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334 G protein-mediated inhibition of phosphoinositide metabolism evoked by metabotropic glutamate receptors in Xenopus oocytes
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333
SPECIES DIFFERENCE OF OPIOID DELTA RECEPTOR-MEDIATED SIGNAL T R A N S D U C T I O N IN SYNAPTIC MEMBRANES. NOBUYUKI FUKUSHIMA, CHIFUMI_ HAYASHI. TAKEAKI MIYAMAE. YOSHIMI MISU A N D H I R O S H I UEDA. Department of P h a r m a c o l o g y . Y o k o h a m a C i t y Universit-v S c h o o l o f M e d i c i n e . Y o k o h a m a 2 3 6 . l a P a n . Opioid delta receptor-mediated effects on G-protein activities in striatal synaptic membranes from rats and guinea pigs were studied. DPDPE/[D-Pen2,D-Pen5]-enkephalin, a selective delta 1 agonist, showed stimulatory effects on low-Km GTPase in rat membranes maximally by 7% of control at 1 laM. DS[ET/[D-Ser2,Thr6]-Leuenkephalin, a selective delta 2 agonist, and DAGO/[D-AIa 2, MePhe 4, GlyS-ol]enkephalin, a selective rt-agonist, stimulated low-Km GTPase activity by 25% and 30% of control at 1 rtM, respectively. In striatal membranes from the guinea pig, both DPDPE and DSLET had no effect on GTPase activity, while DAGO did.
These three
agonist bindings were guanine nucleotide-sensitive and there were no quantitative changes between both preparations. These findings suggest that the domain in the d~lta receptor involved in functional coupling to G-proteins might be different between both preparations.
334
G PROTEIN-MEDIATED INHIBITION OF PHOSPHOINOSITIDE METABOLISM EVOKED BY MEFABOTROPIC GLUTAMATE RECEPTORS IN XENOPUS OOCYTES KOJI NAKAMURA1, TOSHIHIDE NUKADA2, TATSUYAHAGA2 & HIROYUKI SUGIYAMA1,3 tDepartment of ~olecular Biolo~,v, Graduate school of Medical Science. Kvushu.Universitv. Fukuoka 812. Japan. 2Institute of Brain Research. Faculty of Medicine, University of Tokyo. Tokyo 113. Japan. "Department of Biolo~,,: Faculty of Science, Kyushu University Fukuoka 812, Japan. Metabotropic glutamate receptor (mGluR1), when expressed in Xenopus oocytes, activates phospholipase C (PLC) in a G protein-dependent manner. We expressed three different G protein ct subunits together with mGluR1 in oocytes, and examined the effects on the PLC-mediated reaction. The expression of GL~, a bovine version of Gila, potentiated the mGluRl-cvokcd reaction, whereas GLlCt, bovine version of G14ot, storongly suppressed it. The expression of Gsct also suppressed the reaction. We then expressed C_~vf2subunits in addition to the Ge~ subunits. The potentiation by GL2C~or the suppression by GLlO~ was more pronounced in the presence of the Gf~l?2 subunits. In contrast, the suppression by Gso~ was completely reversed by GIMy2. The direct activation of G proteins by the intracellular injection of either fluoride ions or GTP"/S causes similar PLC-mediated reactions. The expression of GL:~Ot, GLlCt or Gset caused potentiation, suppression and no change, respectively, on the fluoride- (or GTPyS-)evoked reactions. ~,~,5concluded that GLZis able to stimulate PLC, whereas there are at least two different mechanisms of inhibition for the PLC-mediated reaction by the G protein ot subunits. One is the competition between the endogenous and exogenous Ga's for the endogenous GfSy to form functional heterotrimers, as exemplified by Gset. The othe mechanism was observed for GL], in which an activated G protein interacts with PLC and thereby inhibits its activity or its activation, at least inXenopus oocytes.
335
CHANGES IN EXPRESSIONS OF INOSITOL 1,4,5-TRISPHOSPHATE RECEPTOR mRNA AND CALBINDIN mRNA IN THE CEREBELLUM OF NEUROLOGICAL MUTANT MICE.
SHIN NAKAGAWA1, .MASAHIKOWATANABE2, TSUKASA KOYAMA1 AND YOSHIRO INOUE2,1Dept. of Psychiatry and Neurology, and 2Dept. of Anatomy, Hokkaido Univ. Sch. of Med. Sapporo 060, Japan. Inositol 1,4,5-trisphosphate receptor (IP3R) and calbindin are known to be specifically expressed in the Purkinje cells within the cerebellum of normal mice. This investigation was to examine the changes in gene expressions of IP3R and calbindin in the cerebellar mutant mice (reeler, weaver, staggerer, pcd) by in situ hybridization with 45-mer antisense oligonucleotide probes. Among the four mutants, marked decreases in expression levels of the IP3R and calbindin mRNAs were observed in the staggerer and pcd mice, whereas the Purkinje cells in the reeler and weaver mice expressed these mRNAs at levels comparable to those in normal mice. In addition, different signal levels of the IP3R mRNA between Purkinje cell subpopulations were remarkable in the staggerer; expressing and non-expressing cells were arranged in several parasagittal bands within the cerebellum. Expression patterns of the calbindin mRNA in the staggerer were almost similar to those of the IP3R mRNA. These findings imply that Purkinje cells in the staggerer would be heterogeneous by the regions of the cerebellum, with regard to regulation of the intracellutar Ca2+ concentration.
333
SPECIES DIFFERENCE OF OPIOID DELTA RECEPTOR-MEDIATED SIGNAL T R A N S D U C T I O N IN SYNAPTIC MEMBRANES. NOBUYUKI FUKUSHIMA, CHIFUMI_ HAYASHI. TAKEAKI MIYAMAE. YOSHIMI MISU A N D H I R O S H I UEDA. Department of P h a r m a c o l o g y . Y o k o h a m a C i t y Universit-v S c h o o l o f M e d i c i n e . Y o k o h a m a 2 3 6 . l a P a n . Opioid delta receptor-mediated effects on G-protein activities in striatal synaptic membranes from rats and guinea pigs were studied. DPDPE/[D-Pen2,D-Pen5]-enkephalin, a selective delta 1 agonist, showed stimulatory effects on low-Km GTPase in rat membranes maximally by 7% of control at 1 laM. DS[ET/[D-Ser2,Thr6]-Leuenkephalin, a selective delta 2 agonist, and DAGO/[D-AIa 2, MePhe 4, GlyS-ol]enkephalin, a selective rt-agonist, stimulated low-Km GTPase activity by 25% and 30% of control at 1 rtM, respectively. In striatal membranes from the guinea pig, both DPDPE and DSLET had no effect on GTPase activity, while DAGO did.
These three
agonist bindings were guanine nucleotide-sensitive and there were no quantitative changes between both preparations. These findings suggest that the domain in the d~lta receptor involved in functional coupling to G-proteins might be different between both preparations.
334
G PROTEIN-MEDIATED INHIBITION OF PHOSPHOINOSITIDE METABOLISM EVOKED BY MEFABOTROPIC GLUTAMATE RECEPTORS IN XENOPUS OOCYTES KOJI NAKAMURA1, TOSHIHIDE NUKADA2, TATSUYAHAGA2 & HIROYUKI SUGIYAMA1,3 tDepartment of ~olecular Biolo~,v, Graduate school of Medical Science. Kvushu.Universitv. Fukuoka 812. Japan. 2Institute of Brain Research. Faculty of Medicine, University of Tokyo. Tokyo 113. Japan. "Department of Biolo~,,: Faculty of Science, Kyushu University Fukuoka 812, Japan. Metabotropic glutamate receptor (mGluR1), when expressed in Xenopus oocytes, activates phospholipase C (PLC) in a G protein-dependent manner. We expressed three different G protein ct subunits together with mGluR1 in oocytes, and examined the effects on the PLC-mediated reaction. The expression of GL~, a bovine version of Gila, potentiated the mGluRl-cvokcd reaction, whereas GLlCt, bovine version of G14ot, storongly suppressed it. The expression of Gsct also suppressed the reaction. We then expressed C_~vf2subunits in addition to the Ge~ subunits. The potentiation by GL2C~or the suppression by GLlO~ was more pronounced in the presence of the Gf~l?2 subunits. In contrast, the suppression by Gso~ was completely reversed by GIMy2. The direct activation of G proteins by the intracellular injection of either fluoride ions or GTP"/S causes similar PLC-mediated reactions. The expression of GL:~Ot, GLlCt or Gset caused potentiation, suppression and no change, respectively, on the fluoride- (or GTPyS-)evoked reactions. ~,~,5concluded that GLZis able to stimulate PLC, whereas there are at least two different mechanisms of inhibition for the PLC-mediated reaction by the G protein ot subunits. One is the competition between the endogenous and exogenous Ga's for the endogenous GfSy to form functional heterotrimers, as exemplified by Gset. The othe mechanism was observed for GL], in which an activated G protein interacts with PLC and thereby inhibits its activity or its activation, at least inXenopus oocytes.
335
CHANGES IN EXPRESSIONS OF INOSITOL 1,4,5-TRISPHOSPHATE RECEPTOR mRNA AND CALBINDIN mRNA IN THE CEREBELLUM OF NEUROLOGICAL MUTANT MICE.
SHIN NAKAGAWA1, .MASAHIKOWATANABE2, TSUKASA KOYAMA1 AND YOSHIRO INOUE2,1Dept. of Psychiatry and Neurology, and 2Dept. of Anatomy, Hokkaido Univ. Sch. of Med. Sapporo 060, Japan. Inositol 1,4,5-trisphosphate receptor (IP3R) and calbindin are known to be specifically expressed in the Purkinje cells within the cerebellum of normal mice. This investigation was to examine the changes in gene expressions of IP3R and calbindin in the cerebellar mutant mice (reeler, weaver, staggerer, pcd) by in situ hybridization with 45-mer antisense oligonucleotide probes. Among the four mutants, marked decreases in expression levels of the IP3R and calbindin mRNAs were observed in the staggerer and pcd mice, whereas the Purkinje cells in the reeler and weaver mice expressed these mRNAs at levels comparable to those in normal mice. In addition, different signal levels of the IP3R mRNA between Purkinje cell subpopulations were remarkable in the staggerer; expressing and non-expressing cells were arranged in several parasagittal bands within the cerebellum. Expression patterns of the calbindin mRNA in the staggerer were almost similar to those of the IP3R mRNA. These findings imply that Purkinje cells in the staggerer would be heterogeneous by the regions of the cerebellum, with regard to regulation of the intracellutar Ca2+ concentration.