Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The orexin-orexin receptor system has been implicated in the regulation of wakefulness/sleep states. Behavioral and psycho-stimulant effects of orexins have also been shown. Mesolimbic dopamine neurons in the ventral tegmental area (VTA) are implicated in the regulation of reward and wakefulness/sleep, In the present study, we examined the effect of orexin-A on cytosolic [Ca2+]i concentration ([Ca2+]) in the isolated rat VTA dopamine neurons. Orexin-A (10-12-10-8 M) concentration dependently increased [Ca2+]i in dopamine-containing neurons. The [Ca2+]i responses to orexin-A were inhibited under Ca2+-free conditions and by blockers of voltage-gated L- and N-type [Ca2+]i channels, nitrendipine and omega-conotoxin, respectively. The [Ca2+]i responses were also abolished by a phosphatidylcholine-specific phospholipase C inhibitor, D609, and a protein kinase C (PKC) inhibitor, calphostin C. A PKC activator, TPA, mimicked orexin-A in increasing [Ca2+]i. These results indicate that orexin-A increases [Ca2+]i in VTA dopamine neurons via phosphatidylcholine-specific PLC- and PKC-mediated activation of L- and N-type Ca2+ channels. This effect may serve as the mechanism by which orexin regulates wakefulness/sleep states and exerts its behavioral and psychostimulant effects.
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PMID:Orexin-a activates phospholipase C- and protein kinase C-mediated Ca2+ signaling in dopamine neurons of the ventral tegmental area. 1143 17

The hypocretin/orexins (Hcrts/ORXs) are peptides produced in neurons in the lateral hypothalamic area that project to neuroendocrine centers in the hypothalamus. Hcrt/ORX receptors are present in the hypothalamus and anterior pituitary gland. We examined the possibility that the Hcrts/ORXs, which we have demonstrated previously to act in the brain to stimulate sympathetic function, could alter stress hormone secretion by a direct pituitary action. In vitro studies revealed a dose-related inhibitory effect of the Hcrts/ORXs on corticotropin-releasing hormone-stimulated ACTH secretion that appeared to be mediated via the orexin-1 receptor and to be expressed at doses (threshold dose 1 nM orexin A) similar to the affinity constant for the receptor. The effect was not due to abrogation of the cAMP response of the corticotroph to corticotropin-releasing hormone and was not pertussis toxin sensitive, suggesting a non-G(i)-mediated mechanism. Instead, a G(q)-mediated signaling mechanism was indicated by the ability of protein kinase C blockade with calphostin C to reverse the inhibitory action of orexin A. Orexin A and orexin B did not significantly alter basal ACTH secretion in vitro and did not alter basal or releasing factor-stimulated secretion of luteinizing hormone, prolactin, thyroid-stimulating hormone or growth hormone from cells harvested from male or random-cycle female donors. Our data suggest a direct, pituitary action of the Hcrts/ORXs to modulate the endocrine response to stress and identify the potential cellular mechanism of a unique biological action of the peptides in the anterior pituitary gland.
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PMID:Hypocretin/orexin suppresses corticotroph responsiveness in vitro. 1155 21

Orexins A and B are hypothalamic peptides, that act through two receptor subtypes, called OX1-R and OX2-R. OX1-R selectively binds orexin A, whereas OX2-R is nonselective for both orexins. High levels of OX1-R mRNA and low levels of OX2-R mRNA have been previously detected in the human adrenal cortex and medulla. Here we demonstrated by RT-PCR the expression of the OX2-R, but not the OX1-R, gene in 10 benign secreting pheochromocytomas. Both orexins A and B stimulated catecholamine secretion from pheochromocytoma slices; the maximal effective concentration was 10(-8) mol/liter. Orexins A and B (10(-8) mol/liter) increased IP3, but not cAMP production, by tumor slices, and the effect was blocked by the PLC inhibitor U-73122. The catecholamine response to 10(-8) mol/liter orexins A and B was abolished by either U-73122 or the PKC antagonist calphostin C and was unaffected by the adenylate cyclase inhibitor SQ-22536 and the PKA inhibitor H-89. Collectively, these findings suggest that orexins stimulate catecholamine secretion from human pheochromocytomas, acting through OX2-R coupled to the PLC-PKC signaling pathway.
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PMID:Human pheochromocytomas express orexin receptor type 2 gene and display an in vitro secretory response to orexins A and B. 1160 May 47

Orexins are recently discovered neuropeptides that play an important role in the regulation of hormone secretion, and their receptors have been recently demonstrated in the pituitary. The effects of orexin-A on voltage-gated Ca2+ currents and GH release in primary cultured ovine somatotropes were examined. The expression of orexin-1 receptor was demonstrated by RT-PCR in ovine somatotropes, from which Ca2+ currents were also isolated as L, T, and N currents. Application of orexin-A (100 nM) significantly and reversibly increased only the L current, and coadministration of orexin-A and GHRH (10 nM) showed an additive effect on this current, but no effect of orexin-A was observed on either T or N current. Furthermore, the orexin-A-induced increase in the L current was completely abolished by the inhibition of protein kinase C (PKC) activity using calphostin C (100 nM), phorbal 12,13-dibutyrate pretreatment (0.5 micro M) for 16 h or specific PKC inhibitory peptide PKC(19-36) (1 mM). However, the increase in L current by orexin-A was sustained when cells were preincubated with a specific protein kinase A blocker H89 (1 micro M) or a specific intracellular Ca2+ store depleting reagent thapsigargin (1 micro M). Finally, orexin-A alone did not significantly increase GH release, but coadministration of orexin-A and GHRH showed a synergistic effect on GH secretion in vitro. Our results therefore suggest that orexin-A may play an important role in regulating GHRH-stimulated GH secretion through the enhancement of the L-type Ca2+ current and the PKC-mediated signaling pathway in ovine somatotropes.
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PMID:Orexin-A augments voltage-gated Ca2+ currents and synergistically increases growth hormone (GH) secretion with GH-releasing hormone in primary cultured ovine somatotropes. 1244 88

Ghrelin is a newly discovered peptide that is released from the stomach and from neurons in the hypothalamic arcuate nucleus (ARC) and potently stimulates growth hormone release and food intake. Neuropeptide-Y (NPY) neurons in the ARC play an important role in the stimulation of food intake. The present study aimed to determine whether ghrelin directly activates NPY neurons and, if so, to explore its signaling mechanisms. Whether the neurons that respond to ghrelin could be regulated by orexin and leptin was also examined. We isolated single neurons from the ARC of rats and measured the cytosolic Ca(2+) concentration ([Ca(2+)](i)) with fura-2 fluorescence imaging. Ghrelin (10(-12) to 10(-8) mol/l) concentration-dependently increased [Ca(2+)](i), which occurred in 35% of the ARC neurons. Approximately 80% of these ghrelin-responsive neurons were proved to be NPY-containing by immunocytochemical staining, and 58% of them were glucose-sensitive neurons as judged by their responses to lowering glucose concentrations. The [Ca(2+)](i) responses to ghrelin were markedly attenuated by inhibitors of protein kinase A (PKA) but not protein kinase C and by a blocker of N-type but not L-type Ca(2+) channels. Orexin increased [Ca(2+)](i) and leptin attenuated ghrelin-induced [Ca(2+)](i) increases in the majority (80%) of ghrelin-responsive NPY neurons. These results demonstrate that ghrelin directly interacts with NPY neurons in the ARC to induce Ca(2+) signaling via PKA and N-type Ca(2+) channel-dependent mechanisms. The integration of stimulatory effects of ghrelin and orexin and inhibitory effect of leptin may play an important role in the regulation of the activity of NPY neurons and thereby feeding.
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PMID:Ghrelin directly interacts with neuropeptide-Y-containing neurons in the rat arcuate nucleus: Ca2+ signaling via protein kinase A and N-type channel-dependent mechanisms and cross-talk with leptin and orexin. 1266 66

Orexins, orexigenic neuropeptides, are secreted from lateral hypothalamus and orexin receptors are expressed in the pituitary. Since growth hormone (GH) secreted from pituitary is integrally linked to energy homeostasis and metabolism, we studied the effect of orexin-B on voltage-gated Ca(2+) currents and the related signalling mechanisms in primary cultured ovine somatotropes using whole-cell patch-clamp techniques. With a bath solution containing TEA-Cl (40 mM) and Tetrodotoxin (TTX) (1 microM), three subtypes of Ca(2+) currents, namely the long-lasting (L), transient (T), and N currents, were isolated using different holding potentials (-80 and -30 mV) in combination with specific Ca(2+) channel blockers (nifedipine and omega-conotoxin). About 75% of the total current amplitude was contributed by the L current, whereas the N and T currents accounted for the rest. Orexin-B (1-100 nM) dose-dependently and reversibly increased only the L current up to approximately 125% of the control value within 4-5 min. Neither a specific protein kinase A (PKA) blocker (H89, 1 microM) nor an inhibitory peptide (PKI, 10 microM) had any effect on the increase in L current by orexin-B. The orexin-B-induced increase in the L current was abolished by concurrent treatment with calphostin C (Cal-C, 100 nM), protein kinase C (PKC) inhibitory peptide (PKC(19-36), 1 microM), or by pretreatment with phorbol-12,13-dibutyrate (PDBu) (0.5 microM) for 16 h (a downregulator of PKC). Orexin-B also increased in vitro GH secretion in a dose-dependent manner. We conclude that orexin-B increases the L-type Ca(2+) current and GH secretion through orexin receptors and PKC-mediated signalling pathways in ovine somatotropes.
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PMID:Orexin-B augments voltage-gated L-type Ca(2+) current via protein kinase C-mediated signalling pathway in ovine somatotropes. 1267 48

Orexin-A has recently been identified as a new hypothalamic peptide working as a mediator in the regulation of feeding behavior and sleep control. To determine the role of orexin-A in peripheral metabolic processes, we examined direct effects of orexin-A on catecholamine synthesis and secretion in cultured bovine adrenal medullary cells. Incubation of cells with orexin-A (100 pM) for 20 min caused a small but significant increase in 14C-catecholamine synthesis from [14C]tyrosine, but not from L-3,4-dihydroxyphenyl[3-14C]alanine. Orexin-A (100 pM) potentiated the stimulatory effects of acetylcholine (0.3 mM) on 14C-catecholamine synthesis. Orexin-A significantly increased tyrosine hydroxylase activity, which was evident at 1 pM and maximal at 100 pM. 4 beta-Phorbol-12 beta-myristate-13 alpha-acetate, an activator of protein kinase C, did not enhance the stimulatory effects of orexin-A on tyrosine hydroxylase activity, while H-7 and staurosporine, inhibitors of protein kinase C, nullified the effects of orexin-A. Orexin-A had little effect on catecholamine secretion from the cells. Orexin receptor 1 (OX(1)R) but not orexin receptor 2 (OX(2)R) mRNA was detected in bovine adrenal medullary cells by reverse transcriptase-polymerase chain reaction. These findings suggest that orexin-A activates tyrosine hydroxylase and then stimulates catecholamine synthesis, probably via activation of the OX(1)R-protein kinase C pathway in adrenal medullary cells.
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PMID:Stimulation of catecholamine synthesis by orexin-A in bovine adrenal medullary cells through orexin receptor 1. 1281 74

Orexins, orexigenic neuropeptides, have recently been discovered in lateral hypothalamus and play an important role in the regulation of pituitary hormone secretion. Two subtypes of orexin receptors (orexin-1 and orexin-2) have been demonstrated in pituitaries. In this experiment, the effects of orexins on voltage-gated Ca2+ currents and the GH release in primary cultured ovine somatotropes were examined. Voltage-gated Ca2+ currents were isolated in ovine somatotropes as L, T, and N currents using whole-cell patch-clamp techniques and specific Ca2+ channel blocker and toxin. Application of orexin-A or orexin-B (100 nM) significantly, dose-dependently, and reversibly increased only nifedipine-sensitive L-type Ca2+ current. Inhibitors of PKC (calphostin C, PKC inhibitory peptide) but not inhibitors of PKA (H89, PKA inhibitory peptide) cancelled the increase in the L current by orexins. Co-administration of orexin-A and GHRH (10 nM) showed an additive effect on the L current. Specific intracellular Ca2+-store-depleting reagent, thapsigargin (1 microM), did not affect the orexin-induced increase in the L current. Orexin-B alone slightly increased GH release and co-administration of orexin-A and GHRH synergistically stimulated GH secretion in vitro. It is therefore suggested that orexins may play an important role in regulating GHRH-stimulated GH secretion through an increase in the L-type Ca2+ current and the PKC-mediated signaling pathways in ovine somatotropes.
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PMID:The in vitro regulation of growth hormone secretion by orexins. 1461 Feb 99

Dysfunction of the hypocretin/orexin (Hcrt/Orx) peptide system is closely linked to the sleep disorder narcolepsy, suggesting that it is also central to the normal regulation of sleep and wakefulness. Indeed, Hcrt/Orx peptides produce long-lasting excitation of arousal-related neurons, including those in the laterodorsal tegmentum (LDT) and the dorsal raphe (DR), although the mechanisms underlying these actions are not understood. Since Hcrt/Orx mobilizes intracellular calcium ([Ca(2+)](i)) in cells transfected with orexin receptors and since receptor-mediated Ca(2+) transients are ubiquitous signaling mechanisms, we investigated whether Hcrt/Orx regulates [Ca(2+)](i) in the LDT and DR. Changes in [Ca(2+)](i) were monitored by fluorescence changes of fura-2 AM loaded cells in young mouse brain slices. We found Hcrt/Orx (Orexin-A, 30-1,000 nM) evoked long-lasting increases in [Ca(2+)](i) with differing temporal profiles ranging from spiking to smooth plateaus. A fragment of Hcrt/Orx (16-33) failed to evoke changes in [Ca(2+)](i) and changes were not blocked by TTX or ionotropic glutamate receptor antagonists, suggesting they resulted from specific activation of postsynaptic orexin receptors. Unlike orexin receptor-transfected cells, Hcrt/Orx-responses were not attenuated by depletion of Ca(2+) stores with cyclopiazonic acid (CPA; 3-30 microM), thapsigargin (3 microM), or ryanodine (20 microM), although store-depletion by either CPA or ryanodine blocked Ca(2+) mobilization by the metabotropic glutamate receptor agonist (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (trans-ACPD; 30 microM). In contrast, Hcrt/Orx responses were strongly attenuated by lowering extracellular Ca(2+) ( approximately 20 microM) but were not inhibited by concentrations of KB-R7943 (10 microM) selective for blockade of sodium/calcium exchange. Nifedipine (10 microM), inhibited Hcrt/Orx responses but was more effective at abolishing spiking than plateau responses. Bay K 8644 (5-10 microM), an L-type calcium channel agonist, potentiated responses. Finally, responses were attenuated by inhibitors of protein kinase C (PKC) but not by inhibitors of adenylyl cyclase. Collectively, our findings indicate that Hcrt/Orx signaling in the reticular activating system involves elevation of [Ca(2+)](i) by a PKC-involved influx of Ca(2+) across the plasma membrane, in part, via L-type calcium channels. Thus the physiological release of Hcrt/Orx may help regulate Ca(2+)-dependent processes such as gene expression and NO production in the LDT and DR in relation with behavioral state. Accordingly, the loss of Hcrt/Orx signaling in narcolepsy would be expected to disrupt calcium-dependent processes in these and other target structures.
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PMID:Hypocretin/orexin peptide signaling in the ascending arousal system: elevation of intracellular calcium in the mouse dorsal raphe and laterodorsal tegmentum. 1499 52

Orexin-A and -B (hypocretin-1 and -2) have been implicated in the stimulation of feeding. Here we show the effector neurons and signaling mechanisms for the orexigenic action of orexins in rats. Immunohistochemical methods showed that orexin axon terminals contact with neuropeptide Y (NPY)- and proopiomelanocortin (POMC)-positive neurons in the arcuate nucleus (ARC) of the rats. Microinjection of orexins into the ARC markedly increased food intake. Orexins increased cytosolic Ca(2+) concentration ([Ca(2+)](i)) in the isolated neurons from the ARC, which were subsequently shown to be immunoreactive for NPY. The increases in [Ca(2+)](i) were inhibited by blockers of phospholipase C (PLC), protein kinase C (PKC) and Ca(2+) uptake into endoplasmic reticulum. The stimulation of food intake and increases in [Ca(2+)](i) in NPY neurons were greater with orexin-A than with orexin-B, indicative of involvement of the orexin-1 receptor (OX(1)R). In contrast, orexin-A and -B equipotently attenuated [Ca(2+)](i) oscillations and decreased [Ca(2+)](i) levels in POMC-containing neurons. These effects were counteracted by pertussis toxin, suggesting involvement of the orexin-2 receptor and Gi/Go subtypes of GTP-binding proteins. Orexins also decreased [Ca(2+)](i) levels in glucose-responsive neurons in the ventromedial hypothalamus (VMH), a satiety center. Leptin exerted opposite effects on these three classes of neurons. These results demonstrate that orexins directly regulate NPY, POMC and glucose-responsive neurons in the ARC and VMH, in a manner reciprocal to leptin. Orexin-A evokes Ca(2+) signaling in NPY neurons via OX(1)R-PLC-PKC and IP(3) pathways. These neural pathways and intracellular signaling mechanisms may play key roles in the orexigenic action of orexins.
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PMID:Orexins (hypocretins) directly interact with neuropeptide Y, POMC and glucose-responsive neurons to regulate Ca 2+ signaling in a reciprocal manner to leptin: orexigenic neuronal pathways in the mediobasal hypothalamus. 1506 49


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