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Biomedical subjects

M E Reith

Publications and source records attributed to M E Reith.

At least 109 records · Page 6Linked to original sources
PubMed1989

Phosphoinositide hydrolysis induced by depolarization and sodium channel activation in mouse cerebrocortical slices.

Carbachol, a muscarinic receptor agonist and the sodium channel-activating agents, scorpion venom, veratridine, batrachotoxin and aconitine, were shown to stimulate the formation of [3H]inositol phosphates in [3H]inositol-labelled miniprisms, obtained from the cerebral cortex of the mouse. The inositol response to the Na+ channel-activating agents was inhibited by the sodium channel blocker tetrodotoxin (TTX), while the response induced by carbachol was partially resistant to TTX. The response to scorpion venom and the TTX-insensitive portion of the response to carbachol was additive, indicating different mechanisms. The presence of high potassium (K+) induced hydrolysis of inositide in a TTX-insensitive manner and was not additive with that resulting from sodium channel activators, thus indicating a common mechanism. The addition of large concentrations of magnesium to block the release of acetylcholine, did not inhibit the inositol response to high K+ or to veratridine. Calcium channel blockers such as nickel or cobalt, or the dihydropyridine calcium (Ca2+) channel activator BAY K 8644 and the calcium channel blocker nifedipine, nimodipine or PN-200 110 had little effect. Monensin, a sodium ionophore, stimulated the turnover of phosphatidylinositol at non-depolarizing concentrations and the omission of Na+ ions inhibited the response to sodium channel agents and to high K+. Thus, membrane potential and gradients of K+, Na+ and Ca2+ are all important factors determining the final effect on the turnover of phosphatidylinositol. The data are consistent with a model in which all these factors impinge on the Na+/Ca2+ exchanger regulating internal Ca2+ that, in turn, activates phospholipase C.

Animals↗
PubMed1989

Oxidative metabolism of cocaine: comparison of brain and liver.

Norcocaine (NC) and N-hydroxynorcocaine (NHNC), products of the oxidative metabolism of cocaine, were examined in plasma, brain, and liver of mice injected intraperitoneally with cocaine. Plasma levels of NHNC were altered in vivo by inhibiting esterase activity with diazinon and chloral hydrate or activating esterase activity with phenobarbital, and activating the microsomal P-450 system with phenobarbital. Changes in plasma concentrations of NHNC resulted in similar changes in brain, which were often different from those in liver. After intracisternal administration of cocaine to mice, no appreciable amount of NC or NHNC could be detected in brain; the same results were obtained upon intracisternal and intraventricular administration to rats. Microsomal preparations from mouse brain were found to be considerably less active than those from liver in converting NC to NHNC. We conclude that the cerebral oxidative metabolism of cocaine is not appreciable and that most of the NC and NHNC found in the brain after systemic cocaine administration is derived from plasma rather than formed centrally by brain microsomal enzymes.

Animals↗
PubMed1988

Uptake of [3H]serotonin into plasma membrane vesicles from mouse cerebral cortex.

Preparations of plasma membrane vesicles were used as a tool to study the properties of the serotonin transporter in the central nervous system. The vesicles were obtained after hypotonic shock of synaptosomes purified from mouse cerebral cortex. Uptake of [3H]serotonin had a Na+-dependent and Na+-independent component. The Na+-dependent uptake was inhibited by classical blockers of serotonin uptake and had a Km of 63-180 nM, and a Vmax of 0.1-0.3 pmol mg-1 s-1 at 77 mM Na+. The uptake required the presence of external Na+ and internal K+. It required a Na+ gradient ([Na+]out greater than [Na+]in) and was stimulated by a gradient of K+ ([K+]in greater than [K+]out). Replacement of Cl- by other anions (NO2-, S2O3-(2-)) reduced uptake appreciably. Gramicidin prevented uptake. Although valinomycin increased uptake somewhat, the membrane potential per se could not drive uptake because no uptake was observed when a membrane potential was generated by the SCN- ion in the absence of internal K+ and with equal [Na+] inside and outside. The increase of uptake as a function of [Na+] indicated a Km for Na+ of 118 mM and a Hill number of 2.0, suggesting a requirement of two sodium ions for serotonin transport. The present results are accommodated very well by the model developed for porcine platelet serotonin transport (Nelson, P. J., and Rudnick, G. (1979) J. Biol. Chem. 254, 10084-10089), except for the number of sodium ions that are required for transport.

Animals↗
PubMed1988

Interaction of yohimbine with batrachotoxinin binding to mouse brain sodium channels.

To study the local anesthetic properties of yohimbine in more detail, its effect was examined in vitro on the scorpion toxin-enhanced specific binding of [3H]batrachotoxinin A 20-alpha-benzoate [( 3H]BTX-B) to the gating complex in sodium channel preparations from mouse brain cortex. Both equilibrium and kinetic experiments were carried out. Yohimbine inhibited the specific binding of [3H]BTX-B in the vesicular preparation with an IC50 value of 2.2 X 10(-5) M. This is about one order of magnitude higher than the concentration required for antagonism via the alpha 2-adrenoceptors; however, yohimbine is 7-fold more potent in inhibiting [3H]BTX-B binding than lidocaine. In a concentration-dependent manner, yohimbine increased the dissociation constant (Kd) of high-affinity [3H]BTX-B binding without changing the maximal binding capacity (Bmax). The dissociation rate constant was not affected by yohimbine, suggesting competitive inhibition as opposed to the action of local anesthetics involving an allosteric action via receptor sites distinct from the BTX site. Alpha 2-adrenoceptors are apparently not involved because clonidine and alpha-methyl-noradrenaline had no appreciable effect on [3H]BTX-B binding and did not antagonize the inhibitory effect of yohimbine. The present findings indicate a mechanism of local anesthetic action of yohimbine that differs from that of other local anesthetics such as tetracaine and lidocaine involving direct binding to the BTX site, thereby stabilizing a non-permeable form of the sodium channel.

Adrenergic alpha-Agonists↗
PubMed1988

The mode of action of ethanol on batrachotoxinin-A benzoate binding to sodium channels in mouse brain cortex.

Since ethanol has local anesthetic activity its effect was examined in vitro on the scorpion toxin-enhanced specific binding of [3H]batrachotoxinin A 20-alpha-benzoate ([3H]BTX-B) to the gating complex in sodium channel preparations from mouse brain cortex by equilibrium and kinetic experiments. Ethanol inhibited the specific binding of [3H]BTX-B in a vesicular preparation with an IC50 value of 310 mM and a Hill number of 1.0. Ethanol increased the equilibrium dissociation constant of batrachotoxin in a concentration dependent manner without changing the maximal binding capacity and decreased the half-time of the aconitine-induced dissociation of [3H]BTX-B. Thus, ethanol acts as an apparent competitive inhibitor, allosterically affecting the [3H]BTX-B binding, like other local anesthetics. Results of competition experiments in the presence of different concentrations of ethanol and fixed concentration of tetracaine (and vice versa) are consonant with an interaction of ethanol and tetracaine with the same binding sites. Experiment carried out at 32, 37 and 42 degrees C indicated that the effect of ethanol is not mimicked by increasing the temperature.

Animals↗
PubMed1988

Evidence that there is no direct correlation between alpha 2-adrenoceptor antagonism and inhibition of voltage-dependent sodium channels.

Electrophysiological and biochemical evidence suggests that the voltage-dependent sodium channel is the site of local anesthetic action, and that there is pharmacological similarity between alpha-adrenoceptors and Na+-channels. Yohimbine, a non-selective alpha 2-adrenoceptor antagonist, with a structure similar to that of cocaine affects the sodium channel by a mechanism different from that of other local anesthetics including cocaine. Some structural analogues of yohimbine -berbane compounds- were found to be potent and selective alpha 2-adrenoceptor antagonists. In this work the local anesthetic properties of two berbane compounds (6c and 6d (CH-38083) from the paper of Vizi, Toth, Somogyi, Szabo, Harsing and Szantay, 1987) were examined and compared to those of yohimbine in vitro on scorpion venom-enhanced specific binding of [3H]batrachotoxinin A 20-alpha-benzoate [( 3H]BTX-B) to the voltage-sensitive sodium channel and on the veratridine-induced depolarization measured by the uptake of [3H]trimethylphenylphosphonium ion [( 3H]TPMP+) in mouse brain cortex. Both of the compounds inhibited the [3H]BTX-B binding with an IC50 of (approximately) 150 microM, which is more than four orders of magnitude higher than the concentration required for antagonism of a presynaptic alpha 2-adrenoceptor (7 nM). They are 15 times less potent in inhibiting [3H]BTX-B binding and 2.5 times less potent in inhibiting veratridine-induced depolarization than yohimbine.

Adrenergic alpha-Antagonists↗
PubMed1988

Metaphit prevents locomotor activation induced by various psychostimulants and interferes with the dopaminergic system in mice.

Metaphit, an isothiocyanate analog of phencyclidine and a proposed phencyclidine receptor acylator, inactivated the carrier involved in the neuronal uptake of dopamine in in vitro experiments with preparations of the striatum in the mouse. In ex vivo experiments 2 and 24 hr after the intravenous administration of metaphit, no changes were observed either in the binding of [3H]cocaine to striatal membranes or in the uptake of [3H]dopamine into synaptosomes or slices. In in vivo experiments 24 hr after pretreatment with metaphit, selective labelling of uptake sites for dopamine in the striatum of the mouse with [3H]GBR 12935 was unaffected. In these in vivo experiments, however, metaphit antagonized the locomotor stimulation induced by blockers of the uptake of dopamine (methylphenidate, mazindol, cocaine, GBR 12909) but not that induced by drugs that affect locomotion by other mechanisms (amphetamine, phencyclidine). Twenty-four hours after treatment with metaphit there was an increase in homovanillic acid in all regions of the brain studied (striatum, olfactory tubercle, cerebral cortex). There was no effect of metaphit on the disappearance rate of 3,4-dihydroxyphenylacetic acid and homovanillic acid from the striatum during the inhibition of monoamine oxidase with pargyline. If the increase in homovanillic acid reflected a greater rate of dopamine catabolism in metaphit-treated mice, it could explain the lack of locomotor stimulation of blockers uptake of the dopamine in these animals, resulting from a rapid breakdown of extracellularly accumulated dopamine.

Amphetamine↗
PubMed1988

Isolation, sequence analysis, and transcriptional studies of the flavodoxin gene from Anacystis nidulans R2.

The nonheme, iron-sulfur protein ferredoxin is the terminal constituent of the photosynthetic electron transport chain. Under conditions of iron stress, many cyanobacteria and eucaryotic algae replace ferredoxin with the flavoprotein flavodoxin. The gene for flavodoxin was cloned from the cyanobacterium Anacystis nidulans R2 by using three mixed oligonucleotide probes derived from the partial Synechococcus sp. strain PCC 6301 amino acid sequence. Nucleotide sequence analysis revealed a 513-base-pair open reading frame with a deduced amino acid sequence having homology to other long-chain flavodoxins. Assuming proteolytic cleavage of the initial methionine residue, the molecular weight of the A. nidulans R2 flavodoxin is 18,609. Southern blot hybridization under conditions of reduced stringency detected only one copy of the flavodoxin sequence in the A. nidulans R2 genome. Northern (RNA) blot hybridization analyses by using cloned flavodoxin gene probes indicated that no transcripts are detectable under conditions of iron saturation. However, under iron-deficient growth conditions the flavodoxin gene appeared to be transcribed as part of a larger operon. The operon yielded at least three transcripts. The first was of approximately 1,100 bases (designated RNA 1) and terminated immediately upstream from the 5' end of the flavodoxin open reading frame. A second, less abundant transcript of approximately 1,900 bases (designated RNA 2) encoded all of RNA 1 as well as the flavodoxin polypeptide. Analysis indicated that both transcripts initiate in close proximity to each other. A third, minor transcript of approximately 1,100 bases (designated RNA 3) was detectable downstream of the flavodoxin gene sequence. Addition of iron-stressed A. nidulans R2 cells resulted in almost total loss of detectable mRNA transcripts within 60 min of the addition. The ferredoxin gene transcript has previously been characterized as a monocistronic message of approximately 430 bases (M. E. Reith, D. E. Laudenbach, and N. A. Straus, J. Bacteriol. 168: 1319-1324, 1986). Here we show that the ferredoxin message is detectable under all iron regimes tested is quantitatively unaffected by decreases in iron availability to the cells.

Amino Acid Sequence↗
PubMed1987

Binding sites for [3H]tetracaine in synaptosomal sodium channel preparations from mouse brain.

The present study was an attempt to answer the question whether the local anesthetic [3H]tetracaine labels sodium channels in mouse brain synaptosomes. Binding of [3H]tetracaine had a Kd of 0.19 microM and a Bmax ranging from 3.7 to 5.2 pmol/mg of protein. Local anesthetics other than tetracaine and cocaine-related compounds inhibited [3H]tetracaine binding with Hill numbers between 0.3 and 0.6. Initiation of the dissociation of [3H]tetracaine binding by an excess of unlabeled tetracaine resulted in non-linear curves. These results are consonant with site heterogeneity, negative cooperativity, or complexities arising from the use of synaptosomal preparations instead of broken membranes. There were important differences between the absolute values of the potency of various local anesthetic drugs and cocaine-related compounds in inhibiting binding of [3H]tetracaine and those in inhibiting binding of [3H]batrachotoxinin A 20-alpha-benzoate. Although there were some effects of manipulating the state of the channel with activating toxins on the apparent rate of association and dissociation of [3H]tetracaine binding, the equilibrium binding was not much affected by the toxins. The results indicate that further characterization is necessary before accepting [3H]tetracaine binding as a valid tool for studying sodium channels.

Anesthetics, Local↗
PubMed1987

Nicotine-induced changes in the metabolism of specific brain proteins.

The effect of acute and chronic nicotine on the metabolism of specific brain proteins was examined by measuring incorporation of labeled valine into protein, with densitometric scanning of proteins resolved by gel electrophoresis. Acute and chronic administration of nicotine (0.4 mg/kg per 30 min for 2 hours, s.c., or 0.5 mg/kg per 30 min for 5 days (Alzet mini-pump implanted subcutaneously] reduced incorporation of [14C]valine administered by approximately 6-7%. The results with chronic nicotine administration indicated a lack of tolerance for this effect of nicotine. Mecamylamine, a nicotinic ganglionic antagonist, does not seem to block the inhibition of protein synthesis. Small increases in protein content were observed in a high- and a low-molecular-weight region of SDS-polyacrylamide gel, used to separate proteins from newborn brain. In adult brain after chronic nicotine administration, selective increases and a decrease were seen in selective bands. Results are consonant with selective effects of nicotine on the synthesis or degradation of specific brain proteins.

Animals↗
PubMed1987

Effects of caffeine on monoaminergic systems in mouse brain.

Effects of caffeine on monoamine systems in the mouse brain were studied in three lines of experiments. First, concentrations of 10(-7) M to 10(-2) M of caffeine were tested for their potency in inhibiting the carriers involved in the neuronal uptake of dopamine, norepinephrine, and serotonin. The IC50 of caffeine in inhibiting the first two carriers was approximately 10(-2) M, and that in inhibiting the serotonin was 2 x 10(-3) M. Second, concentrations of 10(-5) M to 10(-3) M of caffeine were tested for their potency in affecting the in vitro KC1-induced release of [3H]dopamine from dopamine terminals in the striatum and from norepinephrine terminals in the hypothalamus, and the release of [3H]serotonin from serotonin terminals in the striatum. Little or no effect was observed. Third, caffeine was administered for 3 weeks to mice via their drinking water at 73, 123, and 162 mg/kg per day. No changes were found in their D2-dopaminergic, 5-HT2-serotonergic, or alpha 1-adrenergic receptors in the striatum or cerebral cortex as compared with animals on normal drinking water. All these neurochemical results are consonant with the interpretation of behavioral studies suggesting that caffeine is an only mildly stimulatory drug that should not be grouped with other psycho-stimulant drugs such as amphetamine and cocaine that do affect monoamine systems in the brain.

Animals↗
PubMed1987

Pharmacokinetics of systemically administered cocaine and locomotor stimulation in mice.

Cocaine and its metabolites were measured in plasma and brain from mice injected i.p. with cocaine and monitored for spontaneous locomotor behavior. Cocaine concentrations in the brain reached peak values within 5 min after administration of cocaine. At all time points between 5 and 60 min the concentrations of cocaine in the brain were 7-fold higher, on the average, than those in plasma. The opposite was true for the concentrations of benzoylecgonine; brain to plasma ratios of benzoylecgonine were approximately 0.1 from 5 to 30 min after i.p. cocaine injection. After i.p. injection of either 10 or 25 mg/kg of cocaine, cocaine disappeared from plasma and brain with a half-life of 16 min and benzoylecgonine disappeared from plasma with a half-life of 62 min. There was good correspondence between locomotor stimulation and concentration of cocaine in the brain measured at 12, 22 and 32 min after i.p. administration of 25 mg/kg of cocaine. Among individual animals there was a significant correlation between their locomotor stimulation and their brain cocaine concentration, indicating that differences in cocaine levels in the brain between animals contribute to their different behavioral response; however, the correlation analysis also indicated the role of other factors determining the locomotor response to cocaine.

Animals↗
PubMed1987

Cocaine disposition in the brain after continuous or intermittent treatment and locomotor stimulation in mice.

Intermittent s.c. and i.p. injections of cocaine (20 mg/kg; total 430 mg/kg for each animal) for 18 days resulted in locomotor stimulation of mice upon challenge with cocaine on the 25th or 26th day, compared with no locomotor stimulation in a saline-pretreated group. In contrast to the sensitization by intermittent cocaine administration, tolerance was found upon challenge after continuous administration of cocaine by minipumps (25 mg/kg/day; total 450 mg/kg for each animal) on a similar schedule. No differences were found between the sensitized and tolerant groups in the levels of cocaine and benzoylecgonine in plasma and brain 12 min after i.p. administration of a challenge dose of cocaine, suggesting that in these chronic experiments the changes in the locomotor response are not accounted for by dispositional effects. In contrast, in animals treated daily for 2 or 3 days i.p. with cocaine and challenged with cocaine 1 day later, there was both a greater locomotor stimulation and a higher level of brain cocaine than in saline-pretreated animals, suggesting a dispositional effect. Among individual animals there was a positive correlation between their locomotor stimulation by the challenge dose and their brain cocaine concentration.

Animals↗
PubMed1986

Effect of repeated administration of various doses of cocaine and WIN 35,065-2 on locomotor behavior of mice.

Cocaine and a cocaine analog, WIN 36,065-2, were administered daily for 3 or 4 days. Both compounds developed sensitization to their locomotor stimulatory effects, with similarly shaped dose-response curves. Dosages giving optimal sensitization were 25 mg/kg per day for cocaine and 3 mg/kg for WIN 35,065-2. At 40 mg/kg, cocaine produced less stimulation of locomotion on day 3 than on day 1. With doses of 6 and 10 mg/kg of WIN 35,065-2, tolerance to locomotor stimulation was observed. Monitoring by an observer revealed that after 40 mg/kg of cocaine the animals spent about the same amount of time expressing non-ambulatory behavior such as biting, sniffing, rearing, or resting on day 3 as on day 1. Therefore, in the 3 day period there was no increase in non-ambulatory behavior that could have produced the decrease in locomotor stimulation measured with the activity monitor. We assessed the possible involvement of the local anesthetic action of cocaine-like compounds in inhibiting locomotion by measuring the hypomotive effect over a 3 day period by repeated administration of the local anesthetic drugs tetracaine, lidocaine, norcocaine and benzoylpseudotropine. These compounds had the same hypomotive effects on day 1 and 3, suggesting that the decrements in locomotor stimulation observed with daily high doses of cocaine and WIN 35,065-2 involve mechanisms other than their local anesthetic action. The present results indicate that the size of the dose of a psychostimulant is a crucial variable in dosage schedules for studying the sensitization to its effect on locomotor behavior.

Anesthetics, Local↗
PubMed1986

Effect of serotonin on the dissociation of high-affinity binding of [3H]imipramine in mouse cerebral cortex.

A low concentration of norzimelidine can be used to selectively measure the rate of dissociation of [3H]imipramine from high-affinity binding sites in the mouse cerebral cortex. Serotonin attenuates the dissociation initiated by norzimelidine. There is no difference between the half-time of dissociation in the presence of both serotonin and norzimelidine and that in the presence of serotonin alone. These effects are similar to but much less pronounced than those observed in the platelet.

Animals↗