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PubMed2025

100+ years of phase variation: the premier bacterial bet-hedging phenomenon.

Stochastic, reversible switches in the expression of Salmonella flagella variants were first described by Andrewes in 1922. Termed phase variation (PV), subsequent research found that this phenomenon was widespread among bacterial species and controlled expression of major determinants of bacterial-host interactions. Underlying mechanisms were not discovered until the 1970s/1980s but were found to encompass intrinsic aspects of DNA processes (i.e. DNA slippage and recombination) and DNA modifications (i.e. DNA methylation). Despite this long history, discoveries are ongoing with expansions of the phase-variable repertoire into new organisms and novel insights into the functions of known loci and switching mechanisms. Some of these discoveries are somewhat controversial as the term 'PV' is being applied without addressing key aspects of the phenomenon such as whether mutations or epigenetic changes are reversible and generated prior to selection. Another 'missing' aspect of PV research is the impact of these adaptive switches in real-world situations. This review provides a perspective on the historical timeline of the discovery of PV, the current state-of-the-art, controversial aspects of classifying phase-variable loci and possible 'missing' real-world effects of this phenomenon.

Gene Expression Regulation, Bacterial↗

PubMed2005

Folate, homocysteine, and arsenic metabolism in arsenic-exposed individuals in Bangladesh.

Chronic exposure to arsenic is occurring throughout South and East Asia due to groundwater contamination of well water. Variability in susceptibility to arsenic toxicity may be related to nutritional status. Arsenic is methylated to monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA) via one-carbon metabolism, a biochemical pathway that is dependent on folate. The majority of one-carbon metabolism methylation reactions are devoted to biosynthesis of creatine, the precursor of creatinine. Our objectives of this cross-sectional study were to characterize the relationships among folate, cobalamin, homocysteine, and arsenic metabolism in Bangladeshi adults. Water arsenic, urinary arsenic, urinary creatinine, plasma folate, cobalamin, and homocysteine were assessed in 1,650 adults; urinary arsenic metabolites were analyzed for a subset of 300 individuals. The percentage of DMA in urine was positively associated with plasma folate (r = 0.14, p = 0.02) and negatively associated with total homocysteine (tHcys; r = -0.14, p = 0.01). Conversely, percent MMA was negatively associated with folate (r = -0.12, p = 0.04) and positively associated with tHcys (r = 0.21, p = 0.0002); percent inorganic arsenic (InAs) was negatively associated with folate (r = -0.12, p = 0.03). Urinary creatinine was positively correlated with percent DMA (r = 0.40 for males, p < 0.0001; 0.25 for females, p = 0.001), and with percent InAs (r = -0.45 for males, p < 0.0001; -0.20 for females, p = 0.01). Collectively, these data suggest that folate, tHcys, and other factors involved in one-carbon metabolism influence arsenic methylation. This may be particularly relevant in Bangladesh, where the prevalence of hyperhomocysteinemia is extremely high.

Adult↗

PubMed2002

[Aspects of exposure to methyl-methacrylate of varying intensity in an organism and on population levels].

Studies of mortality among 3588 workers who contacted variable doses of methylmetacrilate (MMA) revealed that higher intensity of the exposure led to increased risk of death with lung and colon cancer. Immune state studies in 157 workers proved significant immune disorders due to MMA levels several times exceeding the MAC. Experiments evaluating immune functions under exposure to MMA levels equal to or under the MAC demonstrated mostly adaptational changes.

Adult↗

PubMedDate not supplied

The role of ring D in the antitumour antibiotic streptonigrin: metal complexation, DNA binding and topoisomerase inhibition by ABC ring analogues of streptonigrin.

interaction of 7-amino-2-(6'-carboxy-2'-pyridyl)-6-methoxy-5,8-quinolinedione, an ABC ring analogue of the antitumour antibiotic streptonigrin, with zinc(II), oligonucleotides and DNA in the presence of zinc(II), and on the relaxation of DNA by topoisomerase II, has been studied. This ligand contains the key functional groups present in streptonigrin required for biological activity, but lacks the phenolic ring D which confers optical activity on streptonigrin. Variable temperature NMR experiments showed that in the presence of zinc(II) triflate, the methyl ester of the ligand forms a mixture of 1:1 and 1:2 metal:ligand bipyridyl complexes, whose relative stabilities are temperature dependent. Titrations of the water-soluble ligand with zinc(II) nitrate at room temperature showed that the predominant species present in aqueous solution at physiological pH is the 1:1 bipyridyl complex. The interaction of the ligand with the hexanucleotides d(GCATGC)2 and d(ATGCAT)2 was studied by 1H- and 31P-NMR spectroscopy. In the presence of 1 equiv of zinc(II) nitrate and 1 equiv of the ligand, small changes in chemical shifts of the proton resonances associated with the purine resonances were detected consistent with a weak interaction of the zinc(II) complex of the ligand with the oligonucleotides, possibly via a groove binding mechanism. UV-VIS titrations showed a weak interaction of the ligand with calf thymus DNA and poly(dG-dC)2 in the presence of zinc(II) but negligible interaction with poly(dA-dT)2. Gel electrophoresis experiments showed that, in contrast to streptonigrin, the ligand did not inhibit the relaxation of plasmid DNA by human topoisomerase II. These results show that the interaction of the ABC ligand with zinc(II), oligonucleotides, DNA and topoisomerase II is different to streptonigrin and hence the design of biologically active ABC ring analogues of streptongrin that operate via different mechanisms should be possible.

Antibiotics, Antineoplastic↗

PubMed1996

Preferential loss of expression of p16(INK4a) rather than p19(ARF) in breast cancer.

The tumor suppressor p16(INK4a) has been shown to be inactivated in numerous cancer lines and primary tumors. Recently, we reported loss of heterozygosity of the region in which p16(INK4a) is located in more than one-half of primary breast tumors. However, mutational analysis of these same tumors revealed mutation of p16(INK4a) to be infrequent. Other possible modes of inactivation, such as de novo methylation and homozygous deletion, have since been shown to occur in numerous neoplasias. Furthering the complexity of this locus, a transcript overlapping the p16(INK4a) coding sequence and encoding a novel peptide with growth-suppressive activity, p19(ARF), has been described. To clearly elucidate the target of aberrations affecting this subchromosomal region and approximate frequency in breast cancer, we performed a comprehensive study including p16 deletion analysis by means of interphase chromosomal fluorescence in situ hybridization, methylation analysis of the first exon encoding p16(INK4a) (exon 1alpha), mutational analysis of exon 1beta by single-strand conformational polymorphism analysis of p19(ARF) transcripts, and expression of both alpha and beta transcripts by reverse transcription PCR. Homozygous deletion of p16, as determined by interphase chromosomal fluorescence in situ hybridization, was observed in 3 of 18 (17%) tumors analyzed, whereas de novo methylation of exon 1alpha was observed in an additional 17% (4 of 23). Reduced expression of p16(INK4a) was observed in 11 tumors (48%), including all those in which homozygous deletion or complete methylation was observed. No mutations of exon 1 beta were detected, and expression of its transcript was variable, with 13% demonstrating decreased expression and 17% demonstrating overexpression. These results further support p16(INK4a) as a target of inactivation in the 9p21 region for breast cancer.

Breast Neoplasms↗

PubMed2001

Calpha methylation in molecular recognition. Application to substance P and the two neurokinin-1 receptor binding sites.

Two binding sites NK-1M (major, more abundant) and NK-1m (minor) are associated with the neurokinin-1 receptor. For the first time with a bioactive peptide, the Calpha methylation constraint, shown to be a helix stabiliser in model peptides, was systematically used to probe the molecular requirements of NK-1M and NK-1m binding sites and the previously postulated bioactive helical conformation of substance P (SP). Seven Calpha methylated analogues of the undecapeptide SP (from position 5-11) have been assayed for their affinities and their potencies to stimulate second messenger production. The consequences of Calpha methylation on the structure of SP have been analysed by circular dichroism and nuclear magnetic resonance combined with restrained molecular dynamics. The decreased potencies of six out of these seven Calpha methylated SP analogues do not allow the identification of any clear-cut differences in the structural requirements between the two binding sites. Strikingly, the most active analogue, [alphaMeMet5]SP, leads to variable subnanomolar affinity and potency when interacting with the NK-1m binding site. The conformational analyses show that the structural consequences associated with Calpha methylation of SP are sequence dependent. Moreover, a single Calpha methylation is not sufficient by itself to drastically stabilize a helical structure even pre-existing in solution, except when Gly9 is substituted by an alpha-aminoisobutyric acid. Furthermore, Calpha methylation of residues 5 and 6 of SP in the middle of the postulated helix does not stabilize, but decreases (to different extents) the stability of the helical structure previously observed in the 4-8 domain of other potent SP analogues.

Animals↗

PubMed1993

Histamine N-methyltransferase: inhibition by monoamine oxidase inhibitors.

Histamine N-methyltransferase (HNMT) catalyzes the N tau-methylation of histamine. N tau-Methylhistamine can then undergo oxidation catalyzed by the mitochondrial enzyme monoamine oxidase (MAO). Addition of an MAO inhibitor such as pargyline to tissue preparations can increase the HNMT activity assayed --presumably as a result of inhibition of N tau-methylhistamine metabolism by MAO. However, pargyline-dependent "activation" of HNMT may also occur in tissue preparations that lack mitochondria. Our experiments were performed to determine whether MAO inhibitors, like many other amine compounds, could directly increase the activity of partially purified HNMT, and, if so, to study the mechanism of activation. Human kidney HNMT was partially purified by sequential ion exchange and gel filtration chromatography. The activity of the purified HNMT was increased approximately 50% in the presence of pargyline. However, enzyme kinetic experiments showed that pargyline, like many other amines, was a competitive inhibitor of HNMT. Apparent activation of the enzyme resulted from sequential shifts of histamine substrate curves to higher Vmax values in the presence of increasing concentrations of pargyline. Other acetylenic MAO inhibitors, clorgyline and the two stereoisomers of deprenyl, were also competitive inhibitors of purified human kidney HNMT. Inhibition kinetic experiments performed in the presence of varying concentrations of histamine demonstrated that Kis values for pargyline, clorgyline, (R)-deprenyl and (S)-deprenyl were 0.126, 0.144, 0.217, and 0.627 mM, respectively. When the concentration of the cosubstrate for the reaction, S-adenosyl-L-methionine, was varied in the presence of variable concentrations of pargyline, inhibition of HNMT by pargyline was noncompetitive with regard to the methyl donor, with Kii and Kis values of 1.23 and 0.95 mM, respectively. Finally, several amine compounds related structurally to pargyline were also found to be inhibitors of HNMT.

Enzyme Activation↗

PubMed1990

A study in rat brain cortex synaptic vesicles of endogenous ligands for N-methyl-D-aspartate receptors.

The presence of endogenous ligands for the N-methyl-D-aspartate receptor was looked for in highly purified rat brain cortex synaptic vesicles, the contents of which were extracted and fractionated by gel filtration on Sephadex G-10, or by three different high-voltage electrophoresis procedures. The presence of endogenous ligands was detected by their ability to compete with 50 nM L-[3H]glutamate for binding to whole rat brain N-methyl-D-aspartate receptors. The receptor preparations used were those present in purified postsynaptic densities, in which the quisqualate receptors were blocked by 10 microM quisqualate. Synaptic vesicles had a high content of N-methyl-D-aspartate receptor ligands, which on fractionation always coincided with glutamate or aspartate. A variable and very small amount of a highly acidic endogenous ligand was also found. The latter substance did not coincide in the electrophoresis with homocysteic, cysteic, quinolinic, cysteine sulphinic or homocysteine sulphinic acids, or with N-acetyl-aspartyl-glutamic acid, S-sulphocysteine or sulphoserine. We also found that a single centrifugation, in 0.25 M sucrose, 25 mM Tris-citrate, pH 7.1, of purified synaptic vesicles, at 135,000 gmax for 45 min, led to a 51% loss of endogenous glutamate, but did not change their aspartate content. Thus, in uncentrifuged vesicles the glutamate/aspartate ratio was 9.4, while in centrifuged ones the ratio was 3.9 ATP markedly enhanced L-[3H]glutamate uptake into synaptic vesicles, but did not change the binding of L-[3H]aspartate. Differences in labelled aspartate and glutamate efflux from the vesicles were also found.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗

PubMed1988

Structural studies of O6-methyldeoxyguanosine and related compounds: a promutagenic DNA lesion by methylating carcinogens.

O6-Methylation of guanine residues in DNA can induce mutations by formation of base mispairing due to the deprotonation of N(1). The electronic, geometric and conformational properties of three N(9)-Substituted O6-methylguanine derivatives, O6-methyldeoxyguanosine (O6mdGuo), O6-methylguanosine (O6mGuo) and O6, 9-dimethylguanine (O6mdGua), were investigated by X-ray and/or NMR studies. O6mdGuo crystallizes in the monoclinic space group P2(1) with cell parameters a = 5.267(1), b = 19.109(2), c = 12.330(2) A, beta = 92.45(1) degrees, V = 1239.8(3) A3, z = 4 (two nucleosides per asymmetric unit), and O6mGua in the monoclinic space group P2(1)/n with cell parameters a = 10.729(2), b = 7.640(1) c = 10.216(1) A, beta = 92.17(2) degrees, V = 836.7(2) A3, z = 4. The geometry and conformation of O6-methylguanine moieties observed in both crystals and are very similar. Furthermore, the molecular dimensions of the O6methylguanine residue resemble more closely those of adenine than those of guanine. The methoxy group is coplanar with the purine ring, the methyl group being cis to N(1). The conformation of O6-methylguanine nucleosides is variable. The glycosidic conformation of O6mdGuo is anti for molecule (a) and high-anti for molecule (b) in the crystal, while that of O6mGuo is syn [Parthasarathy, R & Fridey, S. M. (1986) Carcinogenesis 7, 221-227]. The sugar ring pucker of O6mdGuo is C(2')-endo for molecule (a) and C(1')-exo for molecule (b). The C(4')-C(5') exocyclic bond conformation in O6mdGuo is gauche- for molecule (a) but trans for molecule (b), in contrast with gauche+ for O6mGuo. The hydrogen bonds exhibited by O6-methylguanine derivatives differ from those in guanine derivatives; the amino N(2) and ring N(3) and N(7) atoms of O6-methylguanine residues are involved in hydrogen bonding. 1H-NMR data for O6mdGuo and O6mdGuo reveal the predominance of a C(2')-endo type sugar puckering. In O6mdGuo, however, a contribution of a C(1')-exo sugar puckering is significant. The NOE data also indicate that O6mdGuo molecules exist with nearly equal population for anti (including high anti) and syn glycosidic conformations. These observations and their biological implications are discussed.

Alkylating Agents↗

PubMed1981

[Limited accessibility of DNA methylation sites for bacterial methylases M. Eco RII and M.Eco dam in chromatin at different levels of organization].

The bacterial methylases M. Eco RII and M. Eco dam can methylate DNA in rat liver chromatin to form the 5-methylcytosine (m5C) and N6-methyladenine (m6A) residues, respectively. The CH3-accepting capacity of DNA in chromatin (mono- and dinucleosomes, mono- and dinucleomers) is 15 - 30 times less than that of free total DNA in rat liver. Such a low level of DNA methylation in chromatin in vitro suggests that the accessibility and recognition of methylation sites by DNA-methylases are decreased in comparison with free DNA both in the core-particle DNA and in the internucleosomal DNA. The degree of DNA methylation in chromatin particles depends on the ionic strength and Mg2+; when the former is decreased from 0.515 down to 0.176, the DNA methylation by both enzymes is increased 2-fold. An addition of Mg2+ (1 - 2 mM) decreases the CH3-accepting capacity of nucleomeric DNA, that of nucleosomal DNA remains unchanged. Thus, the accessibility of DNA for methylases is variable depending on the conformational changes of chromatin. The values of the m6A to m5C ratio for free and nucleosomal DNAs formed by methylation with a methylation of nucleomeric DNA, i. e. 1.01, 0.92 and 0.51, respectively. As Mg/4 concentration rises, the m6A/m5C ratio for nucleosomal and nucleomeric DNA is increased. It seems therefore that at different levels of organization and upon certain conformation changes the number and, probably, the nature of exposed DNA methylation sites in chromatin are different. Bacterial DNA-methylases can be used as an effective probe for a fine analysis of chromatin ultrastructure, in particular at its different functional states.

Animals↗

PubMed2004

Potential application of tissue Doppler imaging to assess regional left ventricular diastolic function in patients with hypertrophic cardiomyopathy: comparison with 123I-beta-methyl iodophenyl pentadecanoic acid myocardial scintigraphy.

BACKGROUND: Tissue Doppler imaging (TDI) has been utilized to evaluate left ventricular myocardial dysfunction in patients with hypertrophic cardiomyopathy (HCM); however, no clear explanation for the abnormality of TDI variables has been forthcoming. HYPOTHESIS: Peak negative myocardial velocity gradient (MVG) derived from TDI may correlate with a disorder of fatty acid metabolism in patients with HCM. METHODS: Tissue Doppler imaging and 123I-beta-methyl iodophenyl pentadecanoic acid (123I-BMIPP) myocardial scintigraphy were performed in 15 patients with asymmetric septal hypertrophy (mean age 47 +/- 18 years) and in 12 healthy controls (mean age 43 +/- 10 years). RESULTS: In early 123I-BMIPP images, accumulation defects were observed in the ventricular septum in 12 patients and in the posterior wall in 8 patients with HCM. Peak negative MVG in the ventricular septum (1.1 +/- 0.5 vs. 2.8 +/- 0.5, p < 0.0001) and posterior wall (5.2 +/- 1.4 vs. 6.7 +/- 0.8, p < 0.01 ) was significantly lower in the HCM group than in the controls; also, these parameters were significantly lower in patients with than in those without a defect in the region in question. The peak negative MVG in the ventricular septum and posterior wall correlated inversely with the washout rate in all subjects. CONCLUSIONS: Peak negative MVG according to TDI is related to disorder of fatty acid metabolism in the regional left ventricular myocardium of patients with HCM.

Cardiomyopathy, Hypertrophic↗

PubMed1996

L-arginine augments nitric oxide production and mesenteric blood flow in ovine endotoxemia.

We studied the effects of administrating the nitric oxide synthase inhibitor, NG-nitro-L-arginine methyl ester (L-NAME), or the nitric oxide precursor, L-arginine, on hemodynamic variables and serum nitrate concentrations in an anesthetized ovine model of endotoxemia to assess the effects on regional visceral blood flow and to determine whether L-arginine availability limits nitric oxide production. Animals received Escherichia coli endotoxin (2 micrograms/kg) followed 2 h later by L-NAME (25 mg/kg), L-arginine (0.575 g/kg), or saline administered over 1 h followed by an infusion of the same dose over 8 h (n = 6 per group). Renal and mesenteric blood flow were measured by placement of electromagnetic flow probes, and serum nitrate concentrations were determined using vanadium III chloride or nitrate reductase reduction to nitric oxide or nitrite, respectively. The results showed L-NAME significantly increased systemic vascular resistance (P < 0.01), decreased serum nitrate concentrations (P < 0.05), and caused a transient reduction in mesenteric blood flow (P < 0.05). L-Arginine caused a reduction in systemic vascular resistance (P < 0.01), increased mesenteric blood flow (P < 0.001) and conductance (P < 0.05). There were no significant changes in renal arterial blood flow in either group. We conclude that the availability of L-arginine limits nitric oxide production in endotoxemia and, furthermore, that L-arginine administration in this model causes significant mesenteric vasodilatation. L-NAME administration had only limited effect on visceral blood flow despite a marked increase in systemic vascular resistance and a reduction in nitric oxide production.

Animals↗

PubMed2025

Quorum sensing and DNA methylation play active roles in clinical Burkholderia phase variation.

Phenotypic diversity in bacteria often results from adaptation to changing environmental conditions, exemplified by variable colony morphotypes. In Burkholderia pseudomallei, discrete genomic alterations and modulation of gene expression facilitate adaptation. Adapted variants of species within the Burkholderia cepacia complex (Bcc) often lose the pC3 virulence megaplasmid, impacting their colony morphology and their production of virulence factors. In this study, we characterize variants arising in Burkholderia ambifaria clinical isolates using proteomics and phenotypic tests and show that some of them have retained the pC3, indicating a distinct phase variation mechanism at play in this Bcc species. Interestingly, variants of B. ambifaria strains CEP0996 (pC3-null) and HSJ1 (pC3-positive) still share similarities in phenotypes controlled by the Cep quorum-sensing (QS) system. We further investigated the role of QS in B. ambifaria HSJ1 phase variation and confirmed that the Cep QS system is important for the emergence of variants. Given that DNA methylation is a key epigenetic factor regulating virulence factors in Burkholderia cenocepacia, we hypothesized that adenosine DNA methylation also governs phase variation in B. ambifaria HSJ1. By deleting the genes encoding putative adenosine DNA methyltransferases, we discovered that an orphan type II DNA methyltransferase inhibits the emergence of phase variants. This study is the first to demonstrate that quorum sensing and adenosine DNA methylation are two antagonistic systems independently controlling phase variation in B. ambifaria.IMPORTANCESome Burkholderia species are pathogenic to plants, animals, or humans. In immunocompromised individuals, and people suffering from cystic fibrosis, infection from the Burkholderia cepacia complex (Bcc) can lead to "cepacia syndrome." In northern Australia and southeast Asia, melioidosis caused by Burkholderia pseudomallei is prevalent among native population, particularly among people with diabetes, chronic lung or kidney disease or alcoholism. Burkholderia's phenotypic plasticity, including colony morphotype variation (CMV), enables rapid adaptation to diverse environments, enhancing its survival and pathogenicity. This study reveals phase variation as a new CMV mechanism within the Bcc group and is the first to report that quorum sensing and DNA methylation are involved in phase variation. Understanding the underlying mechanisms of CMV could lead to the development of targeted therapies against these highly antibiotic-tolerant bacteria.

Quorum Sensing↗

PubMed2006

Prediction of gas chromatographic retention of polyunsaturated fatty acid methyl esters.

Multivariate regression models were applied to predict retention indices as equivalent chain lengths (ECL) for methylene-interrupted polyunsaturated fatty acids. Simple molecular descriptors, the chain length, the number of double bonds and the position of the double bond system, were used as predictors. The merits of different variable combinations were evaluated. For general models, it was necessary to include the distance from the double bond system to both the carbonyl group (Delta-position) and the methyl end of the fatty acid (n-position). The best accuracy was found for models including higher order terms of Delta and n. For models restricted to n-3 and n-6 isomers, it was not necessary to include the n-position among the variables. The highest residuals for the most accurate models were below 0.06 ECL units, and root mean square error of prediction was below 0.030. The ECL data was achieved by three different temperature programs on a cyanopropyl column.

Chromatography, Gas↗

PubMedDate not supplied

Repair-modification and evolution of the eukaryotic genome organization.

For a complete reconstruction of the damaged unmethylated islands, in theory, the conventional excision-repair is sufficient. For a complete reconstruction of the damaged methylated domains, a coupling has to take place involving the excision-repair (able to reestablish their ATGC-language) plus the DNA-methylase (able to reestablish their modified ATGC5mC-language). This coupling, defined as "repair-modification," is essentially functioning during the S-phase, because the DNA-polymerase beta (pol beta) is active during the whole cell cycle, whereas the DNA-methylase (met) is active in S and appears to be repressed or inactive during the major part of G1 and during the phases G2 and M. Consequently, after damage, some silent genes might become expressed during these phases, if it is true that DNA methylation is inversely proportional to transcription. Repair-modification should, therefore, exert a continuous differential pressure on evolution of given parts of the genome, when they are methylated to a different extent. According to Darwinian concepts, repair-modification would lead to a high variability, especially of uncoding DNA sequences (if hypermethylated), whereas on the basis of this variability, selection might favor transposition of specific regulatory elements into given transcriptional units. In these, the conservative nature of the coding elements (if unmethylated) would obviously be ensured by the conventional excision-repair.

Animals↗

PubMed1981

Hexose transport in Novikoff rat hepatoma cells. A simple carrier with directional symmetry, but variable relative mobilities of loaded and empty carrier.

The kinetics of transport of the non-metabolizable hexose, 3-O-methyl-D-glucose, have been measured in Novikoff rat hepatoma cells by both zero-trans entry and equilibrium exchange procedures. Transport conformed to a simple carrier model which operates symmetrically with respect to direction, but with greater mobility of the loaded than of the empty carrier. Although a complete kinetic description of the transporter can, in theory, be obtained by application of integrated equations describing the time course of substrate equilibrium across the membrane beginning from the zero-trans situation, statistical analysis of hypothetical data indicated that directional asymmetry or differential mobilities of loaded and empty carrier cannot be discerned reliably from such data alone. The difference in mobility of loaded and empty carrier, apparent in a comparison of zero-trans entry and exchange data, ranged from 1.5--7-fold in different batches of cells. It is concluded that the magnitude of the difference is not an inherent property of the transporter, but is determined physiologically, and may be involved in regulation of hexose transport.

Animals↗

PubMed1998

3-Alkyl ethers of clocinnamox: delayed long-term mu-antagonists with variable mu efficacy.

In recent years there has been considerable interest in the relationship between clocinnamox (C-CAM) and its methyl ether methoclocinnamox (MC-CAM). While C-CAM appears to be an insurmountable mu-antagonist, MC-CAM has been shown to be a potent partial agonist at mu-opioid receptors. To further investigate this relationship we prepared other ethers of C-CAM and evaluated these in opioid receptor binding assays and in vivo in mouse antinociceptive assays and in morphine-dependent monkeys. In opioid binding assays, the ethers were generally mu-selective with affinity equivalent to that of C-CAM itself. Although they displayed little or no efficacy in vitro, some of the ethers showed substantial agonist activity in the in vivo antinociceptive tests. Two of the ethers, the propargyl ether 7 and the cyclopropylmethyl ether 5, were chosen for more detailed analysis in vivo. 7 was shown to have significant mu-agonist character and was able to substitute for morphine in morphine-dependent monkeys. Interestingly, when this agonist effect abated, 7 displayed long-lasting mu-antagonism. In contrast, 5 displayed little agonist activity in vivo and was characterized as a potent, long-acting mu antagonist. Although further work is needed to determine whether metabolism is a crucial factor in determining the pharmacological profile of these ethers, it is clear that 3-O-alkylation is a useful means of varying the mu efficacy displayed by this class of acyl-substituted 14-aminomorphinones. MC-CAM itself has generated considerable interest as a potential pharmacotherapy for opiate abuse. These analogues with differing mu efficacy but retaining the long-lasting mu-antagonist effects provide further opportunities for the development of treatment drugs.

Animals↗

PubMed1992

Blockade of conditioned taste aversion by scopolamine and N-methyl scopolamine: associative conditioning, not amnesia.

The anticholinergic, scopolamine, consistently disrupts one-trial passive avoidance conditioning but the effects of such drugs on one-trial conditioned taste aversion (CTA) are variable and contradictory. In the present study, treatment of rats with scopolamine impaired the suppression of sucrose intake by post-ingestion administration of lithium chloride (LiCl) in a two-bottle choice test. A similar effect was obtained by using N-methyl scopolamine which penetrates the brain only to a limited degree on acute administration. The blockade of CTA could be prevented in three ways: (i) by exposing the rats to sucrose only on the training day, (ii) by pre-exposing the rats to both sucrose and scopolamine, and (iii) by using a less palatable sucrose/ascorbate mixture. The results demonstrate that the effect of scopolamine on taste aversion is not mediated by the central nervous system, and can be modified by altering the novelty and relative salience of the taste conditioned stimulus. These experiments suggest that conditioned associations between taste and LiCl, and scopolamine and LiCl may underlie the blockade of CTA by scopolamine.

Amnesia↗