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Biomedical subjects

V M Wiegant

Publications and source records attributed to V M Wiegant.

At least 91 records · Page 5Linked to original sources
PubMed1993

Topography and characteristics of specific binding sites for non-opioid gamma-type endorphins in the rat brain as studied by autoradiography with [35S]Met-desenkephalin-gamma-endorphin.

An in vitro autoradiographic study was performed to characterize specific rat brain binding sites for non-opioid neuroleptic-like gamma-type endorphins, using [35S]Met-des-enkephalin-gamma-endorphin ([35S]Met-DE gamma E; [35]S-beta-endorphins(5-17)) with high specific activity as radioligand. The binding sites appeared to be confined to rat forebrain regions, e.g., orbital cortex, frontal cortex, cingulate cortex, piriform cortex, nucleus accumbens, amygdala, mediodorsal nucleus of the thalamus and arcuate and periventricular nuclei of the hypothalamus. These regions are part of the mesocorticolimbic feedback circuit. Densitometric analysis of the autoradiographs revealed that the density of the binding sites was highest in the mediodorsal nucleus of the thalamus and the amygdala. Concentration-dependent displacement of [35S]Met-DE gamma E (500 pM) with DE gamma E yielded an IC50 of 0.6 nM whereas DE alpha E (beta-endorphin(6-16)) had an IC50 of 210 nM. Various endorphins, sharing the gamma-endorphin C terminus, displaced [35S]Met-DE gamma E to the same extent as non-labelled DE gamma E (at 10(-6) M) whereas non-endorphin peptides did not show displacing capacity. Possible relationships of the binding sites with opioid receptors were investigated. DAMGO (mu) and DPDPE (delta) displaced [35S]Met-DE gamma E to some extent at 10(-6) M whereas U69,593 (kappa) was inactive, suggesting that the binding sites for gamma-type endorphins may resemble mu- and delta-opioid receptors in some aspects. Similarly, relationships with dopamine receptors were investigated. Haloperidol partially displaced [35S]Met-DE gamma E whereas sulpiride, SKF38,393 and 3-PPP at 10(-6) M did not induce significant displacement. Thus, binding sites are distinct from dopamine receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Amygdala↗
PubMed1993

Lesions of the suprachiasmatic nucleus indicate the presence of a direct vasoactive intestinal polypeptide-containing projection to gonadotrophin-releasing hormone neurons in the female rat.

In non-seasonal breeders like the rat, the influence of the suprachiasmatic nucleus (SCN) on reproduction is most clearly expressed in the female. Complete lesions of the SCN induce persistent oestrus (anovulation) in intact female rats, whereas oestrogen implantation in ovariectomized rats results in daily luteinizing hormone surges. Vasoactive intestinal polypeptide (VIP), a peptide synthesized in cell bodies of the SCN, inhibits the increase in pulsatile luteinizing hormone release observed in ovariectomized female rats. In search of the anatomical basis for these observations, the present study employs an immunocytochemical double staining for VIP and gonadotrophin-releasing hormone (GnRH) at the light microscopical level. It was demonstrated that approximately 45% of the GnRH positive neurons in the diagonal band of Broca, the preoptic and anterior hypothalamic area of female rats are innervated by VIP-containing processes. To investigate whether these VIP-containing fibres represent a direct projection of the SCN to the GnRH system, unilateral thermic SCN lesions were made. Lesions that unilaterally destroyed the majority of the VIP synthesizing cells in the SCN resulted in at least a 50% decrease of the VIP innervation of GnRH cell bodies at the lesioned side compared to the intact side. Lesions not affecting the VIP synthesizing cell population in the SCN did not change the percentage of GnRH neurons innervated by VIP-containing fibres, while partial lesions resulted in intermediate effects. These results indicate that the majority of the light microscopical VIP-containing input on GnRH neurons in the hypothalamus is derived from the SCN.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Evidence for a role of protein kinase C substrate B-50 (GAP-43) in Ca(2+)-induced neuropeptide cholecystokinin-8 release from permeated synaptosomes.

To study the involvement of the protein kinase C (PKC) substrate B-50 [also known as growth-associated protein-43 (GAP-43), neuromodulin, and F1] in presynaptic cholecystokinin-8 (CCK-8) release, highly purified synaptosomes from rat cerebral cortex were permeated with the bacterial toxin streptolysin O (SL-O). CCK-8 release from permeated synaptosomes, determined quantitatively by radioimmunoassay, could be induced by Ca2+ in a concentration-dependent manner (EC50 of approximately 10(-5) M). Ca(2+)-induced CCK-8 release was maximal at 10(-4) M Ca2+, amounting to approximately 10% of the initial 6,000 +/- 550 fmol of CCK-8 content/mg of synaptosomal protein. Only 30% of the Ca(2+)-induced CCK-8 release was dependent on the presence of exogenously added ATP. Two different monoclonal anti-B-50 antibodies were introduced into permeated synaptosomes to study their effect on Ca(2+)-induced CCK-8 release. The N-terminally directed antibodies (NM2), which inhibited PKC-mediated B-50 phosphorylation, inhibited Ca(2+)-induced CCK-8 release in a dose-dependent manner, whereas the C-terminally directed antibodies (NM6) affected neither B-50 phosphorylation nor CCK-8 release. The PKC inhibitors PKC19-36 and 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7), which inhibited B-50 phosphorylation in permeated synaptosomes, had no effect on Ca(2+)-induced CCK-8 release. Our data strongly indicate that B-50 is involved in the mechanism of presynaptic CCK-8 release, at a step downstream of the Ca2+ trigger. As CCK-8 is stored in large dense-cored vesicles, we conclude that B-50 is an essential factor in the exocytosis from this type of neuropeptide-containing vesicle.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗
PubMed1993

Nasal administration of an ACTH(4-9) peptide analogue with dimethyl-beta-cyclodextrin as an absorption enhancer: pharmacokinetics and dynamics.

1. The systemic absorption and the neurotrophic effect of the metabolically stabilized ACTH (4-9) analogue, Org2766, were investigated following intranasal (i.n.) administration. 2. Without additives the nasal bioavailability of the peptide was in the order of 15 and 10% in rats and rabbits, respectively. The absorption could be improved by addition of a variety of absorption enhancers to the nasal preparation. The beta-cyclodextrin derivative, dimethyl-beta-cyclodextrin (DM beta CD) at a concentration of 5% (w/v) improved the absorption in rats about 5 fold from 13 +/- 4% (mean +/- s.d.) for administration of the peptide alone to 65 +/- 21%, and in rabbits 1 to 2 fold, from 10 +/- 6% to 17 +/- 8%. 3. The increased permeability of the rat nasal mucosa for Org2766 caused by DM beta CD in rats reversed substantially within 1 h. However, the nasal absorption had not yet completely returned to the level without enhancer. 4. S.c. administered Org2766 accelerated the functional recovery from peripheral nerve damage in rats. However, the peptide did not facilitate nerve repair following i.n. administration with DM beta CD, in spite of the fact that Org2766 was well absorbed. I.v. injection of Org2766 was also ineffective.

Absorption↗
PubMed1993

Analysis of proopiomelanocortin (POMC) messenger ribonucleic acid and POMC-derived peptides in human peripheral blood mononuclear cells: no evidence for a lymphocyte-derived POMC system.

A number of recent studies suggest that cells of the immune system, e.g. peripheral blood mononuclear cells (PBMC), can synthesize and process POMC and secrete POMC-derived peptides, such as ACTH and endorphins, upon immune and hormonal challenges. From this, it has been proposed that POMC-derived peptides originating from lymphoid cells can function as hormones, for instance in a lymphoid-adrenal axis. In view of the important physiological implications of this proposal, the present study was designed to investigate the expression of the POMC gene in human PBMC and the production by these cells of alpha-, beta-, and gamma-endorphins (alpha E, beta E, and gamma E) peptides that are established end products of the posttranslational processing of POMC. PBMC of individual donors were used uncultured (fresh cells) or cultured for 24 and 48 h in the presence and absence of Concanavalin-A (Con-A), bacterial lipopolysaccharide, phytohemagglutinin, or CRH, and vasopressin, conditions that reportedly stimulate POMC activity in those cells, to investigate the presence of POMC transcripts by analysis of total RNA with Northern blotting and the reverse transcriptase polymerase chain reaction (RT-PCR). Large scale preparations containing over 10(9) cells (fresh, cultured with and without Con-A) originating from several donors were examined for the presence of POMC transcripts by analysis of poly(A)+ RNA on Northern blots and for the presence of alpha E, beta E, and gamma E by gel filtration over Sephadex G-75 and reverse phase HPLC, followed by assay of the fractions in four endorphin RIA systems with different specificities. On the Northern blots of total RNA, no POMC transcripts were detectable. In poly(A)+ RNA preparations, no full-length POMC mRNA was found, and it was estimated that the concentration of POMC mRNA, if present, was below approximately 0.005 transcript/cell in Con-A-stimulated cells and still lower in unstimulated cells. In accord with literature data, an 800- to 900-nucleotide POMC transcript was detected in cultured PBMC, and the levels of this transcript were stimulated by Con-A. In all samples analyzed with RT-PCR, a transcript spanning most of exons 2 and 3 was detectable only on Southern blots of the RT-PCR product, but not on agarose gels stained with ethidium bromide. Chromatographic analysis of endorphin immunoreactivities in cell extracts revealed no qualitative differences between the immunoreactive profiles of fresh PBMC or PBMC cultured with or without Con-A.(ABSTRACT TRUNCATED AT 400 WORDS)

Base Sequence↗
PubMed1992

Characterization of the release of Met-enkephalin from isolated nerve terminals: release kinetics and cation-dependence.

The release of the neuropeptide Met-enkephalin (Met-ENK) from isolated nerve terminals (synaptosomes) of the rat forebrain was characterized with respect to the subcellular distribution, the release upon addition of various stimulatory agents, the release kinetics, the cation-dependence of release and the relationship between Met-ENK release and elevations of the intraterminal free Ca(2+)-concentration ([Ca]i). A highly specific radioimmunoassay was developed for determination of Met-ENK (H-Tyr-Gly-Gly-Phe-Met-OH). Truncated and elongated forms of Met-ENK, Leu-enkephalin, beta-endorphin and dynorphin displayed negligible cross-reactivity. Met-ENK-like immunoreactivity (Met-ENK-LI) is enriched in the purified synaptosomal fraction of rat forebrain homogenates and is released in a strictly Ca(2+)-dependent manner upon chemical depolarization or stimulation with the Ca2+ ionophore, ionomycin. A correlation exists between the release of Met-ENK-LI and the elevations of [Ca]i. Barium ions are able to replace Ca2+ in triggering Met-ENK-LI release. The release of Met-ENK-LI is initiated within 20 s after depolarization and is terminated after 3-5 min, although depolarization and [Ca]i elevation are maintained. At this time, > 90% of the initial Met-ENK-LI is still present inside the synaptosomes. Repolarization and renewed stimulation again evokes Ca(2+)-dependent release of this retained Met-ENK-LI. It is concluded that Met-ENK release from isolated nerve terminals is exocytotic, and that exocytosis is terminated by a regulatory mechanism in synaptosomes after 3-5 min of depolarization, a process which can be reversed by repolarization.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1992

Tetanus toxin and botulinum toxins type A and B inhibit glutamate, gamma-aminobutyric acid, aspartate, and met-enkephalin release from synaptosomes. Clues to the locus of action.

Tetanus toxin (100 nM) when preincubated with guinea pig cerebrocortical synaptosomes for 45 min reduces the final extent of the KCl-evoked, Ca(2+)-dependent, glutamate transmitter release to 30% of non-intoxicated controls. Similarly, 100 nM Botulinum neurotoxins, types A and B, preincubated for 90 min inhibit release to 45-60% of non-intoxicated controls. The toxins preferentially attenuate a slow phase of KCl-evoked glutamate release which may be associated with synaptic vesicle mobilization. Tetanus toxin additionally inhibits the release of aspartate, gamma-aminobutyric acid and met-enkephalin from the same preparation. Since amino acids and neuropeptides are released by distinct mechanisms, this indicates that the toxin affects a step common to both exocytotic pathways. When Ba2+ (which does not interact with calmodulin) is substituted for Ca2+, the control KCl-evoked release of each transmitter is unaffected and tetanus toxin is still inhibitory. Taken together these results implicate a calmodulin-independent locus (or loci) of action common to small- and large-dense-core vesicles and associated with vesicle transport.

Amino Acids↗
PubMed1992

Endorphin-like immunoreactivities in uncultured and cultured human peripheral blood mononuclear cells.

It has been reported that cells of the immune system produce and release considerable amounts of pro-opiomelanocortin (POMC) -derived peptides in response to coculture with a variety of stimulatory agents. The present study investigated whether extracts of human peripheral blood mononuclear cells (PBMC) contain immunoreactivity for beta-endorphin (beta E) and related peptides. Using four endorphin RIA systems with different specificities, extracts of freshly isolated PBMC and PBMC cultured in the presence or absence of mitogens or of corticotropin releasing factor (CRF) and vasopressin (VP), were analyzed. With a radioimmunoassay (RIA) system directed to the midportion of beta E, immunoreactivity (MP beta E-IR) was readily detectable, although the concentration was extremely low (ca. 200 pg/10(7) cells). beta E immunoreactivity (beta E-IR) and alpha-endorphin immunoreactivity (alpha E-IR), as determined in C-terminally directed RIA systems, were present in even lower concentrations. gamma-Endorphin immunoreactivity (gamma E-IR) was hardly detectable. Of subsets enriched in T-cells, B-cells or monocytes, the highest concentration of MP beta E-IR was detected in extracts of monocytes. Coculture of PBMC with the mitogen Concanavalin A (Con A) or Phytohaemagglutinin (PHA) increased the amount of MP beta E-IR in extracts of the cells. No increase in alpha E-IR, however, was detected, whereas beta E-IR was only increased in extracts of cells cultured in the presence of Con A. No increase, in any of the immunoreactivities, was observed in extracts of PBMC cultured with bacterial lipopolysaccharide (LPS) or with the combination of CRF and VP, both stimuli that have been reported to induce POMC peptides in cultured PBMC. The present data show that human PBMC contain endorphin-like immunoreactivity, but in very small amounts. The extremely low concentrations and the ineffectiveness of LPS and the combination of CRF and VP to increase the endorphin-like immunoreactivity raise questions about the reported capacity of PBMC to synthesize POMC-derived peptides.

Cells, Cultured↗
PubMed1992

Fc-mediated nonspecific staining of the porcine brain with rabbit antisera in immunocytochemistry is prevented by pre-incubation of the sera with proteins A and G.

Nonspecific staining was detected in immunocytochemical procedures on the porcine hypothalamus with rabbit antisera, irrespective of the antigen specificity of the sera, in magnocellular neurons of the paraventricular (PVN) and supraoptic nuclei (SON), and in the vasopressin- and oxytocin-containing nucleus (VON). The present study was designed to test the hypothesis that this staining is mediated by the Fc portion of rabbit immunoglobulins. Rabbit antisera against neuropeptides localized predominantly outside the PVN, SON, and VON were employed in combination with different detection methods. The intensity of the nonspecific staining varied depending on the antiserum and persisted after pre-absorption of the antisera with their homologous peptides. Nonspecific staining and antigen-specific staining were differentially affected by the method of tissue fixation. The nonspecific staining could be prevented by preincubation of the antisera with proteins A and G, which left the antigen-specific staining intact, whereas additional preabsorption with homologous peptide abolished all staining. These observations suggest that the Fc region of IgGs is indeed involved in the nonspecific staining. On press-blots of homogenates from SON tissue subjected to isoelectric focusing, one band in the low-pH region was found with all antisera. Pre-incubation of the antisera with protein A abolished the staining of this band but did not affect staining of antigen-specific bands. Pre-incubation with proteins A and G is proposed as a routine control to check for nonspecific staining mediated by the Fc region of IgGs in immunocytochemical procedures, particularly those that employ rabbit sera in porcine brain.

Animals↗
PubMed1991

The grasping response in rats: interaction between gamma-type endorphins and haloperidol.

gamma-Type endorphins mimic neuroleptics in inducing a grasping response in rats. It was studied whether the haloperidol-induced grasping response was altered after blockade of gamma-type endorphin activity in the rat brain. To achieve this blockade rats were injected i.c.v. with gamma-endorphin antiserum or with a monoclonal anti-idiotype desenkephalin-gamma-endorphin antibody, which may bio-inactivate the gamma-type endorphins or block the putative receptors for gamma-type endorphins, respectively. The results showed that both treatments attenuated the haloperidol-induced grasping response, particularly 3 h after haloperidol treatment. The influence of these antibodies appeared to be specific, since other sera were without effect. Thus there may be an interaction between the endogenous gamma-type endorphin activity and the haloperidol-induced grasping response.

Animals↗
PubMed1991

[Stress].

Any change in the social or physical environment that threatens the needs or interests of a mammal, induces a state of tension ('stress') in the animal. Driven by this tension, the animal changes its behaviour. At the same time, alterations occur in bodily functions. All organs and regulatory systems (central and peripheral nervous systems, endocrine and immune systems) participate in these behaviour-physiological response and are affected in their activities. The character of the reactions (e.g. fight, flight, passivity) is mainly determined by the animals appraisal of the environmental change and of the possible impact of its response. Aim of the reactions is neutralising the threat and gaining control over the environment (adaptation). When this is achieved, the tension recedes. In case the response is unsuccessful, the state of tension persists ('chronic stress'). Prolonged exposure of brain and/or bodily functions to stress can result in pathological states.

Animals↗
PubMed1991

Effect of anaesthetics on the release of beta-endorphin-immunoreactivity in rat plasma.

Analgesia and anaesthesia produced by fentanyl, urethane and ether, but not pentobarbital, occurred concomitantly with an increase in the concentration of plasma beta-endorphin like immunoreactivity (BEIR), probably of pituitary origin. This increase was not associated with significant changes in pituitary or brainstem beta-endorphin content. Pretreatment with naloxone caused a reduction in plasma BEIR increase following Hypnorm, ether and urethane; and in the analgesia following Hypnorm and urethane. Pentobarbital, alone or in combination with naloxone, did not increase the concentration of plasma beta-endorphin. These results may indicate participation of endogenous opioids in the mechanism of action of urethane.

Analgesics↗
PubMed1991

Differential effects of emotional and physical stress on the central and peripheral secretion of neurohypophysial hormones in male rats.

The effects of various stressful conditions on the levels of oxytocin (OT) and vasopressin (VP) in plasma and cisternal cerebrospinal fluid (CSF) of male rats were investigated. Three experimental models were used: exposure to a novel environment for 5 min, immobilization for 15 min, and ether inhalation for 10 min resulting in anaesthesia. Novelty and immobilization induced a slight but significant increase in OT levels in the CSF immediately after the stress. The effect of ether was considerably more pronounced. The concentration of VP in the CSF was elevated only by ether stress. In plasma, the level of OT was increased immediately following immobilization and ether stress but not after novelty stress, whereas VP only showed a delayed response 20 min after immobilization. These results indicate a rapid preferential release of OT in the periphery in response to physical and pharmacological stress. In addition, they provide evidence that release of OT into the CSF is triggered by physical, pharmacological as well as emotional stress, while the central release of VP is rather resistant to emotional stress. The data suggest that OT is a stress hormone in the central nervous system.

Animals↗
PubMed1991

Differential release of amino acids, neuropeptides, and catecholamines from isolated nerve terminals.

We have investigated transmitter release from small and large dense-core vesicles in nerve terminals isolated from guinea pig hippocampus. Small vesicles are found in clusters near the active zone, and large dense-core vesicles are located at ectopic sites. The abilities of Ca2+ channel activation and uniform elevation of Ca2+ concentration (with ionophores) to evoke secretion of representative amino acids, catecholamines, and neuropeptides were compared. For a given increase in Ca2+ concentration, ionophore was less effective than Ca2+ channel activation in releasing amino acids, but not in releasing cholecystokinin-8. Titration of the average Ca2+ concentration showed that the Ca2+ affinity for cholecystokinin-8 secretion was higher than that for amino acids. Catecholamine release showed intermediate behavior. It is concluded that neuropeptide release is triggered by small elevations in the Ca2+ concentration in the bulk cytoplasm, whereas secretion of amino acids requires higher elevations, as produced in the vicinity of Ca2+ channels.

Amino Acids↗
PubMed1991

Characterization of the release of cholecystokinin-8 from isolated nerve terminals and comparison with exocytosis of classical transmitters.

In the present study, the release of the neuropeptide cholecystokinin-8 (CCK) from purified nerve terminals (synaptosomes) of the rat hippocampus was characterized with respect to the subcellular distribution, the release upon addition of various agents, the release kinetics, the Ca2+ and ATP dependence of release, and the relationship between CCK release and elevations of intraterminal free Ca2+ concentration ([Ca]i). These characteristics were compared with those for the release of classical transmitters in similar preparations. CCK-like immunoreactivity (CCK-LI) is enriched in the purified synaptosomal fraction of hippocampus homogenates and released in a strictly Ca2(+)-dependent manner upon chemical depolarization, addition of 4-aminopyridine, or stimulation with the Ca2+ ionophore ionomycin. The presence of Ca2+ in the medium significantly stimulates the basal efflux of CCK-LI from synaptosomes. The release upon stimulation develops gradually in time with no significant release in the first 10 s and levels off after 3 min of depolarization. At this time, a large amount of CCK-LI is still present inside the synaptosomes. A correlation exists between the release of CCK-LI and the elevations of [Ca]i. The release of CCK-LI is decreased, but not blocked, upon ATP depletion. These characteristics markedly differ from those for classical transmitters, which show a fast component of Ca2(+)-dependent (exocytotic) release, an absolute dependence on cellular ATP, and no marked stimulation of basal efflux in the presence of Ca2+.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗
PubMed1990

Effects of chronic treatment with haloperidol and bromocriptine on the processing of beta-endorphin to gamma- and alpha-endorphin in discrete regions of the rat pituitary gland and brain.

beta-Endorphin is the putative precursor molecule of gamma- and alpha-endorphin. To investigate whether long-term changes in the activity of cells producing beta-endorphin are paralleled by alterations in the enzymatic processing of beta-endorphin, the effects of chronic treatment of rats with dopamine (DA) receptor ligands were examined on the content of immunoreactivity of beta-, gamma- and alpha-endorphin of dissected regions of the pituitary gland and the brain. Treatment with the DA receptor antagonist, haloperidol, resulted in a significant increase in the concentration of immunoreactivity for beta-, gamma-, and alpha-endorphin in the neurointermediate lobe, and of beta-endorphin in the anterior lobe of the pituitary gland. Levels of immunoreactivity of alpha-melanotropin and beta-endorphin in plasma were elevated, but those of corticosterone were decreased. This indicates that, in the intermediate lobe, both the biosynthetic and the secretory activity of cells producing beta-endorphin had increased, whereas in the anterior lobe, the secretory activity of beta-endorphin cells had decreased. No effects were observed on the ratios beta-endorphin/gamma-endorphin and beta-endorphin/alpha-endorphin in the intermediate lobe. In the anterior lobe however, the ratio beta-endorphin/alpha-endorphin had significantly increased. The effects of chronic treatment with the DA receptor agonist, bromocriptine, on levels of hormones in pituitary and plasma were opposite to those induced by haloperidol. In the brain, treatment with haloperidol selectively increased the content of immunoreactivity for beta-, gamma- and alpha-endorphin of the hypothalamus and the hippocampus and did not affect levels of peptides in the other regions of the brain studied.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Isoproterenol-stimulated release of beta-endorphin and related peptides from the rat pituitary neurointermediate lobe in vitro: evidence for preferential release of certain molecular forms of beta-endorphin.

The intermediate lobe of the pituitary gland synthesizes the multifactorial precursor molecule pro-opiomelanocortin (POMC), from which, through a process of post-translational enzymatic processing, beta-endorphin-(1-31) (beta E) and a variety of N alpha-acetylated and C-terminally shortened forms of this peptide are generated. Using an in vitro superfusion system, the release of these endorphins from intact rat neurointermediate lobes (NILs) was investigated under basal and isoproterenol (ISO) stimulated conditions. Superfusion of NILs with the beta-adrenergic agonist ISO (30 min pulse) resulted in a rapid, sustained and concentration-dependent stimulation of the release of beta E-like immunoreactivity (beta E-IR) over basal as determined with an antiserum directed against the C-terminus of the beta E- (1-31) sequence (10(-6) M: + 145%; 10(-7) M: + 73%; 10(-8) m: + 41%). The release of N(alpha)-acetylated-endorphin-like immunoreactivity (AcE-IR) was stimulated to a similar extent. These effects of ISO were antagonized by the competitive alpha-adrenoceptor antagonist propranolol in a concentration-dependent manner, indicating the involvement of alpha-adrenoceptors. The beta-related peptides released from the NILs under basal and ISO-stimulated conditions were further characterized, based on their retention times in a reversed-phase HPLC system and their reactivity with specific antisera recognizing respectively the midportion of beta E, the N-terminus of acetylated endorphins, the C-terminus of tau-endorphin (beta E-(1-17); tau E), or the C-terminus of alpha-endorphin (beta E-(1-16); alpha E). In HPLC fractionated superfusates 10 peaks were resolved that reacted with the midportion beta E antiserum. In superfusates collected under basal conditions, three major peaks possessed chromatographical and immunological characteristics of Ac beta E-(1-26), Ac beta E- (1-27) Ac beta E-(1-31). In addition, a prominent peak was found eluting around the retention time of beta E-(1-31), that contained both acetylated and non-acetylated material. Six smaller peaks were observed, with the characteristics of beta E-(1-26) and beta E-(1-27) (these peptides were not resolved with the HPLC system used), Ac tau E, tau E, Aa alpha E, and des-tyrosine-alpha E (DT alpha E), respectively. In superfusates collected during superfusion of NILs with ISO (10(-6) M) all peaks were increased. However, those eluting as beta E-(1-31), beta E-(1-26)/beta E-(1-27), Ac beta E-(1-26) and Ac tau E appeared to be preferentially stimulated.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetylation↗