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H Higuchi

Publications and source records attributed to H Higuchi.

At least 145 records · Page 8Linked to original sources
PubMed1996

Neuron-specific expression of a chicken gicerin cDNA in transient transgenic zebrafish.

Gicerin, a novel cell adhesion molecule which belongs to the immunoglobulin superfamily, is expressed temporally and spatially in the developing chick brain and retina. The previous in vitro experiments using transfected cells showed that gicerin can function as a cell adhesion molecule which has both homophilic and heterophilic binding activities. For the in vivo analyses of gicerin in neural development, we tried to utilize a zebrafish system, a vertebrate suitable for studying early development. We generated transient transgenic animals by microinjecting DNA constructs into zebrafish embryos. Chicken gicerin, under control of the neurofilament gene promoter, was preferentially expressed in neuronal cells and gicerin-expressing neurons exhibited a fasciculation formation with neighboring gicerin-positive axons, which may be partly due to homophilic cell adhesion activity of gicerin. These experimental results suggest that this fast and efficient transgenic animal system is useful for studying the functional roles of neuron-specific genes during the development.

Animals↗
PubMed1996

BDNF increases the expression of neuropeptide Y mRNA and promotes differentiation/maturation of neuropeptide Y-positive cultured cortical neurons from embryonic and postnatal rats.

The effects of neurotrophic factor on the expression of neuropeptide Y (NPY) mRNA and on morphology of NPY-immunoreactive neurons were investigated. Brain-derived neurotrophic factor (BDNF) increased the expression of NPY mRNA in cultured cortical neurons from both embryonic and postnatal rats. BDNF also increased the number of NPY neurons. Furthermore, multipolar neurites from NPY neurons were observed in cultures treated with BDNF, whereas only monopolar and bipolar neurites were observed in control cultures. These results suggest that BDNF not only increases the expression of NPY mRNA but also promotes the differentiation/maturation of NPY ergic neurons both in number and morphology. NPY expression was strongly increased by neurotrophin-4/5 similarly to BDNF and neurotrophin-3 evoked a slight increase. In contrast, basic fibroblast growth factor, cilliary neurotrophic factor and interferon-gamma had no effect on NPY expression.

Animals↗
PubMed1996

Multiple- and single-molecule analysis of the actomyosin motor by nanometer-piconewton manipulation with a microneedle: unitary steps and forces.

We have developed a new technique for measurements of piconewton forces and nanometer displacements in the millisecond time range caused by actin-myosin interaction in vitro by manipulating single actin filaments with a glass microneedle. Here, we describe in full the details of this method. Using this method, the elementary events in energy transduction by the actomyosin motor, driven by ATP hydrolysis, were directly recorded from multiple and single molecules. We found that not only the velocity but also the force greatly depended on the orientations of myosin relative to the actin filament axis. Therefore, to avoid the effects of random orientation of myosin and association of myosin with an artificial substrate in the surface motility assay, we measured forces and displacements by myosin molecules correctly oriented in single synthetic myosin rod cofilaments. At a high myosin-to-rod ratio, large force fluctuations were observed when the actin filament interacted in the correct orientation with a cofilament. The noise analysis of the force fluctuations caused by a small number of heads showed that the myosin head generated a force of 5.9 +/- 0.8 pN at peak and 2.1 +/- 0.4 pN on average over the whole ATPase cycle. The rate constants for transitions into (k+) and out of (k-) the force generation state and the duty ratio were 12 +/- 2 s-1, and 22 +/- 4 s-1, and 0.36 +/- 0.07, respectively. The stiffness was 0.14 pN nm-1 head-1 for slow length change (100 Hz), which would be approximately 0.28 pN nm-1 head-1 for rapid length change or in rigor. At a very low myosin-to-rod ratio, distinct actomyosin attachment, force generation (the power stroke), and detachment events were directly detected. At high load, one power stroke generated a force spike with a peak value of 5-6 pN and a duration of 50 ms (k(-)-1), which were compatible with those of individual myosin heads deduced from the force fluctuations. As the load was reduced, the force of the power stroke decreased and the needle displacement increased. At near zero load, the mean size of single displacement spikes, i.e., the unitary steps caused by correctly oriented myosin, which were corrected for the stiffness of the needle-to-myosin linkage and the randomizing effect by the thermal vibration of the needle, was approximately 20 nm.

Actomyosin↗
PubMed1996

Rat Kupffer cell-derived nitric oxide suppresses proliferation and induces apoptosis of syngeneic hepatoma cells.

BACKGROUND & AIMS: Evidence increasingly indicates that nitric oxide plays an important role in antitumor mechanisms. The aim of this study was to investigate the role of NO in the mechanisms regulating the proliferation and death of hepatoma cells cocultured with Kupffer cells. METHODS: Kupffer cells were isolated from male Wistar rats and cocultured with rat hepatoma AH70 cells. Proliferation was determined by calculating the number of total and 5-bromodeoxyuridine-positive AH70 cells. Apoptosis was assessed by electron-microscopic and fluorescence-microscopic observations and in situ nick end labeling method. Immunofluorescence and in situ hybridization studies were performed to investigate the induction of inducible NO synthase (iNOS). RESULTS: Kupffer cells reduced proliferation and induced apoptosis of AH70 cells, which were attenuated by the NO synthesis inhibitors NG-monomethyl-L-arginine and aminoguanidine. Increased inductions of iNOS messenger RNA and iNOS were observed in Kupffer cells cocultured with AH70 cells. Addition of monoclonal antibody directed against either rat CD18 or intercellular adhesion molecule 1 also attenuated the increased NO production of Kupffer cells and the alterations of AH70 cells. CONCLUSIONS: Kupffer cell-derived NO suppresses proliferation and induces apoptosis of hepatoma cells. The CD18 intercellular adhesion molecule 1-dependent adhesive interaction with hepatoma cells triggers NO production by Kupffer cells.

Animals↗
PubMed1996

Biosynthesis of B2-integrin, intracellular calcium signalling and functional responses of normal and CD18-deficient bovine neutrophils.

1Biosynthesis of CD11/CD18 in bovine leucocytes, intracellular Ca2+ ([Ca2+]i) signalling, chemiluminescent responses and membrane fluidity of neutrophils and the effects of D-mannose on neutrophils from control heifers and a heifer with bovine leucocyte adhesion deficiency (BLAD) were measured. The synthesis of CD11/CD18 complex was clearly detected in leucocytes from a normal heifer, but not in a BLAD-affected heifer. The transient phase of increased [Ca2+]i was clearly detected in neutrophils from a heifer with BLAD stimulated with opsonised zymosan, aggregated bovine immunoglobulin G or concanavalin A, whereas the sustained phase was deficient or significantly decreased compared with control heifers. [Ca2+]i signalling of neutrophils from control heifers and a heifer with BLAD stimulated with phorbol myristate acetate via an 11b/CD18-independent pathway showed no transient phase, and the subsequent increase in [Ca2+]i was almost identical in neutrophils from affected and control heifers. [Ca2+]i concentration and chemiluminescent responses of neutrophils from a control heifer were clearly decreased by treatment with anti-CD18 and anti-IgG antibodies. No differences in membrane fluidity were detected between neutrophils derived from control and CD18-deficient cattle. D-mannose binds mainly to Fc rather than CD18 receptors, and decreased Agg-IgG induced [Ca2+]i and the chemiluminescent response of neutrophils. The [Ca2+]i responses and Agg-IgG induced chemiluminescent responses of neutrophils from control heifers and a BLAD-affected heifer were inhibited by D-mannose. The characteristic changes of [Ca2+]i signalling and functional responses of B2-integrin-deficient neutrophils were demonstrated.

Animals↗
PubMed1996

Ca2+/calmodulin-dependent transcriptional activation of neuropeptide Y gene induced by membrane depolarization: determination of Ca(2+)- and cyclic AMP/phorbol 12-myristate 13-acetate-responsive elements.

Membrane depolarization stimuli (high potassium concentration and veratridine) increased neuropeptide Y (NPY) mRNA abundance time-dependently, without a change in beta-actin mRNA level, in NG108-15 and PC12 cells. Although the induction by veratridine was blocked completely by tetrodotoxin, the induction by potassium was suppressed minimally. Voltage-dependent Ca channel blockers and calmodulin antagonists inhibited the increases by both depolarization stimuli completely, suggesting involvement of Ca2+/calmodulin-dependent kinases (CaM kinases). Transient assay using chloramphenicol acetyltransferase reporter genes containing the rat NPY gene promoter indicated that membrane depolarization and Ca entry stimulate transcription of the NPY gene. The depolarization-induced transactivation was also blocked by CaM kinase inhibitors. The 200-bp 5'-upstream region (-344/-145) was localized as a Ca2+/ calmodulin-responsive element (CaMRE), which confers depolarization-induced transactivation. It is interesting that this CaMRE did not contain the canonical Ca-responsive elements such as CRE, SRE, NF-AT, or the C/EBP beta-binding site and was separated from a 64-bp cyclic AMP/ phorbol 12-myristate 13-acetate-responsive element (-144/-81). These findings suggested that membrane depolarization regulates the NPY gene transcription positively through the unique CaMRE by activation of CaM kinases following Ca entry through L-type Ca channels.

Animals↗
PubMed1996

Role in nitric oxide in Kupffer cell-mediated hepatoma cell cytotoxicity in vitro and ex vivo.

The metabolic change in tumor cells (AH70, a rat hepatoma cell line) cocultured with isolated rat Kupffer cells were visualized and analyzed by a laser scanning confocal imaging system. When AH70 cells were cocultured with Kupffer cells, fluorescence intensity of rhodamine 123 (Rh123) decreased, indicating the reduction of mitochondrial function. The reduction in Rh123 was eliminated by NG-monomethyl-L-arginine (L-NMMA), an analogue of L-arginine, suggesting the involvement of nitric oxide (NO). Two hour after the cells were cocultured, membrane compromised AH70 cells which were observed as propidium from 2.8% to 25%. This increase was also attenuated by L-NMMA, suggesting that Kupffer cell-mediated injury of tumor cells largely depends on NO. The concentrations of NO-2 + NO-3 in the culture medium markedly increased after coculture of AH70 cells with Kupffer cells. Moreover, NO synthase (NOS) activity in Kupffer cells significantly increased after coculture. These in vitro results suggest that NO mediates Kupffer cell-induced tumor cell damage characterized by reduced mitochondrial function and diminished barrier function. In the ex vivo study of the perfused liver to which AH70 cells were injected via the catheter inserted into the portal vein, some AH70 cells were arrested in the upper stream of sinusoid and the fluorescence intensity of Rh123 in adherent AH70 cells decreased in a time-dependent manner within 2 hours. The number of PI-positive AH70 cells also increased 2 hours after the injection of AH70 cells. These changes were inhibited by either administration of N omega-L-nitroarginine-methylester (L-NAME) to perfusate or pretreatment of the rat liver with GdCl3, which is known to deplete Kupffer cell function. Thus, the present study suggests that NO from Kupffer cells induces mitochondrial dysfunction in tumor cells followed by membrane barrier dysfunction in the liver sinusoid.

Animals↗
PubMed1996

alpha 4 integrin plays a critical role in early stages of T lymphocyte migration in Peyer's patches of rats.

Lymphocyte recirculation through the blood flow circuit and lymphoid organs is important for the maintenance of immune defense, and is defined as lymphocyte homing. During the homing process, several adhesion molecules have been postulated to play an important role in lymphocyte recruitment from the vascular space. In the present study, we investigated the effect of a novel mAb against rat alpha 4 integrin (MR alpha 4-1) on the interaction of T lymphocytes with postcapillary venules (PCV) and their subsequent migration into the interstitium of Peyer's patches, using intravital video microscopy. T lymphocytes collected from intestinal lymph were labeled with a fluorochrome carboxyfluorescein diacetate succinimidyl ester and were than injected into the jugular vein of recipient rats. The microvasculature in the ileal Peyer's patches of recipient rats was observed sequentially by intravital fluorescence microscopy. In controls, lymphocytes exhibited rolling behavior which was followed by firm adhesion to the endothelium of PCV. The density of sticking lymphocytes gradually increased during the first 30 min. These initial interactions of lymphocytes with the PCV (rolling and adherence) were drastically inhibited by treatment with MR alpha 4-1, both when MR alpha 4-1 was preinfused into rats and when T cells were preincubated in vitro with MR alpha 4-1 before administration. MR alpha 4-1 also significantly inhibited the transendothelial migration of T lymphocytes, associated by the ratio of migration to adherence. However, once T lymphocytes migrated into the interstitium, treatment with MR alpha 4-1 did not affect the pattern of travel of these lymphocytes in the interstitium or their transport into the microlymphatics in the parafollicular area. Therefore, we conclude that alpha 4 integrins play a critical role in the rolling and sticking of T cells and their transendothelial migration in PCV of Peyer's patches.

Animals↗
PubMed1996

Kupffer cell-mediated cytotoxicity against hepatoma cells occurs through production of nitric oxide and adhesion via ICAM-1/CD18.

Rat Kupffer cell (KC)-mediated cytotoxicity against both the syngeneic hepatoma cell line AH70 and hepatocytes was evaluated by changes in mitochondrial function, and the possible role of ICAM-1/CD18 in the interaction between the cells was studied. Rhodamine 123 fluorescence, a marker of the mitochondrial membrane potential, decreased in AH70 cells after co-culture with CK, while that in hepatocytes was unchanged by co-culture. This decrease was blocked by anti-ICAM-1 anti-CD18 and the inhibition of nitric oxide synthesis. Cytometric studies demonstrated that ICAM-1 expression on AH70 cells increased after addition of IFN-gamma, IL-1beta, tumor necrosis factor (TNF)-alpha or KC, while in hepatocytes ICAM-1 was not increased. Anti-ICAM-1 pretreatment inhibited the increase in ICAM-1 expression and the decrease in rhodamine 123 fluorescence on AH70 cells after co-culture with KC. CD18 on KC was increased only after co-culture with AH70. TNF-alpha but not IFN-gamma was detected in the supernatant of co-culture between KC and AH70 cells, and this production was partially inhibited by anti-ICAM-1 and anti-CD18. The activity of inducible nitric oxide synthase in Kupffer cells and the levels of nitrites and nitrates in the co-culture supernatant increased over time, and this increase was attenuated either by addition of NO synthesis inhibitors, anti-ICAM-1 or anti-CD18. These results indicate that the rat KC causes mitochondrial dysfunction in cancer cells via the production of NO and cell-to-cell adhesion via ICAM-1/CD18 has an important role in this cytotoxic process.

Animals↗
PubMed1996

Functional characteristics of enhanced Fc receptor expression of beta 2 integrin-deficient bovine mononuclear phagocytes.

Fc receptor expression, cytoplasmic Ca2+ signaling, chemiluminescent (CL) response, and electron spin resonance (ESR) combined with spin trapping of blood mononuclear phagocytes from control heifers and a heifer with leukocyte adhesion deficiency (LAD) were evaluated to elucidate the relationships between complement receptor type 3 (CR3) and Fc receptor expression and their functional responses. The mean fluorescence intensity of fluorescein isothiocyanate (FITC)-conjugated anti-bovine IgG bound to mononuclear phagocytes from the heifer with LAD was 1.8-fold higher than that of control heifers. The mean increments of cytoplasmic Ca2+ concentrations of mononuclear phagocytes from the heifer with LAD stimulated with OPZ, Agg-IgG, and PMA were 39.4 (P < 0.05), 118, and 71.6% compared with those of control heifers. A 1.27-fold increase in the CL response relative to control heifers was detected when mononuclear phagocytes from the heifer with LAD were stimulated with Agg-IgG. The OPZ-induced CL response of mononuclear phagocytes from the heifer with LAD was significantly (P < 0.05) decreased, whereas the PMA-induced CL response was similar to that of control heifers. The ESR spectrum of mononuclear phagocytes from the heifer with LAD was increased when stimulated with Agg-IgG, and was impaired when stimulated by OPZ compared with that of control heifers. The ESR spectrum of mononuclear phagocytes stimulated with PMA was similar in control heifers and the heifer with LAD. Fc receptors on mononuclear phagocytes from the heifer with LAD were enhanced, and their cytoplasmic Ca2+ signaling, CL response, and ESR-spin trapping when stimulated with Agg-IgG and OPZ appeared to be associated with enhanced Fc receptors.

Animals↗
PubMed1996

Adhesiveness for extracellular matrices and lysosomal enzyme release from normal and beta 2 integrin-deficient bovine neutrophils.

The adhesiveness of control and CD18-deficient bovine neutrophils on culture plates precoated with collagen I, collagen IV, fibronectin and laminin was measured to evaluate the possible factors for adherence to extracellular matrices. The release of N-acetyl-beta-D-glucosaminidase (NAGase) from control and CD18-deficient neutrophils stimulated with complement receptor type 3 (CR3) or Fc receptor dependent stimuli was also evaluated. The adhesive activities of CD18-deficient neutrophils to collagen I, collagen IV and fibronectin were significantly diminished (P < 0.05); however, similar adhesion to laminin was observed in CD18-deficient neutrophils and control neutrophils. The adhesive activity of control neutrophils on uncoated plates increased 2.5 times (P < 0.05) with the presence of PMA. The mean activities for NAGase release from CD18-deficient neutrophils stimulated with opsonized zymosan and aggregated bovine immunoglobulin G (Agg-IgG) were 46.7 and 82.7% that of the control neutrophils, respectively. The Agg-IgG-induced NAGase release from control and CD18-deficient neutrophils was eliminated by H7, a protein kinase C inhibitor. These results support that an association between CR3 and Fc receptors on neutrophils appears to play an essential role in neutrophil functions.

Acetylglucosaminidase↗
PubMed1996

Effect of chronic ethanol feeding on Kupffer cell-mediated antitumor cell activity.

We have previously reported that the Kupffer cell has antitumor activity through mitochondrial damage to tumor cells by nitric oxide production. In this study, the effect of chronic ethanol feeding on antihepatoma cell activity of the Kupffer cell was examined in rats. Male rats of the Wistar strain were fed ethanol chronically for 8 weeks by liquid diets. Kupffer cells were isolated from the control rat or the ethanol-fed rat, and cocultured with AH 70 cells, a rat hepatoma cell line. Fluorescence of rhodamine 123 or propidium iodide was observed as indicators of the mitochondrial damage or cell membrane injury, respectively, by a laser scanning confocal microscopy. Mitochondrial damage of AH 70 cells as indicated by reduction of rhodamine 123 fluorescence was smaller by the coculture with Kupffer cell from the ethanol rat than that from the control. Cell membrane barrier dysfunction of AH 70 cell was less frequently observed with the Kupffer cell from ethanol-fed rats. A metabolite of nitric oxide (nitrite and nitrate) was less in the cultured medium with the ethanol Kupffer cell than with the control Kupffer cell. Ca2+ mobilization, which induces inducible nitric oxide synthase and observed by the fluorescence of fluo-3, in Kupffer cells cocultured with AH 70 cells was suppressed in ethanol-fed rats. These result suggests that chronic ethanol feeding suppresses antitumor cell activity of Kupffer cell through the impairment of Ca2+ mobilization and nitric oxide production.

Animals↗
PubMed1996

Effect of chronic ethanol feeding on nitric oxide synthesis by rat Kupffer cells.

Kupffer cells contribute to the important role of the liver defense mechanism through nitric oxide (NO) production. In this study, the effect of chronic ethanol administration on the ability of Kupffer cells to synthesize and release NO was investigated after stimulation with lipopolysaccharide (LPS). Male Wistar rats were chronically fed ethanol for 8 weeks according to the method described by DeCarli and Lieber et al. (J Nutr.91:331-336, 1967). Kupffer cells were isolated and cultured with LPS (1 micrograms/ml) for 24 hr. The levels of nitrite and nitrate, metabolites of NO, were determined in the culture medium, NO synthase (NOS) activity in Kupffer cells was determined by the method that measures conversion of [14C]arginine into [14C]citrulline. In control rats, a significant increase of nitrite and nitrate levels in culture medium was observed after LPS treatment. The magnitude of this increase was significantly smaller in chronic ethanol-fed rats. When the activity of NOS was determined, inducible NOS (iNOS) activity was higher than that of constitutive NOS, and LPS administration produced a significant elevation of iNOS activity in both control and chronic ethanol-fed rats. However, the elevation of iNOS activity by LPS stimulation was diminished by chronic ethanol administration. Distribution of iNOS in Kupffer cells as determined by an immunofluorescence method using a laser scanning confocal image system showed a lower expression of iNOS in chronic ethanol-fed rats even in the presence of LPS. These results demonstrate that the excessive production of NO by increased iNOS activity in Kupffer cells is diminished by chronic ethanol administration.

Animals↗
PubMed1996

Ethanol-induced oxidative stress in the liver.

Oxygen stress is well recognized to be a key step in the pathogenesis of ethanol-associated liver injury. Ethanol administration induces an increase in lipid peroxidation either by enhancing the production of oxygen-reactive species and/or by decreasing the level of endogenous antioxidants. Numerous experimental studies have emphasized the role of the ethanol-inducible cytochrome P-450 in the microsomes, as well as the molybdo-flavoenzymes xanthine oxidase in the cytosol. This review shows the putative role of ethanol-induced disturbances in iron metabolism in relation to iron as a prooxidant factor. Ethanol administration also affects the mitochondrial free radical generation. Although many previous studies suggest a role for active oxygens in ethanol-induced mitochondrial dysfunction in hepatocytes, the detailed mechanism of ethanol-induced oxidative stress on mitochondria remains to be clarified further. Studies of our laboratory using a confocal laser scanning microscopic system strongly suggest that active oxidants produced during ethanol metabolism modulate mitochondrial energy synthesis in isolated and cultured hepatocytes. In addition, our investigations implicate endogenous glutathione-glutathione peroxidase system and catalase as important antioxidants and cytoprotective machinery in the hepatocyte mitochondria exposed to ethanol. The fluorographic investigations using the confocal laser scanning microscopy may be useful to extend our knowledge and provide a new view about ethanol-associated oxidative stress and metabolic changes in hepatocytes.

Animals↗
PubMed1996

Endotoxin stimulates lymphocyte-endothelial interactions in rat intestinal Peyer's patches and villus mucosa.

Although lymphocyte-endothelial cell interactions represent a key step in controlling the recruitment of lymphocytes into gut-associated tissues, its dynamic process in microvessels of lymphoid (Peyer's patches) and nonlymphoid (villus) regions of the small bowel remains poorly understood. We monitored the migration of fluorescence-labeled T lymphocytes into normal and lipopolysaccharide (LPS)-inflamed rat intestinal microvessels using intravital microscopy. In Peyer's patches, T lymphocytes selectively adhered to postcapillary venules, although such selectivity was not observed in submucosal venules of villi. T lymphocytes exhibited rolling behavior followed by firm adhesion in microvessels of both the Peyer's patches and the villi, with both types of adhesive interaction being mediated by alpha 4-integrins. The enhanced rolling and adherence of lymphocytes observed in Peyer's patches and submucosal venules of villi of LPS-treated rats were preceded by a reduction in shear rate and were mediated largely by alpha 4-integrins and partly by beta 2-integrins. In capillaries of intestinal mucosa, lymphocyte adherence occurred without rolling and was independent of alpha 4-integrins. LPS also significantly increased adherence of lymphocytes to villus capillaries, which was not mediated by either alpha 4- or beta 2-integrin. These observations demonstrate significant heterogeneity of lymphocyte-endothelial cell interactions within different regions of the intestinal mucosa.

Animals↗
PubMed1996

Roles of ET-1 in endotoxin-induced microcirculatory disturbance in rat small intestine.

The major objective of this study was to investigate whether endothelin-1 (ET-1) plays a significant role in endotoxin-induced microcirculatory disturbances of the intestinal mucosa. Submucosal microvessels of the rat ileum were observed by intravital microscopy with a high-speed video camera system. Preceding the apparent intestinal mucosal damage, red blood cell (RBC) velocity was significantly decreased 30 min after endotoxin treatment in both arterioles and venules. The number of leukocytes sticking to submucosal venules was significantly increased at 30 min. BQ-123, an ETA-receptor antagonist, significantly attenuated the decrease in RBC velocity and also prevented an increase in leukocyte sticking as well as the subsequent mucosal damage induced by endotoxin. The ET-1 concentrations began to be elevated in plasma at 15 min and in the mucosa at 30 min and subsequently further increased in a time-dependent manner. A significant decrease in calcium-dependent nitric oxide synthase activity and significant increases in the concentration of platelet-activating factor (PAF) were demonstrated in the intestinal mucosa after endotoxin treatment. BQ-123 also significantly attenuated these changes. We concluded that the increased ET-1 production in intestinal mucosa induced by endotoxin stimulation could lead to leukocyte sticking and decreased RBC velocity in the intestinal microcirculatory beds via ETA receptors, which are closely related to increased production of PAF and decreased synthesis of constitutive nitric oxide.

Animals↗
PubMed1996

Growth hormone receptor gene is expressed in neuropeptide Y neurons in hypothalamic arcuate nucleus of rats.

GH feeds back on the hypothalamus and regulates its own secretion. We have previously shown that systemic administration of GH induces expression of the c-fos gene, a marker of neuronal activity, on the hypothalamic neuropeptide Y(NPY) and somatostatin neurons in rats. We argued that if GH were to act directly on NPY neurons, NPY neurons should express the GH receptor (GHR) gene. To test this hypothesis, coronal sections of the medial basal hypothalamus from adult male Wistar rats were processed by double label in situ hybridization using a 35S-labeled NPY complementary RNA probe and a digoxigenin-labeled GHR complementary RNA probe. In the medial basal hypothalamus, NPY messenger RNA (mRNA) was observed in the arcuate nucleus (ARC) and the dorsomedial nucleus. The majority (95%) of NPY mRNA-containing cells in the ARC expressed the GHR gene, whereas no NPY mRNA-containing cells in the dorsomedial nucleus expressed the GHR gene. These findings suggest that NPY neurons in the ARC mediate the feedback effect of GH on the hypothalamus.

Animals↗