Donepezil for the symptomatic treatment of patients with mild to moderate Alzheimer's disease: a meta-analysis of individual patient data from randomised controlled trials

Background: The objective was to evaluate the efficacy and tolerability of donepezil (5 and 10 mg/day) compared with placebo in alleviating manifestations of mild to moderate Alzheimer's disease (AD). Method: A systematic review of individual patient data from Phase II and III double-blind, randomised, placebo-controlled studies of up to 24 weeks and completed by 20 December 1999. The main outcome measures were the ADAS-cog, the CIBIC-plus, and reports of adverse events. Results: A total of 2376 patients from ten trials were randomised to either donepezil 5 mg/day (n = 821), 10 mg/day (n = 662) or placebo (n = 893). Cognitive performance was better in patients receiving donepezil than in patients receiving placebo. At 12 weeks the differences in ADAS-cog scores were 5 mg/day-placebo: - 2.1 [95% confidence interval (CI), - 2.6 to - 1.6; p < 0.001], 10 mg/day-placebo: - 2.5 ( - 3.1 to - 2.0; p < 0.001). The corresponding results at 24 weeks were - 2.0 ( - 2.7 to - 1.3; p < 0.001) and - 3.1 ( - 3.9 to - 2.4; p < 0.001). The difference between the 5 and 10 mg/day doses was significant at 24 weeks (p = 0.005). The odds ratios (OR) of improvement on the CIBIC-plus at 12 weeks were: 5 mg/day-placebo 1.8 (1.5 to 2.1; p < 0.001), 10 mg/day-placebo 1.9 (1.5 to 2.4; p < 0.001). The corresponding values at 24 weeks were 1.9 (1.5 to 2.4; p = 0.001) and 2.1 (1.6 to 2.8; p < 0.001). Donepezil was well tolerated; adverse events were cholinergic in nature and generally of mild severity and brief in duration. Conclusion: Donepezil (5 and 10 mg/day) provides meaningful benefits in alleviating deficits in cognitive and clinician-rated global function in AD patients relative to placebo. Increased improvements in cognition were indicated for the higher dose. Copyright © 2004 John Wiley & Sons, Ltd.

Identification of mutations conferring insecticide-insensitive AChE in the cotton-melon aphid, Aphis gossypii Glover

We have identified two mutations in the ace1 gene of Aphis gossypii that are associated with insensitivity of acetylcholinesterase (AChE) to carbamate and organophosphate insecticides. The first of these, S431F (equivalent to F331 in Torpedo californica), is associated with insensitivity to the carbamate insecticide pirimicarb in a range of A. gossypii clones. The S431F mutation is also found in the peach-potato aphid, Myzus persicae (Sulzer), and a rapid RFLP diagnostic allows the identification of individuals of both aphid species with a resistant genotype. This diagnostic further revealed the presence of S431 in several other pirimicarb-susceptible aphid species. The serine at this position in the wild-type enzyme has only been reported for aphids and provides a molecular explanation of why pirimicarb has a specific aphicidal action. A less specific insensitivity to a wide range of carbamates and organophosphates is associated with a second mutation, A302S (A201 in T. californica).

G protein ss gamma subunits mediate presynaptic inhibition of transmitter release from rat superior cervical ganglion neurones in culture

The activation of presynaptic G protein-coupled receptors (GPCRs) is widely reported to inhibit transmitter release; however, the lack of accessibility of many presynaptic terminals has limited direct analysis of signalling mediators. We studied GPCR-mediated inhibition of fast cholinergic transmission between superior cervical ganglion neurones (SCGNs) in culture. The adrenoceptor agonist noradrenaline (NA) caused a dose-related reduction in evoked excitatory postsynaptic potentials (EPSPs). NA-induced EPSP decrease was accompanied by effects on the presynaptic action potential (AP), reducing AP duration and amplitude of the after-hyperpolarization (AHP), without affecting the pre- and postsynaptic membrane potential. All effects of NA were blocked by yohimbine and synaptic transmission was reduced by clonidine, consistent with an action at presynaptic alpha 2-adrenoceptors. NA-induced inhibition of transmission was sensitive to pre-incubation of SCGNs with pertussis toxin (PTX), implicating the involvement of G alpha(i)/(o)beta y subunits. Expression of G alpha transducin, an agent which sequesters G protein beta gamma (G beta y) subunits, in the presynaptic neurone caused a time-dependent attenuation of NA-induced inhibition. Injection of purified G beta gamma subunits into the presynaptic neurone inhibited transmission, and also reduced the AHP amplitude. Furthermore, NA-induced inhibition was occluded by pre-injection of G beta gamma subunits. The Ca2+ channel blocker Cd2+ mimicked NA effects on transmitter release. Cd2+, NA and G beta gamma subunits also inhibited somatic Ca2+ current. In contrast to effects on AP-evoked transmitter release, NA had no clear action on AP-independent EPSPs induced by hypertonic solutions. These results demonstrate that G beta gamma subunits functionally mediate inhibition of transmitter release by alpha 2-adrenoceptors and represent important regulators of synaptic transmission at mammalian presynaptic terminals.

The cognitive neurochemistry of nicotine

The rational for this review is to provide a coherent formulation of the cognitive neurochemistry of nicotine, with the aim of suggesting research and clinical applications. The first part is a comprehensive review of the empirical studies of the enhancing effects of nicotine on information processing, especially those on attentional and mnemonic processing. Then, these studies are put in the context of recent studies on the neurochemistry of nicotine and cholinergic drugs, in general. They suggest a positive effect of nicotine on processes acting on encoded material during the post acquisition phase, the process of consolidation. Thus, the involvement of nicotinic receptors in mnemonic processing is modulation of the excitability of neurons in the hippocampal formation to enable associative processing.

The 'glial' glutamate transporter, EAAT2 (Glt-1) accounts for high affinity glutamate uptake into adult rodent nerve endings

The excitatory amino acid transporters (EAAT) removes neurotransmitters glutamate and aspartate from the synaptic cleft. Most CNS glutamate uptake is mediated by EAAT2 into glia, though nerve terminals show evidence for uptake, through an unknown transporter. Reverse-transcriptase PCR identified the expression of EAAT1, EAAT2, EAAT3 and EAAT4 mRNAs in primary cultures of mouse cortical or striatal neurones. We have used synaptosomes and glial plasmalemmal vesicles (GPV) from adult mouse and rat CNS to identify the nerve terminal transporter. Western blotting showed detectable levels of the transporters EAAT1 (GLAST) and EAAT2 (Glt-1) in both synaptosomes and GPVs. Uptake of [3H]D-aspartate or [3H]L-glutamate into these preparations revealed sodium-dependent uptake in GPV and synaptosomes which was inhibited by a range of EAAT blockers: dihydrokainate, serine-o-sulfate, l-trans-2,4-pyrrolidine dicarboxylate (PDC) (+/-)-threo-3-methylglutamate and (2S,4R )-4-methylglutamate. The IC50 values found for these compounds suggested functional expression of the 'glial, transporter, EAAT2 in nerve terminals. Additionally blockade of the majority EAAT2 uptake sites with 100 micro m dihydrokainate, failed to unmask any functional non-EAAT2 uptake sites. The data presented in this study indicate that EAAT2 is the predominant nerve terminal glutamate transporter in the adult rodent CNS.

The SV2A ligand levetiracetam inhibits presynaptic Ca2+ channels via an intracellular pathway

Levetiracetam (LEV) is a prominent antiepileptic drug (AED) which binds to neuronal synaptic vesicle glycoprotein 2A (SV2A) protein and has reported effects on ion channels, but retains a poorly-defined mechanism of action. Here, we investigate inhibition of voltage-dependent Ca2+ (CaV) channels as a potential mechanism by which LEV imparts effects on neuronal activity. We used electrophysiological methods to investigate the effects of LEV on cholinergic synaptic transmission and CaV channel activity in superior cervical ganglion neurons (SCGNs). In parallel, we investigated effects of the LEV ‘inactive’ R-enantiomer, UCB L060. Thus, LEV, but not UCB L060 (each 100 μM), inhibited synaptic transmission between SCGNs in long-term culture in a time-dependent manner, significantly reducing excitatory postsynaptic potentials (EPSP) following ≥30 min application. In isolated SCGNs, LEV pretreatment (≥1 h), but not acute (5 min) application, significantly inhibited whole-cell IBa amplitude. In current clamp recordings, LEV reduced the amplitude of the afterhyperpolarizing potential (AHP) in a Ca2+-dependent manner, but also increased action potential (AP) latency in a Ca2+-independent manner, suggesting further mechanisms associated with reduced excitability. Intracellular LEV application (4-5 min) caused a rapid inhibition of IBa amplitude to an extent comparable to that seen following extracellular LEV pretreatment ( ≥ 1 h). Neither pretreatment nor intracellular application of UCB L060 produced any inhibitory effects on IBa amplitude. These results identify a stereospecific intracellular pathway by which LEV inhibits presynaptic CaV channels; resultant reductions in neuronal excitability are proposed to contribute to the anticonvulsant effects of LEV.

Expression and function of the bile acid receptor GpBAR1 (TGR5) in the murine enteric nervous system

BACKGROUND: Bile acids (BAs) regulate cells by activating nuclear and membrane-bound receptors. G protein coupled bile acid receptor 1 (GpBAR1) is a membrane-bound G-protein-coupled receptor that can mediate the rapid, transcription-independent actions of BAs. Although BAs have well-known actions on motility and secretion, nothing is known about the localization and function of GpBAR1 in the gastrointestinal tract. METHODS: We generated an antibody to the C-terminus of human GpBAR1, and characterized the antibody by immunofluorescence and Western blotting of HEK293-GpBAR1-GFP cells. We localized GpBAR1 immunoreactivity (IR) and mRNA in the mouse intestine, and determined the mechanism by which BAs activate GpBAR1 to regulate intestinal motility. KEY RESULTS: The GpBAR1 antibody specifically detected GpBAR1-GFP at the plasma membrane of HEK293 cells, and interacted with proteins corresponding in mass to the GpBAR1-GFP fusion protein. GpBAR1-IR and mRNA were detected in enteric ganglia of the mouse stomach and small and large intestine, and in the muscularis externa and mucosa of the small intestine. Within the myenteric plexus of the intestine, GpBAR1-IR was localized to approximately 50% of all neurons and to >80% of inhibitory motor neurons and descending interneurons expressing nitric oxide synthase. Deoxycholic acid, a GpBAR1 agonist, caused a rapid and sustained inhibition of spontaneous phasic activity of isolated segments of ileum and colon by a neurogenic, cholinergic and nitrergic mechanism, and delayed gastrointestinal transit. CONCLUSIONS & INFERENCES: G protein coupled bile acid receptor 1 is unexpectedly expressed in enteric neurons. Bile acids activate GpBAR1 on inhibitory motor neurons to release nitric oxide and suppress motility, revealing a novel mechanism for the actions of BAs on intestinal motility.

Endogenous abscisic acid and protein contents during seed development of Araucaria angustifolia

This paper describes a proteome analysis and changes in endogenous abscisic acid (ABA) contents during seed development of Araucaria angustifolia (Bert.) O. Ktze. Megagametophytes and embryonic axis tissues exhibited a similar ABA variation pattern during seed development, reaching maximum values at the pre-cotyledonary stage. The embryonic axis protein content increased until the cotyledonary stage with following stabilization at mature seed. The two-dimensional electrophoresis at the torpedo developmental stage showed approximately 230 polypeptides against 340 in the mature stage. Peptide mass fingerprinting analyses identified three polypeptides, corresponding to an AtSAC4, a late embryogenesis abundant (LEA) and a storage protein, respectively.

Structural aspects of the zygotic embryogenesis of Acca sellowiana (O. Berg) Burret (Myrtaceae)

(Structural aspects of the zygotic embryogenesis of Acca sellowiana (O. Berg) Burret (Myrtaceae)). Acca sellowiana has anatropous, bitegmic and crassinucellate ovules. The outer and inner integuments are double-layered except in the micropyle, where they are composed of more layers; the micropyle is zig-zag shaped. The egg apparatus lies at the micropylar pole, and the zynergids present a conspicuous filiform apparatus. The antipodal cells are present in the chalazal region, persisting before the occurrence of double fertilization. The zygote is visible 21 days after pollination; nuclear endosperm is already present. The first mitotic division of the zygote occurs at 24(th) day. The globular, cordiform and torpedo embryo stages can be seen at 30, 45 and 60 days after pollination, respectively. The mature embryo characterized by the presence of a well-developed hypocotyl-radicular axis with two fleshy and folded cotyledons was observed 120 days after pollination. Endosperm is absent in the seeds, and the embryo has spiral form, characteristic of Myrtinae. The zygotic embryology studies of A. sellowiana indicate that this species has embryological characteristics which are in agreement with those reported for Myrtaceae (Myrteae, Myrtinae), and also broaden the knowledge about the sexual reproduction of this native species, whose commercial cultivation has been growing.

Changes in soluble carbohydrates and starch amounts during somatic and zygotic embryogenesis of Acca sellowiana (Myrtaceae)

Comparative analysis of zygotic and somatic embryogenesis of Acca sellowiana showed higher amounts of sucrose, fructose, raffinose, and myo-inositol in zygotic embryos at different developmental stages than in corresponding somatic ones. These differences were mostly constant. In general, glucose levels were significantly lower than the other soluble carbohydrates analyzed, showing minor variation in each embryo stage. Despite the presence of sucrose in the culture medium, its levels conspicuously diminished in somatic embryos compared with the zygotic ones. Raffinose enhanced parallel to embryo development, regardless of its zygotic or somatic origin. Analysis of the soluble carbohydrate composition of mature zygotic cotyledon used as explant pointed out fructose, glucose, myo-inositol, sucrose, and raffinose as the most important. Similar composition was also found in the corresponding somatic cotyledon. Total soluble carbohydrates varied inversely, decreasing in zygotic embryos and increasing in somatic embryos until the 24th d, at which time they increased rapidly about sixfold in zygotic embryos until the 27th d, a period coinciding with the zygotic proembryos formation. Such condition seems to reflect directly the variation of endogenous sucrose level, mainly because glucose and fructose diminished continuously during this time period. This means that, in terms of soluble sugars, zygotic embryo formation occurred under a situation represented by high sucrose amounts, simultaneously with low fructose and glucose levels, while in contrast, somatic embryo formation took place under an endogenous sugar status characterized by a substantial fructose enhancement. Starch levels increased continuously in zygotic embryos and decreased in somatic ones, the reverse to what was found in fructose variation. Starch accumulation was significantly higher in somatic torpedo and cotyledonary embryos than in the corresponding zygotic ones.

Thyroid hormones alter the adenine nucleotide hydrolysis in adult rat blood serum

The effects of ATP, ADP, and adenosine in the processes of platelet aggregation, vasodilatation, and coronary flow have been known for many years. The sequential hydrolysis of ATP to adenosine by soluble nucleotidases constitutes the main system for rapid inactivation of circulating adenine nucleotides. Thyroid disorders affect a number of biological factors including adenosine levels in different fractions. Then, we intend to investigate if the soluble nucleotidases responsible for the ATP, ADP, and AMP hydrolysis are affected by variations in the thyroid hormone levels in blood serum from adult rats. Hyperthyroidism was induced by daily intraperitoneal injections of L-thyroxine (T4) (2.5 and 10.0 mu g/100 g body weight, respectively) for 7 or 14 days. Hypothyroidism was induced by thyroidectomy and methimazole (0.05%) added to their drinking water during 7 or 14 days. The treatments efficacy was confirmed by determination of hemodynamic parameters and cardiac hypertrophy evaluation. T4 treatment predominantly inhibited, and hypothyroidism (14 days after thyroidectomy) predominantly increased the ATP, ADP, and AMP hydrolysis in rat blood serum. These results suggest that both excess and deficiency of thyroid hormones can modulate the ATP diphosphohydrolase and 5`-nucleotidase activities in rat blood serum and consequently modulate the effects mediated by these enzymes and their products in vascular system. (C) 2010 International Union of Biochemistry and Molecular Biology, Inc.

L-NAME-treatment alters ectonucleotidase activities in kidney membranes of rats

Aims: To investigate the effect of N omega-Nitro-L-arginine methyl ester CL-NAME) treatment, known to induce a sustained elevation of blood pressure, on ectonucleotidase activities in kidney membranes of rats. Main methods: L-NAME (30 mg/kg/day) was administered to Wistar rats for 14 days in the drinking water. Enzyme activities were determined colorimetrically and their gene expression patterns were analyzed by semi-quantitative RT-PCR. The metabolism of ATP and the accumulation of adenosine were evaluated by HPLC in kidney membranes from control and hypertensive rats. PKC phosphorylation state was investigated by Western blot. Key findings: We observed an increase in systolic blood pressure from 115 +/- 12 mmHg (control group) to 152 18 mmHg (L-NAME-treated group). Furthermore, the hydrolysis of ATP, ADP, AMP, and p-Nph-5`TMP was also increased (17%, 35%, 27%, 20%, respectively) as was the gene expression of NTPDase2, NTPDase3 and NPP3 in kidneys of hypertensive animals. Phospho-PKC was increased in hypertensive rats. Significance: The general increase in ATP hydrolysis and in ecto-5`-nucleotidase activity suggests a rise in renal adenosine levels and in renal autoregulatory responses in order to protect the kidney against the threat presented by hypertension. (C) 2010 Elsevier Inc. All rights reserved.

Central mechanisms involved in pilocarpine-induced pressor response

Pilocarpine (cholinergic muscarinic agonist) injected peripherally may act centrally to produce pressor responses; in the present study, using c-fos immunoreactive expression, we investigated the forebrain and brainstem areas activated by pressor doses of intravenous (i.v.) pilocarpine. In addition, the importance of vasopressin secretion and/or sympathetic activation and the effects of lesions in the anteroventral third ventricle (AV3V) region in awake rats were also investigated. In male Holtzman rats, pilocarpine (0.04 to 4 mu mol/kg b.w.) i.v. induced transitory hypotension followed by long lasting hypertension. Sympathetic blockade with prazosin (1 mg/kg b.w.) i.v. or AV3V lesions (1 day) almost abolished the pressor response to i. v. pilocarpine (2 mu mol/kg b.w.), whereas the vasopressin antagonist (10 mu g/kg b.w.) i.v. reduced the response to pilocarpine. Pilocarpine (2 and 4 mu mol/kg b.w.) i.v. increased the number of c-fos immunoreactive cells in the subfornical organ, paraventricular and supraoptic nuclei of the hypothalamus, organ vasculosum of the lamina terminalis, median preoptic nucleus, nucleus of the solitary tract and caudal and rostral ventrolateral medulla. These data suggest that i.v. pilocarpine activates specific forebrain and brainstem mechanisms increasing sympathetic activity and vasopressin secretion to induce pressor response. (C) 2011 Elsevier B.V. All rights reserved.

Discrete retinal input to the parabrachial complex of a new-world primate, the common marmoset (Callithrix jacchus)

Traditional retinal projections target three functionally complementary systems it) the brain of mammals: the primary visual system, the visuomotor integration systems and the circadian timing system. In recent years, studies in several animals have been conducted to investigate the retinal projections to these three systems, despite some evidence of additional targets. The aim of this study was to disclose a previously unknown connection between the retina and the parabrachial complex of the common marmoset, by means of the intraocular injection of cholera toxin Subunit b. A few labeled retinal fibers/terminals that are detected in the medial parabrachial portion of the marmoset brain show clear varicosities, Suggesting terminal fields. Although the possible role of these projections remains unknown, they may provide a modulation of the cholinergic parabrachial neurons which project to the thalamic dorsal lateral geniculate nucleus. (c) 2008 Elsevier Ireland Ltd. All rights reserved.

The Snake Venom Peptide Bj-PRO-7a Is a M1 Muscarinic Acetylcholine Receptor Agonist

Proline-rich peptides from Bothrops jararaca venom (Bj-PRO) were characterized based on the capability to inhibit the somatic angiotensin-converting enzyme. The pharmacological action of these peptides resulted in the development of Captopril, one of the best examples of a target-driven drug discovery for treatment of hypertension. However, biochemical and biological properties of Bj-PROs were not completely elucidated yet, and many recent studies have suggested that their activity relies on angiotensin-converting enzyme-independent mechanisms. Here, we show that Bj-PRO-7a (