Biomedical subjects

F Izumi

Publications and source records attributed to F Izumi.

At least 19 recordsLinked to original sources

PubMed2001

Stimulation of catecholamine synthesis in cultured bovine adrenal medullary cells by leptin.

Recently, we characterized leptin receptors in bovine adrenal medullary cells (Yanagihara et al. 2000). Here we report the stimulatory effect of leptin on catecholamine synthesis in the cells. Incubating cells with leptin (10 nM) for 20 min increased the synthesis of 14C-catecholamines from [14C]tyrosine, but not from L-3,4-dihydroxyphenyl [3-14C]alanine. The stimulation of catecholamine synthesis in the cells by leptin was associated with the phosphorylation and activation of tyrosine hydroxylase, the rate-limiting enzyme of catecholamine biosynthesis. The incubation of cells with leptin resulted in a rapid activation of the mitogen-activated protein kinases (MAPKs). An inhibitor of MAPK kinase, U0126, nullified the stimulatory effect of leptin on the synthesis of 14C-catecholamines. Leptin potentiated the stimulatory effect of acetylcholine on 14C-catecholamine synthesis, whereas leptin failed to enhance the phosphorylation and activation of tyrosine hydroxylase induced by acetylcholine. These findings suggest that leptin stimulates catecholamine synthesis via the activation of tyrosine hydroxylase by two different mechanisms, i.e., one is dependent on tyrosine hydroxylase phosphorylation mediated through the MAPK pathway and the second is independent of enzyme phosphorylation.

Acetylcholine↗

PubMed2001

Dual phases of functional change in norepinephrine transporter in cultured bovine adrenal medullary cells by long-term treatment with clozapine.

The effects of long-term treatment with clozapine, a prototype of atypical antipsychotic drugs, on the functional activity, synthesis and mRNA of norepinephrine (NE) transporter were examined in bovine adrenal medullary cells in culture. Treatment of cells with clozapine at 0.1-3.0 microM concentrations produced dual phases of changes in [(3)H]NE uptake, i.e. the first phase showed a decrease in [(3)H]NE uptake at 2-48 h, and the following phase showed an increase in uptake at 72-168 h. Treatment with clozapine for 6 h decreased V(max) to 40% of the control without changing the K(m) value for [(3)H]NE uptake. However, treatment with clozapine for 96 h increased V(max) by 56% over the control without a change in K(m). Scatchard plot analysis of [(3)H]desipramine (DMI) binding to membranes isolated from cells treated with clozapine for 6 h revealed a decrease in B(max) without any change in K(d); in contrast, treatment with clozapine for 96 h caused an increase in B(max) without any change in K(d). Both actinomycin D and cycloheximide, which are inhibitors of protein synthesis, suppressed the clozapine (96 h)-induced increase in [(3)H]NE uptake. Treatment of cells with clozapine for 12-96 h increased the level of NE transporter mRNA in a concentration-dependent manner (0.3-3.0 microM). These findings suggest that treatment of cells with clozapine results in the down-regulation and subsequent up-regulation of NE transporter. The latter change may be caused by the synthesis of new proteins of NE transporter via an increase in its mRNA.

Adrenal Medulla↗

PubMed2000

Characterization and functional role of leptin receptor in bovine adrenal medullary cells.

We report here the characterization and functional roles of the leptin receptor (ObR) in bovine adrenal medullary cells. The plasma membranes isolated from bovine adrenal medulla showed a single class of specific binding sites of (125)I-leptin with an apparent K(d) of 6.6 nM and B(max) of 62 fmol/mg protein. ObRa but not ObRb mRNA was detected in bovine adrenal medulla by reverse transcriptase-polymerase chain reaction. Incubation of cultured adrenal medullary cells with leptin (3-30 nM) for 20 min resulted in a significant increase in [(14)C]catecholamine synthesis from [(14)C]tyrosine without any change in catecholamine secretion. These findings suggest that leptin stimulates catecholamine synthesis through its receptors in bovine adrenal medullary cells.

Adrenal Medulla↗

PubMed2000

Inhibitory effects of clozapine and other antipsychotic drugs on noradrenaline transporter in cultured bovine adrenal medullary cells.

The effects of clozapine and other antipsychotic drugs on noradrenaline (NA) transport were examined in cultured bovine adrenal medullary cells and in transfected Xenopus laevis oocytes expressing the bovine NA transporter. Incubation of adrenal medullary cells with clozapine (30-1000 ng/ml) inhibited desipramine (DMI)-sensitive uptake of [3H]NA in a concentration-dependent manner (IC50=110 ng/ml or 336 nM). Other antipsychotic drugs such as haloperidol, chlorpromazine, and risperidone also decreased [3H]NA uptake (IC50= 144, 220, and 210 ng/ml or 383, 690, and 512 nM, respectively). Eadie-Hofstee analysis showed that clozapine reduced V(max) of uptake of [3H]NA and increased K(m). Furthermore, clozapine inhibited specific binding of [3H]DMI to plasma membranes isolated from bovine adrenal medulla (IC50=48 ng/ml or 146 nM). Scatchard plot analysis of [3H]DMI binding revealed that clozapine decreased both B(max) and K(d). Other antipsychotic drugs, including haloperidol, chlorpromazine, and risperidone, also reduced [3H]DMI binding to the membranes. In transfected Xenopus oocytes expressing the bovine NA transporter, clozapine inhibited [3H]NA uptake in a concentration-dependent manner similar to that observed in adrenal medullary cells. These results suggest that clozapine and haloperidol directly inhibit transport of NA by acting on the site of an NA transporter that influences both substrate transport and binding of tricyclic antidepressants.

Adrenal Medulla↗

PubMed2000

Inhibition by neuromuscular blocking drugs of norepinephrine transporter in cultured bovine adrenal medullary cells.

UNLABELLED: Pancuronium stimulates the cardiovascular system, whereas vecuronium, a derivative of pancuronium, has far fewer effects. The inhibition of norepinephrine transporter (NET) in the sympathetic nervous system may partly account for the stimulatory actions of pancuronium. To investigate the mechanism of action of pancuronium on NET, we examined the effects of pancuronium on NET activity by using cultured bovine adrenal medullary cells and compared pancuronium with other neuromuscular blocking drugs. Pancuronium (1-300 microM) inhibited desipramine-sensitive [(3)H]norepinephrine (NE) uptake in a concentration-dependent manner. Vecuronium (100-300 microM) and d-tubocurarine (300 microM) also decreased [(3)H]NE uptake but were less potent than pancuronium at clinical concentrations. Succinylcholine had little effect on [(3)H]NE uptake. Saturation analysis showed that pancuronium and vecuronium reduced an apparent maximum velocity (V(max)) of [(3)H]NE uptake without altering Michaelis-Menten constant, indicating noncompetitive inhibition. Pancuronium did not inhibit the specific binding of [(3)H]desipramine to plasma membranes isolated from bovine adrenal medulla. A protein kinase C inhibitor, GF109203X, did not affect the inhibition of [(3)H]NE uptake by pancuronium. Pancuronium enhanced the inhibition of NET induced by ketamine. These results suggest that pancuronium, with clinically relevant concentrations, inhibits NET activity by interacting with a site distinct from the recognition site for NE and the desipramine binding site on the transporter. IMPLICATIONS: In this study, pancuronium inhibited norepinephrine uptake and was the most potent of the neuromuscular blocking drugs we tested, including pancuronium, vecuronium, d-tubocurarine, and succinylcholine. Pancuronium may affect the sympathetic nervous system by inhibiting the activity of the presynaptic norepinephrine transporter at clinically relevant concentrations.

Adrenal Medulla↗

PubMed2000

Dual effects of intravenous anesthetics on the function of norepinephrine transporters.

BACKGROUND: Norepinephrine transporters (NETs) terminate the neuronal transmission of norepinephrine, which is released from noradrenergic neurons. To investigate the interaction with NET, the authors examined the effects of short- and long-term treatment with anesthetics on the activity and mRNA level of NET. METHODS: To assay [3H]norepinephrine uptake, bovine adrenal medullary cells in culture were incubated with [3H]norepinephrine in the presence of intravenous anesthetics, including propofol, thiamylal, and diazepam. To study the direct interaction between the anesthetics and NET, the effect of propofol on the binding of [3H]desipramine to the plasma membrane was examined. To study the long-term effect of anesthetics, [3H]norepinephrine uptake by cells pretreated with propofol for 6-24 h and [3H]desipramine binding after pretreatment for 12 h were measured. Simultaneously, we examined the effect of anesthetics on the expression of NET mRNA using the reverse transcriptase-polymerase chain reaction. RESULTS: All of the intravenous anesthetics inhibited [3H]norepinephrine uptake in a concentration-dependent manner. The active concentrations of propofol (1-3 microm) and thiamylal (< or = 30 microm) were similar to those encountered clinically. The kinetic analysis revealed that all the anesthetics noncompetitively inhibited [3H]norepinephrine uptake. Propofol inhibited [3H]desipramine binding with a potency similar to that observed in [3H]norepinephrine uptake. Scatchard analysis showed that propofol competitively inhibited [3H]desipramine binding. On the other hand, long-term treatment of cells with propofol (10 microm) enhanced the NET functional activity and [3H]desipramine binding, and also increased the level of NET mRNA. CONCLUSIONS: These results suggest that intravenous anesthetics have a dual effect on NET; short-term treatment causes inhibition, whereas long-term treatment leads to up-regulation. The interaction of intravenous anesthetics with NET may modulate the neuronal transmission of norepinephrine during anesthesia.

Adrenal Medulla↗

PubMed2000

ATP activates DNA synthesis by acting on P2X receptors in human osteoblast-like MG-63 cells.

In human osteoblast-like MG-63 cells, extracellular ATP increased [(3)H]thymidine incorporation and cell proliferation and synergistically enhanced platelet-derived growth factor- or insulin-like growth factor I-induced [(3)H]thymidine incorporation. ATP-induced [(3)H]thymidine incorporation was mimicked by the nonhydrolyzable ATP analogs adenosine 5'-O-(3-thiotriphosphate) and adenosine 5'-adenylylimidodiphosphate and was inhibited by the P2 purinoceptor antagonist suramin, suggesting involvement of P2 purinoceptors. The P2Y receptor agonist UTP and UDP and a P2Y receptor antagonist reactive blue 2 did not affect [(3)H]thymidine incorporation, whereas the P2X receptor antagonist pyridoxal phosphate-6-azophenyl-2',4-disulfonic acid inhibited ATP-induced [(3)H]thymidine incorporation, suggesting that ATP-induced DNA synthesis was mediated by P2X receptors. RT-PCR analysis revealed that MG-63 cells expressed P2X(4), P2X(5), P2X(6), and P2X(7), but not P2X(1), P2X(2), and P2X(3), receptors. In fura 2-loaded cells, not only ATP, but also UTP, increased intracellular Ca(2+) concentration, and inhibitors for several Ca(2+)-activated protein kinases had no effect on ATP-induced DNA synthesis, suggesting that an increase in intracellular Ca(2+) concentration is not indispensable for ATP-induced DNA synthesis. ATP increased mitogen-activated protein kinase activity in a Ca(2+)-independent manner and synergistically enhanced platelet-derived growth factor- or insulin-like growth factor I-induced kinase activity. Furthermore, the mitogen-activated protein kinase kinase inhibitor PD-98059 totally abolished ATP-induced DNA synthesis. We conclude that ATP increases DNA synthesis and enhances the proliferative effects of growth factors through P2X receptors by activating a mitogen-activated protein kinase pathway.

Adenosine Triphosphate↗

PubMed1999

Evidence that multiple P2X purinoceptors are functionally expressed in rat supraoptic neurones.

1. The expression, distribution and function of P2X purinoceptors in the supraoptic nucleus (SON) were investigated by reverse transcription-polymerase chain reaction (RT-PCR), in situ hybridization, and Ca2+-imaging and whole-cell patch-clamp techniques, respectively. 2. RT-PCR analysis of all seven known P2X receptor mRNAs in circular punches of the SON revealed that mRNAs for P2X2, P2X3, P2X4, P2X6 and P2X7 receptors were expressed in the SON, and mRNAs for P2X3, P2X4 and P2X7 were predominant. 3. In situ hybridization histochemistry for P2X3 and P2X4 receptor mRNAs showed that both mRNAs were expressed throughout the SON and in the paraventricular nucleus (PVN). 4. ATP caused an increase in [Ca2+]i in a dose-dependent manner with an ED50 of 1.7 x 10-5 M. The effects of ATP were mimicked by ATPgammaS and 2-methylthio ATP (2MeSATP), but not by AMP, adenosine, UTP or UDP. alphabeta-Methylene ATP (alphabetaMeATP) and ADP caused a small increase in [Ca2+]i in a subset of SON neurones. 5. The P2X7 agonist 2'- & 3'-O-(4-benzoylbenzoyl)-ATP (BzATP) at 10-4 M increased [Ca2+]i, but the potency of BzATP was lower than that of ATP. In contrast, BzATP caused a more prominent [Ca2+]i increase than ATP in non-neuronal cells in the SON. 6. The effects of ATP were abolished by extracellular Ca2+ removal or by the P2 antagonist pyridoxal phosphate-6-azophenyl-2',4'-disulphonic acid (PPADS), and inhibited by extracellular Na+ replacement or another P2 antagonist, suramin, but were unaffected by the P2X7 antagonist oxidized ATP, and the inhibitor of Ca2+-ATPase in intracellular Ca2+ stores cyclopiazonic acid. 7. Two patterns of desensitization were observed in the [Ca2+]i response to repeated applications of ATP: some neurones showed little or moderate desensitization, while others showed strong desensitization. 8. Whole-cell patch-clamp analysis showed that ATP induced cationic currents with marked inward rectification. The ATP-induced currents exhibited two patterns of desensitization similar to those observed in the [Ca2+]i response. 9. The results suggest that multiple P2X receptors, including P2X3, are functionally expressed in SON neurones, and that activation of these receptors induces cationic currents and Ca2+ entry. Such ionic and Ca2+-signalling mechanisms triggered by ATP may play an important role in the regulation of SON neurosecretory cells.

Adenosine↗

PubMed1999

Modification of low density lipoprotein potentiates its inhibitory effect on catecholamine secretion in cultured bovine adrenal medullary cells.

Low density lipoprotein (LDL) and lipoprotein(a) suppress catecholamine secretion in cultured adrenal medullary cells. Modification of LDL by oxidation or acetylation potentiates various atherogenic actions of LDL. In the present study, we investigated whether the modification of LDL influences catecholamine secretion in cultured bovine adrenal medullary cells. The exposure of LDL to CuSO4 caused a time-dependent oxidation of LDL. Maximal oxidation of LDL was observed after exposure to CuSO4 for 24 h. Native LDL inhibited catecholamine secretion induced by carbachol to 68.5% of control. Oxidized LDL caused further inhibition of carbachol-evoked secretion to 37.6% of control. Acetylated LDL inhibited it to 41.0% of control. There was a good correlation between the extent of LDL oxidation and the inhibition of catecholamine secretion. These results suggest that oxidation or acetylation of LDL augments its inhibitory effect on the secretion of catecholamines. Since catecholamines are a risk factor of atherosclerosis, the inhibitory effect by such modified LDL may be a mechanism inhibiting atherosclerotic progression.

Adrenal Medulla↗

PubMed1999

Cyclic AMP-dependent synthesis and release of adrenomedullin and proadrenomedullin N-terminal 20 peptide in cultured bovine adrenal chromaffin cells.

Adrenomedullin and proadrenomedullin N-terminal 20 peptide are peptides with multiple physiological functions and are most abundant in adrenal medulla. We studied whether the cAMP-dependent pathway is involved in the regulation of synthesis and release of adrenomedullin and proadrenomedullin N-terminal 20 peptide in cultured bovine adrenal chromaffin cells. Exposure of the cells to dibutyryl cAMP (dbcAMP) increased a progressive accumulation of immunoreactive-adrenomedullin and immunoreactive-proadrenomedullin N-terminal 20 peptide in the extracellular medium, while reciprocally decreasing their cellular content in a time-dependent manner. The decrease of levels of both peptides in the cells was much greater in extent than the increase of the peptides in the medium. H89, an inhibitor of cAMP-dependent protein kinase attenuated these changes, induced by dbcAMP. The resulting changes by dbcAMP and H89 were similar to those of chromogranin B, a marker peptide of chromaffin granule. Northern blot analysis showed that the mRNA encoding these peptides, detected as a band of 1.6 kb, was decreased by the treatment with dbcAMP. The effect of dbcAMP on mRNA was attenuated by H89, and was reversible as the decreased mRNA level caused by dbcAMP could be returned to control levels by culturing cells after removal of dbcAMP. These results suggest that the cAMP-dependent protein kinase pathway stimulates the release of adrenomedullin and proadrenomedullin N-terminal 20 peptide, whereas it lowers synthesis of these peptides via the reduction of their transcript level.

Adrenal Medulla↗

PubMed1999

Involvement of PDGF in pressure-induced mesangial cell proliferation through PKC and tyrosine kinase pathways.

In glomerular hypertension, mesangial cells (MC) are subjected to at least two physical forces: mechanical stretch and high transmural pressure. Increased transmural pressure, as well as mechanical stretch, promotes MC proliferation, which may enhance glomerulosclerosis. The exact mechanism of this effect is not fully understood. We examined the effects of transmural pressure alone on cell proliferation and DNA synthesis and investigated the role of platelet-derived growth factor (PDGF) and basic fibroblast growth factor (bFGF), candidates for mediation of glomerular diseases, in the pressure-induced events. Pressure was applied to cultured MC placed in a sealed chamber using compressed helium gas. Application of pressure resulted in a time-dependent ( approximately 2 h) and pressure level-dependent (approximately 80 mmHg) increase in cell number (1.4-fold) and [(3)H]thymidine incorporation (2.7-fold). Pressure-induced DNA synthesis was significantly suppressed by inhibitors of phospholipase C (2-nitro-4-carboxyphenyl-N, N-diphenylcarbamate), protein kinase C [1-(5-isoquinolinylsulfonyl)-2-methylpiperazine and chelerythrine], or tyrosine kinases (genistein). Pressure caused a rapid but transient formation of inositol 1,4,5-trisphosphate, which was blocked by the phospholipase C inhibitor. Pressure also promoted a rapid increase in tyrosine kinase activity. Pressure increased mRNA levels of PDGF-B, with a peak at 6 h, but not those of PDGF-A or bFGF. Pressure-induced DNA synthesis was partially inhibited by a neutralizing anti-PDGF antibody but not by an antibody against bFGF or nonimmune IgG. Our results indicated that pressure by itself increases DNA synthesis and proliferation of cultured rat MC possibly through activation of protein kinase C and tyrosine kinases, and PDGF-B could be partially involved in these pathways.

Animals↗

PubMed1999

Adrenomedullin inhibits transmural pressure induced mesangial cell proliferation through activation of protein kinase A.

Adrenomedullin (AM), a hypotensive peptide isolated from human pheochromocytoma, inhibits the proliferation of mesangial cells (MC) induced by mitogens such as platelet-derived growth factor. Quite recently, we have demonstrated that transmural pressure applied to cultured MC increased DNA synthesis and cell proliferation through protein kinase C and tyrosine kinase pathways. However, the modulatory effect of AM on pressure-induced cell proliferation is as yet unknown. In the present study, we examined the effect of AM on transmural pressure-induced DNA synthesis in cultured rat MC. Pressure was applied to cells placed in a sealed chamber using compressed helium. Application of pressure resulted in an increase in [(3)H]thymidine incorporation (approximately 2.0-fold). AM clearly inhibited pressure-induced DNA synthesis in a concentration-dependent manner. This inhibition was paralleled by an increase in cellular cAMP levels evoked by AM. Forskolin and dibutyryl cAMP mimicked the inhibitory effect of AM. The protein kinase A inhibitor H-89 significantly attenuated the effect of AM. Human AM(22-52)-NH(2), a putative AM receptor antagonist, reversed the inhibitory effects of AM more potently than did human CGRP(8-37), a calcitonin gene related peptide receptor antagonist. Our results suggest that AM, by acting mainly on AM-sensitive receptors, inhibits pressure-induced DNA synthesis in cultured rat MC through activation of protein kinase A. AM may play a protective role against MC proliferation in certain pathological conditions.

Adrenomedullin↗

PubMed1999

Propofol inhibits muscarinic acetylcholine receptor-mediated signal transduction in Xenopus Oocytes expressing the rat M1 receptor.

The effects of propofol, 2,6-diisopropylphenol, an intravenous general anesthetic, on signal transduction mediated by the rat M1 muscarinic acetylcholine (ACh) receptor (M1 receptor) were examined in electrophysiological studies by analyzing receptor-stimulated, Ca2+-activated Cl--current responses in the Xenopus oocyte expression system. In oocytes expressing the M1 receptor, ACh induced the Ca2+-activated C1- current, in a dose-dependent manner (EC50= 114 nM). Propofol (5-50 microM) reversibly and dose-dependently inhibited induction of the Ca2+-activated Cl- current by ACh (100 nM) (IC50=5.6 microM). To determine a possible site affected by propofol in this signal transduction, we tested the effects of this anesthetic (10 microM) on the activation of current by injection of CaCl2 and aluminum fluoride (AlF4-). Propofol did not affect activation of the current by the intracellular injected Ca2+, or activation of the current by the intracellular injected AlF4-. These results indicate that propofol does not affect G protein, the inositol phosphate turnover, release of Ca2+ from Ca2+ store or the Ca2+-activated Cl- channel. Propofol apparently inhibits the M1 receptor-mediated signal transduction at the receptor site and/or the site of interaction between the receptor and associated G protein.

Acetylcholine↗

PubMed1999

Prostaglandin E2 induces Ca2+ release from ryanodine/caffeine-sensitive stores in bovine adrenal medullary cells via EP1-like receptors.

Prostaglandin E2 (PGE2) causes Ca2+ release from intracellular Ca2+ stores and stimulates phosphoinositide metabolism in bovine adrenal medullary cells. These results have been interpreted as PGE2 induces Ca2+ release from inositol trisphosphate (IP3)-sensitive stores. However, we have recently shown that pituitary adenylate cyclase-activating polypeptide (PACAP), bradykinin, and angiotensin II release Ca2+ from caffeine/ryanodine-sensitive stores, although they cause a concomitant increase of intracellular IP3. In light of these results, the mechanism of PGE2-induced Ca2+ release was investigated in the present study. PGE2 dose-dependently caused a transient but consistent Ca2+ release from internal Ca2+ stores. The PGE2-induced Ca2+ release was unaffected by cinnarizine, a blocker of IP3-induced Ca2+ release. By contrast, it was potently inhibited by prior application of caffeine and ryanodine. Although IP3 production in response to PGE2 was abolished by the phospholipase C inhibitor U-73122, Ca2+ release in response to PGE2 was unaffected by U-73122. The PGE2-induced Ca2+ release was unaffected by Rp-adenosine 3',5'-cyclic monophosphothioate, an inhibitor of protein kinase A, and forskolin, a cyclic AMP (cAMP)-elevating agent, did not cause Ca2+ release. The EP1 agonist 17-phenyl-trinorPGE2 and the EP1/EP3 agonist sulprostone mimicked the Ca(2+)-releasing effects of PGE2, whereas the EP2 agonist butaprost or the EP2/EP3 agonist misoprostol caused little or no Ca2+ release. The EP1 antagonist SC-51322 significantly suppressed the Ca2+ release response induced by PGE2, whereas the EP4 antagonist AH-23828B had little effect. These results suggest that PGE2, acting on EP1-like receptors, induces Ca2+ release from ryanodine/caffeine-sensitive stores through a mechanism independent of IP3 and cAMP and that PGE2 may share the same mechanism with PACAP and the other peptide ligands in causing Ca2+ release in bovine adrenal medullary cells.

Adrenal Medulla↗

PubMed1999

Coupling of the expressed cannabinoid CB1 and CB2 receptors to phospholipase C and G protein-coupled inwardly rectifying K+ channels.

Signaling of the cannabinoid CB1 and CB2 receptors through phospholipase C (PLC) and G protein-coupled inwardly rectifying K+ channels (GIRK) was studied after their expression in COS7 cells and Xenopus oocytes. The CB1 or CB2 receptor was co-expressed with alpha subunits of the Galphaq family (Galphaq, Galpha11, Galpha14, Galpha15 and Galpha16) in COS7 cells. Receptor-dependent activation of PLC was observed after co-expressing the CB1 receptor with Galpha14, Galpha15 or Galpha16 but not with Galphaq or Galpha11. Co-expression of Gbeta1 and Ggamma2 abolished the activation, indicating that the activation was mediated by Galpha. PLC activation was not observed when the CB2 receptor was expressed alone or co-expressed with any of the above Galpha subunits. Coupling to GIRK was observed with both CB1 and CB2 receptors after expression in Xenopus oocytes. Significantly larger currents were induced when the receptor was co-expressed with both GIRK1 and GIRK4 than with either GIRK alone. Co-expression of Galpha transducin with the receptor significantly reduced the K+ currents, indicating that GIRK activation was mediated by Gbetagamma but not by Galpha. These findings suggest two new signaling pathways for the cannabinoid receptors.

Animals↗

PubMed1998

Adrenomedullin increases intracellular Ca2+ and inositol 1,4,5-trisphosphate in human oligodendroglial cell line KG-1C.

The effects of adrenomedullin (AM), a hypotensive peptide, were investigated in cultured human oligodendroglial cell line KG-1C. Human AM increased the intracellular Ca2+ concentration ([Ca2+]i) at concentrations greater than 10(-7) M. Human calcitonin gene-related peptide (CGRP), a peptide structurally related to AM, also increased [Ca2+]i with a potency similar to that of AM. AM increased [Ca2+]i in the absence of extracellular Ca2+. Further, AM increased inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) level in a concentration-dependent manner similar to that of AM-induced [Ca2+]i, suggesting that AM-induced elevation of [Ca2+]i is due to Ca2+ release from Ins(1,4,5)P3-sensitive stores. AM (10(-9) to 10(-6) M) increased cAMP in a concentration-dependent manner. Forskolin also increased cAMP, but did not mimic the [Ca2+]i-raising effect of AM. These findings suggest that functional AM receptors are present in oligodendroglial KG-1C cells and that AM increases [Ca2+]i through a mechanism independent of cAMP.

Adrenomedullin↗

PubMed1998

Activation of inwardly rectifying K+ channels by GABA-B receptors expressed in Xenopus oocytes.

In Xenopus oocytes coinjected with poly(A)+ RNA derived from the rat cerebellum and cRNAs for the cloned G protein-gated inwardly rectifying K+ channel (GIRK), GIRK1 and GIRK2, the GABA-B agonist baclofen elicited inwardly rectifying K+ currents. The inward K+ currents elicited by baclofen were inhibited by the selective GABA-B antagonists 2-OH saclofen and CGP 35348, and by the GIRK inhibitor Ba2+. In contrast, baclofen caused no currents in oocytes injected with the cerebellar poly(A)+ RNA alone, the poly(A)+ RNA and cRNA for GIRK1 or GIRK2, or only cRNAs for GIRK1 and GIRK2. These findings indicate that GABA-B receptors in the rat cerebellum were functionally expressed in Xenopus oocytes and activated the cloned GIRKs composed of GIRK1 and GIRK2 as heteromultimers.

Animals↗

PubMed1998

Stimulatory effect of lymphocyte-derived factor on catecholamine efflux from cultured bovine adrenal medullary cells.

The effects of lymphocytes and their conditioned medium on catecholamine efflux and uptake were examined in cultured bovine adrenal medullary cells. Co-culture of adrenal medullary cells with lymphocytes for 3 days caused an increase in appearance of catecholamines in the culture medium. Treatment of adrenal medullary cells with a conditioned medium prepared from lymphocytes also enhanced the appearance of catecholamines in culture medium in time- (8-48 h) and concentration-dependent manners. Heat treatment of the conditioned medium at 60 and 100 degrees C for 10 min reduced its stimulatory effect to 59 and 20% of control, respectively. After gel filtration on a Sephadex G-25 column or dialysis (<8 kDa molecular mass cutoff), the stimulatory activity of the conditioned medium was found in a high molecular fraction. The conditioned medium had little effect on the activity of lactate dehydrogenase in the medium of cultured adrenal medullary cells and on desipramine-sensitive [3H]norepinephrine uptake by the cells. These findings suggest that lymphocytes release a heat-sensitive factor(s) (molecular mass of more than 8 kDa) which increases efflux of catecholamines from cultured adrenal medullary cells.

Adrenal Medulla↗