Early Anglo-Norman recipes in ms London, B.L. Royal 12 C XIX.
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History of Pharmacy↗
Biomedical subjects
T Hunt
Publications and source records attributed to T Hunt.
The use of single-stranded DNA and RNase H to promote quantitative 'hybrid arrest of translation' of mRNA/DNA hybrids in reticulocyte lysate cell-free translations.
Single-stranded cDNA clones complementary to the 5' end of TMV RNA have been used to explore the conditions necessary for efficient 'hybrid arrest of translation' in the reticulocyte lysate. It is shown that incubations of 20 minutes at 60 degrees in 0.1 M KCl are sufficient to give almost complete arrest of translation using a clone complementary to the 5'-non-coding region and first 171 coding nucleotides of TMV RNA. However, hybrids with DNA complementary to regions of the mRNA downstream of the first AUG gave variable and in some cases almost no arrest of translation in the reticulocyte lysate unless they were first digested with RNase H. A simple and rapid method for giving complete and highly specific arrest of translation of particular mRNAs in complex mixtures has been developed using both cDNA clones and synthetic oligodeoxynucleotides in conjunction with RNase H digestion. Evidence is presented that suggests that 'hybrid arrest of translation' in the wheat-germ cell-free system is primarily due to the action of RNase H. When a reticulocyte lysate was doped with 20 U/ml of RNase H, its ability to translate unannealed mRNA was unaffected but it translated DNA/RNA hybrids extremely poorly.
Cell-Free System↗
Differential effects of isolation housing on the conditioned place preference produced by cocaine and amphetamine.
Rats were obtained at 21 days of age and were housed either in isolation or in groups of 4 for 6 weeks. They were then tested for their sensitivity to cocaine HCl (0.31, 0.62, 1.25 or 2.5 mg/kg) or d-amphetamine SO4 (0.031, 0.062, 0.125, 0.25 or 0.5 mg/kg) using a modified place preference paradigm. The isolated rats were insensitive to cocaine in this paradigm whereas the group-housed animals showed peak effects at the lowest dose of this drug. In contrast, there was no difference in sensitivity to amphetamine as a function of housing conditions. These data strengthen the notion that the effects of the early environment on drug sensitivity in the adult are specific to certain classes of drugs. Further, these data lend support to the notion that the effects of cocaine and amphetamine in the place preference paradigm are mediated by different neural systems.
Amphetamine↗
Differential accumulation of ribonucleotide reductase subunits in clam oocytes: the large subunit is stored as a polypeptide, the small subunit as untranslated mRNA.
Within minutes of fertilization of clam oocytes, translation of a set of maternal mRNAs is activated. One of the most abundant of these stored mRNAs encodes the small subunit of ribonucleotide reductase (Standart, N. M., S. J. Bray, E. L. George, T. Hunt, and J. V. Ruderman, 1985, J. Cell Biol., 100:1968-1976). Unfertilized oocytes do not contain any ribonucleotide reductase activity; such activity begins to appear shortly after fertilization. In virtually all organisms, this enzyme is composed of two dissimilar subunits with molecular masses of approximately 44 and 88 kD, both of which are required for activity. This paper reports the identification of the large subunit of clam ribonucleotide reductase isolated by dATP-Sepharose chromatography as a relatively abundant 86-kD polypeptide which is already present in oocytes, and whose level remains constant during early development. The enzyme activity of this large subunit was established in reconstitution assays using the small subunit isolated from embryos by virtue of its binding to the anti-tubulin antibody YL 1/2. Thus the two components of clam ribonucleotide reductase are differentially stored in the oocyte: the small subunit in the form of untranslated mRNA and the large subunit as protein. When fertilization triggers the activation of translation of the maternal mRNA, the newly synthesized small subunit combines with the preformed large subunit to generate active ribonucleotide reductase.
Animals↗
Inhibition of rat mixed lymphocyte pancreatic islet cultures with anti-Ia immunotoxin.
Studies reported here indicate that an anti-Ia immunotoxin can eliminate the allostimulatory subpopulation of cells present within the islets of Langerhans without damaging the hormone-secreting cells. Such studies made use of an in vitro correlate of transplantation rejection, the mixed lymphocyte islet cell (MLIC) reaction. Using the MLIC, it was demonstrated that an anti-Ia immunotoxin removed cells capable of stimulating the MLIC in a dose-dependent fashion without altering the hormone-secreting functions of the remaining cells when challenged with glucose and theophylline. These studies suggest the feasibility of using such anti-Ia immunotoxins in islet allograft transplantation models to circumvent problems inherent in complement-mediated cytotoxicity, a previously documented effective form of inducing islet allotransplantation tolerance.
Animals↗
The medical recipes in MS Royal 5 E.vi.
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History of Medicine↗
A novel approach to the isolation of rabbit reticulocyte haem-controlled eIF-2 alpha protein kinase.
A new strategy for the purification of rabbit reticulocyte haem-controlled eIF-2 alpha kinase is described, based on the fact that this kinase can be self-phosphorylated in several sites. Incubation of partially purified kinase with ATP changes its behaviour on anion exchangers sufficiently to separate it from almost all contaminating proteins.
Animals↗
Identification of the stable free radical tyrosine residue in ribonucleotide reductase. A sequence comparison.
The small subunit of ribonucleoside diphosphate reductase contains a unique tyrosine radical and a binuclear iron center. An alignment of different primary structures of the small subunit in Escherichia coli, the marine mollusc Spisula solidissima, Epstein Barr and Herpes simplex viruses shows that regions comprising residues 115-122, 204-212 and 234-241 (in E.coli numbering) are strikingly similar and are likely to be recognized as functionally important. Two of 16 tyrosine residues and 2 of 8 histidine residues are conserved. We propose that Tyr-122 is responsible for radical stabilization and that His-118 and His-241 together with Glu-115 and Asp-237 or Glu-238 are ligands of the iron center.
Animals↗
Double-stranded DNA induces the phosphorylation of several proteins including the 90 000 mol. wt. heat-shock protein in animal cell extracts.
Double-stranded DNA (dsDNA) induces the transfer of phosphate from ATP to several proteins in extracts of widely divergent eukaryotic cells. Extracts of HeLa cells, rabbit reticulocytes, Xenopus eggs and Arbacia eggs all show dsDNA-dependent protein phosphorylation. The mechanism is specific for dsDNA and will not respond to either RNA or single-stranded DNA. One of the proteins which is phosphorylated in response to dsDNA has a subunit mol. wt. of 90 000 and has been identified as a heat-shock protein (hsp90). Although mouse cell extracts were shown to contain hsp90, they failed to show a dsDNA-dependent protein phosphorylation. The observation that dsDNA can modulate the phosphorylation of a set of proteins raises the possibility that dsDNA may play a role as a cellular regulatory signal.
Animals↗
Involvement of dopamine in the aversive stimulus properties of cocaine in rats.
Previous studies of cocaine self-administration have demonstrated central dopaminergic involvement in cocaine's positive reinforcing properties. The present study reports the ability of pimozide, a dopamine receptor antagonist, to attenuate a conditioned taste aversion induced by repeated injections of cocaine. Rats placed on a daily water deprivation schedule were subsequently presented with a novel saccharin taste in their drinking fluid immediately followed by administration of four 9 mg/kg injections of cocaine spaced at 20 min intervals. These animals exhibited a reduction in saccharin intake on subsequent presentations. Animals pretreated with pimozide 90 min prior to the saccharin-cocaine pairings failed to show this reduction. In a second experiment using an identical procedure, repeated injections of lithium chloride were shown to induce a CTA both in pimozide-pretreated and control animals. The results of these two experiments are consistent with the notion that a functional relationship may exist between neurochemical mechanisms underlying both the aversive (CTA-inducing) and positive reinforcing properties of self-administered drugs such as cocaine.
Animals↗
An examination of heroin conditioning in preferred and nonpreferred environments and in differentially housed mature and immature rats.
The study addressed two issues. First, we examined the effectiveness of heroin as a conditioning agent in a preferred environment using a place preference paradigm. Four daily injections of 80 micrograms/kg (SC) of heroin HCl were paired with environments that rats initially found to be either preferred or non-preferred. In subsequent tests, only those that had experienced the drug effects in the non-preferred environment increased the percentage of time spent in that environment. Rats conditioned in the test chamber that was initially preferred failed to increase the amount of time spent in that chamber post-conditioning. These results suggest that the conditioned place preference paradigm does not simply assess the rewarding consequence of heroin injections. We also examined the effects of grouped and isolation housing conditions on the heroin-produced conditioned place preference. Rats were housed under these conditions either immediately post weaning or at 120 days of age. There was a difference between the magnitude of the place preference produced by 20 micrograms/kg heroin in the isolated but not in the group housed rats. When isolated at weaning the rats were less sensitive to the drug than were rats isolated at maturity. These data are discussed with particular reference to the development of the endogenous opioid system.
Aging↗
Aversive stimulus properties of morphine: evaluation using the drug preexposure conditioned taste aversion paradigm.
Interpretation of the finding that positive-reinforcing drugs such as morphine also possess possible aversive properties, as revealed by their ability to induce a conditioned taste aversion (CTA), remains problematic. This issue was addressed in the present study using the drug preexposure CTA paradigm. Water-deprived rats were given noncontingent preexposure to one of three doses of morphine (2.5, 5.0, or 15.0 mg/kg) or drug vehicle. Subsequently, animals in each of these preexposure groups were presented with a novel 0.1% saccharin-flavored solution followed immediately by injection with one of the same three morphine doses or drug vehicle. This procedure was repeated at 5-day intervals until six saccharin presentations had been performed. Results indicated that while the three morphine doses were differentially potent as taste aversion-conditioning agents, they were equipotent as preexposure agents serving to disrupt CTA. These data suggest that preexposure to morphine's predominantly positive-reinforcing (and non-CTA-inducing) properties is sufficient for preexposure disruption of subsequent morphine CTA. A second experiment indicated that the minimal effective preexposure dose is between 0.3 and 1.25 mg/kg of morphine. It is suggested that an important commonality may exist between the discriminative stimulus properties of morphine as a CTA-inducing agent and as a positive reinforcer.
Animals↗
Further evaluation of morphine aversion: maintenance of a taste aversion using a low, nonaversive morphine dose.
Previously, in an investigation of morphine-conditioned taste aversion (CTA), we found that limited preexposure to a low, nonaversive (non-CTA-inducing) dose of morphine (2.5 mg/kg) was as effective as preexposure to a higher, CTA-inducing dose (15 mg/kg) in blocking the formation of a subsequent morphine CTA. In the present study, we examined the capacity of this low, 2.5-mg/kg morphine dose to maintain a CTA initially induced by the 15-mg/kg dose. A standard CTA procedure was used. Results indicated that rats given three initial taste-drug pairings with 15 mg/kg morphine followed on subsequent pairing days by treatment with the low, non-CTA-inducing, 2.5-mg/kg dose continued to exhibit a strong CTA over 8 pairing days. A similar pattern was observed for animals continuing to receive taste-drug pairings with the 15-mg/kg dose. Animals receiving only one taste-drug pairing with the 15-mg/kg dose, followed on subsequent pairing days by 2.5-mg/kg conditioning, failed to show such a pattern of CTA. An intermediate CTA pattern was seen with animals conditioned with 15, 10, 5, and repeated 2.5-mg/kg doses over consecutive pairing days. These data suggest that exposure to a low dose of morphine, with no apparent CTA-inducing properties, is sufficient to maintain a previously established morphine taste aversion. Potential implications for understanding the apparent discriminative complexity of morphine's motivational properties are discussed.
Animals↗
The small subunit of ribonucleotide reductase is encoded by one of the most abundant translationally regulated maternal RNAs in clam and sea urchin eggs.
In both clam oocytes and sea urchin eggs, fertilization triggers the synthesis of a set of proteins specified by stored maternal mRNAs. One of the most abundant of these (p41) has a molecular weight of 41,000. This paper describes the identification of p41 as the small subunit of ribonucleotide reductase, the enzyme that provides the precursors necessary for DNA synthesis. This identification is based mainly on the amino acid sequence deduced from cDNA clones corresponding to p41, which shows homology with a gene in Herpes Simplex virus that is thought to encode the small subunit of viral ribonucleotide reductase. Comparison with the B2 (small) subunit of Escherichia coli ribonucleotide reductase also shows striking homology in certain conserved regions of the molecule. However, our attention was originally drawn to protein p41 because it was specifically retained by an affinity column bearing the monoclonal antibody YL 1/2, which reacts with alpha-tubulin (Kilmartin, J. V., B. Wright, and C. Milstein, 1982, J. Cell Biol., 93:576-582). The finding that this antibody inhibits the activity of sea urchin embryo ribonucleotide reductase confirmed the identity of p41 as the small subunit. The unexpected binding of the small subunit of ribonucleotide reductase can be accounted for by its carboxy-terminal sequence, which matches the specificity requirements of YL 1/2 as determined by Wehland et al. (Wehland, J., H. C. Schroeder, and K. Weber, 1984, EMBO [Eur. Mol. Biol. Organ.] J., 3:1295-1300). Unlike the small subunit, there is no sign of synthesis of a corresponding large subunit of ribonucleotide reductase after fertilization. Since most enzymes of this type require two subunits for activity, we suspect that the unfertilized oocytes contain a stockpile of large subunits ready for combination with newly made small subunits. Thus, synthesis of the small subunit of ribonucleotide reductase represents a very clear example of the developmental regulation of enzyme activity by control of gene expression at the level of translation.
Amino Acid Sequence↗
Conditioned place preference: an evaluation of morphine's positive reinforcing properties.
The place-preference paradigm was evaluated as a measure of morphine's positive reinforcing properties. Previous place-preference studies obtained a morphine place preference of 26%-63%. In order to examine whether differences in procedure may account for this scatter, the present experiment investigated whether there is any difference in the absolute magnitude of preference when animals are conditioned on their non-preferred side of the box or when animals are randomly assigned to the side of conditioning. Furthermore, the number of conditioning days was extended with 3 intervening test days, and drug doses were doubled following each test day. The results showed no significant difference between conditioning animals on their non-preferred side or randomly assigning them to the side of conditioning. However, by extending the number of conditioning days, as well as by following the drug regimen used, the animals showed a greater magnitude of preference than that observed in previous studies. The implications of these findings for the usage of this paradigm as a measure of morphine's positive reinforcing properties are discussed.
Animals↗
Studies of the kinetics and ionic requirements for the phosphorylation of ribosomal protein S6 after fertilization of Arbacia punctulata eggs.
Fertilization of the eggs of the sea urchin Arbacia punctulata is followed by the phosphorylation of ribosomal protein S6. The increase in phosphorylation starts at the same time that protein synthesis begins to increase, and leads to the appearance of mono-, di-, and triphosphorylated S6 derivatives. Essentially all the S6 is phosphorylated by first cleavage. This phosphorylation requires the occurrence of both the normal Ca2+ transient and the consequent Na+-H+ exchange. Protein synthesis can be partially activated by an increase in intracellular pH brought about by weak bases, but this neither causes S6 phosphorylation, nor the inactivation of the specific S6 phosphatase present in unfertilized Arbacia eggs.
Animals↗
Phosphorylation and the control of protein synthesis.
This paper reviews the evidence that protein synthesis in rabbit reticulocytes is regulated by the reversible phosphorylation of the initiation factor eIF-2 by protein kinases under the control of the cytoplasmic haemin concentration on the one hand, and double-stranded RNA on the other. A molecular mechanism is proposed to account for the observation that inhibition of protein synthesis occurs when considerably less than half the eIF-2 present has been phosphorylated. The question of whether phosphorylation regulates protein synthesis in other types of cell is discussed.
Animals↗