3768
V.Carelli et al./ Bioorg.Med.Chem.Lett.13 (2003) 3765–3769
maximal inhibition=49Æ8%; ꢀlog IC50=5.65Æ0.14),
as previously reported.26 This effect on GABAB recep-
tors subtypes was competitively antagonized by the
central cholinergic neurotransmission through its meta-
bolite 2a.
selective
GABAB
blocker
CGP-52432
Further studies on the pharmacokinetic profile of the
compounds as well as on the paradoxical activity of 2
and 2a are currently in progress.
(pKB=6.56Æ0.10) and not competitively by compound
1 pD02 ¼ 5:59 Æ 0:16. Compounds 2 and 2a were com-
pletely ineffective in this test at concentrations up to
100 mM. GABA dose-dependently contracted the gui-
nea-pig ileal smooth muscle (1–100 mM) (ꢀlog
EC50=5.11Æ0.11), and this GABAA receptor subtype
mediated response was competitively antagonized by
bicuculline (pKB= 5.34Æ0.50). In contrast, compounds
1, 2 and 2a did not affect ileal tone at concentrations up
to 100 mM. Furthermore, compounds 2 and 2a (0.01–
10 mM) were not able to inhibit l-glutamic acid dec-
arboxylase activity in the specific enzymatic test.27
Acknowledgements
This work was supported by 60% funding from
MURST (Italy).
References and Notes
1. Cooper, J. R.; Bloom, F. E.; Roth, R. H. The Biochemical
basis of Neuropharmacology; Oxford University Press: Oxford,
1984; p 133.
2. Frey, H. H.; Pepp, C.; Losher, W. Neuropharmacology
1979, 18, 581.
Discussion
The findings reported in the present paper show that
both the compounds 1 and 2, after peripheral adminis-
tration, entered the brain differently from GABA.28
Indeed, the activities on CNS produced by peripheral
administration of 1 and 2, could occur only if they were
able to cross the blood–brain barrier. Compound 1 was
capable to produce prolongation of the barbiturate
induced hypnosis (Table 1), a typical GABAmimetic
effect, which is indeed associated with an enhancement
in central GABAergic transmission.29
3. Krogsgaard-Larsen, P.; Falch, E.; Larsson, O. M.;
Schousboe, A. Epilepsy Res. 1987, 1, 77.
4. Lewis, P. J.; Richens, A. Brit.J.Clin.Pharmacol.
27 (Suppl.1), 129.
5. Shek, E. Adv.Drug Deliv.Rev. 1994, 14, 227.
1989,
6. Anderson, W. R.; Simpkins, J. W.; Woodard, P. A.; Win-
wood, D.; Stern, W. C.; Bodor, N. Psychopharmacology 1987,
92, 157.
7. Elving, P. J. In Topics in Bioelectrochemistry and Bioener-
getics; Milazzo, G., Ed.; Wiley: New York, 1976; Vol. 1, p 179.
8. Jaegfeldt, H. Bioelectrochem.Bioenerg. 1981, 8, 355.
Prolongation of the duration of hypnosis time was also
exhibited from 1a, the putative oxidative metabolite of 1
(Table 1).
9. Onhishi, Y.; Kitani, M. Bull.Chem.Soc.Jpn.
2674.
10. Ragg, E.; Scaglioni, L.; Mondelli, R.; Carelli, V.; Carelli,
I.; Casini, A.; Finazzi-Agro, A.; Liberatore, F.; Tortorella, S.
Biochem.Biophys.Acta 1991, 1076, 37.
11. Carelli, V.; Liberatore, F.; Casini, A.; Mondelli, R.;
Arnone, A.; Rotilio, G.; Mavelli, I. Biorg.Chem. 1980, 9, 342.
12. Avigliano, L.; Carelli, V.; Casini, A.; Finazzi-Agro, A.;
Liberatore, F. Biochem.J. 1985, 226, 391.
1979, 52,
In this regard, should be noted that 1a, the correspond-
ing brain oxidative metabolite of the dihy-
dronicotinamide 3, is likely to be responsible for the
anxiolytic effects described by Anderson et al.6
13. Avigliano, L.; Carelli, V.; Casini, A.; Finazzi-Agro, A.;
Liberatore, F.; Rossi, A. Biochem.J. 1986, 237, 919.
14. Carelli, V.; Liberatore, F.; Scipione, L.; Impicciatore, M.;
Barocelli, E.; Cardellini, M.; Giorgioni, G. J.Control.Release.
1996, 42, 209.
The GABA derivative 2 gave rise to a CNS effect
opposite to 1 and 1a, because it produced in rats reduc-
tion of hypnosis time, both by icv or iv administration.
The quaternary salt 2a evoked the same arousing effects
only by icv injection; therefore, it is likely that this
compound, resulting from metabolic oxidation of 2 in
the brain, could be the true active principle. The excita-
tory properties of 2 and 2a were confirmed by the con-
15. Woodard, P. A.; Winwood, D.; Brewster, M. F.; Estes,
K. S.; Bodor, N. Drug Des.Deliv. 1990, 6, 15.
16. Matsujama, K.; Yamashita, C.; Noda, A.; Goto, S.; Noda,
H.; Ichimaru, Y.; Gomita, Y. Chem.Pharm.Bull. 1984, 32, 4089.
17. Compound 1: yield 56%; MS (ESI) m/z 649 (M+Na)+.
1H NMR 500 MHz (CDCl3): 6.86 (1H, d, H2, J2,6 =1.3 Hz);
5.88 (1H, dd, H6, J5,6 7.8); 6.98 (1H. bs, NH); 3.36 (1H, d, H4,
J4,5 =4.7 Hz); 4.57 (1H, dd, H5); 7.34 (5H, s, Ar); 5.11 (2H, s,
(d) O-CH2); 2.39 (3H, s, N–CH3); 3.36 (2H, t, (a) N–CH2, Ja,b
=6.7 Hz); 2.46 (2H, t, (c) CH2–CO, Jb,c =7.4Hz); 1.91 (2H,
m, (b), CH2-C). Anal. calcd for C36 H42 N4 O6: C, 68.99; H,
6.75; N, 8.94. Found C, 68.61; H, 6.99; N, 8.63. Compound 2:
vulsive
effects
displayed
after their
central
administration in awake rats (data not shown). These
GABA opposing responses could be ascribed either to
attenuation of the GABAergic function through the
inhibition of GABA receptors or the reduction of
GABA availability.30 A GABAA or GABAB antagonist
activity of 2 and 2a can be ruled out owing to their total
inactivity in guinea-pig isolated ileum tests earlier
described. On the other hand, interference with GABA
cerebral biosynthesis can be excluded because of the
inability of 2 and 2a in blocking l-glutamic acid
decarboxylase (GAD) activity in the in vitro specific
enzymatic test. Most likely, the inhibition exerted by
atropine on arousal effect of compounds 2 and
2a, allows us to hypothesize that 2 enhances the
1
yield 72%; MS (ESI) m/z 491 (M+H)+. H NMR 500 MHz
(D2O): 6.86 (1H, s, H2); 5.03 (1H, d, H6, J5,6 =8.0 Hz); 3.26
(1H, d, H4, J4,5 =4.3 Hz); 4.51 (1H, dd, H5); 2.88 (3H, s, N–
CH3); 3.35 (2H, t, (a) N–CH2, Ja,b =7.0 Hz); 2.37 (2H, t, (c)
CH2–CO, Jb,c =7.6 Hz); 1.96 (2H, m, (b) CH2–C). Compound
2a: yield 1.2 g (80%), mp 95–96 ꢁC. 1H NMR 500 MHz (D2O):
9.21 (1H, s, H2); 8.95 (1H, d, H6, J5,6 =7.8 Hz); 8.83 (1H, d,
H4, J4,5 =5.4 Hz); 8.17 (1H, dd, H5); 4.47 (3H, s, N–CH3);