Search

This article has been officially withdrawn from the journal.

Background We demonstrate a method that includes colocalization studies to analyze cell suspensions after isolation and to characterize 3-dimensional grafts consisting of cells and matrix in vitro and in vivo.Materials and methods Neonatal rat cardiomyocytes were labelled by CFDA-SE after harvest. Cells in the isolated cell suspension, the embodied cells in the seeded scaffolds were characterized measuring features such as viability and distribution of the cell types.Results Selective cell count revealed high yields of viable cardiomyocytes. After seeding cells in collagen matrix, viability of the cells decreased gradually in the time process in vitro. Histology of implanted bioartificial myocardial tissue detected viable cardiomyocytes within the graft.Conclusion Using colocalization histology we could label and track cells within the bioartificial myocardial tissue graft in vitro and post implant and assess viability and distribution.

Neuropeptide Y1–36(NPY1–36) acts through Y1and Y2receptors while the C-terminal NPY fragments NPY18–36and N-acetyl[Leu28,31]pNPY24–36act only through the Y2receptor. We have investigated the effects of intracerebroventricular (i.c.v.) administration of NPY1–36, NPY18–36and N-acetyl[leu28,31]pNPY24–36on LH secretion in the ovariectomised (OVX) ewe. These peptides were administered into a lateral ventricle (LV) or the third ventricle (3V) of OVX ewes during the non-breeding and breeding seasons. Microinjections of NPY were also made into the preoptic area (POA) during both seasons to investigate the effects of NPY at the level of the GnRH cell bodies. Tamed sheep were fitted with 19 gauge guide tubes into the LV, 3V or the septo-preoptic area (POA). Jugular venous blood samples were taken every 10 min for 3 h. Sheep were then given NPY1–36(10 μg), NPY18–36(100 μg) or saline vehicle into the LV; N-acetyl[Leu28,31]pNPY24–36(100 μg), NPY1–36(10 μg or 100 μg), NPY18–36(10 μg or 100 μg) or saline vehicle into the 3V, or NPY1–36(1 μg, 5 μg, 10 μg) into the POA. Blood sampling continued for a further 3 h. LH was measured in plasma by radioimmunoassay.LV or 3V injection of 10 μg NPY1–36caused a small but significant (P<0·025) increase in the interval from the last pre-injection pulse of LH to the first post-injection LH pulse during the breeding season. Other LH pulse parameters were not significantly affected. NPY18–36did not produce any significant change in LH pulsatility when injected into the LV, and neither peptide had any effect on plasma prolactin or GH levels. There was a significant (P<0·01) reduction in LH pulse frequency after 3V injection of 10 μg and 100 μg NPY and 100 μg NPY18–36. Pulse amplitude was reduced by 3V administration of the Y2agonist, N-acetyl[Leu28–31]pNPY24–36 and 100 μ NPY18–36. When the amplitude of the first post-injection LH pulse was analysed, 10 μg NPY also had a significant (P<0·05) suppressive effect. During the non-breeding season, 100 μg NPY1–36(but not 10 μg) decreased (P<0·01) LH pulse frequency. LH pulse amplitude was significantly (P<0·01) decreased by 100 μg NPY18–36. Doses of 10 μg NPY1–36and 100 μg NPY18–36had greater inhibitory effects on pulse frequency during the breeding season but the suppressive effect of 100 μg NPY was similar between seasons. Microinjections of NPY into the POA decreased (P<0·01) average plasma LH levels during the non-breeding season at a dose of 10 μg but did not significantly affect pulse frequency or amplitude.We conclude that a substantial component of the inhibitory action of NPY on LH secretion in the absence of steroids is mediated by the Y2receptor. This inhibition is probably exerted by way of a presynaptic action on GnRH terminals in the median eminence as NPY does not modulate the frequency or amplitude of LH pulses at the level of the GnRH cell bodies in the POA.Journal of Endocrinology(1995) 147,565–579

Neuropeptide Y (NPY) is a 36-amino-acid, C-terminal amidated peptide that is found in bulbospinal pathways and can inhibit the release of the primary afferent C-fiber neurotransmitter, substance P. Based on these observations, the present studies examined the possible antinociceptive effects of this peptide and several NPY fragments after intrathecal administration in rats prepared with chronic intrathecal catheters. In the 52 degrees C hot plate test, NPY produced a dose-dependent elevation in the nociceptive threshold with a median effective dose of 1.1 nmol. The ordering of fragments' activity was: NPY greater than NPY16-36 greater than or equal to NPY19-36 greater than or equal to NPY14-36 greater than or equal to NPY18-36 much greater than NPY1-36-OH = NPY18-36-OH = 0. In the paw pressure test, NPY was not active, even at the highest doses examined (median effective dose greater than 20 nmol), whereas the C-terminal fragments retained their potency and produced significant increases in the pressure required to evoke escape (NPY18-36: median effective dose = 18.7 nmol). The rank ordering of activity in the paw pressure test was: NPY19-36 greater than or equal to NPY14-36 greater than or equal to NPY18-36 greater than or equal to NPY16-36 much greater than NPY = NPY18-36-OH = 0. Peptide YY, human pancreatic polypeptide and avian pancreatic polypeptide behave similarly to NPY.(ABSTRACT TRUNCATED AT 250 WORDS)

Homozygous (di/di) Brattleboro rats have normal hypothalamic levels of oxytocin and neurophysin I, but undetectable levels of neurophysin II and arginine-vasopressin (AVP)1. This defect has been presumed to be at the genomic or transcriptional level, as AVP messenger is reported to be drastically reduced, if not absent, from the hypothalamus of Brattleboro rats2. Recent studies suggest de novo production of various neuropeptides in the mammalian gonad3–7, including AVP7. We report here the detection and localization of immunoreactive (ir) AVP in the luteal cells of adult homozygous Brattleboro rats, and the modulation of this ovarian ir-AVP by gonadotropins. These findings are thus consistent with a tissue-specific defect of AVP expression in the magnocellular neurones of the Brattleboro rat, and suggest that a comparable defect does not occur in the ovaries of such animals.

Previous studies using a variety of opiate ligands have suggested the existence of several subclasses of opiate receptors in crude membrane fractions of rat brain, and a similar diversity in bovine adrenal medulla. To examine the receptor profile of bovine adrenal medulla in detail we have studied the binding of classical ligands for mu (μ), delta (δ) and kappa (k) opiate receptors. [3H]naloxone ([3H]NAL), [3H] morphine ([3H]MOR), [3H]D-Ala2-D-Leu5-enkephalin ([3H]DAL) and [3H]ethyl-ketocyclazocine ([3H]EKCZ) were used as tracers; unlabeled competitors were NAL, MOR, DAL and ketocyclazocine (KCZ). In adrenal medulla [3H]NAL was specifically bound with a hierarchy of displacement NAL > MOR > KCZ >> DAL. No specific binding of [3H]DAL or [3H]EKCZ was found; for [3H]MOR very low levels of binding were seen, with no displacement by NAL or DAL, inconsistent displacement by KCZ and substantial displacement by MOR with an ED50 of 1.5 nM. In parallel studies rat brain membranes bound each labeled ligand with affinity and specificity consistent with previously published reports. Identical results were obtained in membranes from both tissues prepared with a preincubation step including 100 mM Na+, suggesting that the results were not influenced by occupation of binding sites by endogenous ligands. We interpret these data as supporting the existence of opiate receptors of the μ subtype in bovine adrenal medulla. We find, however, no evidence of δ or k sites in this tissue.

Opiate receptor-active peptide fragments (exorphins) have been identified recently in casein1and gluten2hydrolysates, and morphine has been found in bovine and human milk3. To determine whether similar peptides or alkaloids occur in other foodstuffs, we have screened potential sources using a rat brain homogenate assay to detect opiate receptor activity. We report here that instant coffee powders from a variety of manufacturers compete with tiitiated naloxone for binding to opiate receptors in the rat brain membrane preparations, with no significant difference between normal and decaffeinated coffee. The receptor binding activity resembles that seen with opiate antagonists, in that there was no change in the half-maximal effective dose (ED50) in the presence of 100 mM Na+; on bioassay, the activity was similarly shown to be antagonistic and specific for opiate-induced inhibition of twitch. Preliminary characterization of the activity reveals that it has a molecular weight (MW) in the range 1,000–3,500, is heat-stable, ether-extractable, not modified by enzymatic digestion with papain, and clearly separable from caffeine and morphine on TLC. As its concentration in an average cup of coffee is five times the ED50, these data suggest that drinking coffee may be followed by effects mediated via opiate receptors, as well as effects of caffeine.