Frans Vandesande - Academia.edu (original) (raw)

Papers by Frans Vandesande

Research paper thumbnail of Immunocytochemical demonstration of cLHRH-II-containing neurones surrounding the infundibular tract in the quail

Biology of Reproduction, 1998

Research paper thumbnail of Effect of Recombinant Human Igf-I on Growth, Food Conversion and Fat Distribution in Relation to Hormone Plasma Levels in Broiler Chicks

Insulin-like Growth Factor-I (IGF-I) is known to be a growth promoting factor in mammals. This ef... more Insulin-like Growth Factor-I (IGF-I) is known to be a growth promoting factor in mammals. This effect also includes a shift from lipid to protein synthesis and IGF-I therefore has been proposed to function as a lean growth factor. In broilers selection has resulted in fast growing strains which however often show an undesired fat accumulation. In the following study, the effect of IGF-I on body weight, fat deposition and distribution, and on circulating levels of IGF-I, growth hormone (GH), tyroxine (T4) and triiodotyronine (T3) was examined. Different doses of IGF-I (0, 0.03, 0.1 and 0.3 mg/kg BW.day) were administered by mini-osmotic pumps in 40 broiler chicks of a commercial strain between 4 and 6 weeks of age since fat deposition is maximal during this period. Blood samples were taken prior to and 1 and 2 weeks after implantation. Plasma IGF-I, GH, T3 and T4 levels were measured by RIA. At the end of the experimental period all birds were killed, abdominal fat measured, leg and breast muscles dissected and fat extracted. Body weight and relative growth were not affected by IGF-I treatment. However, while fat content in breast and thigh muscles remain unchanged, % abdominal fat was decreased significantly (P < 0.01) with 0.3 mg IGF-I treatment. Plasma IGF-I levels increased, as expected, in a dose-dependent way, while GH did not change with treatment but decreased with increasing age in all groups. A transient decrease in plasma T4 and increase in T3 was observed, at least with the higher IGF-doses. After 2 weeks the effect of IGF on circulating thyroid hormone levels disappeared. Although no effect of IGF-I on body weight or growth rate in commercial broiler chicks was observed, a decrease in abdominal fat indicate a repartition of nutrients available for net energy accretion. The observation that only the abdominal fat depot was affected confirms many other reports indicating a higher flexibility in fat content for depot fat compared to intra-and intermuscular fat. Since T3 is extremely effective in reducing abdominal fat in chickens, the increased T3 levels following IGF-I administration may also be, partly or totally, responsible for the abdominal fat reduction. Further research into causes of the T3-IGF-I interaction and its consequences on animal growth are needed to elucidate this.

Research paper thumbnail of Bovine Neurophysins I, II and C: : New Methods for Their Purification and for the Production of Specific Antibodies

1. Bovine neurophysins were prepared by a modified method, in which a Biogel P-60 column was used... more 1. Bovine neurophysins were prepared by a modified method, in which a Biogel P-60 column was used. This yielded two neurophysin fractions, the first containing neurophysin 1 and small quantities of the other neurophysins, the second containing neurophysin I1 and C, and only traces of ncurophysin I. 2. Antibodies against neurophysin I. I1 and C were prepared by an original method, 5 yg in 100 p1 water of each of the two fractions were applied on a gel slab and separated by iso-electric focusing in a pH gradient 4-6. The separated bands were visualized with 8-aniline-1-naphthalene sulfonic acid, magnesium salt and strips respectively containing neurophysin I, TI or C were cut out. The neurophysin-containing strips were homogenized in complete Freund's adjuvant and injected into rabbits. 3. The specificity of the antisera were tested by immunocytochemistry and by radioimmunoassay. By this latter method, it was determined that cross-reactivity was less than 17". The cross-reaction, observed with the immunohistochemical method could be eliminated by differential absorption. 4. It was found that neurophysin C antisera were undistingiiishable from the neurophysin I1 antisera, while showing little cross-reactivity with the neurophysin I antisera. This suggests that in vivo neurophysin C is not a real neurophysin, or at least, that it is very similar to neurophysin 11. 5. Highly purified bovine neurophysins I, TI and C were prepared by an improved preparative iso-electric focusing method. By modifying a LKB Uniphor electrophoresis apparatus, the elution technique of Svendsen could be applied. This technique makes it possible to elute the proteins without switching off the voltage. The resolution of the technique is close to that offered by analytical gel isoelectric focusing.

Research paper thumbnail of Immunohistochemical localization of corticotropin-releasing factor (CRF)-like immunoreactivity in the hypothalamus of the newt, Triturus cristatus

Neuroscience Letters, 1984

The distribution of corticotropin-releasing factor (CRF) immunoreactivity was investigated in the... more The distribution of corticotropin-releasing factor (CRF) immunoreactivity was investigated in the hypothalamus and preoptic area of the newt by single and double immunocytochemical procedures. CRF immunopositive cell bodies were seen in the preoptic area (from the anterior wall of the preoptic recess to the dorsal parts of the preoptic nucleus) and in the tuberal portions of the posterior hypothalamus. Abundant nerve fibres are seen in the outer zone of the median eminence, while the pars nervosa lacks CRF-immunoreactivity. CRF immunoreactive material is seemingly separared from neurophysins-and somatostatin immunoreactive cell bodies and fibres. Among the numerous research topics concerning the morphology and the biochemistry of neuropeptides, the significance of the comparative aspect was recently stressed [6]. Of particular interest appears to be this kind of approach for the hypothalamus, the evolution of which in hypophysial regulation is widely investigated [1]. The recent identification of ovine corticotropin-releasing factor (CRF) [25, 26] prompted immunohistochemical analyses in the hypothalamus and extrahypothalamic areas of the brain in several mammals [2, 13, 16, 181. Few data are available on the distribution of corticotropin-releasing factor (CRF)like immunoreactivity in submammalian species [3, 17, 28, 29] as compared with the more complete pattern shown in mammals. Following previous immunocytochemical analyses for somatostatin [SRIF] neurophysins (NPH) [7, 8] and substance P (SP) [9], this paper reports the presence of CRF-like material in the preoptic area and in the posterior hypothalamus of the crested newt (Triturus cristatus carnifex Laur.). Normal adult specimens of both sexes of Triturus cristatus (14 animals) were used. The brains, fixed with Zamboni or Susa fluids, were embedded in paraffin and serially cut (sections 5-10 #m in thickness) according to transverse or sagittal planes. The sections were stained for CRF-like immunoreactivity according to: (a) the indirect immunofluorescence (IFL) method [5]; (b) the unlabeled antibody perox

Research paper thumbnail of Fluorescent two-dimensional difference gel electrophoresis and mass spectrometry identify age-related protein expression differences for the primary visual cortex of kitten and adult cat

Journal of Neurochemistry, 2003

The recent introduction of fluorescent two-dimensional difference gel electrophoresis, combined w... more The recent introduction of fluorescent two-dimensional difference gel electrophoresis, combined with mass spectrometry, has greatly simplified the analysis and identification of differentially expressed proteins by eliminating intergel variability. In this report, we describe the successful application of this functional proteomics approach to compare protein expression levels in visual cortical area 17 of adult cats and 30-day-old kittens, in order to identify proteins expressed in an age-related fashion. We identified 16 proteins that were more abundantly expressed in kitten striate cortex and 12 proteins with a pronounced expression in adult cat area 17. Among those isolated from kitten area 17 were proteins related to axon growth and growth cone guidance and to the formation of cytoskeletal filaments. Glial fibrillary acidic protein, as identified in adult cat area 17, has been implicated previously in the termination of the critical period for cortical plasticity in kittens. In situ hybridization experiments for two of the identified proteins, glial fibrillary acidic protein and collapsin response mediator protein 5, confirmed and extended their differential expression to the mRNA level. Our findings show that two-dimensional difference gel electrophoresis combined with mass spectrometry is a powerful approach that permits the identification of small protein expression differences correlated to different physiological conditions.

Research paper thumbnail of Immunoreactive material resembling vertebrate neuropeptides and neurophysins in the brain, suboesophageal ganglion, corpus cardiacum and corpus allatum of the dictyopteran Periplaneta americana L

Cell Tissue Res, 1984

... This may be due to tax-onomic difference or to differences of the antisera used. Other report... more ... This may be due to tax-onomic difference or to differences of the antisera used. Other reports on an oxytocin-like material in inverte-brates apply to the gastropods Achatinafulica (VanNoor-den et al. 1980) and Lymnaea stagnalis (Schot et al. 1981). ...

Research paper thumbnail of Identification and Quantification of Monoaminergic Neuromodulators in the Sub-Cortical Region of Cat Visual Cortex by Microbore Hplced and Protein Assay

Journal of Liquid Chromatography Amp Related Technologies, Mar 5, 2001

Research paper thumbnail of CRF-like immunoreactive system in the quail brain

Journal Fur Hirnforschung, Feb 1, 1986

CITATIONS 18 READS 45 4 authors, including:

Research paper thumbnail of Calcium binding proteins as molecular markers for cat geniculate neurons

Immunocytochemistry revealed that in the cat dorsal lateral geniculate nucleus (dLGN) almost all ... more Immunocytochemistry revealed that in the cat dorsal lateral geniculate nucleus (dLGN) almost all parvalbumin-positive cells are GABAergic and about 56% of the calbindin D-28K (calbindin-immunoreactive neurons are also GABA-positive. On the other hand, in the same nucleus, almost all GABAergic neurons contain parvalbumin, and about 89% of the GABA-immunoreactive neurons contain calbindin. Double-labeling with calbindin and parvalbumin revealed that approximately 50% of the immunoreactive neurons are double-stained. In the PGN, virtually all neurons are GABA and parvalbumin-positive. Only a few scattered cells were also calbindin-immunoreactive. These results show that GABAergic geniculate cells can be differentiated on the basis of their calcium-binding protein immunoreactivity. Four types of immunoreactive cells are described here: (1) cells positive for GABA, parvalbumin and calbindin, (2) cells positive for GABA and parvalbumin, but negative for calbindin, (3) cells negative for GABA and parvalbumin, but positive for calbindin, (4) cells negative for GABA, parvalbumin and calbindin.

Research paper thumbnail of Effect of sensory deafferentation on the GABAergic circuitry of the adult cat visual system

Neuroscience, 1998

ABSTRACT

Research paper thumbnail of Evidence for chicken GH as the only hypophyseal factor responsible for the stimulation of hepatic 5'-monodeiodination activity in the chick embryo

Reproduction, nutrition, development, 1989

The influence of an intravenous injection of chicken growth hormone (cGH), a total chicken pars d... more The influence of an intravenous injection of chicken growth hormone (cGH), a total chicken pars distalis (PD) extract, and a PD extract depleted of cGH by immunoadsorption was studied in the 18-d-old chick embryo. Plasma concentrations of triiodothyronine (T3), thyroxine (T4), and hepatic 5'-monodeiodination (5'-D) activity were measured. An injection of total PD extract raised plasma T3, T4, and 5'-D activity, whereas a PD extract depleted of GH only increased plasma T4. The amount of cGH present in the PD extracts, as measured by homologous cGH radioimmunoassay, increased T3 and raised liver 5'-D, but had no effect on plasma T4. The effect on liver 5'-D was more pronounced with cGH than with a total PD extract, whereas the effect on plasma T3 was somewhat less pronounced. It was concluded that cGH increased the peripheral conversion of T4 into T3 in the chick embryo, whereas a PD extract depleted of cGH was purely thyrotropic. The PD extract also seemed to have...

Research paper thumbnail of Neurotensin-Like immunoreactivity in the pituitary and hypothalamus of bony fishes

Peptides, 1987

Using an antiserum raised against synthetic neurotensin (NT), the distribution of immunoreactivit... more Using an antiserum raised against synthetic neurotensin (NT), the distribution of immunoreactivity in the pituitary and hypothalamus has been examined by immunocytochemistry at light and electron microscope level in a number of species of bony fishes. In most species immunoreactive perikarya were found in the preoptic region of the hypothalamus, with fibres throughout the tuberal hypothalamus and neurohypophysis (neural lobe and median eminence). In the neurohypophysis of teleosts NT-like immunoreactivity was seen in a dense band of fibres bordering the ACTH cells of the rostral pars distalis: absorption controls showed that this was due to the presence of an NT(8-13)-like or xenopsin-like sequence, which, according to electron microscopic observations, was contained in small dense cored vesicles. The antiserum also stained the pituitary ACTH cells of some species, apparently due to cross-reaction with the 17-19 sequence of ACTH. These results suggest that an NT-like peptide may have a role in control of the adenohypophysis in fishes.

Research paper thumbnail of Calbindin D-28K and parvalbumin immunoreactivity is confined to two separate neuronal subpopulations in the cat visual cortex, whereas partial coexistence is shown in the dorsal lateral geniculate nucleus

Neuroscience Letters, 1989

ABSTRACT

Research paper thumbnail of Distribution of immediate early gene zif-268, c-fos, c-jun and jun-D mRNAs in the adult cat with special references to brain region related to vision

Neuroscience Letters, 1994

The distribution of immediate early gene zif-268, c-fos, c-jun and jun-D mRNAs was investigated i... more The distribution of immediate early gene zif-268, c-fos, c-jun and jun-D mRNAs was investigated in the visual cortex, dorsal lateral geniculate nucleus and hippocampus of the adult cat brain with in situ hybridization. In area 17, zif-268, c-jun and jun-D were found predominantly in layers II-III and VI, while c-fos mRNA was abundant in layer VI. In area 18, the zif-268, c-fos and c-jun labelling pattern was identical to that of area 17, this was not true for jun-D. In area 19, only c-jun retained the lamination pattern of areas 17 and 18, while zif-268, c-fos and jun-D were homogeneously distributed. In the dorsal lateral geniculate nucleus, only c-fos and jun-D resulted in labelling. In the pyramidal layer of hippocampus, zif-268 was found in CA1-4, c-jun in CA1-3, and jun-D in CA2-4. In the dentate gyrus, c-jun was abundant, jun-D moderate and zif-268 faint. C-fos labelling was absent in the hippocampal formation.

Research paper thumbnail of Long-term effects of retinal lesions on growth-associated protein 43 (GAP-43) expression in the visual system of adult cats

Neuroscience Letters, 1996

We have investigated the role of growth-associated protein 43 in synaptic reorganization in the v... more We have investigated the role of growth-associated protein 43 in synaptic reorganization in the visual system of adult cats that received binocular central retinal lesions. Different survival times between 3 and 8 months after induction of the lesion were chosen. In the deafferented part of the dorsal lateral geniculate nucleus (dLGN) we found a long-lasting increase in GAP-43 protein, while glial fibfillaty acidic protein (GFAP) immunoreactivity, which initially increased due to the degeneration of retinal ganglion ceils, slowly subsided over this period. In area 17, the pattern of GAP-43 expression did not provide indications for morphological changes in the cortical architecture following retinal lesions.

Research paper thumbnail of Validation of a new antiserum directed towards the synthetic c-terminus of the FOS protein in avian species: immunological, physiological and behavioral evidence

Journal of Neuroscience Methods, 1999

In the past 10 years, the study of the expression of immediate early genes, such as c-fos, in the... more In the past 10 years, the study of the expression of immediate early genes, such as c-fos, in the brain has become a common method for the identification of brain areas involved in the regulation of specific physiological and behavioral functions. The use of this method in avian species has been limited by the paucity of suitable antibodies that cross-react with the FOS protein in birds. We describe in this paper the preparation of an antibody directed against a synthetic fragment of the protein product of the c-fos gene in chickens (Gallus domesticus). We demonstrate that this new antibody can be used in several avian species to study FOS expression induced by a variety of pharmacological, physiological and behavioral stimuli. Western blot studies indicated that this antibody recognizes a protein of the expected size (47 kDa) but also cross reacts to some extent with proteins of lower molecular weight that share sequence homology with FOS (Fos-related antigens). FOS immunocytochemistry was performed with this antibody in four species of birds in three different laboratories utilizing diverse variants of the immunocytochemical procedure. In all cases the antibody provided a reliable identification of the FOS antigen. The new antibody described here appears to be suitable for the study of FOS expression in several different avian species and situations. It is available in substantial amounts and will therefore make it possible to use FOS expression as a tool to map brain activity in birds as has now been done for several years in mammalian species. : S 0 1 6 5 -0 2 7 0 ( 9 9 ) 0 0 0 6 7 -9

Research paper thumbnail of A new alternative for simultaneous immunohistochemical screening of 96 hybridoma clones for tissue-specific antibody production selects a monoclonal antibody to insect corpus cardiacum

Journal of Neuroscience Methods, 1986

In the current search for the elucidation of the true structure of hitherto unidentified &#39... more In the current search for the elucidation of the true structure of hitherto unidentified 'new' insect neuropeptides we designed a novel screening method to facilitate the primary detection of neurone-specific antibody secreting mouse-mouse hybridoma clones obtained after immunization with neuronal tissue homogenates. The present procedure is principally adapted from a conventional immunohistological test and enables one to rapidly screen 96 (and even more) clones at one time for potential secretion of specific antibodies to different tissue compounds, without the necessity of having a purified antigen. It has proved to be sensitive, rapid, practical and reproducible. As such it promises to be very useful to discriminate amongst the wide range of antibodies to various kinds of materials produced by hybridomas by detecting monoclonal antibodies directed against factors contained in well-defined tissues in which one is interested. This paper also reports the successful application of this method to a primary screening of clones producing murine monoclonal antibodies to substances of insect corpora cardiaca (CC), after immunization with crude antigen preparations.

Research paper thumbnail of Development of a novel screening device permitting immunocytochemical screening of numerous culture supernatants during hybridoma production

Journal of Immunological Methods, 1989

Research paper thumbnail of One-step purification of chicken growth hormone from a crude pituitary extract by use of a monoclonal immunoadsorbent

Journal of Endocrinology, 1988

The immunization of mice with an affinity-purified glycoprotein preparation from chicken pituitar... more The immunization of mice with an affinity-purified glycoprotein preparation from chicken pituitary tissue yielded several monoclonal antibodies towards the recently described glycosylated variant of chicken GH. As all these antibodies recognize the classical (non-glycosylated) GH molecule equally well, they provide a suitable tool for the development of both a specific immunoadsorbent and an assay method. This paper deals with the surprising purification power of the immunoadsorbent that was produced with one of the monoclonal antibodies. The resulting preparation was more than 99% pure as assessed by reversed phase high-performance liquid chromatography and sodium dodecyl sulphate-polyacrylamide gel electrophoresis, so that no further purification steps were needed before the determination of the amino acid sequence of the material. The efficiency of the purification protocol as determined by a homologous, monoclonal antibody-based radioimmunoassay was virtually absolute. Moreover, the affinity-purified GH preparation was a mixture representing the multiple molecular forms of pituitary chicken GH, including both oligomeres and glycosylated GH. The purified preparations were finally used to demonstrate the hepatic 5'-monodeiodinase-stimulating activity of GH in the chicken embryo (results not shown), in order to prove that the biological activity of the molecule had not been damaged by elution from the immunoadsorbent.

Research paper thumbnail of Effect of sensory deafferentation on immunoreactivity of GABAergic cells and on GABA receptors in the adult cat visual cortex

The Journal of Comparative Neurology, 1995

To investigate the effects of sensory deafferentation on the cortical GABAergic circuitry in adul... more To investigate the effects of sensory deafferentation on the cortical GABAergic circuitry in adult cats, glutamic acid decarboxylase (GAD) and gamma-aminobutyric acid (GABA) immunoreactivity and GABA receptor binding were studied in the visual cortex of normal cats and compared with cats that had received restricted binocular central lesions of the retina and had survived for 2 weeks postlesion in a normal visual environment. In the visual cortex of lesioned cats, two changes were observed in the number of GAD-immunoreactive elements in the regions affected by the retinal lesions: the number of GAD-positive puncta decreased, whereas that of GAD-immunoreactive somata increased. In contrast, no detectable changes were measured in the number of GABA-immunopositive somata or puncta. At the receptor level, we observed no differences in either the laminar distribution or the affinity of cortical GABAA and GABAB receptors labeled with [3H]-muscimol and [3H]-baclofen, respectively, in the lesioned versus normal cats. We present the hypothesis that sensory deafferentation in these adult cats (1) leads to a reduction of cortical GABAergic inhibition in the deafferented region, and (2) that this decreased inhibition may permit changes in efficiency of synapses and (3) that these changes may represent a first stage of events underlying the retinotopic reorganization preceeding the structural changes.

Research paper thumbnail of Immunocytochemical demonstration of cLHRH-II-containing neurones surrounding the infundibular tract in the quail

Biology of Reproduction, 1998

Research paper thumbnail of Effect of Recombinant Human Igf-I on Growth, Food Conversion and Fat Distribution in Relation to Hormone Plasma Levels in Broiler Chicks

Insulin-like Growth Factor-I (IGF-I) is known to be a growth promoting factor in mammals. This ef... more Insulin-like Growth Factor-I (IGF-I) is known to be a growth promoting factor in mammals. This effect also includes a shift from lipid to protein synthesis and IGF-I therefore has been proposed to function as a lean growth factor. In broilers selection has resulted in fast growing strains which however often show an undesired fat accumulation. In the following study, the effect of IGF-I on body weight, fat deposition and distribution, and on circulating levels of IGF-I, growth hormone (GH), tyroxine (T4) and triiodotyronine (T3) was examined. Different doses of IGF-I (0, 0.03, 0.1 and 0.3 mg/kg BW.day) were administered by mini-osmotic pumps in 40 broiler chicks of a commercial strain between 4 and 6 weeks of age since fat deposition is maximal during this period. Blood samples were taken prior to and 1 and 2 weeks after implantation. Plasma IGF-I, GH, T3 and T4 levels were measured by RIA. At the end of the experimental period all birds were killed, abdominal fat measured, leg and breast muscles dissected and fat extracted. Body weight and relative growth were not affected by IGF-I treatment. However, while fat content in breast and thigh muscles remain unchanged, % abdominal fat was decreased significantly (P < 0.01) with 0.3 mg IGF-I treatment. Plasma IGF-I levels increased, as expected, in a dose-dependent way, while GH did not change with treatment but decreased with increasing age in all groups. A transient decrease in plasma T4 and increase in T3 was observed, at least with the higher IGF-doses. After 2 weeks the effect of IGF on circulating thyroid hormone levels disappeared. Although no effect of IGF-I on body weight or growth rate in commercial broiler chicks was observed, a decrease in abdominal fat indicate a repartition of nutrients available for net energy accretion. The observation that only the abdominal fat depot was affected confirms many other reports indicating a higher flexibility in fat content for depot fat compared to intra-and intermuscular fat. Since T3 is extremely effective in reducing abdominal fat in chickens, the increased T3 levels following IGF-I administration may also be, partly or totally, responsible for the abdominal fat reduction. Further research into causes of the T3-IGF-I interaction and its consequences on animal growth are needed to elucidate this.

Research paper thumbnail of Bovine Neurophysins I, II and C: : New Methods for Their Purification and for the Production of Specific Antibodies

1. Bovine neurophysins were prepared by a modified method, in which a Biogel P-60 column was used... more 1. Bovine neurophysins were prepared by a modified method, in which a Biogel P-60 column was used. This yielded two neurophysin fractions, the first containing neurophysin 1 and small quantities of the other neurophysins, the second containing neurophysin I1 and C, and only traces of ncurophysin I. 2. Antibodies against neurophysin I. I1 and C were prepared by an original method, 5 yg in 100 p1 water of each of the two fractions were applied on a gel slab and separated by iso-electric focusing in a pH gradient 4-6. The separated bands were visualized with 8-aniline-1-naphthalene sulfonic acid, magnesium salt and strips respectively containing neurophysin I, TI or C were cut out. The neurophysin-containing strips were homogenized in complete Freund's adjuvant and injected into rabbits. 3. The specificity of the antisera were tested by immunocytochemistry and by radioimmunoassay. By this latter method, it was determined that cross-reactivity was less than 17". The cross-reaction, observed with the immunohistochemical method could be eliminated by differential absorption. 4. It was found that neurophysin C antisera were undistingiiishable from the neurophysin I1 antisera, while showing little cross-reactivity with the neurophysin I antisera. This suggests that in vivo neurophysin C is not a real neurophysin, or at least, that it is very similar to neurophysin 11. 5. Highly purified bovine neurophysins I, TI and C were prepared by an improved preparative iso-electric focusing method. By modifying a LKB Uniphor electrophoresis apparatus, the elution technique of Svendsen could be applied. This technique makes it possible to elute the proteins without switching off the voltage. The resolution of the technique is close to that offered by analytical gel isoelectric focusing.

Research paper thumbnail of Immunohistochemical localization of corticotropin-releasing factor (CRF)-like immunoreactivity in the hypothalamus of the newt, Triturus cristatus

Neuroscience Letters, 1984

The distribution of corticotropin-releasing factor (CRF) immunoreactivity was investigated in the... more The distribution of corticotropin-releasing factor (CRF) immunoreactivity was investigated in the hypothalamus and preoptic area of the newt by single and double immunocytochemical procedures. CRF immunopositive cell bodies were seen in the preoptic area (from the anterior wall of the preoptic recess to the dorsal parts of the preoptic nucleus) and in the tuberal portions of the posterior hypothalamus. Abundant nerve fibres are seen in the outer zone of the median eminence, while the pars nervosa lacks CRF-immunoreactivity. CRF immunoreactive material is seemingly separared from neurophysins-and somatostatin immunoreactive cell bodies and fibres. Among the numerous research topics concerning the morphology and the biochemistry of neuropeptides, the significance of the comparative aspect was recently stressed [6]. Of particular interest appears to be this kind of approach for the hypothalamus, the evolution of which in hypophysial regulation is widely investigated [1]. The recent identification of ovine corticotropin-releasing factor (CRF) [25, 26] prompted immunohistochemical analyses in the hypothalamus and extrahypothalamic areas of the brain in several mammals [2, 13, 16, 181. Few data are available on the distribution of corticotropin-releasing factor (CRF)like immunoreactivity in submammalian species [3, 17, 28, 29] as compared with the more complete pattern shown in mammals. Following previous immunocytochemical analyses for somatostatin [SRIF] neurophysins (NPH) [7, 8] and substance P (SP) [9], this paper reports the presence of CRF-like material in the preoptic area and in the posterior hypothalamus of the crested newt (Triturus cristatus carnifex Laur.). Normal adult specimens of both sexes of Triturus cristatus (14 animals) were used. The brains, fixed with Zamboni or Susa fluids, were embedded in paraffin and serially cut (sections 5-10 #m in thickness) according to transverse or sagittal planes. The sections were stained for CRF-like immunoreactivity according to: (a) the indirect immunofluorescence (IFL) method [5]; (b) the unlabeled antibody perox

Research paper thumbnail of Fluorescent two-dimensional difference gel electrophoresis and mass spectrometry identify age-related protein expression differences for the primary visual cortex of kitten and adult cat

Journal of Neurochemistry, 2003

The recent introduction of fluorescent two-dimensional difference gel electrophoresis, combined w... more The recent introduction of fluorescent two-dimensional difference gel electrophoresis, combined with mass spectrometry, has greatly simplified the analysis and identification of differentially expressed proteins by eliminating intergel variability. In this report, we describe the successful application of this functional proteomics approach to compare protein expression levels in visual cortical area 17 of adult cats and 30-day-old kittens, in order to identify proteins expressed in an age-related fashion. We identified 16 proteins that were more abundantly expressed in kitten striate cortex and 12 proteins with a pronounced expression in adult cat area 17. Among those isolated from kitten area 17 were proteins related to axon growth and growth cone guidance and to the formation of cytoskeletal filaments. Glial fibrillary acidic protein, as identified in adult cat area 17, has been implicated previously in the termination of the critical period for cortical plasticity in kittens. In situ hybridization experiments for two of the identified proteins, glial fibrillary acidic protein and collapsin response mediator protein 5, confirmed and extended their differential expression to the mRNA level. Our findings show that two-dimensional difference gel electrophoresis combined with mass spectrometry is a powerful approach that permits the identification of small protein expression differences correlated to different physiological conditions.

Research paper thumbnail of Immunoreactive material resembling vertebrate neuropeptides and neurophysins in the brain, suboesophageal ganglion, corpus cardiacum and corpus allatum of the dictyopteran Periplaneta americana L

Cell Tissue Res, 1984

... This may be due to tax-onomic difference or to differences of the antisera used. Other report... more ... This may be due to tax-onomic difference or to differences of the antisera used. Other reports on an oxytocin-like material in inverte-brates apply to the gastropods Achatinafulica (VanNoor-den et al. 1980) and Lymnaea stagnalis (Schot et al. 1981). ...

Research paper thumbnail of Identification and Quantification of Monoaminergic Neuromodulators in the Sub-Cortical Region of Cat Visual Cortex by Microbore Hplced and Protein Assay

Journal of Liquid Chromatography Amp Related Technologies, Mar 5, 2001

Research paper thumbnail of CRF-like immunoreactive system in the quail brain

Journal Fur Hirnforschung, Feb 1, 1986

CITATIONS 18 READS 45 4 authors, including:

Research paper thumbnail of Calcium binding proteins as molecular markers for cat geniculate neurons

Immunocytochemistry revealed that in the cat dorsal lateral geniculate nucleus (dLGN) almost all ... more Immunocytochemistry revealed that in the cat dorsal lateral geniculate nucleus (dLGN) almost all parvalbumin-positive cells are GABAergic and about 56% of the calbindin D-28K (calbindin-immunoreactive neurons are also GABA-positive. On the other hand, in the same nucleus, almost all GABAergic neurons contain parvalbumin, and about 89% of the GABA-immunoreactive neurons contain calbindin. Double-labeling with calbindin and parvalbumin revealed that approximately 50% of the immunoreactive neurons are double-stained. In the PGN, virtually all neurons are GABA and parvalbumin-positive. Only a few scattered cells were also calbindin-immunoreactive. These results show that GABAergic geniculate cells can be differentiated on the basis of their calcium-binding protein immunoreactivity. Four types of immunoreactive cells are described here: (1) cells positive for GABA, parvalbumin and calbindin, (2) cells positive for GABA and parvalbumin, but negative for calbindin, (3) cells negative for GABA and parvalbumin, but positive for calbindin, (4) cells negative for GABA, parvalbumin and calbindin.

Research paper thumbnail of Effect of sensory deafferentation on the GABAergic circuitry of the adult cat visual system

Neuroscience, 1998

ABSTRACT

Research paper thumbnail of Evidence for chicken GH as the only hypophyseal factor responsible for the stimulation of hepatic 5'-monodeiodination activity in the chick embryo

Reproduction, nutrition, development, 1989

The influence of an intravenous injection of chicken growth hormone (cGH), a total chicken pars d... more The influence of an intravenous injection of chicken growth hormone (cGH), a total chicken pars distalis (PD) extract, and a PD extract depleted of cGH by immunoadsorption was studied in the 18-d-old chick embryo. Plasma concentrations of triiodothyronine (T3), thyroxine (T4), and hepatic 5'-monodeiodination (5'-D) activity were measured. An injection of total PD extract raised plasma T3, T4, and 5'-D activity, whereas a PD extract depleted of GH only increased plasma T4. The amount of cGH present in the PD extracts, as measured by homologous cGH radioimmunoassay, increased T3 and raised liver 5'-D, but had no effect on plasma T4. The effect on liver 5'-D was more pronounced with cGH than with a total PD extract, whereas the effect on plasma T3 was somewhat less pronounced. It was concluded that cGH increased the peripheral conversion of T4 into T3 in the chick embryo, whereas a PD extract depleted of cGH was purely thyrotropic. The PD extract also seemed to have...

Research paper thumbnail of Neurotensin-Like immunoreactivity in the pituitary and hypothalamus of bony fishes

Peptides, 1987

Using an antiserum raised against synthetic neurotensin (NT), the distribution of immunoreactivit... more Using an antiserum raised against synthetic neurotensin (NT), the distribution of immunoreactivity in the pituitary and hypothalamus has been examined by immunocytochemistry at light and electron microscope level in a number of species of bony fishes. In most species immunoreactive perikarya were found in the preoptic region of the hypothalamus, with fibres throughout the tuberal hypothalamus and neurohypophysis (neural lobe and median eminence). In the neurohypophysis of teleosts NT-like immunoreactivity was seen in a dense band of fibres bordering the ACTH cells of the rostral pars distalis: absorption controls showed that this was due to the presence of an NT(8-13)-like or xenopsin-like sequence, which, according to electron microscopic observations, was contained in small dense cored vesicles. The antiserum also stained the pituitary ACTH cells of some species, apparently due to cross-reaction with the 17-19 sequence of ACTH. These results suggest that an NT-like peptide may have a role in control of the adenohypophysis in fishes.

Research paper thumbnail of Calbindin D-28K and parvalbumin immunoreactivity is confined to two separate neuronal subpopulations in the cat visual cortex, whereas partial coexistence is shown in the dorsal lateral geniculate nucleus

Neuroscience Letters, 1989

ABSTRACT

Research paper thumbnail of Distribution of immediate early gene zif-268, c-fos, c-jun and jun-D mRNAs in the adult cat with special references to brain region related to vision

Neuroscience Letters, 1994

The distribution of immediate early gene zif-268, c-fos, c-jun and jun-D mRNAs was investigated i... more The distribution of immediate early gene zif-268, c-fos, c-jun and jun-D mRNAs was investigated in the visual cortex, dorsal lateral geniculate nucleus and hippocampus of the adult cat brain with in situ hybridization. In area 17, zif-268, c-jun and jun-D were found predominantly in layers II-III and VI, while c-fos mRNA was abundant in layer VI. In area 18, the zif-268, c-fos and c-jun labelling pattern was identical to that of area 17, this was not true for jun-D. In area 19, only c-jun retained the lamination pattern of areas 17 and 18, while zif-268, c-fos and jun-D were homogeneously distributed. In the dorsal lateral geniculate nucleus, only c-fos and jun-D resulted in labelling. In the pyramidal layer of hippocampus, zif-268 was found in CA1-4, c-jun in CA1-3, and jun-D in CA2-4. In the dentate gyrus, c-jun was abundant, jun-D moderate and zif-268 faint. C-fos labelling was absent in the hippocampal formation.

Research paper thumbnail of Long-term effects of retinal lesions on growth-associated protein 43 (GAP-43) expression in the visual system of adult cats

Neuroscience Letters, 1996

We have investigated the role of growth-associated protein 43 in synaptic reorganization in the v... more We have investigated the role of growth-associated protein 43 in synaptic reorganization in the visual system of adult cats that received binocular central retinal lesions. Different survival times between 3 and 8 months after induction of the lesion were chosen. In the deafferented part of the dorsal lateral geniculate nucleus (dLGN) we found a long-lasting increase in GAP-43 protein, while glial fibfillaty acidic protein (GFAP) immunoreactivity, which initially increased due to the degeneration of retinal ganglion ceils, slowly subsided over this period. In area 17, the pattern of GAP-43 expression did not provide indications for morphological changes in the cortical architecture following retinal lesions.

Research paper thumbnail of Validation of a new antiserum directed towards the synthetic c-terminus of the FOS protein in avian species: immunological, physiological and behavioral evidence

Journal of Neuroscience Methods, 1999

In the past 10 years, the study of the expression of immediate early genes, such as c-fos, in the... more In the past 10 years, the study of the expression of immediate early genes, such as c-fos, in the brain has become a common method for the identification of brain areas involved in the regulation of specific physiological and behavioral functions. The use of this method in avian species has been limited by the paucity of suitable antibodies that cross-react with the FOS protein in birds. We describe in this paper the preparation of an antibody directed against a synthetic fragment of the protein product of the c-fos gene in chickens (Gallus domesticus). We demonstrate that this new antibody can be used in several avian species to study FOS expression induced by a variety of pharmacological, physiological and behavioral stimuli. Western blot studies indicated that this antibody recognizes a protein of the expected size (47 kDa) but also cross reacts to some extent with proteins of lower molecular weight that share sequence homology with FOS (Fos-related antigens). FOS immunocytochemistry was performed with this antibody in four species of birds in three different laboratories utilizing diverse variants of the immunocytochemical procedure. In all cases the antibody provided a reliable identification of the FOS antigen. The new antibody described here appears to be suitable for the study of FOS expression in several different avian species and situations. It is available in substantial amounts and will therefore make it possible to use FOS expression as a tool to map brain activity in birds as has now been done for several years in mammalian species. : S 0 1 6 5 -0 2 7 0 ( 9 9 ) 0 0 0 6 7 -9

Research paper thumbnail of A new alternative for simultaneous immunohistochemical screening of 96 hybridoma clones for tissue-specific antibody production selects a monoclonal antibody to insect corpus cardiacum

Journal of Neuroscience Methods, 1986

In the current search for the elucidation of the true structure of hitherto unidentified &#39... more In the current search for the elucidation of the true structure of hitherto unidentified 'new' insect neuropeptides we designed a novel screening method to facilitate the primary detection of neurone-specific antibody secreting mouse-mouse hybridoma clones obtained after immunization with neuronal tissue homogenates. The present procedure is principally adapted from a conventional immunohistological test and enables one to rapidly screen 96 (and even more) clones at one time for potential secretion of specific antibodies to different tissue compounds, without the necessity of having a purified antigen. It has proved to be sensitive, rapid, practical and reproducible. As such it promises to be very useful to discriminate amongst the wide range of antibodies to various kinds of materials produced by hybridomas by detecting monoclonal antibodies directed against factors contained in well-defined tissues in which one is interested. This paper also reports the successful application of this method to a primary screening of clones producing murine monoclonal antibodies to substances of insect corpora cardiaca (CC), after immunization with crude antigen preparations.

Research paper thumbnail of Development of a novel screening device permitting immunocytochemical screening of numerous culture supernatants during hybridoma production

Journal of Immunological Methods, 1989

Research paper thumbnail of One-step purification of chicken growth hormone from a crude pituitary extract by use of a monoclonal immunoadsorbent

Journal of Endocrinology, 1988

The immunization of mice with an affinity-purified glycoprotein preparation from chicken pituitar... more The immunization of mice with an affinity-purified glycoprotein preparation from chicken pituitary tissue yielded several monoclonal antibodies towards the recently described glycosylated variant of chicken GH. As all these antibodies recognize the classical (non-glycosylated) GH molecule equally well, they provide a suitable tool for the development of both a specific immunoadsorbent and an assay method. This paper deals with the surprising purification power of the immunoadsorbent that was produced with one of the monoclonal antibodies. The resulting preparation was more than 99% pure as assessed by reversed phase high-performance liquid chromatography and sodium dodecyl sulphate-polyacrylamide gel electrophoresis, so that no further purification steps were needed before the determination of the amino acid sequence of the material. The efficiency of the purification protocol as determined by a homologous, monoclonal antibody-based radioimmunoassay was virtually absolute. Moreover, the affinity-purified GH preparation was a mixture representing the multiple molecular forms of pituitary chicken GH, including both oligomeres and glycosylated GH. The purified preparations were finally used to demonstrate the hepatic 5'-monodeiodinase-stimulating activity of GH in the chicken embryo (results not shown), in order to prove that the biological activity of the molecule had not been damaged by elution from the immunoadsorbent.

Research paper thumbnail of Effect of sensory deafferentation on immunoreactivity of GABAergic cells and on GABA receptors in the adult cat visual cortex

The Journal of Comparative Neurology, 1995

To investigate the effects of sensory deafferentation on the cortical GABAergic circuitry in adul... more To investigate the effects of sensory deafferentation on the cortical GABAergic circuitry in adult cats, glutamic acid decarboxylase (GAD) and gamma-aminobutyric acid (GABA) immunoreactivity and GABA receptor binding were studied in the visual cortex of normal cats and compared with cats that had received restricted binocular central lesions of the retina and had survived for 2 weeks postlesion in a normal visual environment. In the visual cortex of lesioned cats, two changes were observed in the number of GAD-immunoreactive elements in the regions affected by the retinal lesions: the number of GAD-positive puncta decreased, whereas that of GAD-immunoreactive somata increased. In contrast, no detectable changes were measured in the number of GABA-immunopositive somata or puncta. At the receptor level, we observed no differences in either the laminar distribution or the affinity of cortical GABAA and GABAB receptors labeled with [3H]-muscimol and [3H]-baclofen, respectively, in the lesioned versus normal cats. We present the hypothesis that sensory deafferentation in these adult cats (1) leads to a reduction of cortical GABAergic inhibition in the deafferented region, and (2) that this decreased inhibition may permit changes in efficiency of synapses and (3) that these changes may represent a first stage of events underlying the retinotopic reorganization preceeding the structural changes.