Biomedical subjects

B Meister

Publications and source records attributed to B Meister.

At least 37 records · Page 2Linked to original sources

PubMed2000

Effective ex vivo generation of granulopoietic postprogenitor cells from mobilized peripheral blood CD34(+) cells.

OBJECTIVE: Neutropenia following high-dose chemotherapy and peripheral blood progenitor cell (PBPC) transplantation might be abrogated by an additional transplantation of ex vivo generated granulopoietic postprogenitor cells (GPPC). Therefore, the ex vivo expansion of CD34(+) PBPC was systematically studied aiming for optimum GPPC production. MATERIALS AND METHODS: CD34(+) PBPC were cultured in serum-free medium comparing different (n = 32) combinations of stem cell factor (S), interleukin 1 (1), interleukin 3 (IL-3) (3), interleukin-6 (6), erythropoietin (E), granulocyte colony-stimulating factor (G), granulate-macrophage colony-stimulating factor (GM), daniplestim (D, a novel IL-3 receptor agonist), and Flt3 ligand (FL) under various culture conditions. Ex vivo generated cells were assessed by flow cytometry, morphology, and progenitor cell assays. RESULTS: Addition of G +/- GM but not GM alone to cultures stimulated with S163E effectively induced the generation of GPPC. GPPC production was maximum after 12 to 14 days. Best expansion rates were observed when cells were cultured at 1.5x10(4)/mL in 21% O(2). Modifications of culture conditions were either less or equally effective (i.e., modification of starting cell concentrations, low oxygen, addition of serum albumin or autologous plasma, repetitive feeding). Comparison of different cytokine combinations revealed that the optimum GPPC expansion cocktail consisted of S6GD+FL (day 12: 130-fold cellular expansion, 32% myeloblasts/promyelocytes, 49.4% myelocytes/metamyelocytes, 12.4% bands/segmented), which furthermore expanded CD34(+) cells (3.4-fold) and clonogenic progenitors (13.4-fold). CONCLUSION: Using the S6DG+FL expansion cocktail, GPPC could be effectively produced ex vivo starting from positively selected CD34 PBPC, possibly enabling amelioration or even abrogation of posttransplant neutropenia.

Adjuvants, Immunologic↗

PubMed2000

Differential sorting of SNAP-25a and SNAP-25b proteins in neuroblastoma cells.

Exocytosis is mediated by high-affinity interactions between different SNARE proteins. The existence of several variants of each SNARE protein suggests that the specificity of fusion may be directed by unique combination of SNARE family members. We examined if two alternatively spliced variants of synaptosomal-associated protein of 25 kD, SNAP-25a and SNAP-25b, possessed distinct cellular distribution if coexpressed within the same neuroblastoma cell. Double-labelling immunofluorescence histochemistry in combination with confocal laser microscopy of individual cell clones revealed a different subcellular localisation pattern for the two SNAP-25 variants. Sucrose density gradient centrifugation of cell homogenates followed by Western blotting showed that the SNAP-25 protein variants associated with intracellular organelles of different density. Taken together, this study shows that two alternatively spliced variants of SNAP-25, differing in only nine amino acids, possess distinct properties at the level of intracellular trafficking, suggesting that the cellular localisation of SNAP-25 protein is regulated at the level of mRNA splicing.

Animals↗

PubMed2000

Down-regulated STAT3 messenger ribonucleic acid and STAT3 protein in the hypothalamic arcuate nucleus of the obese leptin-deficient (ob/ob) mouse.

Leptin is a weight-reducing hormone produced by adipose tissue, which reduces food intake via hypothalamic leptin receptors and the JAK-STAT signaling pathway. In vivo studies have shown that leptin activates specifically STAT3 in the hypothalamus. We have studied the cellular localization of STAT3 messenger RNA (mRNA) and STAT3 protein in the mouse mediobasal hypothalamus using, respectively, in situ hybridization and immunohistochemistry. Strong STAT3 mRNA and STAT3 immunoreactivity was demonstrated in neurons located in the ventral part of the mouse arcuate nucleus. Comparison of STAT3 mRNA levels in the arcuate nucleus of lean control mice and obese leptin-deficient ob/ob mice showed that the levels of STAT3 mRNA in the arcuate nucleus were significantly lower (31% less in ob/ob mice), compared with control mice. Hybridization with a probe specific for STAT3alpha mRNA showed that the down-regulated STAT3 expression in the arcuate nucleus of ob/ob mice is represented by STAT3alpha. There was a marked difference in numbers and intensity of STAT3-immunoreactive cell bodies, with virtually no STAT3-immunoreactive cell bodies in the mediobasal hypothalamus of ob/ob mice, compared with control mice. Direct double-labeling immunofluorescence histochemistry of sections from control mice, combining a goat antiserum raised against a peptide sequence present in all leptin receptor isoforms (Ob-R) or a guinea pig anti-serum generated to a peptide sequence specific for Ob-Rb with rabbit STAT3 antiserum, demonstrated colocalization of STAT3 and Ob-R as well as colocalization of STAT3 and Ob-Rb, in many cell bodies of the arcuate nucleus. The results suggest that circulating leptin acts via leptin receptor-/STAT3-containing neurons in the ventral arcuate nucleus and that congenital leptin deficiency, as seen in obese ob/ob mice, results in a down-regulation of STAT3 mRNA and protein levels.

Animals↗

PubMed2000

Synaptotagmin III isoform is compartmentalized in pancreatic beta-cells and has a functional role in exocytosis.

Synaptotagmin is involved in Ca2+-regulated secretion and has been suggested to serve as a general Ca2+ sensor on the membrane of secretory vesicles in neuronal cells. Insulin exocytosis from the pancreatic beta-cell is an example of a Ca2+-dependent secretory process. Previous studies of pancreatic beta-cells were unable to show presence of synaptotagmin I. We now present biochemical and immunohistochemical data showing that synaptotagmin III is present in pancreatic beta-cells as well as in the insulin-secreting cell line HIT-T15 and in rat insulinoma. By subcellular fractionation, we found synaptotagmin III in high-density fractions together with insulin and secretogranin I, indicating colocalization of synaptotagmin III and insulin in secretory granules. We could also show that blockade of synaptotagmin III by a specific antibody inhibited Ca2+-induced changes in beta-cell membrane capacitance, suggesting that synaptotagmin III is part of the functional protein complex regulating beta-cell exocytosis. The synaptotagmin III antibody did not affect the activity of the voltage-gated L-type Ca2+-channel. These findings are compatible with the view that synaptotagmin III, because of its distinct localization in the pancreatic beta-cell, functionally modulates insulin exocytosis. This indicates that synaptotagmin may have a general role in the regulation of exocytosis not only in neuronal cells but also in endocrine cells.

Animals↗

PubMed1999

Differential expression of SNAP-25a and SNAP-25b RNA transcripts in cranial nerve nuclei.

Synaptosomal-associated protein of 25 kDa (SNAP-25) is involved in the molecular regulation of neurotransmitter release. SNAP-25 exists in two isoforms, SNAP-25a and SNAP-25b, which arise from alternate splicing and which are differentially expressed throughout the nervous system. In situ hybridization was used to examine the presence and subcellular localization of SNAP-25a and SNAP-25b RNA transcript expression in motor and parasympathetic nuclei associated with cranial nerves. SNAP-25a RNA transcripts were strongly expressed in the parasympathetic Edinger-Westphal nucleus and dorsal motor nucleus of the vagus nerve but weakly expressed in motor nuclei such as the oculomotor, trochlear, trigeminal, facial, ambiguus, hypoglossal and accessory nuclei and in motoneurons of mouse lumbar spinal cord. In contrast, SNAP-25b RNA transcripts were not detectable in the Edinger-Westphal nucleus and dorsal motor nucleus of the vagus nerve but were strongly expressed in the oculomotor, trochlear, trigeminal, facial, ambiguus, hypoglossal, and accessory nuclei and in the motoneurons of mouse lumbar spinal cord. In the parasympathetic cranial nerve nuclei, displaying high levels of SNAP-25a RNA transcripts, the labeling was cytoplasmic, whereas the labeling was nuclear in the cranial nerve motor nuclei, displaying lower levels of transcripts. In contrast, labeling of SNAP-25b RNA transcripts was cytoplasmic in cranial nerve motor nuclei and not detectable in parasympathetic cranial nerve nuclei. Possible explanations for the region-specific and differential subcellular localization of SNAP-25a and SNAP-25b RNA transcripts are discussed.

Animals↗

PubMed1999

Differential expression of nerve terminal protein isoforms in VAChT-containing varicosities of the spinal cord ventral horn.

Of the different types of synaptic contacts with the mammalian spinal motoneuron, the synapse made by the cholinergic, so-called C-terminal of unknown origin and function has special morphological characteristics. Thus, in this synapse, there is no postsynaptic density but rather a large subsynaptic cistern in the motoneuron. To see whether this particular arrangement imposes special demands on the transmitter release machinery, we examined the presence of nerve terminal proteins in the C-terminal by using immunohistochemistry. Cholinergic nerve fibers and terminals in the spinal cord ventral horn were identified with an antiserum to the vesicular acetylcholine transporter (VAChT) protein. Immunohistochemistry in combination with confocal laser microscopy showed the presence of synaptosomal-associated protein of 25 kDa (SNAP-25)-, syntaxin-, cysteine string protein (CSP)-, synuclein-, synapsin I-, synapsin I/II-, synaptotagmin I-, synaptotagmin I/II-, synaptophysin-, and synaptobrevin-2-like immunoreactivity (-LI) in VAChT-containing C-terminals. Synaptotagmin III and synaptobrevin 1 could not be demonstrated in this type of terminal. VAChT-containing varicosities in the Renshaw cell area, with a probable origin from motoneuron axon collaterals, exhibited CSP, synapsin I/II, and synaptobrevin-1-LI, but not SNAP-25-, syntaxin-, synuclein-, synapsin I-, synaptotagmin I-, synaptotagmin I/II-, synaptophysin- and synaptobrevin-2-LI. The results suggest a differential content of nerve terminal proteins and their isoforms in cholinergic C-terminals apposing motoneurons and in the Renshaw cell area. It is concluded that C-terminals contain synaptic proteins necessary for fast transmitter release, and their origin should not be the motoneurons themselves.

Animals↗

PubMed1999

Syntaxin 1 interacts with the L(D) subtype of voltage-gated Ca(2+) channels in pancreatic beta cells.

Interaction of syntaxin 1 with the alpha(1D) subunit of the voltage-gated L type Ca(2+) channel was investigated in the pancreatic beta cell. Coexpression of the enhanced green fluorescent protein-linked alpha(1D) subunit with the enhanced blue fluorescent protein-linked syntaxin 1 and Western blot analysis together with subcellular fractionation demonstrated that the alpha(1D) subunit and syntaxin 1 were colocalized in the plasma membrane. Furthermore, the alpha(1D) subunit was coimmunoprecipitated efficiently by a polyclonal antibody against syntaxin 1. Syntaxin 1 also played a central role in the modulation of L type Ca(2+) channel activity because there was a faster Ca(2+) current run-down in cells incubated with antisyntaxin 1 compared with controls. In parallel, antisyntaxin 1 markedly reduced insulin release in both intact and permeabilized cells, subsequent to depolarization with K(+) or exposure to high Ca(2+). Exchanging Ca(2+) for Ba(2+) abolished the effect of antisyntaxin 1 on both Ca(2+) channel activity and insulin exocytosis. Moreover, antisyntaxin 1 had no significant effects on Ca(2+)-independent insulin release trigged by hypertonic stimulation. This suggests that there is a structure-function relationship between the alpha(1D) subunit of the L type Ca(2+) channel and the exocytotic machinery in the pancreatic beta cell.

Animals↗

PubMed1999

Trends in infection morbidity in a pediatric oncology ward, 1986-1995.

BACKGROUND AND PROCEDURE: We retrospectively studied the type, severity, frequency, and outcome of febrile infectious complications in 217 cancer patients receiving cytotoxic chemotherapy (603 episodes) over a 10-year period in a single pediatric institution. RESULTS: A total of 48.8% of the episodes occurred in severely leukopenic patients (WBC < 1.0 x 10(9)/l, absolute neutrophil count < 500 x 10(6)/l). In the second half of the study period febrile episodes occurred at increased frequency. The number of patients with gram-positive isolates in blood cultures increased over the years, most frequently coagulase-negative staphylococci were found. Remarkably, gram-negative bacteria increasingly resistant to the administered first-line antibiotic regimen emerged, necessitating modifications of the antimicrobial strategy every 3 years. Furthermore, Clostridium difficile-associated enterocolitis posed a clinical problem at increasing frequency since 1993. As expected, the speed of leukocyte recovery within 5 days from the onset of a febrile complication had an influence on the outcome of these episodes. CONCLUSIONS: Rapid recovery of the WBC was associated with an excellent prognosis whereas persisting neutropenia was found to be a negative factor associated with fatal outcomes. The fatality rate of all febrile episodes (2.3%) remained the same throughout the study period despite the availability and wider use of recombinant hematopoietic growth factors since 1991.

Adolescent↗

PubMed1999

Expression of vascular endothelial growth factor (VEGF) and its receptors in human neuroblastoma.

Angiogenic factors may play a role in the biology of neuroblastoma, a well vascularised tumour, which frequently spreads haematogenously. Therefore, we analysed expression of vascular endothelial growth factor (VEGF) in six human neuroblastoma cell lines and five primary neuroblastomas. High VEGF levels (1-3 ng/10(6) cells/day) were found in the supernatant of all cell lines examined (SK-N-LO, SK-N-SH, LS, SH-SY5Y, IMR-32, Kelly). VEGF peptide was also detected in tissue homogenates from four of five primary tumours. Reverse transcription-polymerase chain reaction (RT-PCR) revealed that VEGF165 is the major isoform produced by neuroblastomas. In addition, all cell lines and primary tumours expressed the mitogenic VEGF receptor FLK-1, whilst the non-mitogenic receptor FLT-1 was less frequently positive, suggesting that the tyrosine kinase FLK-1 is involved in malignant transformation of neuroblastoma cells. However, neutralising antibodies to VEGF did not inhibit growth of neuroblastoma cell lines, which argues against a role of VEGF as an autocrine growth factor, at least for cell lines in vitro. We conclude that neuroblastoma cells produce VEGF, which may contribute to tumour vascularisation, growth and invasion.

Adolescent↗

PubMed1999

Leptin receptor- and STAT3-immunoreactivities in hypocretin/orexin neurones of the lateral hypothalamus.

Hypocretins/orexins are recently characterized peptides that are synthesized in neurones of the lateral hypohalamus and stimulate food intake in rats. To clarify whether leptin may interact with hypocretin/orexin to reduce ingestive behaviour, the presence of leptin receptor-immunoreactivity in hypocretin/orexin-containing neurones was examined. Many leptin receptor-and hypocretin/orexin-immunoreactive neurones were demonstrated in the lateral hypothalamic area and perifornical region. Both direct double-labelling and elution-restaining methods showed that leptin receptor-immunoreactivity was present in the vast majority of hypocretin/orexin-containing neurones. Immunoreactivity for STAT3, a transcription factor activated by leptin, was also demonstrated in hypocretin/orexin-containing neurones. Isolated hypocretin/orexin cell bodies in the dorsal part of the lateral hypothalamic area and the ventral perifornical region were shown to contain immunoreactivity for galanin, another peptide known to affect feeding. Galanin neurones were also seen to contain leptin receptor-and STAT3-immunoreactivity. Melanin-concentrating hormone (MCH)-containing neurones constituted a cell population within the lateral hypothalamus distinct from the one containing hypocretin/orexin-immunoreactivity, as shown by elution-restaining methodology. The presence of leptin receptor-and STAT3-immunoreactivities in hypocretin/orexin-containing neurones of the lateral hypothalamus suggests that leptin may directly regulate these hypothalamic neurones, most likely via an inhibitory action on hypocretin/orexin expression and/or secretion resulting in reduced food intake.

Animals↗

PubMed1999

The cellular and developmental expression of hrs-2 in rat.

The molecular events underlying vesicular trafficking probably involve the formation and dissolution of protein complexes between integral components of the vesicle and its target membrane. SNAP-25 is associated with the plasma membrane and is a component of a core protein complex thought to be essential for neurotransmitter release. We have previously characterized a protein, hrs-2, that interacts with SNAP-25 and inhibits secretion from permeabilized PC12 cells. The cellular localization and developmental expression patterns of a number of proteins involved in the secretion machinery have been documented. To understand more about the possible cellular role of hrs-2, we have examined hrs-2 distribution, developmental expression and subcellular localization in rat tissues and cell lines. We show herein that the distribution of hrs-2 in brain and periphery parallels that of SNAP-23/25, and that recombinant hrs-2 binds to both SNAP-23 and SNAP-25. Hrs-2 mRNA and protein are found almost ubiquitously in neurons in the brain. Hrs-2 mRNA is expressed in the neural tube at E10 and thereafter mRNA and protein levels remain relatively constant in the whole brain through adulthood. In cultured PC12 cells, endogenous hrs-2 is expressed in the cytoplasm and on the limiting membranes of multivesicular bodies. Overexpression of hrs-2 in mammalian cells results in the appearance of large intracellular compartments that are labelled with hrs-2 antibodies. The wide distribution, the interaction with SNAP-23 and the localization on multivesicular body membranes suggest a general role for hrs-2 in cellular machinery.

Adenosine Triphosphatases↗

PubMed1999

Vascular neuropeptide Y Y1-receptors in the rat kidney: vasoconstrictor effects and expression of Y1-receptor mRNA.

Neuropeptide Y (NPY) -receptor subtypes were studied in the rat kidney in vivo by systemic administration of NPY, the two agonists [Leu(31), Pro(34)]NPY (Y1-receptor agonist) and NPY (13-36) (Y2-receptor agonist), or the Y1-receptor antagonist BIBP 3226. Effects on mean arterial blood pressure (MAP) and renal arterial blood flow were recorded. The Y1-receptor agonist evoked a dose-dependent increase in MAP concomitantly with a reduction in renal blood flow. At the largest dose administered (1.42 pmol/g), the Y1-agonist [Leu(31), Pro(34)] NPY increased MAP by 20 +/- 6 mmHg and reduced the renal vascular conductance by more than 50%. The same dose of the Y2-agonist NPY (13-36) did not evoke any clear-cut effects on the renal blood flow or MAP. Furthermore, administration of the Y1-receptor antagonist BIBP 3226 reduced the NPY-induced renal vasoconstriction, but did not affect the response to angiotensin II or phenylephrine. The effects evoked by 0.71 pmol/g NPY were almost abolished by 3 mg/kg BIBP 3226. In situ hybridization histochemistry was used to study the expression of Y1-receptor mRNA in the developing rat kidney. The levels of Y1-receptor mRNA expression in the vascular smooth muscle of the rat kidney varied at different ages, with low levels at postnatal day 10 and high levels at 20 days and again low levels at 40 days. In summary, the present study show a maturation-specific expression pattern of NPY Y1-receptor mRNA as well as functional effects of vascular NPY receptors of the Y1-subtype in the rat kidney.

Angiotensin II↗

PubMed1999

Isoform-specific exocytotic protein mRNA expression in hypothalamic magnocellular neurons: regulation after osmotic challenge.

Exocytosis is regulated by proteins which interact to promote docking and fusion of secretory granules with the plasma membrane. We have used in situ hybridization to study the mRNA expression for vesicle-associated membrane protein (VAMP) isoforms VAMP-1 and VAMP-2, synaptosomal-associated protein of 25-kDa (SNAP-25) isoforms SNAP-25a and SNAP-25b, mammalian homologue of unc-18 (munc-18) and Hrs-2 in neurosecretory neurons of the magnocellular paraventricular (PVN) and supraoptic (SON) nuclei of normal and osmotically challenged animals. In PVN and SON neurons of normal animals, strong labeling was demonstrated for VAMP-2 and SNAP-25a mRNA, whereas VAMP-1 or SNAP-25b mRNA could not be detected. Salt-loading (2% NaCl as drinking water), an animal model which increases the expression and secretion of hormones from hypothalamic magnocellular neurons, resulted in significantly increased mRNA levels for VAMP-2 (36%, 28%), munc-18 (74%, 68%) and SNAP-25a (59%, 77%) in the PVN and SON, respectively. There was no significant increase in Hrs-2 mRNA levels in the PVN, whereas a significant increase (22%) was observed in the SON. In the posterior pituitary, immunohistochemistry showed a marked decrease in numbers and intensity of vasopressin-immunoreactive (-IR) nerve endings after salt-loading. There were no obvious changes in numbers or intensity of VAMP-2-, munc-18-, Hrs-2- or SNAP-25-IR fibers. Large varicosities containing VAMP-2- and Hrs-2 immunocreactivity were seen in salt-loaded animals. The results show isoform-specific mRNA expression in neurosecretory neurons and an increased mRNA expression of proteins participating in the molecular regulation of exocytosis during an experimental situation characterized by increased secretion.

Animals↗

PubMed1998

[Orexins--new hypothalamic peptides that stimulate appetite].

A family of novel hypothalamus-specific peptides called orexins have recently been discovered and characterized. The orexins stimulate appetite (the greek word orexis means appetite) when given intraventricularly to rats. Their genes are expressed bilaterally in the lateral hypothalamus, a region previously known to regulate food intake. The two peptides, orexin A (33 amino-acids) and orexin B (28 amino acids), are derived from a common prepro-orexin precursor. The peptides bind to specific G-protein-coupled orexin receptors termed OX-1R and OX-2R. The identification of the orexins will increase our understanding on how the brain regulates food intake.

Amino Acid Sequence↗

PubMed1998

Cysteine string protein (CSP) is an insulin secretory granule-associated protein regulating beta-cell exocytosis.

Cysteine string proteins (CSPs) are novel synaptic vesicle-associated protein components characterized by an N-terminal J-domain and a central palmitoylated string of cysteine residues. The cellular localization and functional role of CSP was studied in pancreatic endocrine cells. In situ hybridization and RT-PCR analysis demonstrated CSP mRNA expression in insulin-producing cells. CSP1 mRNA was present in pancreatic islets; both CSP1 and CSP2 mRNAs were seen in insulin-secreting cell lines. Punctate CSP-like immunoreactivity (CSP-LI) was demonstrated in most islets of Langerhans cells, acinar cells and nerve fibers of the rat pancreas. Ultrastructural analysis showed CSP-LI in close association with membranes of secretory granules of cells in the endo- and exocrine pancreas. Subcellular fractionation of insulinoma cells showed CSP1 (34/36 kDa) in granular fractions; the membrane and cytosol fractions contained predominantly CSP2 (27 kDa). The fractions also contained proteins of 72 and 70 kDa, presumably CSP dimers. CSP1 overexpression in INS-1 cells or intracellular administration of CSP antibodies into mouse ob/ob beta-cells did not affect voltage-dependent Ca2+-channel activity. Amperometric measurements showed a significant decrease in insulin exocytosis in individual INS-1 cells after CSP1 overexpression. We conclude that CSP is associated with insulin secretory granules and that CSP participates in the molecular regulation of insulin exocytosis by mechanisms not involving changes in the activity of voltage-gated Ca2+-channels.

Animals↗

PubMed1998

Decreased mRNA levels for exocytotic proteins in the pituitary of aged rats.

Several protein components that are involved in the molecular regulation of transmitter release have been identified in neuronal, neuroendocrine and endocrine tissues. The expression of VAMP-2 (vesicle-associated membrane protein), munc-18 (mammalian homologue of unc-18) and SNAP-25 (synaptosomal-associated protein of 25 kDa) mRNA was studied in the rat anterior and intermediate pituitary gland of adult (2 months) and old (24 months) rats using in situ hybridization. In the pituitary anterior lobe of aged rats, there was a significant decrease in VAMP-2 (33%), munc-18 (17%) and SNAP-25 (20%) mRNA as compared to adult rats. In the intermediate lobe, there was a significant decrease in VAMP-2 (48%) and SNAP-25 (32%) mRNA of aged rats, whereas munc-18 mRNA levels were not significantly changed. Pituitaries from aged rats showed an increase in size which was paralleled by a significant decrease in the number of cells per unit area in the intermediate lobe, whereas the number was unaltered in the anterior lobe. The results suggest a genuine decrease in mRNA for exocytotic protein mRNA in the anterior pituitary, but that part of the decrease in the expression of VAMP-2 and SNAP-25 mRNA in the intermediate lobe can be explained by a decreased number of cells per unit area. The decline in anterior pituitary hormone secretion reported in aged rats appears to be parallelled by a down-regulation in mRNA levels for several proteins involved in the molecular regulation of exocytosis.

Aging↗

PubMed1998

Leptin injected into the fourth ventricle inhibits gastric emptying.

The effect of leptin on gastric emptying of glucose was studied in freely moving rats bearing intragastric fistulas. Leptin (0.39 microg and 3.9 microg) injected into the fourth ventricle inhibited gastric emptying significantly, whereas s.c. administration of leptin (10 microg/kg) had no effect. Leptin receptor immunoreactivity, revealed by an antiserum that recognizes all leptin receptor isoforms, was demonstrated in choline acetyltransferase (ChAT)-containing neurones of the dorsal motor nucleus of the vagus nerve (DMX). The data indicate that leptin acts centrally to suppress gastric emptying possibly via leptin receptors located on cholinergic DMX neurones.

Amino Acid Sequence↗

PubMed1998

Leptin receptor immunoreactivity in chemically defined target neurons of the hypothalamus.

The adipose tissue-derived hormone leptin regulates body weight homeostasis by decreasing food intake and increasing energy expenditure. The weight-reducing action of leptin is thought to be mediated primarily by signal transduction through the leptin receptor (LR) in the hypothalamus. We have used immunohistochemistry to localize LR-immunoreactive (LR-IR) cells in the rat brain using an antiserum against a portion of the intracellular domain of LR that is common to all LR isoforms. The antiserum recognized the short and long isoforms of LR in transfected hematopoietic BaF3 cells. To examine the chemical nature of target cells for leptin, direct double-labeling immunofluorescence histochemistry was applied. The results show extensive distribution of LR-like immunoreactivity (LR-LI) in the brain with positively stained cells present, e.g., in the choroid plexus, cerebral cortex, hippocampus, thalamus, and hypothalamus. In the hypothalamus, strongly LR-IR neurons were present in the supraoptic nucleus (SON) and paraventricular nucleus (PVN), periventricular nucleus, arcuate nucleus, and lateral hypothalamus. Weaker LR-IR neurons were also demonstrated in the lateral and medial preoptic nuclei, suprachiasmatic nucleus, ventromedial and dorsomedial nuclei, and tuberomammillary nucleus. Confocal laser scanning microscopy showed LR-LI in the periphery of individual cells. In magnocellular neurons of the SON and PVN, LR-LI was demonstrated in vasopressin- and oxytocin-containing neurons. In parvocellular neurons of the PVN, LR-LI was demonstrated in many corticotropin-releasing hormone-containing neurons. LR-IR neurons were mainly seen in the ventromedial aspect of the arcuate nucleus, where LR-LI co-localized with neuropeptide Y. In the ventrolateral part of the arcuate nucleus, LR-LI was present in many large adrenocorticotropic hormone-IR proopiomelanocortin-containing neurons and in a few galanin-, neurotensin-, and growth hormone-releasing hormone-containing neurons. In the dorsomedial arcuate nucleus, few tyrosine hydroxylase (dopamine)-containing neurons were seen to have LR-LI. Melanin-concentrating hormone-containing neurons in the lateral hypothalamus had LR-LI. Based on the immunohistochemical results, possible interactions of leptin with brain mechanisms are discussed.

Animals↗