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In this present study, a series of novel (E)-2-benzylidene-N-(3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophen-2-yl)hydrazine-1-carboxamide derivatives against α-glucosidase were designed and synthesized, and their biological activities were evaluated in vitro and in vivo. Most of the designed analogues exhibited better inhibitory activity than the marketed acarbose, especially the most potent compound 7 with an IC 50 value of 9.26 ± 1.84 μM. The direct binding of 7 and 8 with α-glucosidase was confirmed by fluorescence quenching experiments, and the kinetic and molecular docking studies revealed that 7 and 8 inhibited α-glucosidase in a non-competitive manner. Cytotoxicity bioassay indicated compounds 7 and 8 were non-toxic towards LO2 and HepG2 at 100 μM. Furthermore, both compounds were demonstrated to have in vivo hypoglycemic activity by reducing the blood glucose levels in sucrose-treated rats., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

In this paper, the impact of different substitution levels of cress seed gum (CSG, 0, 5, 10, and 15%) and sucrose (SUC, 0, 5, and 10%) on the rheological properties, textural attributes, syneresis, FTIR and microstructure of native wheat starch (NWS, 4%) gel was investigated. According to the rheological tests, the NWS-CSG and NWS-CSG-SUC gels showed thixotropic behavior and all the samples exhibited shear-thinning flow behavior. Increasing the CSG substitution level up to 15% elevated the apparent viscosity, consistency coefficient whereas the SUC substitution with NWS reduced these values. The higher apparent viscosity, consistency coefficient, and stronger pseudoplastic behavior were obtained for NWS-CSG-SUC gel than NWS gel. The addition of CSG greatly decreased hardness and consistency from 140 to 55.5 g and from 6.9 to 3.0 mJ, respectively during storage at 4 °C for 14 days; while in the presence of SUC these values slightly decreased. After storage, syneresis of NWS and NWS-10%SUC gels increased by 46.78% and 32.11%, respectively; whereas it decreased 19.88% for NWS-15%CSG gel. The SEM images showed that the mixed gels had a denser structure with a smaller pore size. The results indicated that CSG had positive effect in modifying the properties of NWS-SUC mixed gels., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Prescription opioids such as oxycodone are highly effective analgesics for clinical pain management, but their misuse and abuse have led to the current opioid epidemic in the United States. In order to ameliorate this public health crisis, the development of effective pharmacotherapies for the prevention and treatment of opioid abuse and addiction is essential and urgently required. In this study, we evaluated-in laboratory rats-the potential utility of VK4-116, a novel and highly selective dopamine D3 receptor (D3R) antagonist, for the prevention and treatment of prescription opioid use disorders. Pretreatment with VK4-116 (5-25 mg/kg, i.p.) dose-dependently inhibited the acquisition and maintenance of oxycodone self-administration. VK4-116 also lowered the break-point (BP) for oxycodone self-administration under a progressive-ratio schedule of reinforcement, shifted the oxycodone dose-response curve downward, and inhibited oxycodone extinction responding and reinstatement of oxycodone-seeking behavior. In addition, VK4-116 pretreatment dose-dependently enhanced the antinociceptive effects of oxycodone and reduced naloxone-precipitated conditioned place aversion in rats chronically treated with oxycodone. In contrast, VK4-116 had little effect on oral sucrose self-administration. Taken together, these findings indicate a central role for D3Rs in opioid reward and support further development of VK4-116 as an effective agent for mitigating the development of opioid addiction, reducing the severity of withdrawal and preventing relapse.

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease. Triacylglycerol accumulation in the liver is a hallmark of NAFLD. Metabolic studies have confirmed that increased hepatic de novo lipogenesis (DNL) in humans contributes to fat accumulation in the liver and to NAFLD progression. Mice deficient in carboxylesterase (Ces)1d expression are protected from high-fat diet-induced hepatic steatosis. To investigate whether loss of Ces1d can also mitigate steatosis induced by over-activated DNL, WT and Ces1d-deficient mice were fed a lipogenic high-sucrose diet (HSD). We found that Ces1d-deficient mice were protected from HSD-induced hepatic lipid accumulation. Mechanistically, Ces1d deficiency leads to activation of AMP-activated protein kinase and inhibitory phosphorylation of acetyl-CoA carboxylase. Together with our previous demonstration that Ces1d deficiency attenuated high-fat diet-induced steatosis, this study suggests that inhibition of CES1 (the human ortholog of Ces1d) might represent a novel pharmacological target for prevention and treatment of NAFLD., (Copyright © 2019 Lian et al.)

Capsaicin causes a burning or spicy sensation when this vanilloid compound comes in contact with trigeminal neurons of the tongue. This compound has low solubility in water, which presents difficulties in examining the psychophysical properties of capsaicin by standard aqueous chemosensory tests. This report describes a new approach that utilizes edible strips for delivering precise amounts of capsaicin to the human oral cavity for examining threshold and suprathreshold amounts of this irritant. When incorporated into pullulan-based edible strips, recognition thresholds for capsaicin occurred over a narrow range, with a mean value near 1 nmol. When incorporated into edible strips at suprathreshold amounts, capsaicin yielded robust intensity values that were readily measured in our subject population. Maximal capsaicin intensity was observed 20 s after strips dissolved on the tongue surface, and then decreased in intensity. Suprathreshold studies showed that complete blockage of nasal airflow diminished capsaicin perception in the oral cavity. Oral rinses with vanillin-linoleic acid emulsions decreased mean intensity values for capsaicin by approximately 75%, but only modestly affected recognition threshold values. Also, oral rinses with isointense amounts of aqueous sucrose and sucralose solutions decreased mean intensity values for capsaicin by approximately 50%. In addition, this decrease in capsaicin intensity following an oral rinse with sucrose was partially reversed by the sweet taste inhibitor lactisole. These results suggest that blockage of nasal airflow, vanillin, sucrose, and sucralose modulate capsaicin perception in the human oral cavity. The results further suggest a chemosensory link between receptor cells that detect sweet taste stimuli and trigeminal neurons that detect capsaicin., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Despite the cost to both individual and society, alcohol use disorders (AUDs) remain a major health risk within society, and both relapse and heavy drinking are still poorly controlled with current medications. Here we demonstrate for the first time that a centrally active and selective negative allosteric modulator for the rat M 5 muscarinic acetylcholine receptor (mAChR), ML375, decreases ethanol self-administration and attenuates cue-induced reinstatement of ethanol seeking in ethanol-preferring (iP) rats. Importantly, ML375 did not affect sucrose self-administration or general locomotor activity indicative of a selective effect on ethanol seeking. Based on the expression profile of M 5 mAChRs in the brain and the distinct roles different aspects of the dorsal striatum have on long-term and short-term ethanol use, we studied whether intra-striatal microinjection of ML375 modulated ethanol intake in rats. We show in iP rats with an extensive history of ethanol intake that intra-dorsolateral (DL), but not intra-dorsomedial, striatal injections of ML375 reduced ethanol self-administration to a similar extent as the nicotinic acetylcholine receptor ligand varenicline, which has preclinical and clinical efficacy in reducing the reinforcing effects of ethanol. These data implicate the DL striatum as a locus for the effects of cholinergic-acting drugs on ethanol seeking in rats with a history of long-term ethanol use. Accordingly, we demonstrate in rats that selectively targeting the M 5 mAChR can modulate both voluntary ethanol intake and cue-induced ethanol seeking and thereby provide direct evidence that the M 5 mAChR is a potential novel target for pharmacotherapies aimed at treating AUDs.

Until now, specific inhibitors of sucrose carriers were not available. This led us to study the properties of the recently synthesized D-glucose-fenpiclonil conjugate (D-GFC). This large amphiphilic glucoside exhibited an extremely low phloem systemicity in contrast to L-amino acid-fenpiclonil conjugates. Using Ricinus seedlings, the effect of D-GFC on 0.5 mM [14C]sucrose (Suc), 3-O-[3H]methylglucose, and [3H]glutamine uptake by cotyledon tissues was compared with that of p-chloromercuribenzenesulfonic acid (PCMBS). D-GFC dramatically inhibited H+-Suc symport at the same concentrations as PCMBS (0.5 and 1 mM), but in contrast to the thiol reagent, it did not affect 3-O-methylglucose and glutamine transport, nor the acidification of the incubation medium by cotyledon tissues. Similarly, 0.5 mM D-GFC inhibited active Suc uptake by Vicia faba leaf tissues and by Saccharomyces cerevisiae cells transformed with AtSUC2, a gene involved in Suc phloem loading in Arabidopsis, by approximately 80%. The data indicated that D-GFC was a potent inhibitor of Suc uptake from the endosperm and of Suc phloem loading. It is the first chemical known to exhibit such specificity, at least in Ricinus, and this property permitted the quantification of the two routes involved in phloem loading of endogenous sugars after endosperm removal., (© The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Although the five basic taste qualities-sweet, sour, bitter, salty and umami-can be recognized by the respective gustatory system, interactions between these taste qualities are often experienced when food is consumed. Specifically, the umami taste has been investigated in terms of whether it enhances or reduces the other taste modalities. These studies, however, are based on individual perception and not on a molecular level. In this study we investigated umami-sweet taste interactions using umami compounds including monosodium glutamate (MSG), 5'-mononucleotides and glutamyl-dipeptides, glutamate-glutamate (Glu-Glu) and glutamate-aspartic acid (Glu-Asp), in human sweet taste receptor hT1R2/hT1R3-expressing cells. The sensitivity of sucrose to hT1R2/hT1R3 was significantly attenuated by MSG and umami active peptides but not by umami active nucleotides. Inhibition of sweet receptor activation by MSG and glutamyl peptides is obvious when sweet receptors are activated by sweeteners that target the extracellular domain (ECD) of T1R2, such as sucrose and acesulfame K, but not by cyclamate, which interact with the T1R3 transmembrane domain (TMD). Application of umami compounds with lactisole, inhibitory drugs that target T1R3, exerted a more severe inhibitory effect. The inhibition was also observed with F778A sweet receptor mutant, which have the defect in function of T1R3 TMD. These results suggest that umami peptides affect sweet taste receptors and this interaction prevents sweet receptor agonists from binding to the T1R2 ECD in an allosteric manner, not to the T1R3. This is the first report to define the interaction between umami and sweet taste receptors.

Background: Orexin (OX) neurons originating in the lateral hypothalamus (LH) are ideally positioned to modulate reward processing as they form connections with several key brain regions known to be involved in the reward pathway. Consistent with these findings, a growing number of studies have implicated the OX system in modulating the rewarding properties of several drugs of abuse, including ethanol (EtOH). However, the role of the OX system in excessive binge-like EtOH intake remains relatively unexplored. Here, we assessed changes in OX immunoreactivity (IR) in the hypothalamus following repeated cycles of binge-like EtOH drinking and assessed the participation of the OX-1 receptor (OX1R) in binge-like EtOH consumption., Methods: The drinking-in-the-dark (DID) paradigm was used to model binge-like EtOH drinking in male C57BL/6J mice. In the first experiment, mice experienced 1 or 3 cycles of binge-like EtOH or sucrose drinking with DID procedures to assess changes in OX IR in distinct subregions of the hypothalamus. Subsequent experiments examined binge-like EtOH and saccharin drinking following peripheral injections of 0.0, 5.0, or 10.0 mg/kg SB-334867 (SB), a selective OX1R antagonist. Finally, mice were given peripheral injections of SB and open-field locomotor activity was measured., Results: Relative to water drinking controls, binge-like consumption of EtOH and sucrose resulted in a marked reduction in OX IR in the LH. Inhibition of the OX1R via SB blunted EtOH and saccharin drinking, but did not alter open-field locomotor activity., Conclusions: Our observed reduction in OX IR in the LH indicates that the OX system in engaged during binge-like consumption of EtOH and sucrose. The observation that inhibition of the OX1R signaling blunted binge-like EtOH, and saccharin drinking suggests that reward-related OX circuits originating in the LH participate in the consumption of salient reinforcers regardless of calories., (Copyright © 2015 by the Research Society on Alcoholism.)

Immune tolerance induction (ITI) can overcome inhibitory factor VIII (FVIII) antibodies in haemophilia A patients receiving FVIII replacement therapy. The objective was to evaluate the use of sucrose-formulated, full-length recombinant FVIII (rFVIII-FS) for ITI therapy. Patients (<8 years at ITI start) with severe haemophilia A and a peak inhibitor titre >5 Bethesda units (BU) who underwent ITI with any rFVIII-FS dose for ≥ 9 months (or until success) were eligible for this retrospective study. Efficacy analyses included descriptions of ITI treatment regimens and outcomes; ITI success was determined solely at the discretion of the investigator. Safety analyses included assessment of adverse events. Of 51 enrolled patients, 32 [high dose (≥ 85 IU kg(-1) day(-1)), n = 21; low dose, n = 11] were eligible for analysis. ITI was successful in 69% (22/32) of patients (high dose, 66.7%; low dose, 72.7%) after a median of 1.4 years (range, 0.1-3.6 years). Influencing factors for ITI success were start of ITI <1 year after inhibitor detection and an inhibitor titre <10 BU at ITI start. All patients successfully tolerized with ITI continued to receive rFVIII-FS prophylaxis as maintenance therapy, with no inhibitor recurrence from the end of ITI until study enrolment. Use of rFVIII-FS for ITI was effective and well tolerated; success rates were similar to those in published studies., (© 2013 Blackwell Publishing Ltd.)

Plant cell shape is determined by the orientation of cellulose microfibrils in the primary cell wall. Consequently, mutations that affect genes encoding the enzymes responsible for the synthesis of cellulose, namely, the cellulose synthase catalytic subunits, can alter cell shape substantially, particularly in the roots of affected plants. The multiple response expansion1 (mre1) mutant of Arabidopsis thaliana results from a point mutation in the AtCESA3 gene, which encodes one of the three isoforms of the cellulose synthase catalytic subunit required for synthesis of cellulose in the primary cell wall. Phenotypic comparison of the mre1 mutant with three other alleles (ectopic lignification1-1, ectopic lignification1-2 and constitutive expression of vsp1) showed that these four alleles form an allelic series with respect to their root phenotypes, with mre1 being the weakest allele identified to date. These analyses demonstrated that sucrose affects a significant alteration of cell shape in the roots of these mutants and likely suppresses root cell division in them as well, and that the chemical aminoisobutyric acid can suppress these effects of sucrose. Interestingly, the cell walls in the roots of these four AtCESA3 alleles contain different percentages of cellulose, and these percentages correlate with the lengths of the roots and cortex cells in these roots when grown on media containing high levels of sucrose.

Food reward is neurologically and psychologically divided into at least two properties; 'liking' and 'wanting'. Although umami taste enhances food palatability, the liking and wanting properties of umami taste, and the underlying neural mechanisms for these properties are not clear. Here, we compared sucrose (0, 10, 30, 120 and 480 mM) and monosodium l-glutamate (MSG; 0, 10, 30, 60 and 120 mM) solutions using a taste reactivity test to evaluate liking, and fixed/progressive-ratio operant licking tasks to evaluate wanting. To determine the underlying neural mechanisms, we also conducted systemic blockade of opioid receptors in both tests. In the taste reactivity test, the hedonic reactions to 30, 60 and 120 mM MSG were greater than those to water (0mM) but lower than those to 480 mM sucrose. In the operant task, the intake, number of licks, and breakpoint to MSG reached peaks at around 60mM but they were lower than those to 30-480 mM sucrose. The systemic naloxone treatment decreased the hedonic responses to MSG and sucrose, and reduced the incentive salience of MSG but not sucrose. These findings indicate that the hedonic response and incentive salience of MSG is lower than those of sucrose when compared at the maximum response and that the incentive salience of MSG is lower than sucrose even where the hedonic response is similar. The present study also suggest that the hedonic response and incentive salience of umami compound is modulated by brain opioid signaling., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Emerging evidence indicates that specific metabotropic glutamate receptors (mGluRs) modulate ethanol self-administration. In general, inhibition of glutamate transmission through blockade of postsynaptic mGluRs, or activation of presynaptic mGluRs, inhibits ethanol self-administration. The goal of this preclinical study was to further characterize mGluR regulation of ethanol self-administration by examining effects of AMN082, an allosteric positive modulator of presynaptic mGluR7 activity. Separate groups of C57BL/6J male mice were trained to self-administer ethanol or sucrose on a fixed-ratio 4 schedule of reinforcement during 1 h sessions. On test days, mice were pretreated with AMN082 (0, 1.0, 3.0, 5.6, or 10 mg/kg) 30 min prior to self-administration sessions. Functional specificity and activity was examined by testing the effects of AMN082 (0-10 mg/kg) on open-field locomotor activity and HPA axis function as measured by plasma corticosterone levels. AMN082 (10 mg/kg) produced a significant reduction in ethanol and sucrose reinforced responding, and inhibited locomotor activity. Plasma corticosterone levels were significantly increased following AMN082 (5.6 and 10 mg/kg) suggesting a dose-dependent dissociation between the behavioral and hormonal effects of the compound. These data suggest that activation of mGluR7 by AMNO82 produces nonspecific reductions in motivated behavior that are associated with negative effects on motor activity.

Plant sucrose transporters (SUTs) are members of the glycoside-pentoside-hexuronide (GPH) cation symporter family (TC2.A.2) that is part of the major facilitator superfamily (MFS). All plant SUTs characterized to date function as proton-coupled symporters and catalyze the cellular uptake of sucrose. SUTs are involved in loading sucrose into the phloem and sink tissues, such as seeds, roots and flowers. Because monocots are agriculturally important, SUTs from cereals have been the focus of recent research. Here we present a functional analysis of the SUT ShSUT1 from sugarcane, an important crop species grown for its ability to accumulate high amounts of sucrose in the stem. ShSUT1 was previously shown to be expressed in maturing stems and plays an important role in the accumulation of sucrose in this tissue. Using two-electrode voltage clamping in Xenopus oocytes expressing ShSUT1, we found that ShSUT1 is highly selective for sucrose, but has a relatively low affinity for sucrose (K(0.5) = 8.26 mM at pH 5.6 and a membrane potential of -137 mV). We also found that the sucrose analog sucralose (4,1',6'-trichloro-4,1',6'-trideoxy-galacto-sucrose) is a competitive inhibitor of ShSUT1 with an inhibition coefficient (K(i)) of 16.5 mM. The presented data contribute to our understanding of sucrose transport in plants in general and in monocots in particular.

The sense of taste provides humans with necessary information about the composition and quality of food. For humans, five basic tastes are readily distinguishable and include sweet, bitter, salty, sour, and savory (or umami). Although each of these qualities has individualized transduction pathways, sweet and umami tastes are believed to share a common receptor element, the T1R3 receptor subunit. The two G-protein-coupled heteromer receptors that comprise an umami stimulus receptor (T1R1-T1R3) and a sweetener receptor (T1R2-T1R3) constitute a potential link between these two qualities of perception. While the role of the individual monomers in each human heteromer has been examined in vitro, very little is known of the implication of this research for human perception, or specifically, how sweet and savory taste perceptions may be connected. Using a psychophysical approach, we demonstrate that lactisole, a potent sweetness inhibitor that binds in vitro to hT1R3, also inhibits a significant portion of the perception of umami taste from monosodium glutamate. Following the molecular logic put forward by Xu et al. (2004, Proc. Natl Acad. Sci. USA, 101, 14258-14263), our psychophysical data support the in vitro hypothesis that the shared T1R3 monomer moderates the activation of both T1R2 and T1R1 in humans and impairs suprathreshold perception, respectively, of sweetness and, to a lesser degree, umaminess in the presence of lactisole.

Traditional intake measures of voluntary consumption of food or fluid from a specific location involve both appetitive and consummatory behaviors. Appetitive behaviors are food finding behaviors displayed by an animal prior to the consumption of the food, whereas consummatory behaviors are the behaviors involved in the actual consumption of the food. Intraoral intake of a fluid can be measured by directly infusing it into the oral cavity of an animal and quantifying the consummatory behaviors. The present study compared the effects of immune activation (lipopolysaccharide, LPS) and toxin (lithium chloride, LiCl)-induced changes on both a traditional intake measure (bottle drinking) and an intraoral intake measure. In Experiment 1, rats were injected intraperitoneally with LPS (200 microg/kg), LiCl (0.15 M, 20 ml/kg) or NaCl vehicle, and voluntary sucrose (0.3 M) intake was monitored for 1 h from a graduated drinking tube. Voluntary intake was again assessed on a second test day, 72 h later under the same conditions. In Experiment 2, a continuous intraoral infusion of sucrose (0.3 M) was given via intraoral cannulae following systemic injections of LPS, LiCl or NaCl vehicle on two different test days, 72 h apart. Rats injected with LiCl displayed reduced sucrose intake on both the voluntary intake measure and the intraoral intake measure relative to controls (P's<.05). The reduced intake observed was of greater magnitude on the second test day of both experiments, consistent with conditioning effects. In contrast, LPS reduced sucrose intake only when assessed with the traditional intake measure. Intraoral sucrose intake remained unchanged relative to controls. The present results provide further evidence that activation of the immune system has adverse effects on the appetitive phase of ingestion, whereas the consummatory aspects are unaffected.

Naloxone, a nonselective opioid antagonist, has been demonstrated to reduce oral self-administration of ethanol (EtOH) in rats. Conflicting conclusions have been drawn about the effects of naloxone on consumption of non-EtOH control liquids. A preliminary meta-analysis found large and homogeneous effects of naloxone on EtOH consumption and heterogeneous effects on the consumption of control liquids. Although many of the authors concluded that their control liquid results were "not significant," when they were combined using meta-analytic techniques, it was apparent that there were some strong, but widely divergent, effect sizes. In the first experiment in the current study, 60 male Sprague-Dawley rats were trained to drink 10% EtOH in tap water over 3 weeks of limited-access sessions. Then, their limited-access consumption was measured in single-bottle tests of four liquids (water, 10% EtOH in water, an isocaloric sucrose solution, and an "equally sweet" saccharin solution) 15 min following an intraperitoneal injection of either saline or 1.0 mg/kg naloxone. Every animal was tested 36 times in a counterbalanced order: three times for each liquid following an injection of naloxone and six times for each liquid following an injection of saline. There were distinct differences in the quantity of each liquid consumed in the saline trials. However, the suppression percentages for each liquid in the naloxone trials were identical ( approximately 50%). There were significant correlations, in the range of.23-.42, between the mean amount of each liquid consumed during saline trials for each animal and the suppression percentage during naloxone trials for the same animal and liquid. When the animals were divided into high, low, and medium drinkers for each liquid, the low drinkers demonstrated a much lower suppression after naloxone treatment than did the other two groups. The data confirm that blockade of opioid receptors suppresses consumption of both EtOH and non-EtOH liquids to a degree that is related to the amount of voluntary, untreated consumption of the liquids.

We recently reported that 6-beta naltrexol, the major metabolite of naltrexone in humans, reduced ethanol consumption in rats. Two new experiments were designed to compare 6-beta naltrexol and naltrexone across three dose levels on an ethanol or sucrose baseline using a limited-access procedure in Wistar rats. The results of Experiment 1 showed that both 6-beta naltrexol and naltrexone reduced ethanol consumption across a range of doses. An in vivo assay showed that naltrexone was approximately 25 times more potent than 6-beta naltrexol at comparable ED50 doses. In addition, there was no indication of systematic development of tolerance to the effect of either drug across the 4 days of drug administration. In Experiment 2, both 6-beta naltrexol and naltrexone reduced the consumption of a sucrose solution using a limited-access procedure. The implications of these data for the development of pharmacotherapeutic agents capable of reducing drinking in recovering alcoholics are discussed.

The use of gas chromatography-mass spectrometry-selected ion monitoring along with a (13)C internal standard has allowed sensitive measurements of the sucrose (Suc) content of individual shoot apices of Fuchsia hybrida. With intact plants, as the photosynthetic irradiance increased, so did shoot apex Suc content, reaching saturation at about 500 micromol m(-2) s(-1). These same plants flowered at the higher irradiances, remaining vegetative in 10-h short days at an irradiance of 230 micromol m(-2) s(-1). The strong correlation (r = 0.93) in these studies between flowering and shoot apex Suc content indicates a role for Suc as a stimulus to flowering in this species. However, Suc is not the long-day (LD) "florigen" of F. hybrida because 2 to 4 LD given as a 14-h low-irradiance photoperiod extension (10-15 micromol m(-2) s(-1)) induced flowering but without increase in shoot apex Suc content. Flowering induced by either pathway, the LD- or the Suc-mediated one, was inhibited by applying gibberellin (GA) to the shoot tip. Such inhibition of flowering by GA, at least for the LD pathway, was associated with a reduced apex Suc content, enhanced elongation of subapical stem tissue, and a reduced import into the shoot apex of leaf-sourced assimilate. Thus, our findings show how GA inhibits flowering of F. hybrida and confirm the importance of nutrient diversion in regulating flowering.

The sucrose (CscB) permease belongs to the oligosaccharide:H(+) symporter family of the Major Facilitator Superfamily and is homologous to the lactose permease from Escherichia coli. Sucrose transport in cells expressing sucrose permease is completely inhibited by N-ethylmaleimide (NEM), suggesting that one or more of the seven native Cys residues may be important for transport. In this paper, each Cys residue was individually replaced with Ser, and transport activity, membrane expression, and NEM sensitivity are documented. All seven single Cys-->Ser mutants are expressed normally in the membrane and catalyze sucrose transport with activities ranging from 80% to 180% of wild type. Six of the seven Ser mutants are completely inactivated by NEM, while Cys122-->Ser permease is insensitive to the sulfhydryl reagent, indicating that NEM inhibition results from alkylation of Cys122. Subsequently, a sucrose permease devoid of Cys residues (Cys-less) was constructed in which all Cys residues were replaced with Ser simultaneously by using a series of overlap-extension PCRs. Membrane expression and kinetic parameters for Cys-less [K(m) 4.8 mM, V(max) 192 nmol min(-1) (mg of protein)(-1)] are essentially identical to those of wild type [K(m) 5.4 mM, V(max) 196 nmol min(-1) (mg of protein)(-1)]. However, Cys-less permease catalyzes sucrose accumulation to steady-state levels that are approximately 2-fold higher than those of wild type. As anticipated, Cys-less permease is completely resistant to NEM inhibition. The observations demonstrate that Cys residues play no functional role in sucrose permease. Furthermore, the approach described to create the Cys-less transporter is generally applicable to other proteins. An application of Cys-less permease in the study of the substrate binding site is presented in the accompanying paper.

Arnica and propolis have been used for thousands of years in folk medicine for several purposes. They possess several biological activities such as anti-inflammatory, antifungal, antiviral and tissue regenerative, among others. Although the antibacterial activity of propolis has already been demonstrated, very few studies have been done on bacteria of clinical relevance in dentistry. Also, the antimicrobial activity of Arnica has not been extensively investigated. Therefore the aim here was to evaluate in vitro the antimicrobial activity, inhibition of adherence of mutans streptococci and inhibition of formation of water-insoluble glucan by Arnica and propolis extracts. Arnica montana (10%, w/v) and propolis (10%, w/v) extracts from Minas Gerais State were compared with controls. Fifteen microorganisms were used as follows: Candida albicans--NTCC 3736, F72; Staphylococcus aureus--ATCC 25923; Enterococcus faecalis--ATCC 29212; Streptococcus sobrinus 6715; Strep. sanguis--ATCC 10556; Strep. cricetus--HS-6; Strep. mutans--Ingbritt 1600; Strep. mutans--OMZ 175; Actinomyces naeslundii--ATCC 12104, W 1053; Act. viscosus OMZ 105; Porphyromonas gingivalis; Porph. endodontalis and Prevotella denticola (the last three were clinical isolates). Antimicrobial activity was determined by the agar diffusion method and the zones of growth inhibition were measured. To assess cell adherence to a glass surface, the organisms were grown for 18 h at 37 degrees C in test-tubes at a 30 degree angle. To assay water-insoluble glucan formation, a mixture of crude glucosyltransferase and 0.125 M sucrose was incubated for 18 h at 37 degrees C in test-tubes at a 30 degree angle. Arnica and propolis extracts (20 microl) were added to these tubes to evaluate the % of inhibition of cell adherence and water-insoluble glucan formation. The propolis extract significantly inhibited all the microorganisms tested (p < 0.05), showing the largest inhibitory zone for Actinomyces spp. The Arnica extract did not demonstrate significant antimicrobial activity. Cell adherence and water-insoluble glucan formation were almost completely inhibited by the propolis extract at a final concentration of 400 microg/ml and 500 microg/ml, respectively. The Arnica extract showed slight inhibition of the adherence of the growing cells (19% for Strep. mutans and 15% for Strep. sobrinus) and of water-insoluble glucan formation (29%) at these same concentrations. Thus, the propolis extract showed in vitro antibacterial activity, inhibition of cell adherence and inhibition of water-insoluble glucan formation, while the Arnica extract was only slightly active in those three conditions.

In bacteria and yeast, glutamine synthetase (GS) expression is tightly regulated by the metabolic status of the cell, both at the transcriptional and posttranscriptional levels. We discuss the relative contributions of light and metabolic cues on the regulation of members of the GS gene family (chloroplastic GS2 and cytosolic GS1) in Arabidopsis. These studies reveal that the dramatic induction of mRNA for chloroplastic GS2 by light is mediated in part by phytochrome and in part by light-induced changes in sucrose (Suc) levels. In contrast, the modest induction of mRNA for cytosolic GS1 by light is primarily mediated by changes in the levels of carbon metabolites. Suc induction of mRNA for GS2 and GS1 occurs in a time- and dose-dependent manner. Suc-induced changes in GS mRNA levels were also observed at the level of GS enzyme activity. In contrast, amino acids were shown to antagonize the Suc induction of GS, both at the level of mRNA accumulation and that of enzyme activity. For GS2, the gene whose expression was the most dramatically regulated by metabolites, we used a GS2 promoter-beta-glucuronidase fusion to demonstrate that transcriptional control is involved in this metabolic regulation. Our results suggest that the metabolic regulation of GS expression in plants is controlled by the relative abundance of carbon skeletons versus amino acids. This would allow nitrogen assimilation into glutamine to proceed (or not) according to the metabolic status and biosynthetic needs of the plant. This type of GS gene regulation is reminiscent of the nitrogen regulatory system in bacteria, and suggests an evolutionary link between metabolic sensing and signaling in bacteria and plants.

From the fresh stem of Stephanotis lutchuensis var. japonica, ten new oleanane-type triterpenoid glycosides, named sitakisosides XI-XX (1-10), were isolated. Their structures were determined on the basis of spectroscopic data and chemical evidence. The results show that all have a 3-O-beta-D-xylopyranosyl(1 --> 6)-beta-D-glucopyranosyl(1 --> 6)- beta-D-glucopyranosyl moiety and the aglycones of sitakisosides XI-XV, XVI and XVII, XVIII and XIX, and XX are sitakisogenin, chichipegenin, marsglobiferin and longispinogenin, respectively. Sitakisosides XI-XIII, XVI and XVIII, having an acyl group, showed antisweet activity.

A role for the NO-cGMP pathway in mediating chemosensory activation of feeding is suggested by intense NADPH diaphorase staining observed in nerve fibers that project from sensory cells in the lips to the CNS and by the presence in the CNS of a NO-activated guanylyl cyclase. In preparations reduced to isolated lips and CNS, intracellular recordings were made from motoneurons driven by the interneurons of the central pattern generator (CPG) for feeding. Fictive feeding in such preparations can be recorded from these motoneurons following the application of sucrose to the lips. Sucrose activation of fictive feeding is inhibited by the NO scavenger hemoglobin, the NO synthase inhibitor N omega-Nitro-L-Arginine Methyl Ester (L-NAME) and by methylene blue, an inhibitor of guanylyl cyclase. Fictive feeding in isolated lip-CNS preparations can be activated without sucrose by superfusion of NO donor molecules such as SNAP and hydroxylamine and by the nonhydrolyzable analog of cGMP, 8-bromo-cGMP. The feeding CPG can also be activated centrally by depolarizing a modulatory interneuron, the slow oscillator (SO). When the CPG is activated in this way, fictive feeding is not susceptible to inhibition by hemoglobin, the most potent of the inhibitors of sucrose-activated fictive feeding. Behavioral experiments on intact snails confirm the findings from in vitro experiments and show that hemoglobin prevents feeding and methylene blue significantly delays the onset of feeding. These results indicate (1) that NO is a putative chemosensory transmitter in the snail L. stagnalis, (2) that the NO-cGMP pathway can mediate chemosensory activation of specific patterns of centrally generated behavior, (3) that NO is not involved in transmission within the central network of neurons responsible for the behavior, and more generally (4) that a freely diffusing and highly reactive gaseous signalling molecule can have restricted and specific behavioral functions.

1. Osmolarity-dependent (osmo-dependent) ionic currents from follicle-enclosed Xenopus oocytes (follicles) were studied using the two-microelectrode voltage-clamp technique, combined with intra-oocyte pressure injection of sucrose or polyethylene glycols (PEGs). 2. Intra-oocyte injections of sucrose or PEG (3-25 nmol) generated inward membrane currents (follicles held at -60 mV) associated with an increase in membrane conductance. These currents were carried mainly by chloride ions (ICl(osm)), and were strongly attenuated by increasing the tonicity of the external medium, or by external application of La3+ (0.1-1 mM). 3. The ability to generate ICl(osm) depended on the molecular weight of the injected PEG. Injections of PEG 200 or 300 generated ICl(osm) in 95% of the follicles tested, PEG 600 generated comparable currents in only 20% of the follicles, while similar injections of PEG 1000 did not elicit ICl(osm). 4. Octanol (1-1.5 mM), a gap junction channel blocker, reversibly inhibited 50-90% of the ICl(osm) generated by injections of sucrose or PEG 300. Moreover, sucrose or PEG injections did not elicit ICl(osm) in defolliculated oocytes. 5. It is concluded that an increase in the internal osmolarity of the follicular cells activates a mechanism, probably involving cellular swelling, which leads to the opening of ICl(osm) channels most probably located in the follicular cell membrane.

From the fresh stem of Stephanotis lutchuensis var. japonica, five new oleanane-type triterpenoid glycosides named sitakisosides VI-X (1-5) were isolated. Their structures were determined on the basis of spectroscopic data and chemical evidence. Sitakisosides VI and VII are 3-O-beta-D-xylopyranosyl(1-->6)-beta-D-glucopyranosyl(1-->6)-beta-D- glucopyranosido-21-O-(6-N-methylanthranilyl)-beta-D-glucopyr anosyl and 3-O-beta-D-xylopyranosyl(1-->6)-beta-D-glucopyranosyl(1-->6)-beta-D- glucopyranosido-21-O-(4-N-methylanthranilyl)-beta-D-glucopyr anosyl sitakisogenin, respectively. Sitakisoside VIII is 3-O-beta-D-xylopyranosyl(1-->6)-beta-D- glucopyranosyl(1-->6)-beta-D-glucopyranosido-21-O-N-methylanthr anilyl- 3 beta,16 beta,21 beta,28-tetrahydroxyolean-12-ene-22-one. Sitakisoside IX is 3-O-beta-D-xylopyranosyl(1-->6)-beta-D-glucopyranosyl(1-->6)-beta-D- glucopyranosido-21-O-(6-N-methylanthranilyl)-beta-D-glucopyr anosyl gymnestrogenin. Sitakisoside X is 3-O-beta-D-xylopyranosyl(1-->6)-beta-D- glucopyranosyl(1-->6)-beta-D-glucopyranosyl longispinogenin.

From the fresh stem of Stephanotis lutchuensis var. japonica, we have isolated five new oleane glycosides named sitakisosides I-V (1-5). Their structures were determined on the basis of spectroscopic data and chemical evidence. Sitakisoside V showed the strongest antisweet activity among sitakisosides I-V.

Pyruvate dehydrogenase complex (PDC) has a key role in the regulation of hepatic lipogenesis by dietary factors. We have investigated the effects of dietary carbohydrate and fat on hepatic PDC. Sucrose-based or starch-based diets were administered for 15 days. A positive correlation between PDC activity and the lipogenic potential of the diet was found. A high-sucrose, fat-free diet caused a 3-fold increase in total activity whereas a high-starch, fat-free diet caused a 1.5-fold increase, as compared with chow-fed rats. Dietary polyunsaturated fat (PUF) caused a marked inhibitory effect on total and active PDC; fish oil being more effective than corn oil. Dietary saturated fat (butter) failed to inhibit the sucrose-induced elevation in total activity, but was almost as effective as fish oil in depressing percent active enzyme. Changes in total PDC activity closely correlated with modifications in the content of enzyme quantitated by immunoblotting, indicating that increased enzyme content and not activation is the predominant mechanism underlying the adaptive response to high-sucrose feeding. This response is suppressed by dietary PUF. Inhibition of hepatic lipogenesis by PUF involves a reduction of PDC content as well as that of several lipogenic enzymes. The relevant mechanisms remain to be established.

The insulin-like effects of vanadate were compared in streptozotocin-induced diabetic rats fed on high starch control and high sucrose diets for a period of six weeks. Diabetic rats in both diet groups were characterized by hypoinsulinemia, hyperglycemia (6.8-7.0 fold increase) and significant decreases (p < 0.001) in the activities of glycogen synthase, phosphorylase and lipogenic enzymes, ATP-citrate lyase, glucose 6-phosphate dehydrogenase and malic enzyme in liver. There were no diet-dependent differences in these abnormalities. However, the insulin-mimetic agent vanadate was more effective in diabetic rats fed sucrose diet as compared to animals fed control starch diet. Vanadate administration resulted in 30% and 64% decreases in plasma glucose levels in diabetic rats fed control and sucrose diets, respectively. The activities of glycogen synthase (active) and phosphorylase (active and total) were restored significantly by vanadate in control (p < 0.05-0.01) and sucrose (p < 0.001) diets fed diabetic rats. This insulin-mimetic agent increased the activities of hepatic lipogenic enzymes in control diet fed rats to 38-47% of normal levels whereas in sucrose fed group it completely restored the activities. Sucrose diet caused a distinct effect on the plasma levels of triacylglycerol (4-fold increase) and apolipoprotein B (2.8-fold increase) in diabetic rats and vanadate supplementation decreased their levels by 65-75%. These data indicate that vanadate exerts insulin-like effects in diabetic rats more effectively in sucrose fed group than the animals fed control diet. In addition, vanadate also prevents sucrose-induced hypertriglyceridemia.

Effects of feeding sucrose rich diet supplemented with and without the insulinmimetic agent vanadate for a period of six weeks were studied in rats. Sucrose diet caused hypertriglyceridemia (140% increase), hyperinsulinemia (120% increase) and significant elevations in the levels of glucose (p < 0.001) and cholesterol (p < 0.05) in plasma as compared to control starch fed rats. Activities of hepatic lipogenic enzymes, ATP-citrate lyase, glucose 6-phosphate dehydrogenase and malic enzyme increased by 100-150% as a result of sucrose feeding. However, glycogen content and the activities of glycogen synthase and phosphorylase in liver remained unaltered in these animals. The plasma levels of triacylglycerols and insulin in the rats fed on vanadate supplemented sucrose diet were 65% and 85% less, respectively as compared to rats on sucrose diet without vanadate. The concentrations of glucose and cholesterol in plasma and the activities of lipogenic enzymes in liver did not show any elevation in sucrose fed rats when supplemented with vanadate. These data indicate that the sucrose diet-induced metabolic aberrations can be prevented by the insulin-mimetic agent, vanadate.

In stationary phase, 95% of the fructosyltransferase (FTase) activity of Streptococcus salivarius ATCC 25975 was found associated with the cells. Within the first 15 min after inoculation into fresh medium, the specific activity of cell-associated FTase decreased by 92% of its initial value. After this period of initial loss, the enzyme was synthesized during exponential growth until a maximum level equivalent to that present before inoculation was obtained. The inactivation of FTase was also demonstrated in a nongrowing system. Washed cell suspensions incubated at 37 degrees C in 200 mM potassium phosphate buffer (pH 6.5) containing 10 microM Cu2+ lost 80 to 95% of their FRase activity after 30 min. This loss could be prevented by the addition of histidine, cysteine, or Ca2+ to the suspension mixture. A factor(s) essential for the inactivation of cell-associated FTase could itself be preferentially inactivated by heating cells at 40 degrees C for periods of up to 3 h, or by storage of cells at 0 to 4 degrees C for several days in a low-ionic-strength, low-pH, potassium phosphate buffer. Treatment of cells with the N-acetylmuramidase enzyme M-1, in the presence of 0.5 M melezitose, resulted in the release of FTase from the cell. The released enzyme was recovered in the supernatant fraction after centrifugation at 160,000 x g for 90 min. Comparison of solubilized active and inactivated FTase preparations by polyacrylamide gel electrophoresis demonstrated that the inactivation of cell-associated FTase activity was associated with the loss of specific protein bands.

Rats with chronic gastric cannulas were intraperitoneally injected with the dopamine receptor antagonist pimozide (0.25 mg/kg) before sham feeding 5, 10, 20, or 40% (w/v) sucrose solutions. Amount of sham intake after control injections increased as a function of sucrose concentration. At each concentration, the rate of sham feeding was greatest during the initial 3 min of sham feeding and subsequently decelerated. Pimozide inhibited sham feeding rate, and the temporal pattern of decreases in sham feeding rate after pimozide were similar to those produced by decreasing the concentration of sucrose sham fed. These data extend previous reports of the inhibitory effect of pimozide on ingestion of sweet fluids by eliminating the possibility that the effect was the result of pimozide facilitating postingestional inhibitory mechanisms. Further, pimozide did not appear to produce fatigue or sedation, or to reduce the rats' motor capacity to sham feed. Therefore, these data are consistent with the hypothesis that central dopaminergic synaptic activity mediates the reinforcing effects of sweet taste that drive sham feeding.

Immune sera to strains of Streptococcus mutans were found to inhibit agglutination of bacterial suspensions to which either high-molecular-weight dextran or sucrose was added. This inhibitory activity was shown to be mediated by antibody of the immunoglobulin G class. A semiquantitative assay was developed which demonstrated cross-inhibition of dextran/sucrose-induced agglutination among several strains of S. mutans. Antiserum to a partially purified glucosyltransferase was found to lack agglutination inhibition activity, consistent with the hypothesis that the dextran-binding antigen detected by the assay is immunologically distinct from the glucan-synthetic enzyme. A model for glucan synthesis and binding consistent with the reported data is described.

Three new sucrose analogs modified at C-3 have been studied as inhibitors and substrates for the glucosyltransferases (glucansucrases) of Streptococcus mutans 6715. Although none of the analogs were found to be substrates for polymer synthesis with either the soluble-polysaccharide producing enzyme, GTF-S, or the insoluble-polysaccharide producing enzyme, GTF-I, 3-deoxysucrose and 3-deoxy-3-fluorosucrose were able to donate glycosyl residues for acceptor reactions with both enzymes. Modification at C-3 considerably decreased the binding at the active site of both enzymes, since all of the analogs had inhibition constants at least one order of magnitude greater than the Km value for sucrose.

In order to assess the effect of the antagonist methyl 4,6-dichloro-4,6-dideoxy-alpha-D-galactopyranoside (MAD-diCl-Gal) upon the gerbil's chorda tympani sucrose taste response, we tested several concentrations of this compound, as well as single concentrations of closely related derivatives, and found that MAD-diCl-Gal was the most potent inhibitor tested. It appears that the inhibition mechanism is very specific. For example, we have found that 2 chlorine atoms at the C-4 and C-6 positions on the glucopyranoside ring are required for inhibition. In addition, with regard to the orientation of the chlorine atoms, the galacto derivative seems to be more potent than the gluco derivative. We have also found that the methyl glycoside is more potent than the free sugar. With regard to the orientation of the methyl group, MAD-diCl-Gal is more potent than its beta-anomer. (Because of this discovery of the methyl group enhancement and orientation effect, we shall discontinue using the acronym diCl-Gal and replace it with the more specific MAD-diCl-Gal.) Of particular significance is the fact that there appears to be a structure-activity relationship between the most active stimulants and inhibitors in that the requirement for an axial orientation at C-1 and the enhancement by the methyl group at that position are the same in both cases. These results suggest that both the stimulator and the antagonist are acting at the same receptor site.(ABSTRACT TRUNCATED AT 250 WORDS)

An oral radiotelemetric technique was used to determine whether a range of efficient buffers had the capacity to control the levels of hydrogen ions within dental plaque in the presence of sucrose. It was found that the buffers failed to control both the intensity and duration of the acid produced.

We have discovered that the gerbil's chorda tympani nerve response to sucrose is suppressed by p-nitrophenyl alpha-D-glucopyranoside (PNP-Glu) and chloramphenicol (CAP). Mixture experiments of PNP-Glu and CAP with sodium chloride, potassium chloride, hydrochloric acid, and sucrose gave rise to the following observations: Neither PNP-Glu nor CAP alone stimulates the gerbil's taste nerve; while the sucrose response is suppressed by these inhibitors, taste responses produced by sodium chloride, potassium chloride, and hydrochloric acid are unaffected by the presence of PNP-Glu or CAP; the antagonisms of PNP-Glu and CAP were surmounted by a high concentration of sucrose; CAP is a more potent antagonist (IC50 = 0.0013 M) than PNP-Glu (IC50 = 0.022 M), and both are more potent than methyl 4,6-dichloro-4,6-dideoxy-alpha-D-galactopyranoside (IC50 = 0.048 M); and sucrose antagonism occurs only when PNP-Glu and CAP are mixed with sucrose. It is short-lived and ceases when the mixtures are rinsed from the gerbil's tongue. Structure-activity studies provided the following information: The alpha anomer of PNP-Glu is a more potent inhibitor than its beta anomer; among the PNP-Glu derivatives tested (p-aminophenyl, p-nitrophenyl, and phenyl) only p-nitrophenyl inhibited; among the nitrophenyl galactosides, the para derivative was more potent than the ortho or meta; and p-nitrophenyl alpha-D-mannopyranoside and p-nitrophenyl alpha-D-galactoside are slightly more potent than PNP-Glu. On the basis of concentration experiments, we believe that the inhibitory mechanisms of PNP-Glu and CAP are different, which suggests the existence of at least 2 sucrose receptor sites.

The effect of inhibition of disaccharidases on the degree of absorption of glucose, lactose, and sucrose was examined utilizing an in vivo model in the rat. Acarbose, a competitive alpha-glucosidase inhibitor was utilized to selectively inhibit small intestinal mucosal enzymes. Adult rats (250-350 g body weight) were the subjects of intraduodenal bolus infusion experiments with either sugar alone or sugar plus acarbose. All sugars were infused at a dose of 0.5 g/kg body weight. Portal venous blood glucose was determined at 30-min intervals from 0 to 150 min. Glucose (monosaccharide) and lactose (beta-galactoside) absorption were not altered by the presence of acarbose. In contrast, sucrose (alpha-glucosidase) absorption was significantly diminished in the presence of acarbose. Sucrose absorption in the presence of increasing acarbose doses (0.7-5.6 mg/kg body weight) was depressed in a dose-dependent fashion. Linear regression analysis revealed a high degree of correlation between residual sucrase activity and area under blood glucose curve (r = 0.9837). Similar degrees of correlation were found between acarbose dose and area under blood glucose curve (r = -0.9322), and between residual sucrase activity and acarbose dose (r = -0.9695). These data confirm that acarbose is a selective alpha-glucosidase inhibitor that does not affect monosaccharidase transport. In the presence of acarbose, alpha-glucosidase absorption is diminished in a dose-dependent fashion. Postprandial glucose rise following an alpha-glucosidase meal seems to be determined, in the presence of graded acarbose inhibition, by residual mucosal alpha-glucosidase activity.

The production of glutamic gamma-semialdehyde, an intermediate in the synthesis of proline, was inhibited in Escherichia coli by physiological concentrations of penicillin. Sucrose (0.6 m) and sodium chloride (0.1 m) prevented penicillin inhibition of glutamic gamma-semialdehyde synthesis. Cells which were in the stationary phase, or which had been permitted to metabolize without growth, were insensitive to the effects of penicillin on glutamic gamma-semialdehyde synthesis.

The addition of 0.08 M sucrose to a culture medium containing Chang-strain human liver cells causes intense cytoplasmic vacuolation. Electron microscopy of these cells grown inferritin, time-lapse cinematography, and radioautography reveal that the vacuoles arise by endocytosis and that the sucrose is taken into the cell and localized in the vacuoles. Tracer studies demonstrate that sucrose-(3)H provides a marker for quantitation of endocytosis and that it neither induces nor stimulates endocytosis. Electron micrographs of vacuolated liver cells show microfilaments in close proximity to the inside of the plasma membrane, in the pseudopodia, and to the cytoplasmic side of the membrane surrounding endocytosis vacuoles. Cytochalasin B (CB), a mold metabolite that inhibits various types of cell motility, has a dose-dependent inhibitory effect on the uptake of sucrose-(3)H by these cells. This inhibition is accompanied by a cessation of the movement of ruffles and pseudopodia on the surface of the cells and the formation of blebs which arise from the cell's surface. These morphological changes are quickly reversible upon removal of CB. Alterations in the appearance and location of microfilaments are also observed in CB-treated cells.