17 research outputs found
The role of Galpha(s) in heterologous sensitization of adenylyl cyclase
No full textAcute activation of Gαi/o-coupled receptors inhibits adenylyl cyclase, whereas persistent activation of Gαi/o-coupled receptors results in a compensatory sensitization of adenylyl cyclase activity following subsequent activation. Several indirect observations have suggested the involvement of enhanced Gαs adenylyl cyclase interactions in the expression of sensitization, however, evidence supporting a direct role for Gαs has not been well established. We have used several genetic approaches to further examine the role of Gαs in heterologous sensitization of adenylyl cyclase. In the first approach, Gαs insensitive mutants of AC1 (F293L and Y973S) were constructed, and these AC1 mutants retained sensitivity to Ca2+ and forskolin activation. Persistent (2 h) activation of the D2 dopamine receptor resulted in a significant augmentation of basal and drug-stimulated AC1 activity, whereas sensitization of AC1 F293L and Y973S was markedly reduced. In the second strategy, the requirement of an intact receptor-Gαs signaling pathway for the expression heterologous sensitization was examined using dominant negative Gαs mutants (α3β5 G226A/A366S or α3β5 G226A/E268A/A366S) that disrupt D1 dopamine receptor activation of AC1. D1 dopamine receptor-Gα s signaling was blocked in the presence of these Gαs mutants, but D2 agonist-induced sensitization of Ca2+-stimulated AC1 activity was not altered. Subsequent studies examined the absolute requirement of Gαs in sensitization of recombinant AC1 and AC5 using a novel Gαs-deficient (GnasE2−/E2− ) cell line. Persistent activation (18 h) of the D2 dopamine receptor markedly enhanced subsequent forskolin and Ca2+-stimulated AC1 activity in GnasE2−/E2− cells, whereas sensitization of forskolin-stimulated AC5 activity occurred only in the presence of transfected Gαs. The physiological significance of D2 dopamine receptor-induced sensitization of adenylyl cyclase was also examined in the present report. D2 dopamine receptors are colocalized with Gαs-coupled A2A adenosine receptors on GABAergic neurons in the striatum and their functional interactions have been implicated in the pharmacotherapy of Parkinson\u27s disease. We have confirmed that acute activation of the D 2 dopamine receptor inhibits A2A adenosine receptor-stimulated cyclic AMP accumulation. In contrast, persistent activation of the D 2 dopamine receptor resulted in sensitization of neuronal A2A adenosine receptor signaling. Taken together, these observations suggest a differential role for Gαs in the expression of heterologous sensitization of adenylyl cyclase, which may be involved in neuropsychiatric disorders associated with increased D2 dopamine receptor signaling
Nigrostriatal dopaminergic deficits and hypokinesia caused by inactivation of the familial parkinsonism-linked gene DJ-1
No full textThe manifestations of Parkinson's disease are caused by reduced dopaminergic innervation of the striatum. Loss-of-function mutations in the DJ-1 gene cause early-onset familial parkinsonism. To investigate a possible role for DJ-1 in the dopaminergic system, we generated a mouse model bearing a germline disruption of DJ-1. Although DJ-1(-/-) mice had normal numbers of dopaminergic neurons in the substantia nigra, evoked dopamine overflow in the striatum was markedly reduced, primarily as a result of increased reuptake. Nigral neurons lacking DJ-1 were less sensitive to the inhibitory effects of D2 autoreceptor stimulation. Corticostriatal long-term potentiation was normal in medium spiny neurons of DJ-1(-/-) mice, but long-term depression (LTD) was absent. The LTD deficit was reversed by treatment with D2 but not D1 receptor agonists. Furthermore, DJ-1(-/-) mice displayed hypoactivity in the open field. Collectively, our findings suggest an essential role for DJ-1 in dopaminergic physiology and D2 receptor-mediated functions
China's Health Reform Update
No full textChina experienced both economic and epistemological transitions within the past few decades, greatly increasing demand for accessible and affordable health care. These shifts put significant pressure on the existing outdated, highly centralized bureaucratic system. Adjusting to growing demands, the government has pursued a new round of health reforms since the late 2000s; the main goals are to reform health care financing, essential drug policies, and public hospitals. Health care financing reform led to universal basic medical insurance, whereas the public hospital reform required more complex measures ranging from changes in regulatory, operational, and service delivery settings to personnel management. This article reviews these major policy changes and the literature-based evidence of the effects of reforms on cost, access, and quality of care. It then highlights the outlook for future reforms. We argue that a better understanding of the unintended consequences of reform policies and of how practitioners' and patients' interests can be better aligned is essential for reforms to succeed.SCI(E)SSCIREVIEW431-4483
Dopaminergic and Behavioral Deficits Caused by Inactivation of the Familial Parkinsonism-Linked Gene DJ-1
No full textThe manifestations of Parkinson's disease are caused by reduced dopaminergic innervation of the striatum. Loss-of-function mutations in the DJ-1 gene cause early-onset familial parkinsonism. To investigate a possible role for DJ-1 in the dopaminergic system, we generated a mouse model bearing a germline disruption of DJ-1. Although DJ-1(-/-) mice had normal numbers of dopaminergic neurons in the substantia nigra, evoked dopamine overflow in the striatum was markedly reduced, primarily as a result of increased reuptake. Nigral neurons lacking DJ-1 were less sensitive to the inhibitory effects of D2 autoreceptor stimulation. Corticostriatal long-term potentiation was normal in medium spiny neurons of DJ-1(-/-) mice, but long-term depression (LTD) was absent. The LTD deficit was reversed by treatment with D2 but not D1 receptor agonists. Furthermore, DJ-1(-/-) mice displayed hypoactivity in the open field. Collectively, our findings suggest an essential role for DJ-1 in dopaminergic physiology and D2 receptor-mediated functions
Adrenergic agonists induce heterologous sensitization of adenylate cyclase in NS20Y-D2L cells
No full textAbstractAdenylate cyclase activity in NS20Y cells expressing D2L dopamine receptors was examined following chronic treatment with norepinephrine and epinephrine. Initial acute experiments revealed that both norepinephrine and epinephrine inhibited forskolin-stimulated cyclic AMP accumulation via D2 receptors. Furthermore, chronic (18 h) activation of D2 dopamine receptors by norepinephrine or epinephrine induced a marked increase (>10-fold) in subsequent forskolin-stimulated cyclic AMP accumulation. This heterologous sensitization of adenylate cyclase activity was blocked by D2 dopamine receptor antagonists and by pertussis toxin pretreatment. In contrast, concurrent activation of Gαs or adenylate cyclase did not appear to alter noradrenergic agonist-induced sensitization
Differential Helical Orientations among Related G Protein-coupled Receptors Provide a Novel Mechanism for Selectivity: STUDIES WITH SALVINORIN A AND THE κ-OPIOID RECEPTOR
No full textSalvinorin A, the active component of the hallucinogenic sage Salvia divinorum, is an apparently selective and highly potent kappa-opioid receptor (KOR) agonist. Salvinorin A is unique among ligands for peptidergic G protein-coupled receptors in being nonnitrogenous and lipid-like in character. To examine the molecular basis for the subtype-selective binding of salvinorin A, we utilized an integrated approach using chimeric opioid receptors, site-directed mutagenesis, the substituted cysteine accessibility method, and molecular modeling and dynamics studies. We discovered that helix 2 is required for salvinorin A binding to KOR and that two residues (Val-108(2.53) and Val-118(2.63)) confer subtype selectivity. Intriguingly, molecular modeling studies predicted that these loci exhibit an indirect effect on salvinorin A binding, presumably through rotation of helix 2. Significantly, and in agreement with our in silico predictions, substituted cysteine accessibility method analysis of helix 2 comparing KOR and the delta-opioid receptor, which has negligible affinity for salvinorin A, revealed that residues known to be important for salvinorin A binding exhibit a differential pattern of water accessibility. These findings imply that differences in the helical orientation of helix 2 are critical for the selectivity of salvinorin A binding to KOR and provide a structurally novel basis for ligand selectivity
Autoxidation of Salvinorin A under Basic Conditions
No full textTreatment of salvinorin A (1a) with KOH in MeOH gave the enedione 3, for which the dienone
structure 7 was recently proposed. Also isolated, after methylation, were the secotriesters 4a−c. A
mechanism for this unusual series of autoxidations is proposed. Surprisingly, 4a showed weak
affinity at the κ-opioid receptor. Divinatorins A−C (2a−c) showed no affinity at opioid receptors.
Attempted reduction of 3 to a novel salvinorin diol (9d) was unsuccessful, but careful deacetylation
of salvinorin C (9a) provided a viable route to this compound. A general method for identifying
salvinorin 8-epimers by TLC is also presented
<i>cis</i>-4-(Piperazin-1-yl)-5,6,7a,8,9,10,11,11a-octahydrobenzofuro[2,3-<i>h</i>]quinazolin-2-amine (A-987306), A New Histamine H<sub>4</sub>R Antagonist that Blocks Pain Responses against Carrageenan-Induced Hyperalgesia
No full textcis-4-(Piperazin-1-yl)-5,6,7a,8,9,10,11,11a-octahydrobenzofuro[2,3-h]quinazolin-2-amine, 4 (A-987306) is a new histamine H4 antagonist. The compound is potent in H4 receptor binding assays (rat H4, Ki = 3.4 nM, human H4 Ki = 5.8 nM) and demonstrated potent functional antagonism in vitro at human, rat, and mouse H4 receptors in cell-based FLIPR assays. Compound 4 also demonstrated H4 antagonism in vivo in mice, blocking H4-agonist induced scratch responses, and showed anti-inflammatory activity in mice in a peritonitis model. Most interesting was the high potency and efficacy of this compound in blocking pain responses, where it showed an ED50 of 42 μmol/kg (ip) in a rat post-carrageenan thermal hyperalgesia model of inflammatory pain
<i>cis</i>-4-(Piperazin-1-yl)-5,6,7a,8,9,10,11,11a-octahydrobenzofuro[2,3-<i>h</i>]quinazolin-2-amine (A-987306), A New Histamine H<sub>4</sub>R Antagonist that Blocks Pain Responses against Carrageenan-Induced Hyperalgesia
No full textcis-4-(Piperazin-1-yl)-5,6,7a,8,9,10,11,11a-octahydrobenzofuro[2,3-h]quinazolin-2-amine, 4 (A-987306) is a new histamine H4 antagonist. The compound is potent in H4 receptor binding assays (rat H4, Ki = 3.4 nM, human H4 Ki = 5.8 nM) and demonstrated potent functional antagonism in vitro at human, rat, and mouse H4 receptors in cell-based FLIPR assays. Compound 4 also demonstrated H4 antagonism in vivo in mice, blocking H4-agonist induced scratch responses, and showed anti-inflammatory activity in mice in a peritonitis model. Most interesting was the high potency and efficacy of this compound in blocking pain responses, where it showed an ED50 of 42 μmol/kg (ip) in a rat post-carrageenan thermal hyperalgesia model of inflammatory pain
Discovery of 4-[(2<i>R</i>,4<i>R</i>)-4-({[1-(2,2-Difluoro-1,3-benzodioxol-5-yl)cyclopropyl]carbonyl}amino)-7-(difluoromethoxy)-3,4-dihydro-2<i>H</i>-chromen-2-yl]benzoic Acid (ABBV/GLPG-2222), a Potent Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Corrector for the Treatment of Cystic Fibrosis
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