Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.3.16 (calcineurin)
 17,112 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The molecular mechanisms underlying regulation of fluid production by secretory epithelia such as the choroid plexus are poorly understood. Two cAMP-regulated inhibitors of protein phosphatase-1, inhibitor-1 (I1) and a dopamine- and cAMP-regulated phosphoprotein, M(r) = 32,000 (DARPP-32), are enriched in the choroid plexus. We show here that these two phosphoproteins are colocalized in choroid plexus epithelial cells. We have developed a novel method for studying the phosphorylation state of DARPP-32 and I1 in intact cells, using a phosphorylation state-specific monoclonal antibody. Several drugs and hormones that are known to alter fluid secretion and that increase cAMP levels (forskolin, isoproterenol, vasoactive intestinal peptide) or cGMP levels (atrial natriuretic peptide) or that may use additional second messenger pathways (5-HT), increase the phosphorylation of I1 and DARPP-32 in rat choroid plexus. In contrast, dopamine does not alter cAMP and cGMP levels, or I1 and DARPP-32 phosphorylation. Our results indicate that DARPP-32, known to be regulated by dopamine in a number of tissues, can be phosphorylated in response to non-dopaminergic factors, including hormones acting through non-cAMP-dependent pathways. Our results also raise the possibility that inhibition of phosphatase-1, as a result of I1 and DARPP-32 phosphorylation, might be part of a final common pathway in the action of several factors that are known or thought to alter cerebrospinal fluid production.
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PMID:Phosphorylation of DARPP-32 and protein phosphatase inhibitor-1 in rat choroid plexus: regulation by factors other than dopamine. 149 46

1. In previous work we have shown that in the snail Helix aspersa neuron F1 carbamylcholine (CCh) and other muscarinic agonists enhance the inward current carried through high voltage-activated Ca2+ channels by Ba2+ (HVA-ICa). It was also found that cyclic nucleotides, inositol trisphosphate or arachidonic acid are not involved in this modulation. Moreover, despite the effect of CCh being blocked by intracellular injection of EGTA, neither protein kinase C nor Ca(2+)-calmodulin-dependent protein kinase II appeared to play a role. 2. In the present paper, the intracellular mechanism of this muscarinic modulation was investigated further by studying the effects of inhibitors of Ser-Thr protein phosphatases (PP) on both the HVA-ICa of neuron F1 and its enhancement by CCh. 3. Intracellular injections in the F1 neuron of either microcystin LR or okadaic acid, both inhibitors of PP1 and PP2A, mimic the action of CCh on the HVA-ICa and occlude the effects of CCh on this current. In contrast, cyclosporin A, an inhibitor of PP2B (calcineurin), affects neither the HVA Ca2+ current itself nor its modulation by CCh. 4. The efficacy of PP inhibitors was tested in F1 neurons in which serotonin (5-HT) induces an inward current involving intracellular increases in cAMP and a protein kinase A-dependent closing of K+ channels. We found that intracellular injection of either microcystin LR or okadaic acid mimicked the 5-HT-induced inward current and occluded the effect of further application of 5-HT.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Enhancement by muscarinic agonists of a high voltage-activated Ca2+ current via phosphorylation in a snail neuron. 765 75

The influence of internal Ca2+ ions has been investigated during intracellular perfusion of isolated neurones from pedal ganglia of Helix pomatia in which serotonin (5-HT) induces a cyclic-adenosine-monophosphate-(cAMP)-dependent enhancement of high-threshold Ca2+ current (ICa). Internal free Ca2+ ([Ca2+]i) was varied between 0.01 and 10 microM by addition of Ca(2+)-EGTA [ethylenebis(oxonitrilo)tetraacetate] buffer. Elevation of [Ca2+]i depressed the 5-HT effect. The dose/effect curve for the Ca2+ blockade had a biphasic character and could be described by the sum of two Langmuir's isotherms for tetramolecular binding with dissociation constants Kd1 = 0.063 microM and Kd2 = 1 microM. Addition of calmodulin (CM) antagonists (50 microM trifluoperazine or 50 microM chlorpromazine), phosphodiesterase (PDE) antagonists [100 microM isobutylmethylxanthine (IBMX) or 5 mM theophylline] and protein phosphatase antagonists [2 microM okadaic acid (OA)] in the perfusion solution caused "anticalcium" action and modified the Ca2+ binding isotherm. Using the effect of OA and IBMX, two components of the total Ca2+ inhibition were separated and evaluated. In the presence of one of these blockers tetramolecular curves with Kd1 = 0.04 microM and Kd2 = 0.69 microM were obtained describing the activation of the retained unblocked enzyme--PDE or calcineurin (CN) correspondingly. The sum of these isotherms gave a biphasic curve similar to that in control. Leupeptin (100 microM), a blocker of Ca(2+)-dependent proteases did not influence the amplitude of 5-HT effect, indicating that channel proteolysis is not involved in the depression. Our findings show that the molecular mechanism of Ca(2+)-induced suppression of the cAMP-dependent upregulation of Ca2+ channels is due to involvement of two Ca(2+)-CM-dependent enzymes: PDE reducing the cAMP level, and CN causing channel dephosphorylation. No other processes are involved in the investigated phenomenon at a Ca2+ concentration of less than or equal to 10 microM.
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PMID:Mechanisms of antagonistic action of internal Ca2+ on serotonin-induced potentiation of Ca2+ currents in Helix neurones. 768 96

Prostaglandin G/H synthase (PGHS) is one of the key enzymes in prostaglandin synthesis. Regulation of the mRNA expression of the two isozymes PGHS-1 and PGHS-2 was investigated in mesangial cells. PGHS-1 was constitutively expressed and not modulated by any of the stimuli used. PGHS-2 was induced by the platelet products serotonin (5-HT) and thromboxane A2 (used as its analogue U46619), but not by ATP. Expression of PGHS protein was regulated correspondingly; whereas PGHS-1 protein was constitutively expressed, PGHS-2 protein was virtually absent in unstimulated cells, but could increasingly be induced by the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), 5-HT, or fetal calf serum. Induction of PGHS-2 mRNA was transient with a peak after 2-3 h. Stimulated mRNA levels persisted for more than 6 h when transcription was inhibited by actinomycin D or when translation was inhibited by cycloheximide. As shown by specific inhibitors, 5-HT signal transduction was mediated by 5-HT2 receptors, which couple to phospholipase C via pertussis toxin-sensitive G-proteins. Induction of PGHS-2 mRNA by 5-HT was dependent on protein kinase C. Down-regulation of the enzyme by prolonged incubation with TPA abolished 5-HT-induced PGHS-2 mRNA expression. Short time activation of protein kinase C by TPA induced PGHS-2 mRNA expression. On the other hand, TPA given immediately before 5-HT decreased the 5-HT-induced PGHS-2 mRNA expression, indicating a negative feedback. The immunosuppressive drug cyclosporin A reduced induction of PGHS-2 mRNA expression by 5-HT, indicating interference with the signaling cascade, most likely with the Ser/Thr phosphatase calcineurin. Involvement of Tyr phosphorylation in 5-HT signaling was shown by the Tyr kinase inhibitor genistein, which inhibited the induction, while the Tyr phosphatase inhibitor vanadate by itself was able to induce PGHS-2 mRNA expression, which was further augmented when vanadate was combined with 5-HT. PGHS-2 mRNA expression is thus tightly regulated in mesangial cells and therefore allows modulation at various levels by physiological and pharmacological stimuli.
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PMID:Signal transduction pathways responsible for serotonin-mediated prostaglandin G/H synthase expression in rat mesangial cells. 808 94

1. We have investigated the mechanism of regulation of 5-HT3 receptor channel sensitivity in voltage-clamped (-80 mV) NG108-15 neuroblastoma cells. 2. The 5-HT-induced inward current activated rapidly. The fast onset was followed by a biphasic decay which was characterized by two time constants, tau 1 (1.1 +/- 0.21s) and tau 2 (8.9 +/- 1.6s), respectively. Brief applications of 5-HT, applied at 2 min intervals, induced a decrease in the amplitude of the 5-HT3 receptor-mediated peak inward currents. 3. Buffering of intracellular calcium with the calcium chelator BAPTA (10 mM) instead of EGTA (10 mM) attenuated the 5-HT-induced loss of responsiveness of 5-HT3 receptors. Omission of calcium from the extracellular medium yielded a similar attenuation of loss of responsiveness. 4. Inclusion of the protein kinase inhibitor, staurosporine (1 microM) or of okadaic acid (1 microM), an inhibitor of protein phosphatases 1 and 2A, in the intracellular buffer solution did not affect 5-HT3 receptor sensitivity. 5. Injection of cyclosporin A-cyclophilin A complex (20 nM), which potently inhibits calcineurin, did not affect the time constants of the biphasic decay of the 5-HT response tau 1 (1.4 +/- 0.28s) and tau 2 (11.3 +/- 1.7s). The complex, however, prevented the loss of 5-HT3, receptor responsiveness upon repeated application of 5-HT. A similar, but weaker effect was observed after intracellular application of the autoinhibitory peptide domain of calcineurin (1 microM). 6. The recovery of desensitized 5-HT3 receptors upon a second application of 5-HT (1 microM) showed a half-life time (tau 1/2) of 2.6 +/- 0.12 min in control cells which was reduced to 1.6 +/- 0.09 min in cells treated with cyclosporin A-cyclophilin A (20 nM) complex. 7. We conclude that calcineurin does not affect the fast decay of the 5-HT3 receptor response but may be involved in a slower process which regulates channel activity.
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PMID:Modulation by calcineurin of 5-HT3 receptor function in NG108-15 neuroblastoma x glioma cells. 884 51

Immunophilins, protein receptors for immunosuppressant drugs such as cyclosporin A and FK506, are enriched far more in the brain than in the immune system. Drug-immunophilin complexes bind to calcineurin, inhibiting its phosphatase activity and leading to immunosuppressant effects. The immunophilin FKBP-12 (FK506 binding protein, 12 kDa) forms a complex with the ryanodine and inositol (1,4,5) trisphosphate (IP3) receptors to regulate their physiological release of intracellular Ca2+. Here, Solomon Snyder and colleagues describe how non-immunosuppressant as well as immunosuppressant immunophilin ligands are neurotrophic for numerous classes of damaged neurones, both in culture systems and intact animals. Their ability to stimulate functional regrowth of damaged sciatic, cortical cholinergic, dopamine and 5-HT neurones may have therapeutic relevance.
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PMID:Neural actions of immunophilin ligands. 950 98

Presynaptic serotonin (5-hydroxytryptamine, 5-HT) transporters (SERTs) mediate antidepressant-sensitive clearance of 5-HT following release. Although we have been aware for decades that SERT-mediated 5-HT clearance can be modulated by exogenous agents including serotonin-selective reuptake inhibitors, amphetamines, and cocaine, we have had little reason to speculate that SERT activity was actively controlled through endogenous pathways. Recent studies indicate that SERTs are likely to be trafficked to specific plasma membrane subdomains to achieve localized clearance of 5-HT, and that the number of SERTs resident in the plasma membrane is controlled through kinase- and phosphatase-linked pathways. In particular, roles for protein kinase C and phosphatase 2A become apparent through studies with enzyme activators and inhibitors in SERT-transfected cells, where SERT proteins are rapidly phosphorylated in parallel with transporter redistribution and loss of functional uptake capacity. Based on our findings, and the studies of others in native tissues and transfected cells, we propose a model whereby SERTs are organized in a macromolecular complex in the plasma membrane that may serve to locate reuptake activity near release sites. Although many elements of this model remain hypothetical, our findings suggest a much more dynamic picture of transporter-mediated 5-HT reuptake than typically described and suggest opportunities both for the development of new SERT regulatory agents and for the identification of regulatory pathways that may be compromised in mental illness.
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PMID:Regulated phosphorylation and trafficking of antidepressant-sensitive serotonin transporter proteins. 969 89

The involvement of adenosine on the development of time-dependent reversal of long-term potentiation (LTP) by low-frequency stimulation (LFS) was investigated at Schaffer collateral-CA1 synapses of rat hippocampal slices. A train of LFS (2 Hz, 10 min, 1200 pulses) had no long-term effects on synaptic transmission but produced lasting depression of previously potentiated responses. This reversal of LTP (depotentiation) was observed when the stimulus was delivered </=3 min after induction of LTP. However, application at 10 min after induction had no detectable effect on potentiation. This time-dependent reversal of LTP by LFS appeared to be mediated by extracellular adenosine, because it was mimicked by bath-applied adenosine and was specifically inhibited by the selective A(1) adenosine receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (100 nM). The effect of adenosine could be mimicked by 5-HT(1A) receptor agonist buspirone, but the LFS-induced depotentiation could not be antagonized by 5-HT(1A) receptor antagonist NAN-190. The source of extracellular adenosine in response to LFS appeared to be attributable to the efflux of cAMP. In addition, this LFS-induced depotentiation was blocked by bath application of adenylyl cyclase activator forskolin or injection of a cAMP analog Sp-adenosine cAMP (10 mM) into postsynaptic neurons. Moreover, the selective protein phosphatase 1 and 2A inhibitors okadaic acid and calyculin A prevented the LFS-induced depotentiation. These results thus suggest that increasing extracellular adenosine appears to underlie the LFS-induced depotentiation via acting on the A(1) receptor subtype to interrupt the cAMP-dependent biochemical processes leading to the LTP expression.
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PMID:A role for extracellular adenosine in time-dependent reversal of long-term potentiation by low-frequency stimulation at hippocampal CA1 synapses. 1055 82

There is growing evidence linking alterations in serotonergic signaling in the prefrontal cortex to the etiology of schizophrenia. Prefrontal pyramidal neurons are richly innervated by serotonergic fibers and express high levels of serotonergic 5-HT(2)-class receptors. It is unclear, however, how activation of these receptors modulates cellular activity. To help fill this gap, whole cell voltage-clamp and single-cell RT-PCR studies of acutely isolated layer V-VI prefrontal pyramidal neurons were undertaken. The vast majority (>80%) of these neurons had detectable levels of 5-HT(2A) or 5-HT(2C) receptor mRNA. Bath application of 5-HT(2) agonists inhibited voltage-dependent Ca(2+) channel currents. L-type Ca(2+) channels were a particularly prominent target of this signaling pathway. The L-type channel modulation was blocked by disruption of G(alphaq) signaling or by inhibition of phospholipase Cbeta. Antagonism of intracellular inositol trisphosphate signaling, chelation of intracellular Ca(2+), or depletion of intracellular Ca(2+) stores also blocked this modulation. Inhibition of the Ca(2+)-dependent phosphatase calcineurin prevented receptor-mediated modulation of L-type currents. Last, the 5-HT(2) receptor modulation was robustly expressed in neurons from Ca(v)1.3 knockout mice. These findings argue that 5-HT(2) receptors couple through G(alphaq) proteins to trigger a phospholipase Cbeta/inositol trisphosphate signaling cascade resulting in the mobilization of intracellular Ca(2+), activation of calcineurin, and inhibition of Ca(v)1.2 L-type Ca(2+) currents. This modulation and its blockade by atypical neuroleptics could have wide-ranging effects on synaptic integration and long-term gene expression in deep-layer prefrontal pyramidal neurons.
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PMID:Stimulation of 5-HT(2) receptors in prefrontal pyramidal neurons inhibits Ca(v)1.2 L type Ca(2+) currents via a PLCbeta/IP3/calcineurin signaling cascade. 1197 86

The aim of the present study was to characterize the influence of the phosphatase type 1 and 2A inhibitor okadaic acid on non-adrenergic, non-cholinergic (NANC) neurotransmission in the rat gastric fundus. Okadaic acid (10-6 M), an inhibitor of protein phosphatases 1 and 2A, did not show any influence on the basal tonus or on a contraction plateau induced by 5-HT (10-7 M) within 30 min of observation. When okadaic acid (10-6 M) was applied 10 min prior to 5-HT (10-7 M), the contraction plateau of serotonin was unchanged. To investigate the inhibitory neurotransmission, the muscle strips were pre-contracted using 5-HT (10-7 M), and inhibitory stimuli were applied at the contraction plateau, which was stable over 30 min. The inhibitory effects of vasoactive intestinal peptide (VIP), nitric oxide (NO) and electrical field stimulation (EFS, 40 V, 0.5 ms, frequencies ranging from 0.5 to 16 Hz) were examined. When okadaic acid (10-6 M) was applied prior to EFS-induced NANC relaxation, significant attenuation of the inhibitory response was demonstrated (16 Hz: control: -92.4 +/- 1.9%; okadaic acid 10-7 M: -60.7 +/- 6.1%; okadaic acid 10-6 M: -25.3 +/- 3.4%; n=11; P < 0.01). By contrast, neither the concentration-dependent inhibitory actions of VIP (10-11-10-8 M) (VIP 10-8 M: -100%; VIP 10-8 M + okadaic acid 10-6 M: -89.9 +/- 8.3%; n=8; n.s) nor that of diethylamine nitric oxide (DEA-NO) (3 x 10-7-10-4 M) (DEA-NO 10-4 M: -95.3 +/- 8.4%; DEA-NO 10-4 M + okadaic acid 10-7 M: -98.3 +/- 6.3%; DEA-NO 10-4 M + okadaic acid 10-6 M: 96.5 +/- 7.6%; n=9; n.s.) on 5-HT induced contraction were altered by pre-incubation with okadaic acid (10-6 M). This is the first report that supports the concept that protein phosphatases 1 and 2A may contribute to the regulation of rat gastric fundus motility. The protein phosphatase inhibitor okadaic acid significantly reduces electrically induced inhibitory NANC responses, while leaving direct muscular effects of the inhibitory NANC neurotransmitters VIP and NO unaffected - suggesting a neural site of action. The potential roles of protein phosphatases on NANC neurotransmission remain to be clarified in detail, as this might offer a new pathway for modulating smooth-muscle function.
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PMID:Okadaic acid inhibits relaxant neural transmission in rat gastric fundus in vitro. 1198 2


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