Biomedical subjects

A Nishi

Publications and source records attributed to A Nishi.

At least 19 recordsLinked to original sources

PubMed2006

Role of adenosine A1 receptors in the modulation of dopamine D1 and adenosine A2A receptor signaling in the neostriatum.

Adenosine is known to modulate the function of neostriatal neurons. Adenosine acting on A(2A) receptors increases the phosphorylation of dopamine- and cAMP-regulated phosphoprotein of M(r) 32 kDa (DARPP-32) at Thr34 (the cAMP-dependent protein kinase [PKA] site) in striatopallidal neurons, and opposes dopamine D2 receptor signaling. In contrast, the role of adenosine A(1) receptors in the regulation of dopamine/DARPP-32 signaling is not clearly understood. Here, we investigated the effect of adenosine A(1) receptors on D(1), D(2) and A(2A) receptor signaling using mouse neostriatal slices. An A(1) receptor agonist, 2-chloro-N(6)-cyclopentyladenosine (100 nM), caused a transient increase, followed by a transient decrease, in DARPP-32 Thr34 phosphorylation. Our data support the following model for the actions of the A(1) receptor agonist. The A(1) receptor-induced early increase in Thr34 phosphorylation was mediated by presynaptic inhibition of dopamine release, and the subsequent removal of tonic inhibition by D(2) receptors of A(2A) receptor/G(olf)/cAMP/PKA signaling. The A(1) receptor-induced late decrease in Thr34 phosphorylation was mediated by a postsynaptic G(i) mechanism, resulting in inhibition of D(1) and A(2A) receptor-coupled G(olf)/cAMP/PKA signaling in direct and indirect pathway neurons, respectively. In conclusion, A(1) receptors play a major modulatory role in dopamine and adenosine receptor signaling.

Adenosine↗

PubMed2004

A network of control mediated by regulator of calcium/calmodulin-dependent signaling.

Calmodulin (CaM) is a major effector for the intracellular actions of Ca2+ in nearly all cell types. We identified a CaM-binding protein, designated regulator of calmodulin signaling (RCS). G protein-coupled receptor (GPCR)-dependent activation of protein kinase A (PKA) led to phosphorylation of RCS at Ser55 and increased its binding to CaM. Phospho-RCS acted as a competitive inhibitor of CaM-dependent enzymes, including protein phosphatase 2B (PP2B, also called calcineurin). Increasing RCS phosphorylation blocked GPCR- and PP2B-mediated suppression of L-type Ca2+ currents in striatal neurons. Conversely, genetic deletion of RCS significantly increased this modulation. Through a molecular mechanism that amplifies GPCR- and PKA-mediated signaling and attenuates GPCR- and PP2B-mediated signaling, RCS synergistically increases the phosphorylation of key proteins whose phosphorylation is regulated by PKA and PP2B.

Animals↗

PubMed2001

Regulation of cyclin-dependent kinase 5 and casein kinase 1 by metabotropic glutamate receptors.

Cyclin-dependent kinase 5 (Cdk5) is a multifunctional neuronal protein kinase that is required for neurite outgrowth and cortical lamination and that plays an important role in dopaminergic signaling in the neostriatum through phosphorylation of Thr-75 of DARPP-32 (dopamine and cAMP-regulated phosphoprotein, molecular mass 32 kDa). Casein kinase 1 (CK1) has been implicated in a variety of cellular functions such as DNA repair, circadian rhythm, and intracellular trafficking. In the neostriatum, CK1 has been found to phosphorylate Ser-137 of DARPP-32. However, first messengers for the regulation of Cdk5 or CK1 have remained unknown. Here we report that both Cdk5 and CK1 are regulated by metabotropic glutamate receptors (mGluRs) in neostriatal neurons. (S)-3,5-dihydroxyphenylglycine (DHPG), an agonist for group I mGluRs, increased Cdk5 and CK1 activities in neostriatal slices, leading to the enhanced phosphorylation of Thr-75 and Ser-137 of DARPP-32, respectively. The effect of DHPG on Thr-75, but not on Ser-137, was blocked by a Cdk5-specific inhibitor, butyrolactone. In contrast, the effects of DHPG on both Thr-75 and Ser-137 were blocked by CK1-7 and IC261, specific inhibitors of CK1, suggesting that activation of Cdk5 by mGluRs requires CK1 activity. In support of this possibility, the DHPG-induced increase in Cdk5 activity, measured in extracts of neostriatal slices, was abolished by CK1-7 and IC261. Treatment of acutely dissociated neurons with DHPG enhanced voltage-dependent Ca(2+) currents. This enhancement was eliminated by either butyrolactone or CK1-7 and was absent in DARPP-32 knockout mice. Together these results indicate that a CK1-Cdk5-DARPP-32 cascade may be involved in the regulation by mGluR agonists of Ca(2+) channels.

Animals↗

PubMed2001

Effects of chronic exposure to cocaine are regulated by the neuronal protein Cdk5.

Cocaine enhances dopamine-mediated neurotransmission by blocking dopamine re-uptake at axon terminals. Most dopamine-containing nerve terminals innervate medium spiny neurons in the striatum of the brain. Cocaine addiction is thought to stem, in part, from neural adaptations that act to maintain equilibrium by countering the effects of repeated drug administration. Chronic exposure to cocaine upregulates several transcription factors that alter gene expression and which could mediate such compensatory neural and behavioural changes. One such transcription factor is DeltaFosB, a protein that persists in striatum long after the end of cocaine exposure. Here we identify cyclin-dependent kinase 5 (Cdk5) as a downstream target gene of DeltaFosB by use of DNA array analysis of striatal material from inducible transgenic mice. Overexpression of DeltaFosB, or chronic cocaine administration, raised levels of Cdk5 messenger RNA, protein, and activity in the striatum. Moreover, injection of Cdk5 inhibitors into the striatum potentiated behavioural effects of repeated cocaine administration. Our results suggest that changes in Cdk5 levels mediated by DeltaFosB, and resulting alterations in signalling involving D1 dopamine receptors, contribute to adaptive changes in the brain related to cocaine addiction.

Animals↗

PubMed2001

XAFS study on active Pr sites in zeolite as a photocatalyst for decomposition of nitrous oxide.

The praseodymium-exchanged mordenite degassed at 873 K acted as a highly active heterogeneous photocatalyst for the stoichiometric decomposition of N2O. Analysis of Pr L3-edge EXAFS revealed that the Pr species were isolated each other in the large channel of zeolite, and that the active Pr species were coordinated by ca. four oxygen atoms and one Al atom. On the other hands, the inactive Pr species are surrounded by ca. 10 oxygens of adsorbed waters. The former low-coordinated Pr species allowed a strong adsorption of N2O, which could be the first step of the decomposition reaction.

Journal Article↗

PubMed2001

Phosphorylation of protein phosphatase inhibitor-1 by Cdk5.

Protein phosphatase inhibitor-1 is a prototypical mediator of cross-talk between protein kinases and protein phosphatases. Activation of cAMP-dependent protein kinase results in phosphorylation of inhibitor-1 at Thr-35, converting it into a potent inhibitor of protein phosphatase-1. Here we report that inhibitor-1 is phosphorylated in vitro at Ser-67 by the proline-directed kinases, Cdk1, Cdk5, and mitogen-activated protein kinase. By using phosphorylation state-specific antibodies and selective protein kinase inhibitors, Cdk5 was found to be the only kinase that phosphorylates inhibitor-1 at Ser-67 in intact striatal brain tissue. In vitro and in vivo studies indicated that phospho-Ser-67 inhibitor-1 was dephosphorylated by protein phosphatases-2A and -2B. The state of phosphorylation of inhibitor-1 at Ser-67 was dynamically regulated in striatal tissue by glutamate-dependent regulation of N-methyl-d-aspartic acid-type channels. Phosphorylation of Ser-67 did not convert inhibitor-1 into an inhibitor of protein phosphatase-1. However, inhibitor-1 phosphorylated at Ser-67 was a less efficient substrate for cAMP-dependent protein kinase. These results demonstrate regulation of a Cdk5-dependent phosphorylation site in inhibitor-1 and suggest a role for this site in modulating the amplitude of signal transduction events that involve cAMP-dependent protein kinase activation.

Animals↗

PubMed2001

Liver abscess caused by Clostridium difficile.

We report the first case of an infected cyst and liver abscess caused by Clostridium difficile. It recurred 11 months later, despite therapy with vancomycin and percutaneous drainage. Administration of metronidazole following percutaneous drainage achieved a favorable outcome.

Aged↗

PubMed2001

Biochemical and genetic analysis of the effects of amylose-extender mutation in rice endosperm.

Biochemical analysis of amylose-extender (ae) mutant of rice (Oryza sativa) revealed that the mutation in the gene for starch-branching enzyme IIb (BEIIb) specifically altered the structure of amylopectin in the endosperm by reducing short chains with degree of polymerization of 17 or less, with the greatest decrease in chains with degree of polymerization of 8 to 12. The extent of such change was correlated with the gelatinization properties of the starch granules, as determined in terms of solubility in urea solution. The ae mutation caused a dramatic reduction in the activity of BEIIb. The activity of soluble starch synthase I (SSI) in the ae mutant was significantly lower than in the wild type, suggesting that the mutation had a pleiotropic effect on the SSI activity. In contrast, the activities of BEI, BEIIa, ADP-Glc pyrophosphorylase, isoamylase, isoamylase, pullulanase, and Suc synthase were not affected by the mutation. Therefore, it is stressed that the function of BEIIb cannot be complemented by BEIIa and BEI. These results strongly suggest that BEIIb plays a specific role in the transfer of short chains, which might then be extended by SS to form the A and B(1) chains of amylopectin cluster in rice endosperm.

1,4-alpha-Glucan Branching Enzyme↗

PubMed2000

Phosphorylation of protein phosphatase-1 inhibitors, inhibitor-1 and DARPP-32, in renal medulla.

Inhibitor-1 and DARPP-32 (dopamine and cAMP-regulated phosphoprotein, Mr 32 kDa) are each phosphorylated by cAMP-dependent protein kinase, resulting in their conversion to potent inhibitors of protein phosphatase-1. Protein phosphatase-1 is involved in the regulation of Na(+) reabsorption from renal tubule by modulating the activity of Na(+),K(+)-ATPase. In this study, we have investigated the regulation of inhibitor-1 and DARPP-32 phosphorylation in slices of renal medulla. Activation of cAMP-dependent protein kinase by forskolin and 8-bromo-cAMP increased the level of phosphorylated inhibitor-1. Okadaic acid (1 microM), used to inhibit protein phosphatase-2A, increased the level of phosphorylated inhibitor-1, but cyclosporin A had no effect. DARPP-32, like inhibitor-1, was phosphorylated by cAMP-dependent protein kinase and dephosphorylated only by protein phosphatase-2A. These data demonstrate that the phosphorylation of inhibitor-1 and DARPP-32 is regulated by the balance of phosphorylation by cAMP-dependent protein kinase and dephosphorylation by protein phosphatase-2A in renal medulla. Furthermore, the phosphorylation step is regulated by pharmacological stimuli such as activation of beta(1)-adrenoceptors and dopamine D1 receptors.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed2000

Amplification of dopaminergic signaling by a positive feedback loop.

Dopamine and cAMP-regulated phosphoprotein of M(r) 32,000 (DARPP-32) plays an obligatory role in most of the actions of dopamine. In resting neostriatal slices, cyclin-dependent kinase 5 (Cdk5) phosphorylates DARPP-32 at Thr-75, thereby reducing the efficacy of dopaminergic signaling. We report here that dopamine, in slices, and acute cocaine, in whole animals, decreases the state of phosphorylation of striatal DARPP-32 at Thr-75 and thereby removes this inhibitory constraint. This effect of dopamine is achieved through dopamine D1 receptor-mediated activation of cAMP-dependent protein kinase (PKA). The activated PKA, by decreasing the state of phosphorylation of DARPP-32-Thr-75, de-inhibits itself. Dopamine D2 receptor stimulation has the opposite effect. The ability of activated PKA to reduce the state of phosphorylation of DARPP-32-Thr-75 is apparently attributable to increased protein phosphatase-2A activity, with Cdk5 being unaffected. Together, these results indicate that via positive feedback mechanisms, Cdk5 signaling and PKA signaling are mutually antagonistic.

Animals↗

PubMed2000

Biliary atresia with extrahepatic biliary cysts--cholangiographic patterns influencing the prognosis.

PURPOSE: Biliary atresia (BA) with extrahepatic biliary cysts (EHBC) has been recognized generally as "correctable" BA, which indicates a good prognosis. The variants of BA with EHBC according to cholangiographic findings and their outcomes were reviewed. METHODS: An EHBC was observed in 8 (20%) of 40 patients with BA who underwent operation at our institute. Intraoperative cholangiographic patterns included visualization of the intrahepatic bile ducts (type I BA with EHBC) in 6 patients and no visualization (type III BA with EHBC) in 2. Intrahepatic biliary cysts (IHBC) and EHBC were observed simultaneously in 2 patients diagnosed at older age. The follow-up periods ranged between 4 months and 20 years. RESULTS: Good bile drainage after a hepaticoenterostomy or portoenterostomy was obtained in all 6 patients with type I BA with EHBC. Two who showed IHBC on intraoperative cholangiography had complications caused by postoperative recurrent cholangitis, which led to a liver transplantation in 1. Revision after the portoenterostomy was required in 2 patients with type III BA with EHBC. One became jaundice free after revision, whereas the other died of hepatic failure without bile drainage. CONCLUSION: Intraoperative cholangiographic findings showing IHBC and type III BA are poor prognostic factors in patients with BA with EHBC.

Biliary Atresia↗

PubMed2000

A 90-kilobase conjugative chromosomal element coding for biphenyl and salicylate catabolism in Pseudomonas putida KF715.

The biphenyl and salicylate metabolic pathways in Pseudomonas putida KF715 are chromosomally encoded. The bph gene cluster coding for the conversion of biphenyl to benzoic acid and the sal gene cluster coding for the salicylate meta-pathway were obtained from the KF715 genomic cosmid libraries. These two gene clusters were separated by 10-kb DNA and were highly prone to deletion when KF715 was grown in nutrient medium. Two types of deletions took place at the region including only the bph genes (ca. 40 kb) or at the region including both the bph and sal genes (ca. 70 kb). A 90-kb DNA region, including both the bph and sal genes (termed the bph-sal element), was transferred by conjugation from KF715 to P. putida AC30. Such transconjugants gained the ability to grow on biphenyl and salicylate as the sole sources of carbon. The bph and sal element was located on the chromosome of the recipient. The bph-sal element in strain AC30 was also highly prone to deletion; however, it could be mobilized to the chromosome of P. putida KT2440 and the two deletion mutants of KF715.

Base Sequence↗

PubMed1999

Phosphorylation of DARPP-32 by Cdk5 modulates dopamine signalling in neurons.

The physiological state of the cell is controlled by signal transduction mechanisms which regulate the balance between protein kinase and protein phosphatase activities. Here we report that a single protein can, depending on which particular amino-acid residue is phosphorylated, function either as a kinase or phosphatase inhibitor. DARPP-32 (dopamine and cyclic AMP-regulated phospho-protein, relative molecular mass 32,000) is converted into an inhibitor of protein phosphatase 1 when it is phosphorylated by protein kinase A (PKA) at threonine 34. We find that DARPP-32 is converted into an inhibitor of PKA when phosphorylated at threonine 75 by cyclin-dependent kinase 5 (Cdk5). Cdk5 phosphorylates DARPP-32 in vitro and in intact brain cells. Phospho-Thr 75 DARPP-32 inhibits PKA in vitro by a competitive mechanism. Decreasing phospho-Thr 75 DARPP-32 in striatal slices, either by a Cdk5-specific inhibitor or by using genetically altered mice, results in increased dopamine-induced phosphorylation of PKA substrates and augmented peak voltage-gated calcium currents. Thus DARPP-32 is a bifunctional signal transduction molecule which, by distinct mechanisms, controls a serine/threonine kinase and a serine/threonine phosphatase.

Animals↗

PubMed1999

Mobile catabolic genes in bacteria.

The recent findings of various mobile catabolic genes have provided some insight into the evolution of microbial degradation systems for xenobiotic compounds. The catabolic genes undergo marked genetic rearrangements due to their presence on transposons or association with mobile genetic elements. Bacterial catabolic transposons fall into three defined structural classes. Class I elements include catabolic genes flanked by two copies of insertion sequences. Class II elements carry short terminal inverted repeats and transpose by the replicative mode in which transposase and resolvase are involved. Conjugative catabolic transposons represent the third class of mobile genetic elements. They carry all the genes required for excision, conjugal transfer to a new host, and integration. This review focuses on the structures, functions and roles of the recently characterized catabolic transposons in bacteria. Also described are the mobile catabolic elements that share structural similarity with the pathogenicity and symbiosis islands.

Journal Article↗

PubMed1999

Engineering a hybrid pseudomonad to acquire 3,4-dioxygenase activity for polychlorinated biphenyls.

We constructed a hybrid strain that acquired 3,4-dioxygenase activity for polychlorinated biphenyls (PCBs). This strain, KF707-D34, possessed a chimeric biphenyl dioxygenase gene, of which a portion of bphA1 (coding for a large subunit of biphenyl dioxygenase) of Pseudomonas pseudoalcaligenes KF707 was replaced with that of a PCB-degrader, Burkholderia cepacia LB400 by homologous recombination. KF707-D34 retained the ability to degrade 4,4'-dichlorobiphenyl via 2,3-dioxygenation in a fashion identical to that of KF707 and gained novel capability to degrade 2,5,4'-trichlorobiphenyl and 2,5,2',5'-tetrachlorobiphenyl via 3,4-dioxygenation in a fashion identical to that of LB400. Sequence analysis of bphA1 from KF707-D34 revealed that three nucleotides in the 3'-terminal region of KF707 bphA1 were changed to correspond to those in LB400 bphA1. The resulting BphA1 protein in KF707-D34 was changed at position 376 from threonine (Thr) to asparagine (Asn). The results demonstrate that a minor alteration of the amino acid sequence in BphA1 improved the PCB degradation capability in biphenyl-utilizing bacteria.

Journal Article↗

PubMed1999

Requirement for DARPP-32 in mediating effect of dopamine D2 receptor activation.

It is well documented that dopamine and dopamine D1 agonists convert the protein phosphatase-1 inhibitor, DARPP-32, from its dephosphorylated, inactive form into its Thr34-phosphorylated, active form, and that these effects on DARPP-32 constitute essential components of the mechanism by which dopamine and D1 agonists achieve their biological effects. In contrast to dopamine and D1 agonists, dopamine D2 agonists dephosphorylate and inactivate DARPP-32. Here we have examined the possibility that the biological effects of dopamine D2 receptor agonists might also involve DARPP-32. For this purpose, we have examined regulation of the activity of the electrogenic ion pump Na+,K+-ATPase, an established target for dopamine signalling. We have found that dopamine D1 agonists and dopamine D2 agonists inhibit Na+,K+-ATPase activity in dissociated cells from the mouse neostriatum and that, in each case, the effect is abolished in cells from mice deficient in DARPP-32. We conclude that DARPP-32 may play an obligatory role in dopaminergic signalling mediated both by D1 receptors and by D2 receptors.

Animals↗

PubMed1999

Role of calcineurin and protein phosphatase-2A in the regulation of DARPP-32 dephosphorylation in neostriatal neurons.

DARPP-32, a dopamine- and cyclic AMP-regulated phosphoprotein of Mr 32 kDa, is phosphorylated on Thr34 by cyclic AMP-dependent protein kinase, resulting in its conversion to a potent inhibitor of protein phosphatase-1 (PP-1). Conversely, Thr34-phosphorylated DARPP-32 is dephosphorylated and inactivated in vitro by calcineurin and protein phosphatase-2A (PP-2A). We have investigated the relative contributions of these protein phosphatases to the regulation of DARPP-32 dephosphorylation in mouse neostriatal slices. Cyclosporin A (5 microM), a calcineurin inhibitor, maximally increased the level of phosphorylated DARPP-32 by 17+/-2-fold. Okadaic acid (1 microM), an inhibitor of PP-1 and PP-2A, had a smaller effect, increasing phospho-DARPP-32 by 5.1+/-1.3-fold. The effect of okadaic acid on DARPP-32 phosphorylation was shown to be due to inhibition of PP-2A activity. Incubation of slices in the presence of cyclosporin A plus either okadaic acid or calyculin A, another PP-1/PP-2A inhibitor, caused a synergistic increase in the level of phosphorylated DARPP-32. The use of Ca2(+)-free/EGTA medium mimicked the effects of cyclosporin A on DARPP-32 phosphorylation, supporting the conclusion that the action of cyclosporin on DARPP-32 phosphorylation was attributable to blockade of the Ca2(+)-dependent activation of calcineurin. The results indicate that calcineurin and PP-2A, but not PP-1, act synergistically to maintain a low level of phosphorylated DARPP-32 in neostriatal slices.

Animals↗

PubMed1999

Regulation of Na+, K+-ATPase isoforms in rat neostriatum by dopamine and protein kinase C.

Our previous studies showed that dopamine inhibits Na+,K+-ATPase activity in acutely dissociated neurons from striatum. In the present study, we have found that in this preparation, dopamine inhibited significantly (by approximately 25%) the activity of the alpha3 and/or alpha2 isoforms, but not the alpha1 isoform, of Na+,K+-ATPase. Dopamine, via D1 receptors, activates cyclic AMP-dependent protein kinase (PKA) in striatal neurons. Dopamine is also known to activate the calcium- and phospholipid-dependent protein kinase (PKC) in a number of different cell types. The PKC activator phorbol 12,13-dibutyrate reduced the activity of Na+,K+-ATPase alpha3 and/or alpha2 isoforms (by approximately 30%) as well as the alpha1 isoform (by approximately 15%). However, dopamine-mediated inhibition of Na+,K+-ATPase activity was unaffected by calphostin C, a PKC inhibitor. Dopamine did not affect the phosphorylation of Na+,K+-ATPase isoforms at the PKA-dependent phosphorylation site. Phorbol ester treatment did not alter the phosphorylation of alpha2 or alpha3 isoforms of Na+,K+-ATPase in neostriatal neurons but did increase the phosphorylation of the alpha1 isoform. Thus, in rat neostriatal neurons, treatment with either dopamine or PKC activators results in inhibition of the activity of specific (alpha3 and/or alpha2) isoforms of Na+,K+-ATPase, but this is not apparently mediated through direct phosphorylation of the enzyme. In addition, PKC is unlikely to mediate inhibition of rat Na+,K+-ATPase activity by dopamine in neostriatal neurons.

Animals↗