associated with a dramatic increase in VIP immunoreactivity in the motor cortex, primarily in bipolar cells. Reverse transcriptase-polymerase chain reaction analysis indicated that VIP mRNA increases transiently by more than twofold, before the increases in peptide immunoreactivity in both the hippocampus and motor cortex. By comparison, another marker of inhibitory interneurons, glutamate decarboxylase, did not change its expression pattern after locomotion. The calcium binding protein, calbindin-D28K, normally expressed in interneurons, was now found also in glial cells of the hippocampus and motor cortex. Another marker of enhanced electrical activity, the immediate early gene, c-Fos, was expressed in pyramidal and granular neurons at 3 hours but not at 6 hours after locomotion. These results suggest that mapping of peptide expression in the brain of a docile, inactive rat may not reflect the real distribution and functions of a peptide in an active animal.

Abstract : The vulnerability of neurons and the irreversibility of loss make discoveries of neuroprotective compounds fundamentally important. Here, the complete coding sequence of a novel protein (828 amino acids, pl 5.99), derived from mouse neuroglial cells, is revealed. The sequence contained (1) a neuroprotective peptide, NAPVSIPQ, sharing structural and immunological homologies with the previously reported, activity‐dependent neurotrophic factor ; (2) a glutaredoxin active site ; and (3) a zinc binding domain. Gene expression was enriched in the mouse hippocampus and cerebellum and augmented in the presence of the neuropeptide vasoactive intestinal peptide, in cerebral cortical astrocytes. In mixed neuron—astrocyte cultures, NAPVSIPQ provided neuroprotection at subfemtomolar concentrations against toxicity associated with tetrodotoxin (electrical blockade), the β‐amyloid peptide (the Alzheimer's disease neurotoxin), N‐methyl‐D‐aspartate (excitotoxicity), and the human immunodeficiency virus envelope protein. Daily NAPVSIPQ injections to newborn apolipoprotein E‐deficient mice accelerated the acquisition of developmental reflexes and prevented short‐term memory deficits. Comparative studies suggested that NAPVSIPQ was more efficacious than other neuroprotective peptides in the apolipoprotein E‐deficiency model. A potential basis for rational drug design against neurodegeneration is suggested with NAPVSIPQ as a lead compound. The relative enrichment of the novel mRNA transcripts in the brain and the increases found in the presence of vasoactive intestinal peptide, an established neuroprotective substance, imply a role for the cloned protein in neuronal function.

The understanding of the molecular mechanisms leading to peptide action entails the identification of a core active site. The major 28-aa neuropeptide, vasoactive intestinal peptide (VIP), provides neuroprotection. A lipophilic derivative with a stearyl moiety at the N-terminal and norleucine residue replacing the Met-17 was 100-fold more potent than VIP in promoting neuronal survival, acting at femtomolar-picomolar concentration. To identify the active site in VIP, over 50 related fragments containing an N-terminal stearic acid attachment and an amidated C terminus were designed, synthesized, and tested for neuroprotective properties. Stearyl-Lys-Lys-Tyr-Leu-NH2 (derived from the C terminus of VIP and the related peptide, pituitary adenylate cyclase activating peptide) captured the neurotrophic effects offered by the entire 28-aa parent lipophilic derivative and protected against beta-amyloid toxicity in vitro. Furthermore, the 4-aa lipophilic peptide recognized VIP-binding sites and enhanced choline acetyltransferase activity as well as cognitive functions in Alzheimer's disease-related in vivo models. Biodistribution studies following intranasal administration of radiolabeled peptide demonstrated intact peptide in the brain 30 min after administration. Thus, lipophilic peptide fragments offer bioavailability and stability, providing lead compounds for drug design against neurodegenerative diseases.

The intracellular stress-induced proteins provide protection against toxic insults. Here, a 60,000-Da heat shock 60 (hsp60)-like protein was detected, with five different antibodies, in conditioned media derived from rat cortical astrocytes and a human neuroblastoma cell line. Extracellular neuroblastoma hsp60-like immunoreactivity was increased 3-fold in the presence of the neuropeptide vasoactive intestinal peptide (VIP) and was augmented 2-fold after temperature elevation. Intracellular hsp60 immunoreactivity was reduced 2-3-fold in the presence of VIP; this reduction was attenuated in the presence of brefeldin A, an inhibitor of protein secretion. In contrast, the activity of lactate dehydrogenase (LDH), an intracellular marker, did not change in the presence of VIP. Essentially no extracellular LDH activity was detected, indicating no cellular damage. A novel aspect for stress proteins having extracellular protective roles is suggested.

A bstract : Vasoactive intestinal peptide has neurotrophic and growth‐regulating properties. As in the case of many neurotrophic molecules, VIP also has neuroprotective properties, including the prevention of cell death associated with excitotoxicity (NMDA), beta‐amyloid peptide, and gp120, the neurotoxic envelope protein from the human immunodeficiency virus. The neurotrophic and neuroprotective properties are mediated in part through the action of glial‐derived substances released by VIP. These substance include cytokines, pro tease nexin I, and ADNF, a novel neuroprotective protein with structural similarities to heat‐shock protein 60. Antiserum against ADNF produced neu ronal cell death and an increase in apoptotic neurons in cell culture. A 14 amino acid peptide (ADNF‐14) derived from ADNF has been discovered that mimics the survival‐promoting action of the parent protein. These studies support the conclusion that VIP, PACAP, and associated molecules are both important regulators of neurodevelopment and strong candidates for therapeutic development for the treatment of neurodegenerative disease.

Activity-dependent neurotrophic factor (ADNF) is a glia-derived protein that is neuroprotective at femtomolar concentrations. A 14-amino acid peptide of ADNF (ADNF-14) has been reported that protects cultured neurons from multiple neurotoxins. Structure-activity relationships of peptides related to ADNF-14 now have been determined. A 9-amino acid core peptide (ADNF-9) has been identified that has greater potency and a broader effective concentration range (10(-16) to 10(-13) M) than ADNF or ADNF-14 in preventing cell death associated with tetrodotoxin treatment of cerebral cortical cultures. Deletions or conservative amino acid substitutions to ADNF-9 resulted in reduced potency, narrower effective concentration range and/or decreased efficacy. Removal of the N-terminal serine or the COOH-terminal isoleucine-proline-alanine from ADNF-9 produced a significant reduction in survival-promoting activity. Comparative studies of ADNF-9 action in mixed (glia plus neurons) vs. glia-depleted neuronal cultures indicated that ADNF-9 can act directly on neurons, although the potency of the peptide was 10,000-fold greater in mixed cultures. Kinetic studies showed that exposure to ADNF-9 for only 2 hr was sufficient to produce a 4-day protection against the cell-killing action of tetrodotoxin. Treatment with bafilomycin A1 (an inhibitor of receptor-mediated endocytosis) for 2 hr prevented the ADNF- and ADNF-9-mediated neuroprotection. ADNF-9, like ADNF-14, was neuroprotective against N-methyl-D-aspartate and the beta-amyloid peptide (amino acids 25-35), and had a much broader range of effective concentrations than ADNF-14. These studies identify ADNF-9 as an attractive lead compound for the development of therapeutic agents against neurodegenerative diseases.

Neuropeptides generally exhibit multiple roles in the maintenance of homeostasis. Classically, neuropeptides were found to exert neurohormonal and neurotransmitter (neuromodulator) effects in the central and peripheral nervous system. It is becoming increasingly apparent that neuropeptides also act as regulators of cell division, differentiation, and survival. In the past we reviewed neuropeptides as growth factors.'Z2 Recent studies indicate that neuronal cells, in response to axotomy, increase expression of neuropeptides that are associated with the promotion of survival and regeneration (see ref. 3 for review). Examples include corticotropin-releasing factor, dynorphin, calcitonin gene-related peptide, cholecystokinin, galanin, vasoactive intestinal peptide (VIP), neuropeptide Y, and others. At the same time, down-regulation of neurotransmitters and substances related to the secretion of neurotransmitters may occur. Similarly, neurotrophins of the family of nerve growth factors (NGF) increase the expression of specific neuropeptides, for example, brain-derived neurotrophic factor enhanced neuropeptide Y and somatostatin levels in cortical neurons, whereas NGF did not.4 In the periph-

Single-neuron actions are the basis of brain function, as clinical sequelae, neuronal dysfunction or failure for most of the central nervous system (CNS) diseases and injuries can be identified via tracing single-neurons. The bulk analysis methods tend to miscue critical information by assessing the population-averaged outcomes. However, its primary requisite in neuroscience to analyze single-neurons and to understand dynamic interplay of neurons and their environment. Microfluidic systems enable precise control over nano-to femto-liter volumes via adjusting device geometry, surface characteristics, and flow-dynamics, thus facilitating a well-defined micro-environment with spatio-temporal control for single-neuron analysis. The microfluidic platform not only offers a comprehensive landscape to study brain cell diversity at the level of transcriptome, genome, and/or epigenome of individual cells but also has a substantial role in deciphering complex dynamics of brain development and brain-related disorders. In this review, we highlight recent advances of microfluidic devices for single-neuron analysis, i.e., single-neuron trapping, single-neuron dynamics, single-neuron proteomics, single-neuron transcriptomics, drug delivery at the single-neuron level, single axon guidance, and single-neuron differentiation. Moreover, we also emphasize limitations and future challenges of single-neuron analysis by focusing on key performances of throughput and multiparametric activity analysis on microfluidic platforms.

Stearyl-Nle17-VIP (SNV) is a novel agonist of vasoactive intestinal peptide (VIP) exhibiting a 100-fold greater potency than the parent molecule and specificity for a receptor associated with neuronal survival. Here, mice deficient in apolipoprotein E (ApoE), a molecule associated with the etiology of Alzheimer's disease, served as a model to investigate the developmental and protective effects of SNV. In comparison to control animals, the deficient mice exhibited (a) reduced amounts of VIP messenger RNA; (b) decreased cholinergic activity (c) significant retardation in the acquisition of developmental milestones: forelimb placing behavior and cliff avoidance behavior; and (d) learning and memory impairments. Daily injections of SNV to ApoE-deficient newborn pups resulted in increased cholinergic activity and marked improvements in the time of acquisition of behavioral milestones, with peptide-treated animals developing as fast as control animals and exhibiting improved cognitive functions after cessation of peptide treatment. Specificity was demonstrated in that treatment with a related peptide (PACAP), pituitary adenylate cyclase-activating peptide, produced only limited amelioration. As certain genotypes of ApoE increase the probability of Alzheimer's disease, early counseling and preventive treatments may now offer an important route for therapeutics design.

The effects of pituitary adenylate cyclase activating polypeptide (PACAP) hybrid, a synthetic antagonist, was investigated on NIH/3T3 cells containing PACAP receptor (R) splice variants (SVs). PACAPhybrid inhibited 125I-PACAP-27 binding to NIH/3T3 cells stably expressing PACAP-R basic, SV-1, SV-2 or SV-3 with an IC50 of 1000 nM. PACAPhybrid antagonized the ability of PACAP-27 to elevate cAMP regardless of the PACAP-R SV used. PACAP was more efficacious at increasing cytosolic Ca2+ in NIH/3T3 cells containing PACAP-R SV-2 than PACAP-R basic, SV-1 or SV-3. PACAPhybrid antagonized the increase in cytosolic Ca2+ caused by PACAP-27 regardless of the PACAP-R SV used. PACAP was more potent at elevating c-fos mRNA using NIH/3T3 cells transfected with PACAP-R SV-2 than PACAP-R basic, SV-1 or SV-3. PACAPhybrid antagonized the increase in c-fos mRNA caused by PACAP-27. These data suggest that PACAPhybrid is a useful PACAP receptor antagonist for PACAP-R SVs.

Annals of the New York Academy of SciencesVolume 814, Issue 1 p. 161-166 Neuropeptides and Neuronal Survival: Neuroprotective Strategy for Alzheimer's Diseasea I. GOZES, Corresponding Author I. GOZES Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, Israele Corresponding author.Search for more papers by this authorA. BARDEA, A. BARDEA Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorM. BECHAR, M. BECHAR Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorO. PEARL, O. PEARL Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. RESHEF, A. RESHEF Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorR. ZAMOSTIANO, R. ZAMOSTIANO Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. DAVIDSON, A. DAVIDSON Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorS. RUBINRAUT, S. RUBINRAUT Department of Organic Chemistry Weizmann Institute of Science Rehovot 76100, IsraelSearch for more papers by this authorE. GILADI, E. GILADI Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorM. FRIDKIN, M. FRIDKIN Department of Organic Chemistry Weizmann Institute of Science Rehovot 76100, IsraelSearch for more papers by this authorD. E. BRENNEMAN, D. E. BRENNEMAN Section on Developmental and Molecular Pharmacology Laboratory of Developmental Neurobiology National Institute of Child Health and Human Development National Institutes of Health Bethesda, Maryland 20892Search for more papers by this author I. GOZES, Corresponding Author I. GOZES Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, Israele Corresponding author.Search for more papers by this authorA. BARDEA, A. BARDEA Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorM. BECHAR, M. BECHAR Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorO. PEARL, O. PEARL Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. RESHEF, A. RESHEF Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorR. ZAMOSTIANO, R. ZAMOSTIANO Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorA. DAVIDSON, A. DAVIDSON Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorS. RUBINRAUT, S. RUBINRAUT Department of Organic Chemistry Weizmann Institute of Science Rehovot 76100, IsraelSearch for more papers by this authorE. GILADI, E. GILADI Department of Clinical Biochemistry Sackler School of Medicine Tel Aviv University Tel Aviv 69978, IsraelSearch for more papers by this authorM. FRIDKIN, M. FRIDKIN Department of Organic Chemistry Weizmann Institute of Science Rehovot 76100, IsraelSearch for more papers by this authorD. E. BRENNEMAN, D. E. BRENNEMAN Section on Developmental and Molecular Pharmacology Laboratory of Developmental Neurobiology National Institute of Child Health and Human Development National Institutes of Health Bethesda, Maryland 20892Search for more papers by this author First published: 17 December 2006 https://doi.org/10.1111/j.1749-6632.1997.tb46154.xCitations: 15 a This research was supported in part by the Fujimoto Corporation and the U.S.-Israel Binational Science Foundation. ‡ This paper was written while I.G. was a scholar-in-residence at the Fogarty International Center for Advanced Study in the Health Sciences, National Institutes of Health, Bethesda, Maryland. Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Citing Literature Volume814, Issue1Neuropeptides in Development and AgingApril 1997Pages 161-166 RelatedInformation

Activity-dependent neurotrophic factor (ADNF) is a glia-derived protein that is neuroprotective at femtomolar concentrations. ADNF is released from astroglia after treatment with 0.1 nM vasoactive intestinal peptide (VIP). To further assess the biological role of ADNF, antiserum was produced following sequential injections of purified ADNF into mice. Anti-ADNF ascites fluid (1:10,000) decreased neuronal survival by 45–55% in comparison to untreated cultures or those treated with control ascites. The neuronal death after anti-ADNF treatment was observed in cultures derived from the spinal cord, hippocampus or cerebral cortex at similar IC50's. Using a terminal deoxynucleotidyl transferase in situ assay to estimate apoptosis in cerebral cortical cultures, anti-ADNF was shown to produce a 70% increase in the number of labeled cells in comparison to controls. In spinal cord cultures, anti-ADNF treatment produced a 20% decrease in choline acetyltransferase activity in comparison to controls. Neuronal cell death produced by the antiserum to ADNF was prevented in cultures co-treated with purified ADNF or ADNF-15, an active peptide derived from the parent ADNF. In vitro binding between the anti-ADNF and ADNF-15 was demonstrated with size exclusion chromatography. Comparative studies with other growth factors (insulin-like growth factor-1, platelet-derived growth factor, nerve growth factor, epidermal growth factor, ciliary neurotrophic growth factor, and neurotrophin-3) demonstrated that only ADNF prevented neuronal cell death associated with electrical blockade. These investigations indicated that an ADNF-like substance was present in cultures derived from multiple locations in the central nervous system and that ADNF-15 exhibited both neuroprotection and immunogenicity. ADNF appears to be both a regulator of activity-dependent neuronal survival and a neuroprotectant. ©1997 Elsevier Science B.V. All rights reserved.

To distinguish vasoactive intestinal peptide (VIP) receptors in the brain-mediating neurotransmission and neurotrophism, potent VIP analogues were designed. Using a single amino acid substitution and the addition of a fatty acyl moiety, an analogue was devised that exhibited both a 100-fold greater potency than VIP and specificity for a VIP receptor associated with neuronal survival. This VIP agonist increased neuronal survival via a cAMP-independent mechanism. Identical chemical modification of a prototype VIP antagonist (Met-Hybrid, Neurotensin6-11-VIP7-28) also resulted in a 100-fold greater potency in blocking VIP-mediated increases in neuronal survival. Blockade of circadian activity rhythms was limited to VIP antagonists that could inhibit VIP-mediated increases in cAMP. These lipophilic peptides provide novel tools in receptor discrimination and drug design.