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A novel glutamate agonist, TAN-950 A, isolated from streptomycetes
471
low incidence of side effects. Gn the other hand, there have been several reports that normal therapeutic doses of ofloxacin can result in central nervous system symptoms. primarily insomnia and convulsions, in some patients (Iiingsl and Mohr, 1985). Oxolinic acid, another quinolone derivative, which has psychostimulant property, was previously observed to induce insomnia in animal experiments. However there are lack of sufficient ofloxacin induced insomnia. Thus the effects of ofloxacin on some sleep parameters were recorded duced sleep in mice. Inbred male albino mice, weighing 20-25 g, were arranged in 3 groups. At zero time, the members of the ofloxacin groups were pretreated by 20 mg or 40 mg of ofloxacin per kg body-weight (Iglesias and Peres, 1986). Control mice received saline only. Fifteen minutes after the first injections each mice in the control and ofloxacin groups were given 35 mg/kg sodium pentobarbital. Injections were done by the intraperitoneal route (0.1 ml/25 g) and the ex t was carried out between 10.00 a.m. and 15.00 p.m. The righting reflex was used to. assess whether or not the animals were “asleep”. Thus following the pentobarbital injection, the time passed for the mouse to be placed on its back and the time period between loss and regaining of the righting reflex were recorded as the sleep latency and sleeping time respectively. Present results showed that ofloxacin had no apparent effect on the sleep latency. However both 20 mg/kg and 40 mg/kg ofloxacin treatments shortened sleeping time. This effect was highly significant (P c 0.02) in the 40 mg/kg ofloxacin treated group and was contrary to the suggestion that the incidence of side effects has lack of correlation with dosage. Recent results obtained in our laboratory also showed that ofloxacin has proconvulsan effect. Ofloxacin potentiates pentetrazol convulsions in mice (unpublished data). Additionally the effects of ofloxacin on natural sleep in mice have being investigated. Although neurochemical data are inadequate, a possible psychostimulant effect due to an interaction between ofloxacin and inhibitory neurotransmission mediated by y-aminobutyric acid (GABA) (Tsuji et al., 1988) may be involved in shortening of sleeping time in pentobarbital-induced sleep. References t&&s, M.C.G. and E.J. Peres, 1986. Infection 14 (Suppl. 4). 266. Jiingst, G. and M. Riidiger, 1987. Drugs 34 (Suppl. 1). 144. Tsuji, A., H. Sato, Y. Kume. I. Tamai. E. Okezaki. 0. Nogata and H. Kate, 1988. Antimicrob. Agents Chemother. 32. 190.
ycetes
A novel glutamate agonist, T Iwama
*, T., Nagai
*, Y., Harada
* *, S., Itoh * * *, K. and Nagaoka
*, A.
* Biology Res. Lab., * * Microbiology Res. Lab. and * ’ * Chemistry Res. Lub.. Takeda Chemical Industries Lta!. Jusohonmachi. Yodogawa-ku. Osaka 532, Japan
Excitatory amino acids (EAA), especially glutamate, are regarded as the main neurotransmitters in the vertebrate central nervous system (CNS). Recently many reports have suggested that they play a key role in memory, learning and even neuronal damage. There are at least three subtypes of EM receptors, that is, NMDA. quisqualate (Q) and kainate (K) receptors, and each has a unique pharmacological profile. Selective agonists and/or antagonists are needed to investigate the physiological function of the EAA receptors and to improve their function. TAN-950 A, (S)-2-amino-3-(2,5-dihydro-5-oxo-4-isoxac acid, is a novel amino acid isolated as a microbial product. This compound has high affinity for three EAA receptors in the binding assay with [3H]-CPP, [‘HI-AMPA and [3H]-kainate. It effectively elicited the firing of the rat hippocampal CA1 neurons in vitro, and this action was antagonized by a selective Q/K antagonist, DNQX. We investigated the structure-activity relationship of TAN-950 A derivatives for EM receptor subtypes. Alkylation, especially methylation at the 3-position of the isoxazole ring (Me-TAN-950 A), slightly reduced the affinity for the three EAA receptors but increased the potency to elicite the neuronal firing. This agonistic effect was antagonized by DNQX but not by selective NMDA antagonists. CPP or AP7. On the other hand, the (R)-enantiomer of TAN-950 A had increased selectivity for the NMDA receptor subtype. The NMDA selectivity was further enhanced by the removal of the methylene group in the amino moiety.
DA ago&tic activity was observed with (R)-MeNor-TAN-950 A (20% enantiomer excess). of TAN-950 A will be good tools for ~;nvestigatingthe ph~a~lo~~ and physiology roles of
urm&ity
relations of TAN-950 A derivatives on EAA receptors. Binding (IC+& CM)
(S)-slutmte TAN-950 A
[31i)-AMPA
f3R)-K&ate
(%q-CPP
Ekcitation of CA1 neuron MEC * (CM)
0.24 0.28 0.30 >I00
0.11 3.6 17 >I00
0.98 19 40 8.3
300 300 1 10
nists Smith *, T.W., Siionds * &i&d8TW
ate rece
*, MA., Blagbrough, IS., Bycroft, B.W., Mather, A.J., Millington, S. and Usherwood * *. P.N.R. Sciences Gmp, IO1Newi~gton Ccnwwny, Lonrion SE1 6BU. Dept. of P~a~~ic~~
and * * Dept. of Zariogy, Universiq of Nottingham University Park, Nottingham NG7 2BD and * Dept. of Phanrurndogv, Schoolof Plronnacy. 29/39 Brunswick Square, London WClN IAX, U.K.
The @mmacological demnstrat& ~~~0~~
investigation of acylated polyamine action on a vertebrate cortical wedge preparation has of L-glutamate receptors at ~crornol~ ~n~nt~tions. Substituted ~~y~n~ (e.g. ~n~~lat~ spermine analogues) were tested initially on the metatboracic retractor unguis nerve-muscle preparations of the locust S&smerca gregcuia. Quis-GluR antagonism was inferred from the depression of the twitch contraction evoked by stimulation (at 0.22 Hz or at 0.6 Hz) of the excitatory motoneurones which innervate the retractor unguis. In some experiments the postjunctional quis-GluR of this muscle were excited by bath application of L-glutamate; in others the receptors channel was opened by endogenous agonist. on ~~~ excitatory amino acid receptor systems were performed with the rat cortical wedge preparation (Harrison and Simmonds, 1985). Control responses to N-methyl-D-aspartate (NMDA) and (DL)-aamino-3-hydroxy-5-methylisoxaxole4propionic acid (AMPA) (2.5 and 5.0 CM) were obtained in Krebs bicarbonate buffer (magnesium free) gassed with oxygen/carbon dioxide (95 : 5). Tetrodotoxin (100 nM) was included to supress epileptiform discharges. Subsequent incubation of the tissue with philantbotoxin-343 (10 PM) (Blagbrough et al., 1989) for 20 mm, followed by addition of AMPA or NMDA, resulted in no detectable ~~go~srn of the amino acid response. However, upon washing out the toxin and repeating the additions of AMPA or NMDA, the responses progressively declined to about 20% and 40% of control, respectively, by the third application. A similar, but less marked effect was seen with N-(4-hydroxyphenylpropanoyl~spermine which significantly antagonised AMPA responses by 23.13, and 19% following application at 1, 10, and 100 FM. The analogue lacking one methylene group, N~~hy~ox~h~yla~tyl~s~~e, was essentially inactive against AMPA, at these concentrations. At low conlotions (1 and 10 FM) neither synthetic toxin had any effect on responses to NMDA. However, silent ~~o~srn was about in the presence of the toxins at higher (100 PM) concentrations, reduction of the responses by 55 and 92% with the phenylpropanoyl and phenylacetyl analogues respectively. After washout of the antagonists there no recovery with two subsequent additions of NNDA. The antagonism by the phenylacetyl analogue of the DA response was not prevented by the prior addition of D-serine (1 mM). Conclusions: The synthetic toxin philanthotoxin-343 and the simpler analogue N~~hydrox~henylprop~oyl~-
low incidence of side effects. Gn the other hand, there have been several reports that normal therapeutic doses of ofloxacin can result in central nervous system symptoms. primarily insomnia and convulsions, in some patients (Iiingsl and Mohr, 1985). Oxolinic acid, another quinolone derivative, which has psychostimulant property, was previously observed to induce insomnia in animal experiments. However there are lack of sufficient ofloxacin induced insomnia. Thus the effects of ofloxacin on some sleep parameters were recorded duced sleep in mice. Inbred male albino mice, weighing 20-25 g, were arranged in 3 groups. At zero time, the members of the ofloxacin groups were pretreated by 20 mg or 40 mg of ofloxacin per kg body-weight (Iglesias and Peres, 1986). Control mice received saline only. Fifteen minutes after the first injections each mice in the control and ofloxacin groups were given 35 mg/kg sodium pentobarbital. Injections were done by the intraperitoneal route (0.1 ml/25 g) and the ex t was carried out between 10.00 a.m. and 15.00 p.m. The righting reflex was used to. assess whether or not the animals were “asleep”. Thus following the pentobarbital injection, the time passed for the mouse to be placed on its back and the time period between loss and regaining of the righting reflex were recorded as the sleep latency and sleeping time respectively. Present results showed that ofloxacin had no apparent effect on the sleep latency. However both 20 mg/kg and 40 mg/kg ofloxacin treatments shortened sleeping time. This effect was highly significant (P c 0.02) in the 40 mg/kg ofloxacin treated group and was contrary to the suggestion that the incidence of side effects has lack of correlation with dosage. Recent results obtained in our laboratory also showed that ofloxacin has proconvulsan effect. Ofloxacin potentiates pentetrazol convulsions in mice (unpublished data). Additionally the effects of ofloxacin on natural sleep in mice have being investigated. Although neurochemical data are inadequate, a possible psychostimulant effect due to an interaction between ofloxacin and inhibitory neurotransmission mediated by y-aminobutyric acid (GABA) (Tsuji et al., 1988) may be involved in shortening of sleeping time in pentobarbital-induced sleep. References t&&s, M.C.G. and E.J. Peres, 1986. Infection 14 (Suppl. 4). 266. Jiingst, G. and M. Riidiger, 1987. Drugs 34 (Suppl. 1). 144. Tsuji, A., H. Sato, Y. Kume. I. Tamai. E. Okezaki. 0. Nogata and H. Kate, 1988. Antimicrob. Agents Chemother. 32. 190.
ycetes
A novel glutamate agonist, T Iwama
*, T., Nagai
*, Y., Harada
* *, S., Itoh * * *, K. and Nagaoka
*, A.
* Biology Res. Lab., * * Microbiology Res. Lab. and * ’ * Chemistry Res. Lub.. Takeda Chemical Industries Lta!. Jusohonmachi. Yodogawa-ku. Osaka 532, Japan
Excitatory amino acids (EAA), especially glutamate, are regarded as the main neurotransmitters in the vertebrate central nervous system (CNS). Recently many reports have suggested that they play a key role in memory, learning and even neuronal damage. There are at least three subtypes of EM receptors, that is, NMDA. quisqualate (Q) and kainate (K) receptors, and each has a unique pharmacological profile. Selective agonists and/or antagonists are needed to investigate the physiological function of the EAA receptors and to improve their function. TAN-950 A, (S)-2-amino-3-(2,5-dihydro-5-oxo-4-isoxac acid, is a novel amino acid isolated as a microbial product. This compound has high affinity for three EAA receptors in the binding assay with [3H]-CPP, [‘HI-AMPA and [3H]-kainate. It effectively elicited the firing of the rat hippocampal CA1 neurons in vitro, and this action was antagonized by a selective Q/K antagonist, DNQX. We investigated the structure-activity relationship of TAN-950 A derivatives for EM receptor subtypes. Alkylation, especially methylation at the 3-position of the isoxazole ring (Me-TAN-950 A), slightly reduced the affinity for the three EAA receptors but increased the potency to elicite the neuronal firing. This agonistic effect was antagonized by DNQX but not by selective NMDA antagonists. CPP or AP7. On the other hand, the (R)-enantiomer of TAN-950 A had increased selectivity for the NMDA receptor subtype. The NMDA selectivity was further enhanced by the removal of the methylene group in the amino moiety.
DA ago&tic activity was observed with (R)-MeNor-TAN-950 A (20% enantiomer excess). of TAN-950 A will be good tools for ~;nvestigatingthe ph~a~lo~~ and physiology roles of
urm&ity
relations of TAN-950 A derivatives on EAA receptors. Binding (IC+& CM)
(S)-slutmte TAN-950 A
[31i)-AMPA
f3R)-K&ate
(%q-CPP
Ekcitation of CA1 neuron MEC * (CM)
0.24 0.28 0.30 >I00
0.11 3.6 17 >I00
0.98 19 40 8.3
300 300 1 10
nists Smith *, T.W., Siionds * &i&d8TW
ate rece
*, MA., Blagbrough, IS., Bycroft, B.W., Mather, A.J., Millington, S. and Usherwood * *. P.N.R. Sciences Gmp, IO1Newi~gton Ccnwwny, Lonrion SE1 6BU. Dept. of P~a~~ic~~
and * * Dept. of Zariogy, Universiq of Nottingham University Park, Nottingham NG7 2BD and * Dept. of Phanrurndogv, Schoolof Plronnacy. 29/39 Brunswick Square, London WClN IAX, U.K.
The @mmacological demnstrat& ~~~0~~
investigation of acylated polyamine action on a vertebrate cortical wedge preparation has of L-glutamate receptors at ~crornol~ ~n~nt~tions. Substituted ~~y~n~ (e.g. ~n~~lat~ spermine analogues) were tested initially on the metatboracic retractor unguis nerve-muscle preparations of the locust S&smerca gregcuia. Quis-GluR antagonism was inferred from the depression of the twitch contraction evoked by stimulation (at 0.22 Hz or at 0.6 Hz) of the excitatory motoneurones which innervate the retractor unguis. In some experiments the postjunctional quis-GluR of this muscle were excited by bath application of L-glutamate; in others the receptors channel was opened by endogenous agonist. on ~~~ excitatory amino acid receptor systems were performed with the rat cortical wedge preparation (Harrison and Simmonds, 1985). Control responses to N-methyl-D-aspartate (NMDA) and (DL)-aamino-3-hydroxy-5-methylisoxaxole4propionic acid (AMPA) (2.5 and 5.0 CM) were obtained in Krebs bicarbonate buffer (magnesium free) gassed with oxygen/carbon dioxide (95 : 5). Tetrodotoxin (100 nM) was included to supress epileptiform discharges. Subsequent incubation of the tissue with philantbotoxin-343 (10 PM) (Blagbrough et al., 1989) for 20 mm, followed by addition of AMPA or NMDA, resulted in no detectable ~~go~srn of the amino acid response. However, upon washing out the toxin and repeating the additions of AMPA or NMDA, the responses progressively declined to about 20% and 40% of control, respectively, by the third application. A similar, but less marked effect was seen with N-(4-hydroxyphenylpropanoyl~spermine which significantly antagonised AMPA responses by 23.13, and 19% following application at 1, 10, and 100 FM. The analogue lacking one methylene group, N~~hy~ox~h~yla~tyl~s~~e, was essentially inactive against AMPA, at these concentrations. At low conlotions (1 and 10 FM) neither synthetic toxin had any effect on responses to NMDA. However, silent ~~o~srn was about in the presence of the toxins at higher (100 PM) concentrations, reduction of the responses by 55 and 92% with the phenylpropanoyl and phenylacetyl analogues respectively. After washout of the antagonists there no recovery with two subsequent additions of NNDA. The antagonism by the phenylacetyl analogue of the DA response was not prevented by the prior addition of D-serine (1 mM). Conclusions: The synthetic toxin philanthotoxin-343 and the simpler analogue N~~hydrox~henylprop~oyl~-