Synthesis and radioligand binding studies of C-5- and C-8-substituted 1-(3,4-dimethoxybenzyl)-2,2-dimethyl-1,2,3,4-tetrahydroisoquinoliniums as SK channel blockers related to N-methyl-laudanosine and N-methyl-noscapine

Article abstract of DOI:10.1021/jm049025p

The synthesis and the ¹²⁵I-apamin binding studies of original C-5- and C-8-substituted 1-(3,4-dimethoxy-benzyl)-2,2-dimethyl-1,2,3,4- tetrahydroisoquinoliniums and 1-(3,4-dimethoxy-benzyl)-6,6-dimethyl-4,5,6,7- tetrahydrothieno[2,3-c]pyridiniums were performed in order to find a reversible and selective SK channel blocker structurally related to N-methyl-laudanosine and N-methyl-noscapine. A bulky alkyl substituent in the C-8 position of the tetrahydroisoquinoline produces a clear increase in the affinity for the apamin sensitive binding sites. The presence of an electron-withdrawing group in the C-5 and C-8 positions is not a suitable substitution for the affinity of drugs structurally related to N-methyl-laudanosine. Thiophenic analogues and 8-methoxy derivatives possess a poor affinity for the apamin sensitive binding sites. Electrophysiological studies performed with the most effective compound showed a blockade of the apamin sensitive afterhyperpolarization in rat dopaminergic neurons.

Full text of DOI:10.1021/jm049025p

4972
J. Med. Chem. 2005, 48, 4972-4982
Synthesis and Radioligand Binding Studies of C-5- and C-8-Substituted
1-(3,4-Dimethoxybenzyl)-2,2-dimethyl-1,2,3,4-tetrahydroisoquinoliniums as SK
Channel Blockers Related to N-Methyl-laudanosine and N-Methyl-noscapine
Amaury Graulich,*,† Jacqueline Scuve´e-Moreau,‡ Vincent Seutin,‡ and Jean-Franc¸ois Lie´geois†,§
Laboratory of Medicinal Chemistry, Natural and Synthetic Drugs Research Center, University of Lie`ge, avenue de l’Hoˆpital,
1 (B36), Laboratory of Pharmacology, Research Center for Cellular and Molecular Neurobiology, University of Lie`ge, avenue de
l’Hoˆpital, 3 (B23), and Laboratory of Physiology, University of Lie`ge, avenue de l’Hoˆpital, 3 (B23), B-4000 Lie`ge 1, Belgium
Received December 1, 2004
The synthesis and the 125I-apamin binding studies of original C-5- and C-8-substituted 1-(3,4-
dimethoxy-benzyl)-2,2-dimethyl-1,2,3,4-tetrahydroisoquinoliniums and 1-(3,4-dimethoxy-ben-
zyl)-6,6-dimethyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridiniums were performed in order to find
a reversible and selective SK channel blocker structurally related to N-methyl-laudanosine
and N-methyl-noscapine. A bulky alkyl substituent in the C-8 position of the tetrahydroiso-
quinoline produces a clear increase in the affinity for the apamin sensitive binding sites. The
presence of an electron-withdrawing group in the C-5 and C-8 positions is not a suitable
substitution for the affinity of drugs structurally related to N-methyl-laudanosine. Thiophenic
analogues and 8-methoxy derivatives possess a poor affinity for the apamin sensitive binding
sites. Electrophysiological studies performed with the most effective compound showed a
blockade of the apamin sensitive afterhyperpolarization in rat dopaminergic neurons.
Introduction
From their biophysical and pharmacological proper-
ties, three families of calcium-activated potassium chan-
nels have been identified. These are called BK, IK, and
SK reflecting their large, intermediate, and low con-
ductances, respectively.1,2
Small conductance Ca2+-activated K+ (SK) channels
Figure 1. Chemical structures of NML and NMN.
underlie the prolonged postspike afterhyperpolarization
(AHP), which plays an important role in modulating the
firing rate and the firing pattern of neurons.3,4 Func-
tional, pharmacological, and structural data have sug-
gested the existence of variants of the SK channel.1
Indeed, three SK channel subtypes have been identified
by DNA cloning, namely, SK1, SK2, and SK3.3 In
contrast to BK channels, SK channels are voltage
insensitive but are activated by an increase in the
intracellular calcium concentration. The distribution of
the SK channel subtypes was investigated in the rat
by using in situ hybridization and immunohistochem-
istry and revealed that SK1 and SK2 subtypes are
mostly expressed in the cortex and hippocampus5 while
SK3 channels expression is higher in subcortical areas,
especially in the monoamine cell regions, e.g., substantia
nigra, dorsal raphe, and locus coeruleus. These features
attract great attention to SK channels as putative tar-
gets for indications in cognitive dysfunction,6-10 neu-
ronal hyperexcitability,11 dopamine-related disorders,12-14
and depression.9
octadecapeptide possesses two arginine residues in
contiguous positions and a rigid cyclic conformation due
to two disulfide bridges.15 Leiurotoxin I isolated from
scorpion Androctonus mauretanicus mauretanicus po-
tently blocks human SK2 and SK3 but not SK1 chan-
nels.16 Dequalinium, a nonpeptidic ligand, presents
some SK channel blocking properties, and extensive
structure-activity relationships studies led to UCL
compounds.17 Furthermore, N-methyl-bicuculline was
reported to potentiate burst firing in dopaminergic
neurons by blocking the apamin sensitive Ca2+-acti-
vated K+ current.18 However, the GABAA antagonist
activity of bicuculline quaternary salts is a serious
drawback, so more selective compounds are needed.
Therefore, we decided to develop new SK blockers
structurally close to N-methyl-bicuculline. First, studies
started with N-methyl-laudanosine (NML)19 and more
recently with N-methyl-noscapine (NMN) (Figure 1).
Unlike apamin, these two molecules possessing medium
potency blocking properties are quickly reversible.20
Different drugs structurally related to these compounds
were synthesized and evaluated by using an in vitro
binding assay in order on the one hand to find drugs
with better affinity and selectivity and on the other
hand to increase our knowledge of the pharmacophore.
So far, the most potent SK channel blockers are
venom toxins such as apamin and leiurotoxin I. Apamin
is extracted from honey bee Apis mellifera venom. This
* To whom correspondence should be addressed. Tel: ++32(0)43 66
43 77. Fax: ++32(0)43 66 43 62. E-mail: A.Graulich@student.ulg.ac.be.
† Laboratory of Medicinal Chemistry.
‡ Laboratory of Pharmacology.
§ Laboratory of Physiology.
10.1021/jm049025p CCC: $30.25 © 2005 American Chemical Society
Published on Web 07/01/2005

N-Methyl-laudanosine and N-Methyl-noscapine
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 15 4973
Scheme 1. Synthesis of 1 and 8-Halogenated
Scheme 3. Synthesis of 8-Substituted Isoquinolinesa by
the Hendrickson Modification of the Pomeranz-Fritsch
Procedureb
Isoquinolinesa
a Key: (i) NBS, H2SO4, -23 to -25 °C. (ii) KNO3, H2SO4, room
temperature. (iii) H2, 10 bar, Pd/CaCO3, MeCOONH4, MeOH, room
temperature. (iv) NaNO2, HX, 0-5 °C. (v) CuX, HX.
a Thieno[2,3-c]pyridines 9e,f were synthesized following the
same chemical pathway. bKey: (i) NH2CH2CH(OMe)2, ArMe,
Dean-Stark trap, reflux. (ii) ClCOOEt, THF, -10 °C. (iii) P(OMe)3,
THF, room temperature. (iv) TiCl4, CH2Cl2, room temperature.
Scheme 2. Preparation of 7a
Scheme 4. Synthesis of 8a
a Key: (i) H2, 10 bar, Pd/C, MeOH, room temperature. (ii)
NaNO2, HCl, 0-5 °C. (iii) CuCl, HCl.
Chemistry
a Key: (i) Mg, Et2O, room temperature. (ii) PhNdCHOEt, Et2O,
Substituted isoquinolines were first prepared. 5-Bro-
moisoquinoline (1) was obtained by bromination using
NBS in concentrated H2SO4 (Scheme 1). The selectivity
of this reaction was highly dependent on the reaction
temperature, which should be kept between -25 and
-22 °C.21 8-Aminoisoquinoline (3) was obtained by
selective nitration in the C-8 position of 1 to afford
5-bromo-8-nitroisoquinoline (2) (Scheme 1).22 Then,
catalytic reduction of 2 with palladized calcium carbon-
ate in MeOH, in the presence of ammonium acetate,
gave only a moderate yield of 3.22 5-Aminoisoquinoline
(6) was obtained by catalytic hydrogenation of 5-ni-
troisoquinoline with palladized charcoal in MeOH
(Scheme 2). Diazotization of 3 and 6 was then achieved
to afford 8-bromoisoquinoline (4), 8-chloroisoquinoline
(5), and 5-chloroisoquinoline (7) by using the Sandmeyer
reaction (Schemes 1 and 2).
The syntheses of 8-alkylisoquinoline (9a-c), 8-meth-
oxyisoquinoline (9d),23 and thieno[2,3-c]pyridine ana-
logues (9e,f)24 were carried out by using a modified
procedure of the Pomeranz-Fritsch synthesis. An aryl-
aldehyde was condensed with aminoacetaldehyde di-
methyl acetal to afford a Schiff base. The corresponding
imine was then protected by reaction with ethyl chlo-
roformate. The resulting acyliminium undergoes a nu-
cleophilic addition by trimethyl phosphite to form a
carbamate-phosphonate intermediate, which was di-
rectly cyclized in the presence of titanium tetrachloride
in refluxing CH2Cl2 (Scheme 3). For the synthesis of
8-isopropyl-isoquinoline (9d), 2-isopropylbenzaldehyde
(8) was prepared by reaction of ethyl N-phenylformimi-
date with Grignard reagent from o-iodocumene and then
hydrolyzed to afford the appropriate benzaldehyde (8)
(Scheme 4).25
room temperature. (iii) H3O+, reflux.
intermediates were usually synthesized from the ap-
propriate nitrogen heterocycles and acyl chlorides in the
presence of a cyanide source.26 So, the isoquinolines
react with benzoyl chloride and the resulting acylimin-
ium undergoes a nucleophilic addition by cyanide to
afford the appropriate Reissert compounds (10a-h).27
This reaction was carried out with trimethylsilyl cya-
nide in anhydrous CH2Cl2 and gave good yields (Scheme
5). The thienopyridines were converted into Reissert
compounds (10i,j) by the same chemical procedure, but
ethyl chloroformate was used as an acylating agent
instead of benzoyl chloride (Scheme 6). The yields are
generally superior.28 All Reissert compounds (10a-j)
were then deprotonated by sodium hydride in DMF. The
resulting Reissert anions were alkylated by using 3,4-
dimethoxybenzyl chloride. Then, the alkylated Reissert
compounds were hydrolyzed to 1-(3,4-dimethoxybenzyl)-
isoquinolines (11a-h) and 7-(3,4-dimethoxybenzyl)-
thieno[2,3-c]pyridines (11i,j) (Schemes 5 and 6).
Compounds 11a-j were methylated by methyl iodide
in refluxing MeCN. Because of their chemical lability,
the resulting N-methylisoquinoliniums were directly
reduced to N-methyl-1,2,3,4-tetrahydroisoquinolines
(12a-j) by using an excess of sodium borohydride in
MeOH. A further methylation of compounds 12a-j gave
the quaternary ammoniums 13a-j by using methyl
iodide in refluxing MeCN.
Results
The binding data are summarized in Tables 1 and 2.
First, the compounds were screened at a concentration
of 10 µM. Then, the affinities were determined for the
drugs that displaced at least 20% of the radioligand.
In our conditions, iodinated apamin has a Kd of 2.02
( 0.31 pM and the affinity of apamin is equal to 3.8 pM.
After the synthesis of these isoquinolines (1, 4, 5, 7,
and 9a-d), the alkylation in the C-1 position was
performed by using the Reissert compounds. These

4974 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 15
Graulich et al.
Scheme 5. Synthesis of 1-(3,4-Dimethoxybenzyl)-2,2-dimethyl-1,2,3,4-tetrahydroisoquinoliniums from Isoquinoline
Derivativesa
a Key: (i) BzCl, (Me)3SiCN, CH2Cl2, 30 °C. (ii) NaH, DMF, -10 °C. (iii) 3,4-(MeO)2C6H3CH2Cl, DMF, -10 °C. (iv) 50% NaOH, H2O-
EtOH, reflux. (v) MeI, MeCN, reflux. (vi) NaBH4, MeOH, room temperature. (vii) MeI, MeCN, reflux.
Scheme 6. Synthesis of 7-(3,4-Dimethoxybenzyl)-6,6-dimethyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridiniums from
Thieno[2,3-c]pyridine Analoguesa
a Key: (i) ClCOOEt, (Me)3SiCN, CH2Cl2, 30 °C. (ii) NaH, DMF, -10 °C. (iii) 3,4-(MeO)2C6H3CH2Cl, DMF, -10 °C. (iv) 50% NaOH,
H2O-EtOH, reflux. (v) MeI, MeCN, reflux. (vi) NaBH4, MeOH, room temperature. (vii) MeI, MeCN, reflux.

N-Methyl-laudanosine and N-Methyl-noscapine
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 15 4975
Table 1. Physical Data and Percentage of 125I-Apamin
Displaced of NML, NMN, and Analogues (12a-j and 13a-j) for
Rat Cortical Apamin Sensitive Sites
Figure 2. Effect of increasing concentrations of compound
13c on the AHP of midbrain dopaminergic neurons. Total
blockade of the AHP is obtained with 100 µM compound 13c.
Each trace is the mean of four sweeps. Action potentials are
truncated.
preparation are 5.5, 2.6, and 1.6 µM, respectively, for
i
Me (13a), Et (13b), and Pr (13c) derivatives.
yield
mp
Compound 13d substituted with a methoxy group in
the C-8 position has no significant activity (percentage
of displaced iodinated apamin, 9%) so its affinity was
not determined as mentioned above.
b
compd R1 R2 R3 (%)
(°C)
formula
anal.
%
NML
NMN
216-218 C22H30NO4I
23H26NO7I
C, H, N 57
C, H, N 27
C
12a
12b
12c
12d
12e
12f
H
H
H
H
H
H
Me
Et
98 133-135 C20H26NO2Cl‚H2O C, H, N 10
Brominated analogues in the C-5 (13e) and C-8
positions (13g) possess an affinity for the apamin
sensitive binding sites of 4.9 and 6.8 µM, respectively.
Chlorinated analogues in the C-5 (13f) and C-8 positions
(13h) have an affinity for the apamin sensitive binding
sites of 7.6 and 9.3 µM, respectively. Finally, thiophenic
analogues displace the radioligand by 15 and 18% for
13i and 13j, respectively. In electrophysiological experi-
ments, compound 13c blocks the apamin sensitive AHP
in dopaminergic neurons with an IC50 equal to 22 ( 6
µM (n ) 3) (Figure 2).
97 135-136 C21H28NO2Cl
99 204-205 C22H30NO2Cl
97 148-149 C24H29NO7
C, H, N
C, H, N 10
C, H, N
C, H, N 10
2
iPr
OMe
Br
1
42 166-167 C19H23NO2BrCl
68 158-159 C19H23NO2Cl2
52 187-189 C19H23NO2BrCl
72 198-200 C19H23NO2Cl2
Cl
C, H, N
C, H, N
C, H, N
5
1
8
2
1
12g
12h
12i
Br H
Cl H
H
88
97-99
C17H22NO2SCl‚H2O C, H, N
12j
Me 86 165-169 C18H24NO2SCl‚
C, H, N
1/4H2O
13a
13b
13c
13d
13e
13f
H
H
H
H
H
H
Me
91 208-209 C21H28NO2I
89 190-191 C22H30NO2I
85 238-240 C23H32NO2I
88 194-195 C21H28NO3I
82 216-217 C20H25NO2BrI
77 220-221 C20H25NO2ClI
83 166-167 C20H25NO2BrI
85 136-138 C20H25NO2ClI
C, H, N 24
C, H, N 44
C, H, N 55
Et
iPr
OMe
Br
C, H, N
9
C, H, N 26
C, H, N 23
C, H, N 29
C, H, N 27
C, H, N 15
C, H, N 18
Cl
Discussion
13g
13h
13i
Br H
Cl H
First, it was demonstrated that N-methyl-bicuculline
was a reversible blocker of the apamin sensitive AHP
in dopaminergic neurons with a poor selectivity.29 Then,
NML was prepared and tested for its blockade of the
apamin sensitive AHP. This compound revealed an
interesting binding profile19 and a very quickly revers-
ible effect on the apamin sensitive AHP,20 but after
extensive biological evaluation, a non-SK-mediated ef-
fect on serotoninergic neurons of the dorsal raphe has
been detected with this compound.20 NMN was also
evaluated in in vitro binding and electrophysiological
experiments. It was found that this drug also has a
quickly reversible effect on SK channels but a lower
affinity than NML.20 In a previous study, it was shown
that a 3,4-dimethoxybenzyl substituent in the C-1
position appears more effective than a benzyl group.30
So, different compounds structurally close to the NML
and NMN template were synthesized and evaluated by
radioligand binding studies. In our binding experiments,
iodinated apamin and apamin have an affinity in the
same range than the results previously described.31,32
In the chemistry part, the synthesis of different
isoquinolines has allowed the preparation of several
original Reissert compounds (10 a-f,h,j). The alkylation
of these Reissert compounds also affords original 1-(3,4-
dimethoxybenzyl)isoquinolines (11a-h,j). Finally, al-
most all tested compounds (12a-h,j and 13a-j) are
described for the first time.
H
80 216-217 C18H24NO2SI
13j
Me 83 139-140 C19H26NO2SI
a See also the Experimental Section. b % of 125I-apamin dis-
placed at 10 µM.
Table 2. Binding Affinities of NML, NMN, and Analogues
(13a-j) for Rat Cortical Apamin Sensitive Sites
a
compd
R1
R2
R3
%
IC50 (µM)
8.7 ( 2.3
Ki (µM)
NML
NMN
13a
13b
13c
13d
13e
13f
13g
13h
13i
57
1.6 ( 0.4
3.6 ( 1.3
5.5 ( 0.7
2.6 ( 0.7
1.6 ( 0.5
27 14.1 ( 2.6
24 28.4 ( 3.3
44 13.6 ( 3.9
H
H
H
H
H
H
Br
Cl
Me
Et
iPr
OMe
Br
Cl
55
9
8.6 ( 2.5
26 35.2 ( 7.0
23 50.0 ( 2.1
29 25.4 ( 2.0
27 39.6 ( 7.1
15
6.8 ( 1.2
9.3 ( 0.9
4.9 ( 0.4
7.6 ( 1.2
H
H
H
Me 18
13j
apamin
20.4 ( 5.7 pM 3.8 ( 1.1 pM
a % of 125I-apamin displaced at 10 µM.
Racemic NML and its enantiomers have an affinity
(Ki) for the apamin sensitive sites of ∼1.5 µM.
Compounds possessing an alkyl group in the C-8
position (13a-c) displace 24, 44, and 55% for Me (13a),
Et (13b), and Pr (13c) derivatives, respectively. The
affinities for apamin sensitive sites of the rat cortex
i

4976 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 15
Graulich et al.
In vitro binding results show that the quaternization
of the compound is an important parameter for the
apamin sensitive sites affinity. Actually, all SK channel
blockers have at least one ionized or ionizable function
as found in animal toxins such as apamin, leiurotoxin,
and tityus κ possessing one or two arginine resi-
dues.16,33,34 Synthetic blockers as UCL compounds pos-
sessing two quinolinium nuclei also have this structural
characteristic35 that seems to be indispensable for the
affinity on apamin sensitive binding sites.
Experimental Section
Chemistry. Melting points were determined on a Bu¨chi-
Tottoli capillary melting point apparatus in an open capillary
and are uncorrected. NMR spectra were recorded on a Bruker
Advance 500 spectrometer at 500 MHz. IR spectra were
performed on a Perkin-Elmer FTIR-1750 spectrometer. IR
spectra were measured using KBr disks. Only significant
bands from IR are reported. Elemental analyses were deter-
mined using a Carlo-Erba elemental analyzer CHNS-O model
EA1108, and the results are within 0.4% of the theorical
values. Mass spectra were recorded on a QTOF II (Micromass,
Manchester, United Kingdom) spectrometer with electrospray
mode. All starting materials and reagents were obtained from
Aldrich Chemical Co. and were used without further purifica-
tion. Separations by column chromatography were carried out
using Merck Kieselgel 60 (230-400 mesh). Concentration and
evaporation refer to removal of volatile materials under
reduced pressure (10-15 mmHg at 30-50 °C) on a Buchi
Rotavapor.
For alkylated compounds in the C-8 position, the
results demonstrate clearly that the affinity of the
compound increases with the bulkiness of the substitu-
ent in the C-8 position [5.5, 2.6, and 1.6 µM, respectively,
i
for Me (13a), Et (13b), and Pr (13c) derivatives]. So,
the presence of a bulky group in the C-8 position of the
isoquinoline is important for the affinity of this series
of compounds. Such an increase of affinity is particularly
attractive for further modulations. The 8-isopropyl
derivative (13c) has an affinity equal to the lead
compound, NML, although not possessing a 6,7-dimeth-
oxy group.
8-Aminoisoquinoline (3). Catalytic reduction of 2 with
palladized calcium carbonate in MeOH, in the presence of
ammonium acetate, gave a moderate yield of 3.22 Recrystal-
lization from petroleum ether 100-140 gave 3 as a cream solid;
yield, 53%; mp 167-168 °C. IR (KBr): 3147, 1570, 824 cm-1
.
1H NMR (CDCl3): δ 4.36 (s, 2H), 6.79 (d, 1H, J ) 8.0 Hz),
7.21 (d, 1H, J ) 8.0 Hz), 7.45 (t, 1H, J ) 8.0 Hz), 7.54 (d, 1H,
J ) 5.7 Hz), 8.45 (d, 1H, J ) 5.7 Hz), 9.29 (s, 1H). Anal.
(C9H8N2) C, H, N.
NMN, a quickly reversible ligand possessing a slightly
lower affinity than NML, has a methoxy group in the
C-8 position, but this is not always a favorable substitu-
tion as the results show that the 8-methoxy derivative
(13d), which has a very low affinity for the apamin
sensitive sites. So, the presence of an electronegative
atom in this part of the molecule was not favorable as
found with the halogenated analogues (13e-h).
The presence of a halogen in the C-5 and C-8 positions
(13e-h) is not favorable for the affinity on apamin
sensitive binding sites. However, results show that
compounds halogenated in the C-5 position (13g,h) have
a higher affinity than the corresponding compound
halogenated in the C-8 position (13e,f). In this halogen-
ated series, brominated analogues (13e,g) have a slightly
better affinity for apamin sensitive binding sites than
the chlorinated analogues (13f,h). So, the presence of
an electronegative atom is mainly unfavorable in the
C-8 position of the molecule.
Thiophenic analogues (13i,j) have a very low affinity,
and this bioisosteric modulation appears to be inef-
ficient, but chemical development is needed in order to
obtain more precision about the impact of bioisosteric
replacements of 1,2,3,4-tetrahydroisoquinoline on the
affinity of these molecules for the apamin sensitive
binding sites.
To demonstrate the blocking potential of these drugs
on SK channels, the most effective compound of this
series, compound 13c, was evaluated by electrophysi-
ological experiments. This compound blocked the apam-
in sensitive AHP in dopaminergic neurons with an IC50
of 22 ( 6 µM (n ) 3), which is quite equivalent to that
of NML (15 ( 1 µM)19 in similar conditions.
8-Bromoisoquinoline (4). Compound 3 was diazotized and
then treated with a solution of freshly prepared cuprous
bromide in excess of hydrobromic acid to afford 4.22 Recrys-
tallization from n-hexane gave 4 as a white solid; yield, 45%;
mp 75-77 °C. IR (KBr): 1545, 830 cm-1. 1H NMR (CDCl3): δ
7.51 (t, 1H, J ) 7.9 Hz), 7.60 (d, 1H, J ) 5.7 Hz), 7.77 (d, 1H,
J ) 7.9 Hz), 7.83 (d, 1H, J ) 7.9 Hz), 8.60 (d, 1H, J ) 5.7 Hz),
9.61 (s, 1H). Anal. (C9H6NBr) C, H, N.
8-Chloroisoquinoline (5). Compound 3 was diazotized and
then treated with a solution of cuprous chloride in excess of
hydrochloric acid to afford 5.22 Recrystallization from n-hexane
gave 5 as a white solid; yield, 49%; mp 53-54 °C. IR (KBr):
1
1552, 833 cm-1. H NMR (CDCl3): δ 7.58 (t, 1H, J ) 7.6 Hz),
7.61-7.64 (t, 2H), 7.72 (d, 1H, J ) 7.6 Hz), 8.60 (d, 1H, J )
5.7 Hz), 9.66 (s, 1H). Anal. (C9H6NCl) C, H, N.
5-Aminoisoquinoline (6). Catalytic reduction of 5-ni-
troisoquinoline with palladized charcoal in MeOH gave 6.22
Recrystallization from n-hexane gave 6 as a white solid; yield,
83%; mp 126-127 °C. IR (KBr): 3173, 1582, 799 cm-1. 1H NMR
(CDCl3): δ 4.19 (s, 2H), 6.93 (t, 1H, J ) 4.6 Hz), 7.37-7.40 (d,
2H), 7.55 (d, 1H, J ) 5.9 Hz), 8.47 (d, 1H, J ) 5.9 Hz), 9.16 (s,
1H). Anal. (C9H8N2) C, H, N.
5-Chloroisoquinoline (7). Compound 6 was diazotized and
then treated with a solution of cuprous chloride in excess of
hydrochloric acid to afford 7.22 Recrystallization from n-hexane
gave 7 as a white solid; yield, 53%; mp 69-71 °C. IR (KBr):
1
1580, 824 cm-1. H NMR (CDCl3): δ 7.52 (t, 1H, J ) 7.8 Hz),
7.76 (d, 1H, J ) 7.8 Hz), 7.89 (d, 1H, J ) 7.8 Hz), 8.00 (d, 1H,
J ) 5.9 Hz), 8.63 (d, 1H, J ) 5.9 Hz), 9.25 (s, 1H). Anal. (C9H6-
NCl) C, H, N.
2-Isopropylbenzaldehyde (8). The reaction of ethyl N-
phenylformimidate with Grignard reagent from o-iodoc-
umene25 affords after purification a colorless oil. This crude
oil is used without further purification for the preparation of
8-isopropylisoquinoline; yield, 52%. IR (C2Cl4): 2968, 1706
cm-1
.
Permanent ionization gives to these quaternary am-
moniums an interesting solubility in aqueous media for
the in vitro test. Nevertheless, this characteristic is a
major drawback for the in vivo experiments. In fact, the
interest for targeting SK channels is mostly turned
toward the central nervous system. So, a nonpeptidic
and nonquaternized compound would be of great inter-
est for the progress of in vivo experiments in the field
of these ion channels.
8-Methylisoquinoline (9a). A solution of o-tolualdehyde
(5.3 g; 44.1 mmol) and aminoacetaldehyde dimethyl acetal (4.8
mL; 44.1 mmol) in toluene (50 mL) was refluxed for 3 h using
a Dean-Stark trap. After the solvent was removed, the oil was
dissolved in dry THF (30 mL) and ethyl chloroformate (4.2 mL;
44.1 mmol) was added dropwise at -10 °C. After 5 min under
stirring, the cooling bath was removed and trimethyl phosphite
(6.7 mL; 56.3 mmol) was added at room temperature. The
solution was evaporated under reduced pressure after 20 h.
To remove traces of trimethyl phosphite, toluene was added

N-Methyl-laudanosine and N-Methyl-noscapine
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 15 4977
and evaporated twice. The resulting oil was dissolved in dry
CH2Cl2 (50 mL), and titanium tetrachloride (30 mL; 264 mmol)
was added. The mixture was heated under reflux in a dry
atmosphere for 24 h. The reaction medium was poured in a
mixture of ice (200 g) and NH4OH (100 mL). The suspension
was filtered, and the TiO2 precipitate was rinsed with CHCl3
(3 × 50 mL). The organic layers were collected and extracted
with 1 N aqueous HCl (2 × 50 mL).The acidic layer was
washed with CH2Cl2 (10 mL) and was then basified with NH4-
OH. The suspension was extracted with CH2Cl2 (3 × 50 mL).
The organic solution was dried over anhydrous MgSO4 and
evaporated under reduced pressure. The crude product was
purified by sublimation to afford 8-methylisoquinoline (3.9 g)
2-Benzoyl-1-cyano-8-ethyl-1,2-dihydroisoquinoline(10b).
Compound 10b was prepared according to the same chemical
procedure as described for compound 10a using compound 9b
as the starting material; yield, 64%; mp 95-97 °C. IR (KBr):
2232, 1660, 1625, 1343 cm-1. 1H NMR (CDCl3): δ 1.36 (t, 3H,
J ) 6.9 Hz), 2.83 (d, 2H, J ) 6.2 Hz), 6.08 (d, 1H, J ) 6.9 Hz),
6.61 (br s, 1H), 6.81 (br s, 1H), 7.08 (d, 1H, J ) 7.6 Hz), 7.23
(d, 1H, J ) 7.6 Hz), 7.35 (t, 1H, J ) 7.6 Hz), 7.48 (t, 2H, J )
7.5 Hz), 7.56 (t, 1H, J ) 7.5 Hz), 7.61 (d, 2H, J ) 7.5 Hz).
Anal. (C19H16N2O) C, H, N.
2-Benzoyl-1-cyano-8-isopropyl-1,2-dihydroisoquino-
line (10c). Compound 10c was prepared according to the same
chemical procedure as described for compound 10a using
compound 9c as the starting material; yield, 63%; mp 150-
1
as an oil; yield, 62%. IR (C2Cl4): 3058, 1624, 1578 cm-1. H
NMR (CDCl3): δ 2.77 (s, 3H), 7.37 (d, 1H), 7.54-7.67 (m, 3H),
152 °C. IR (KBr): 2225, 1671, 1634, 1340 cm-1 1H NMR
.
8.54 (d, 1H), 9.45 (s, 1H); m/z 144 (MH+).
(CDCl3): δ 1.37 (d, 6H, J ) 6.8 Hz), 3.29 (br s, 1H), 6.09 (d,
1H, J ) 6.7 Hz), 6.61 (br s, 1H), 6.94 (br s, 1H), 7.07 (d, 1H, J
) 7.6 Hz), 7.33 (d, 1H, J ) 7.6 Hz), 7.39 (t, 1H, J ) 7.6 Hz),
7.48 (t, 2H, J ) 7.5 Hz), 7.56 (t, 1H, J ) 7.5 Hz), 7.61 (d, 2H,
J ) 7.5 Hz). Anal. (C20H18N2O) C, H, N.
8-Ethylisoquinoline (9b). Compound 9b was prepared
according to the same chemical pathway as described for
compound 9a using 2-ethylbenzaldehyde as the starting mate-
1
rial; yield, 66%. IR (C2Cl4): 2972, 1622, 1575 cm-1. H NMR
(CDCl3): δ 1.41 (t, 3H, J ) 7,5 Hz), 3.20 (q, 2H, J ) 7,5 Hz),
7.41 (d, 1H), 7.57-7.67 (m, 3H), 8.53 (d, 1H), 9.51 (s, 1H); m/z
158 (MH+).
2-Benzoyl-1-cyano-8-methoxy-1,2-dihydroisoquino-
line (10d). Compound 10d was prepared according to the
same chemical procedure as described for compound 10a using
compound 9d as the starting material; yield, 60%; mp 130-
8-Isopropylisoquinoline (9c). Compound 9c was prepared
according to the same chemical pathway as described for
compound 9a using compound 8 as the starting material; yield,
132 °C. IR (KBr): 2232, 1665, 1635, 1337 cm-1 1H NMR
.
(CDCl3): δ 3.95 (s, 3H), 5.99 (d, 1H, J ) 6.5 Hz), 6.62 (br s,
1H), 6.81 (d, 1H, J ) 7.9 Hz), 6.85 (br s, 1H), 6.88 (d, 1H, J )
7.9 Hz), 7.35 (t, 1H, J ) 7.9 Hz), 7.48 (t, 2H, J ) 7.5 Hz), 7.55
(t, 1H, J ) 7.5 Hz), 7.60 (d, 2H, J ) 7.5 Hz). Anal. (C18H14N2O2)
C, H, N.
1
65%. IR (C2Cl4): 2968, 1621, 1575 cm-1. H NMR (CDCl3): δ
1.46 (d, 6H), 3.91 (m, 1H), 7.52 (dd, 1H), 7.63-7.68 (m, 3H),
8.53 (d, 1H), 9.60 (s, 1H); m/z 172 (MH+).
8-Methoxyisoquinoline (9d). Compound 9d was prepared
according to the same chemical pathway as described for
compound 9a using o-anisaldehyde as the starting material;
2-Benzoyl-8-bromo-1-cyano-1,2-dihydroisoquinoline
(10e). Compound 10e was prepared according to the same
chemical procedure as described for compound 10a using
compound 4 as the starting material; yield, 86%; mp 149-
1
yield, 20%. H NMR (CDCl3): δ 4.03 (s, 3H, OCH3), 6.89 (d,
1H), 7.36 (d, 1H), 7.57-7.59 (m, 2H), 8.53 (d, 1H), 9.63 (s, 1H);
m/z 160 (MH+).
150 °C. IR (KBr): 2239, 1662, 1628, 1341 cm-1 1H NMR
.
Thieno[2,3-c]pyridine (9e). Compound 9e was prepared
according to the same chemical pathway as described for
compound 9a using 2-thiophenecarboxaldehyde as the starting
material. The crude product was purified by sublimation to
afford a white solid; yield, 28%; mp 54-55 °C. IR (KBr): 1577,
(CDCl3): δ 6.01 (d, 1H, J ) 6.7 Hz), 6.66 (br s, 1H), 6.85 (br s,
1H), 7.16 (d, 1H, J ) 7.6 Hz), 7.27 (t, 1H, J ) 7.6 Hz), 7.49 (t,
2H, J ) 7.5 Hz), 7.53-7.59 (m, 2H), 7.61 (d, 2H, J ) 7.5 Hz).
Anal. (C17H11N2OBr) C, H, N.
2-Benzoyl-8-chloro-1-cyano-1,2-dihydroisoquinoline
(10f). Compound 10f was prepared according to the same
chemical procedure as described for compound 10a using
compound 5 as the starting material; yield, 83%; mp 144-
1
719 cm-1. H NMR (CDCl3): δ 7.43 (d, 1H, J ) 5.4 Hz), 7.48
(d, 1H, J ) 5.4 Hz), 7.80 (d, 1H, J ) 5.4 Hz), 8.44 (d, 1H, J )
5.4 Hz), 9.11 (s, 1H); m/z 136 (MH+). Anal. (C7H5NS) C, H, N.
2-Methylthieno[2,3-c]pyridine (9f). Compound 9f was
prepared according to the same chemical pathway as described
for compound 9a using 5-methyl-2-thiophenecarboxaldehyde
as the starting material. The resulting crude product was
purified by sublimation to afford a white solid; yield, 17%; mp
52-54 °C. IR (KBr): 1577, 848 cm-1. 1H NMR (CDCl3): δ 2.62
(s, 3H), 6.98 (s, 1H), 7.50 (d, 1H, J ) 5.3 Hz), 8.40 (d, 1H, J )
5.3 Hz), 8.97 (s, 1H); m/z 150 (MH+). Anal. (C8H7NS) C, H, N.
145 °C. IR (KBr): 2239, 1663, 1629, 1343 cm-1 1H NMR
.
(CDCl3): δ 6.03 (d, 1H, J ) 7.1 Hz), 6.68 (br s, 1H), 6.91 (br s,
1H), 7.13 (d, 1H, J ) 7.0 Hz), 7.33-7.38 (m, 2H), 7.49 (t, 2H,
J ) 7.5 Hz), 7.58 (t, 1H, J ) 7.5 Hz), 7.62 (d, 2H, J ) 7.5 Hz).
Anal. (C17H11N2OCl) C, H, N.
2-Benzoyl-5-bromo-1-cyano-1,2-dihydroisoquinoline
(10g). Compound 10g was prepared according to the same
chemical procedure as described for compound 10a using
compound 1 as the starting material; yield, 87%; mp 177-
2-Benzoyl-1-cyano-8-methyl-1,2-dihydroisoquinoline
(10a). Anhydrous aluminum chloride (10 mg) was added to a
stirred solution of 8-methoxyisoquinoline 9a (2.25 g; 15.7
mmol) and trimethylsilyl cyanide (3.9 mL; 31.4 mmol) in
anhydrous CH2Cl2 (50 mL) at room temperature. Then, benzoyl
chloride (3.6 mL; 31.4 mmol) was added dropwise to the stirred
solution over a course of 5 min. The mixture was warmed to
30 °C if no exotherm began after the addition of benzoyl
chloride. After it was stirred for a further 3 h period, water
(50 mL) was added and stirring was continued for 30 min. The
organic layer was collected and washed successively with 1 N
aqueous HCl (2 × 50 mL), water (50 mL), 1 N aqueous NaOH
(2 × 50 mL), and finally water (50 mL). The organic solution
was dried over anhydrous MgSO4 and evapored under reduced
pressure to give an oil, which was triturated with Et2O (20
mL) resulting in crystallization. The solid was collected,
washed with small volumes of Et2O, and dried (3.7 g); yield,
178 °C. IR (KBr): 2232, 1669, 1623, 1347 cm-1 1H NMR
.
(CDCl3): δ 6.42 (d, 1H, J ) 7.8 Hz), 6.55 (br s, 1H), 6.74 (br d,
1H, J ) 7.8 Hz), 7.21 (t, 1H, J ) 7.7 Hz), 7.31 (d, 1H, J ) 7.7
Hz), 7.50 (t, 2H, J ) 7.5 Hz), 7.57-7.65 (m, 4H). Anal.
(C17H11N2OBr) C, H, N.
2-Benzoyl-5-chloro-1-cyano-1,2-dihydroisoquinoline
(10h). Compound 10h was prepared according to the same
chemical procedure as described for compound 10a using
compound 7 as the starting material; yield, 85%; mp 175-
177 °C. IR (KBr): 2240, 1668, 1625, 1347 cm-1 1H NMR
.
(CDCl3): δ 6.44 (d, 1H, J ) 7.8 Hz), 6.55 (br s, 1H), 6.74 (br d,
1H, J ) 7.8 Hz), 7.26-7.30 (m, 2H), 7.45-7.51 (m, 3H), 7.57-
7.62 (m, 3H). Anal. (C17H11N2OCl) C, H, N.
7-Cyano-6-ethoxycarbonyl-6,7-dihydrothieno[2,3-c]py-
ridine (10i). Compound 10i was prepared according to the
same chemical procedure as described for compound 10a using
compound 9e as the starting material and using ethyl chlo-
roformate instead of benzoyl chloride. Recrystallization from
Et2O/n-hexane gave 10i as a white solid; yield, 84%; mp 86-
87 °C. IR (KBr): 2232, 1705, 1617, 1330 cm-1. Anal. (C11H10N2-
O2S) C, H, N.
85%; mp 149-151 °C. IR (KBr): 2232, 1663, 1634, 1339 cm-1
.
1H NMR (CDCl3): δ 2.48 (s, 3H), 6.01 (d, 1H, J ) 7.3 Hz),
6.58 (br s, 1H), 6.69 (br s, 1H), 7.03 (d, 1H, J ) 7.6 Hz), 7.16
(d, 1H, J ) 7.6 Hz), 7.27 (t, 1H, J ) 7.6 Hz), 7.45 (t, 2H, J )
7.6 Hz), 7.54 (t, 1H, J ) 7.6 Hz), 7.59 (d, 2H, J ) 7.6 Hz).
Anal. (C18H14N2O) C, H, N.

4978 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 15
Graulich et al.
7-Cyano-6-ethoxycarbonyl-2-methyl-6,7-dihydrothieno-
[2,3-c]pyridine (10j). Compound 10j was prepared according
to the same chemical procedure as described for compound 10a
using compound 9f as the starting material and using ethyl
chloroformate instead of benzoyl chloride. Recrystallization
from Et2O/n-hexane gave 10j as a white solid; yield, 74%; mp
98-100 °C. IR (KBr): 2232, 1712, 1633, 1328 cm-1. Anal.
(C12H12N2O2S) C, H, N.
(dd, 1H, J ) 0.9 and 7.6 Hz), 8.52 (d, 1H, J ) 5.5 Hz). Anal.
(C18H16NO2Br) C, H, N.
8-Chloro-1-(3,4-dimethoxy-benzyl)-isoquinoline (11f).
Compound 11f was prepared according to the same chemical
procedure as described for compound 11a using compound 10f
as the starting material. Recrystallization from petroleum
ether 100-140 gave 11f as a cream solid; yield, 96%; mp 103-
1
105 °C. IR (KBr): 1544, 1514, 842 cm-1. H NMR (CDCl3): δ
3.79 (s, 3H), 3.81 (s, 3H), 5.06 (s, 2H), 6.60 (dd, 1H, J ) 0.8
and 8.2 Hz), 6.73 (d, 1H, J ) 8.2 Hz), 6.75 (s, 1H) 7.50 (t, 1H,
J ) 7.7 Hz), 7.58 (d, 1H, J ) 5.6 Hz), 7.79 (d, 1H, J ) 7.7 Hz),
7.92 (d, 1H, J ) 7.7 Hz), 8.52 (d, 1H, J ) 5.6 Hz). Anal. (C18H16-
NO2Cl) C, H, N.
1-(3,4-Dimethoxy-benzyl)-8-methyl-isoquinoline (11a).
A solution of 10a (1.9 g; 6.89 mmol) and of 3,4-dimethoxybenzyl
chloride (1.3 g; 6.89 mmol) in DMF (15 mL) was added
dropwise to a stirred suspension of sodium hydride (0.2 g; 8.33
mmol) in DMF (30 mL) at -10 °C. The content was stirred for
4 h and poured into ice-cold water (200 mL). The creamy solid
was filtered off. After it was dried, the solid was hydrolyzed
by treatment with 50% NaOH in a 1:1 EtOH-water solution
at reflux. After removal of EtOH, the crude residue was
dissolved in ArMe (50 mL) and water (50 mL). The organic
layer was collected, washed with water (50 mL), and then
extracted with 1 N aqueous HCl (2 × 50 mL). The acidic layers
were basified with concentrated NH4OH and finally extracted
with CH2Cl2 (3 × 30 mL). The organic layers were dried over
anhydrous MgSO4 and evaporated under reduced pressure to
afford a white solid, which recrystallized from petroleum ether
100-140 (1.8 g); yield, 90%; mp 100-102 °C. IR (KBr): 1560,
1518, 844 cm-1. 1H NMR (CDCl3): δ 2.83 (s, 3H), 3.73 (s, 3H),
3.79 (s, 3H), 4.79 (s, 2H), 6.43 (dd, 1H, J ) 0.7 and 8.2 Hz),
6.62 (d, 1H, J ) 0.7 Hz), 6.70 (d, 1H, J ) 8.2 Hz), 7.32 (d, 1H,
J ) 7.5 Hz), 7.48 (t, 1H, J ) 7.5 Hz), 7.55 (d, 1H, J ) 5.5 Hz),
7.67 (d, 1H, J ) 7.5 Hz), 8.46 (d, 1H, J ) 5.5 Hz). Anal. (C19H19-
NO2) C, H, N.
5-Bromo-1-(3,4-dimethoxy-benzyl)-isoquinoline (11g).
Compound 11g was prepared according to the same chemical
procedure as described for compound 11a using compound 10g
as the starting material. Recrystallization from petroleum
ether 100-140 gave 11g as a cream solid; yield, 90%; mp 86-
1
87 °C. IR (KBr): 1576, 1517, 804 cm-1. H NMR (CDCl3): δ
3.79 (s, 3H), 3.81 (s, 3H), 4.62 (s, 2H), 6.75 (s, 2H), 6.81 (s,
1H), 7.38 (t, 1H, J ) 8.2 Hz), 7.92-7.95 (t, 2H), 8.16 (d, 1H, J
) 8.2 Hz), 8.60 (d, 1H, J ) 6.0 Hz). Anal. (C18H16NO2Br) C, H,
N.
5-Chloro-1-(3,4-dimethoxy-benzyl)-isoquinoline (11h).
Compound 11h was prepared according to the same chemical
procedure as described for compound 11a using compound 10h
as the starting material. Recrystallization from petroleum
ether 100-140 gave 11h as a cream solid; yield, 94%; mp 105-
107 °C. IR (KBr): 1578, 1510, 832, 804 cm-1 1H NMR
.
(CDCl3): δ 3.79 (s, 3H), 3.81 (s, 3H), 4.62 (s, 2H), 6.76 (s, 2H),
6.81 (s, 1H), 7.44 (t, 1H, J ) 8.2 Hz), 7.72 (d, 1H, J ) 8.2 Hz),
7.97 (d, 1H, J ) 6.0 Hz), 8.11 (d, 1H, J ) 8.2 Hz), 8.61 (d, 1H,
J ) 6.0 Hz). Anal. (C18H16NO2Cl) C, H, N.
1-(3,4-Dimethoxy-benzyl)-8-ethyl-isoquinoline (11b).
Compound 11b was prepared according to the same chemical
procedure as described for compound 11a using compound 10b
as the starting material. Recrystallization from petroleum
ether 100-140 gave 11b as a cream solid; yield, 83%; mp 74-
7-(3,4-Dimethoxy-benzyl)-thieno[2,3-c]pyridine (11i).
Compound 11i was prepared according to the same chemical
procedure as described for compound 11a using compound 10i
as the starting material. Recrystallization from petroleum
ether 100-140 gave 11i as a cream solid; yield, 46%; mp 99-
1
76 °C. IR (KBr): 1558, 1514, 844 cm-1. H NMR (CDCl3): δ
1.36 (t, 3H, J ) 7.4 Hz), 3.18 (q, 2H, J ) 7.4 Hz), 3.75 (s, 3H),
3.80 (s, 3H), 4.76 (s, 2H), 6.46 (dd, 1H, J ) 0.7 and 8.2 Hz),
6.64 (s, 1H), 6.70 (d, 1H, J ) 8.2 Hz), 7.41 (d, 1H, J ) 7.1 Hz),
7.54-7.59 (m, 2H), 7.70 (d, 1H, J ) 8.1 Hz), 8.49 (d, 1H, J )
5.5 Hz). Anal. (C20H21NO2) C, H, N.
1
101 °C. IR (KBr): 1575, 1513, 828 cm-1. H NMR (CDCl3): δ
3.82 (s, 3H), 3.83 (s, 3H), 4.38 (s, 2H), 6.79 (d, 1H, J ) 8.7
Hz), 6.92-6.94 (d, 2H), 7.35 (d, 1H, J ) 5.4 Hz), 7.58 (d, 1H,
J ) 5.5 Hz), 7.63 (d, 1H, J ) 5.4 Hz), 8.47 (d, 1H, J ) 5.5 Hz).
Anal. (C16H15NO2S) C, H, N.
1-(3,4-Dimethoxy-benzyl)-8-isopropyl-isoquinoline (11c).
Compound 11c was prepared according to the same chemical
procedure as described for compound 11a using compound 10c
as the starting material. Recrystallization from petroleum
ether 100-140 gave 11c as a cream solid; yield, 78%; mp 95-
7-(3,4-Dimethoxy-benzyl)-2-methyl-thieno[2,3-c]pyri-
dine (11j). Compound 11j was prepared according to the same
chemical procedure as described for compound 11a using
compound 10j as the starting material. Recrystallization from
petroleum ether 100-140 gave 11j as a cream solid; yield, 44%;
1
96 °C. IR (KBr): 1557, 1512, 856 cm-1. H NMR (CDCl3): δ
mp 118-120 °C. IR (KBr): 1576, 1510, 846 cm-1 1H NMR
.
1.25 (d, 6H, J ) 6.7 Hz), 3.76 (s, 3H), 3.82 (s, 3H), 4.00
(multiplet, 1H, J ) 6.7 Hz), 4.73 (s, 2H), 6.55 (dd, 1H, J ) 0.8
and 8.2 Hz), 6.69 (s, 1H), 6.74 (d, 1H, J ) 8.2 Hz), 7.56-7.62
(m, 3H), 7.68 (dd, 1H, J ) 1.4 and 7.7 Hz), 8.47 (d, 1H, J )
5.4 Hz). Anal. (C21H23NO2) C, H, N.
(CDCl3): δ 2.59 (s, 3H), 3.82 (s, 3H), 3.83 (s, 3H), 4.30 (s, 2H),
6.78 (d, 1H, J ) 8.6 Hz), 6.0 (s, 2H), 6.98 (s, 1H), 7.40 (d, 1H,
J ) 5.4 Hz), 8.40 (d, 1H, J ) 5.5 Hz). Anal. (C17H17NO2S) C,
H, N.
1-(3,4-Dimethoxy-benzyl)-8-methyl-2-methyl-1,2,3,4-
tetrahydroisoquinoline hydrochloride (12a). A solution
of compound 11a (1.2 g; 4.2 mmol) in MeCN (10 mL) was
refluxed with an excess of methyl iodide (1.0 mL; 16 mmol).
After 2 h, Et2O was added resulting in a rapid crystallization
of a yellow solid. The precipitate was filtered off, washed with
Et2O (2 × 10 mL), dried, and used without further purification.
Under an inert atmosphere, NaBH4 was added to a solution
of the yellow compound (1.85 g; 4.2 mmol) in MeOH (50 mL)
at room temperature. After 15 min, MeOH was removed under
reduced pressure and the crude residue was dissolved in a 1
N aqueous HCl (100 mL). The acidic layer was washed with
Et2O (3 × 20 mL) and then basified with NH4OH. The
suspension was extracted with CH2Cl2 (3 × 30 mL). The
organic layers were collected, dried over anhydrous MgSO4,
and evaporated under reduced pressure to afford a colorless
oil, which was isolated as a hydrochloride salt and further
recrystallizated from THF/Et2O (1.51 g); yield, 98%; mp 133-
1-(3,4-Dimethoxy-benzyl)-8-methoxy-isoquinoline (11d).
Compound 11d was prepared according to the same chemical
procedure as described for compound 11a using compound 10d
as the starting material. Recrystallization from petroleum
ether 100-140 gave 11d as a cream solid; yield, 93%; mp 83-
1
84 °C. IR (KBr): 1560, 1513, 838 cm-1. H NMR (CDCl3): δ
3.79 (s, 3H), 3.81 (s, 3H), 3.89 (s, 3H), 4.82 (s, 2H), 6.69 (dd,
1H, J ) 1.4 and 8.2 Hz), 6.72 (d, 1H, J ) 8.2 Hz), 6.81 (s, 1H),
6.86 (d, 1H, J ) 8.0 Hz), 7.36 (d, 1H, J ) 8.0 Hz), 7.48 (d, 1H,
J ) 5.6 Hz), 7.53 (t, 1H, J ) 8.0 Hz), 8.46 (d, 1H, J ) 5.6 Hz).
Anal. (C19H19NO3) C, H, N.
8-Bromo-1-(3,4-dimethoxy-benzyl)-isoquinoline (11e).
Compound 11e was prepared according to the same chemical
procedure as described for compound 11a using compound 10e
as the starting material. Recrystallization from petroleum
ether 100-140 gave 11e as a cream solid; yield, 74%; mp 112-
1
114 °C. IR (KBr): 1537, 1513, 839 cm-1. H NMR (CDCl3): δ
135 °C. IR (KBr): 2465, 1593, 1518, 1268 cm-1 1H NMR
.
3.78 (s, 3H), 3.82 (s, 3H), 5.15 (s, 2H), 6.57 (dd, 1H, J ) 0.9
and 8.2 Hz), 6.72-6.74 (d, 2H), 7.42 (t, 1H, J ) 7.6 Hz), 7.58
(d, 1H, J ) 5.5 Hz), 7.79 (dd, 1H, J ) 0.9 and 7.6 Hz), 7.92
(CDCl3): δ 1.70 (s, 3H), 2.71 (s, 3H), 3.03-3.15 (m, 3H), 3.64
(s, 3H), 3.80 (s, 3H), 3.81-3.85 (m, 3H), 4.51 (d, 1H, J ) 5.4

N-Methyl-laudanosine and N-Methyl-noscapine
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 15 4979
Hz), 6.43 (d, 1H, J ) 1.7 Hz), 6.57 (dd, 1H, J ) 1.7 and 8.1
Hz), 6.67 (d, 1H, J ) 8.1 Hz), 7.00 (d, 1H, J ) 7.5 Hz), 7.04 (d,
1H, J ) 7.5 Hz), 7.20 (t, 1H, J ) 7.5 Hz), 13.02 (br s, 1H).
Anal. (C20H26NO2Cl‚H2O) C, H, N.
as a white solid; yield, 52%; mp 187-189 °C. IR (KBr): 1594,
1519, 1269 cm-1. 1H NMR (CDCl3): δ 2.84 (d, 3H, J ) 4.9 Hz),
2.94-3.03 (m, 2H), 3.18 (dd, 1H, J ) 5.8 and 18.7 Hz), 3.41
(m, 1H), 3.69 (m, 1H), 3.71 (s, 3H) 3.86 (s, 3H), 4.04 (dd, 1H,
J ) 2.7 and 13.0 Hz), 4.26 (d, 1H, J ) 8.6 Hz), 6.40 (d, 1H, J
) 7.9 Hz), 6.59 (dd, 1H, J ) 1.7 and 8.1 Hz), 6.64 (d, 1H, J )
1.7 Hz), 6.75 (d, 1H, J ) 8.1 Hz), 6.99 (t, 1H, J ) 7.9 Hz), 7.58
(d, 1H, J ) 7.9 Hz), 13.42 (br s, 1H). Anal. (C19H23NO2BrCl)
C, H, N.
1-(3,4-Dimethoxy-benzyl)-8-ethyl-2-methyl-1,2,3,4-tet-
rahydroisoquinoline hydrochloride (12b). Compound 12b
was prepared according to the same chemical procedure as
described for compound 12a using compound 11b as the
starting material. Recrystallization from MeCOOEt/n-hexane
gave 12b as a white solid; yield, 97%; mp 135-136 °C. IR
(KBr): 2566, 1590, 1519, 1265 cm-1. 1H NMR (CDCl3): δ 0.98
(t, 3H, J ) 7.5 Hz), 1.95 (m, 2H, J ) 7.5 Hz), 2.75 (d, 3H, J )
5.9 Hz), 3.05-3.18 (m, 4H), 3.66 (s, 3H), 3.83-3.88 (m, 5H),
4.57 (d, 1H, J ) 5.9 Hz), 6.40 (d, 1H, J ) 1.7 Hz), 6.57 (dd,
1H, J ) 1.7 and 8.2 Hz), 6.69 (d, 1H, J ) 8.2 Hz), 7.07 (d, 2H,
J ) 7.7 Hz), 7.28 (t, 1H, J ) 7.7 Hz), 13.09 (br s, 1H). Anal.
(C21H28NO2Cl) C, H, N.
1-(3,4-Dimethoxy-benzyl)-8-isopropyl-2-methyl-1,2,3,4-
tetrahydroisoquinoline hydrochloride (12c). Compound
12c was prepared according to the same chemical procedure
as described for compound 12a using compound 11c as the
starting material. Recrystallization from EtOH/Et2O gave 12c
as a white solid; yield, 99%; mp 204-205 °C. IR (KBr): 2522,
2437, 1590, 1519, 1265 cm-1. 1H NMR (CDCl3): δ 0.87 (d, 3H,
J ) 6.8 Hz), 0.95 (d, 3H, J ) 6.8 Hz), 2.33 (m, 1H, J ) 6.8
Hz), 2.74 (d, 3H, J ) 4.9 Hz), 3.07-3.18 (m, 4H), 3.65 (s, 3H),
3.81-3.90 (m, 5H), 4.67 (d, 1H, J ) 6.1 Hz), 6.42 (d, 1H, J )
1.7 Hz), 6.57 (dd, 1H, J ) 1.7 and 8.1 Hz), 6.69 (d, 1H, J ) 8.1
Hz), 7.06 (d, 1H, J ) 7.7 Hz), 7.17 (d, 1H, J ) 7.7 Hz), 7.31 (t,
1H, J ) 7.7 Hz), 13.14 (br s, 1H). Anal. (C22H30NO2Cl) C, H,
N.
5-Chloro-1-(3,4-dimethoxy-benzyl)-2-methyl-1,2,3,4-tet-
rahydroisoquinoline hydrochloride (12h). Compound 12h
was prepared according to the same chemical procedure as
described for compound 12a using compound 11h as the
starting material. Recrystallization from MeCOOEt/n-hexane
gave 12h as a white solid; yield, 72%; mp 198-200 °C. IR
(KBr): 1593, 1518, 1267 cm-1 1H NMR (CDCl3): δ 2.85 (d,
.
3H, J ) 4.9 Hz), 2.95-3.06 (m, 2H), 3.21 (dd, 1H, J ) 5.8 and
18.8 Hz), 3.42 (m, 1H), 3.69 (m, 1H), 3.77 (s, 3H) 3.86 (s, 3H),
4.03 (dd, 1H, J ) 2.8 and 13.0 Hz), 4.29 (d, 1H, J ) 8.4 Hz),
6.36 (d, 1H, J ) 7.9 Hz), 6.59 (dd, 1H, J ) 1.7 and 8.1 Hz),
6.65 (d, 1H, J ) 1.7 Hz), 6.74 (d, 1H, J ) 8.1 Hz), 7.06 (t, 1H,
J ) 7.9 Hz), 7.39 (d, 1H, J ) 7.9 Hz), 13.40 (br s, 1H). Anal.
(C19H23NO2Cl2) C, H, N.
7-(3,4-Dimethoxy-benzyl)-6-methyl-4,5,6,7-tetrahy-
drothieno[2,3-c]pyridine hydrochloride (12i). Compound
12i was prepared according to the same chemical procedure
as described for compound 12a using compound 11i as the
starting material. Recrystallization from THF/n-hexane gave
12i as a white solid; yield, 88%; mp 97-99 °C. IR (KBr): 2407,
1592, 1517, 1263 cm-1. 1H NMR (CDCl3): δ 1.84-3.06 (m, 5H),
3.20-3.34 (m, 3H), 3.79-3.85 (m, 7H), 4.51 and 5.06 (br d,
1H), 6.75-6.99 (m, 4H), 7.18-7.26 (m, 1H), 13.24 and 13.43
(br s, 1H). Anal. (C17H22NO2SCl‚H2O) C, H, N.
1-(3,4-Dimethoxy-benzyl)-8-methoxy-2-methyl-1,2,3,4-
tetrahydroisoquinoline fumarate (12d). Compound 12d
was prepared according to the same chemical procedure as
described for compound 12a using compound 11d as the
starting material, but the oil was isolated as a fumarate salt.
Recrystallization from MeCOOEt gave 12d as a white solid;
yield, 97%; mp 148-149 °C. IR (KBr): 3432, 1591, 1516, 1262
7-(3,4-Dimethoxy-benzyl)-2,6-dimethyl-4,5,6,7-tetrahy-
drothieno[2,3-c]pyridine hydrochloride (12j). Compound
12j was prepared according to the same chemical procedure
as described for compound 12a using compound 11j as the
starting material. Recrystallization from EtOH/Et2O gave 12j
as a white solid; yield, 86%; mp 165-167 °C. IR (KBr): 2534,
cm-1 1H NMR (DMSO): δ 2.33 (s, 3H), 2.44-2.48 (m, 1H),
.
1
2.70-2.86 (m, 4H), 3.24 (m, 1H), 3.67 (s, 3H), 3.71 (s, 3H), 3.79
(s, 3H), 4.01 (dd, 1H, J ) 3.8 and 7.3 Hz), 6.60 (s, 2H), 6.69-
6.72 (t, 3H), 6.81 (d, 2H, J ) 7.9 Hz), 7.13 (t, 1H, J ) 7.9 Hz).
Anal. (C24H29NO7) C, H, N.
1593, 1518, 1261 cm-1. H NMR (CDCl3): δ 2.35 and 2.38 (s,
3H), 2.80-2.95 (m, 5H), 3.19-3.41 (m, 3H), 3.77-3.89 (m, 7H),
4.40 and 4.99 (br d, 1H), 6.47 (s, 1H), 6.79-7.00 (m, 3H), 13.15
and 13.37 (br s, 1H). Anal. (C18H24NO2SCl‚1/4H2O) C, H, N.
8-Bromo-1-(3,4-dimethoxy-benzyl)-2-methyl-1,2,3,4-tet-
rahydroisoquinoline hydrochloride (12e). Compound 12e
was prepared according to the same chemical procedure as
described for compound 12a using compound 11e as the
starting material. Recrystallization from MeCOOEt/n-hexane
gave 12e as a white solid; yield, 42%; mp 166-167 °C. IR
1-(3,4-Dimethoxy-benzyl)-2,2,8-trimethyl-1,2,3,4-tet-
rahydroisoquinolinium iodide (13a). A solution of com-
pound 12a (0.36 g; 1.1 mmol) in MeCN (10 mL) was refluxed
with an excess of methyl iodide (0.5 mL; 8 mmol). After 4 h,
the solvent was removed under reduced pressure, and the
white residue was recrystallized from EtOH/Et2O (0.45 g);
1
(KBr): 2519, 2429, 1592, 1517, 1273 cm-1. H NMR (CDCl3):
yield, 91%; mp 208-209 °C. IR (KBr): 1516, 1263 cm-1 1H
.
δ 2.72 (d, 3H, J ) 4.9 Hz), 2.90-2.99 (m, 2H), 3.14 (m, 1H),
3.59 (dd, 1H, J ) 4.6 and 14.8 Hz), 3.63-3.68 (m, 1H), 3.72-
3.77 (m, 4H), 3.86 (s, 3H), 4.74 (br s, 1H), 6.72-6.73 (d, 2H),
6.81 (dd, 1H, J ) 1.7 and 8.2 Hz), 7.16 (d, 1H, J ) 7.7 Hz),
7.23 (t, 1H, J ) 7.7 Hz), 7.57 (d, 1H, J ) 7.7 Hz), 13.10 (br s,
1H). Anal. (C19H23NO2BrCl) C, H, N.
NMR (CDCl3): δ 2.02 (s, 3H), 3.08-3.24 (m, 3H), 3.40 (s, 3H),
3.52 (dd, 1H, J ) 4.7 and 15.4), 3.64 (s, 3H), 3.74 (m, 1H),
3.80 (s, 3H), 3.85 (s, 3H), 3.93 (m, 1H), 5.49 (t, 1H, J ) 5.8
Hz), 6.26 (d, 1H, J ) 1.7 Hz), 6.62 (dd, 1H, J ) 1.7 and 8.1
Hz), 6.70 (d, 1H, J ) 8.1 Hz), 7.01 (d, 1H, J ) 7.6 Hz), 7.04 (d,
1H, J ) 7.6 Hz), 7.21 (t, 1H, J ) 7.6 Hz). Anal. (C21H28NO2I)
C, H, N.
8-Chloro-1-(3,4-dimethoxy-benzyl)-2-methyl-1,2,3,4-tet-
rahydroisoquinoline hydrochloride (12f). Compound 12f
was prepared according to the same chemical procedure as
described for compound 12a using compound 11f as the
starting material. Recrystallization from MeCOOEt/n-hexane
gave 12f as a white solid; yield, 68%; mp 158-159 °C. IR
1-(3,4-Dimethoxy-benzyl)-2,2-dimethyl-8-ethyl-1,2,3,4-
tetrahydroisoquinolinium iodide (13b). Compound 13b
was prepared according to the same chemical procedure as
described for compound 13a using compound 12b as the
starting material. Recrystallization from EtOH/Et2O gave 13b
as a white solid; yield, 89%; mp 190-191 °C. IR (KBr): 1518,
(KBr): 1592, 1517, 1274 cm-1 1H NMR (CDCl3): δ 2.72 (d,
.
1
3H, J ) 4.8 Hz), 2.90-3.01 (m, 2H), 3.14 (m, 1H), 3.57-3.63
(m, 2H), 3.72-3.76 (m, 4H), 3.83 (s, 3H), 4.75 (br s, 1H), 6.72
(d, 1H, J ) 8.2 Hz), 6.74 (d, 1H, J ) 1.6 Hz), 6.80 (dd, 1H, J
) 1.6 and 8.2 Hz), 7.12 (d, 1H, J ) 7.8 Hz), 7.30 (t, 1H, J )
7.8 Hz), 7.38 (d, 1H, J ) 7.8 Hz), 13.08 (br s, 1H). Anal. (C19H23-
NO2Cl2) C, H, N.
1263 cm-1. H NMR (CDCl3): δ 1.13 (t, 3H, J ) 7.5 Hz), 2.17
(m, 1H, J ) 7.5 Hz), 2.52 (m, 1H, J ) 7.5 Hz), 3.12-3.27 (m,
3H), 3.42 (s, 3H), 3.52 (dd, 1H, J ) 4.7 and 14.7), 3.64 (s, 3H),
3.75 (m, 1H), 3.82 (s, 3H), 3.88 (s, 3H), 3.97 (m, 1H), 5.44 (t,
1H, J ) 5.6 Hz), 6.21 (d, 1H, J ) 1.7 Hz), 6.62 (dd, 1H, J )
1.7 and 8.1 Hz), 6.73 (d, 1H, J ) 8.1 Hz), 7.07 (d, 1H, J ) 7.6
Hz), 7.12 (d, 1H, J ) 7.6 Hz), 7.31 (t, 1H, J ) 7.6 Hz). Anal.
(C22H30NO2I) C, H, N.
5-Bromo-1-(3,4-dimethoxy-benzyl)-2-methyl-1,2,3,4-tet-
rahydroisoquinoline hydrochloride (12g). Compound 12g
was prepared according to the same chemical procedure as
described for compound 12a using compound 11g as the
starting material. Recrystallization from MeCOOEt gave 12g
1-(3,4-Dimethoxy-benzyl)-2,2-dimethyl-8-isopropyl-
1,2,3,4-tetrahydroisoquinolinium iodide (13c). Compound
13c was prepared according to the same chemical procedure

4980 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 15
Graulich et al.
as described for compound 13a using compound 12c as the
starting material. Recrystallization from EtOH/Et2O gave 13c
as a white solid; yield, 85%; mp 238-240 °C. IR (KBr): 1519,
1269 cm-1. 1H NMR (CDCl3): δ 0.92 (d, 3H, J ) 6.7 Hz), 1.13
(d, 3H, J ) 6.7 Hz), 2.88 (m, 1H, J ) 7.5 Hz), 3.13-3.28 (m,
3H), 3.40 (s, 3H), 3.56 (dd, 1H, J ) 4.4 and 14.6), 3.64 (s, 3H),
3.73 (m, 1H), 3.81 (s, 3H), 3.94-3.96 (d, 4H), 5.59 (t, 1H, J )
5.6 Hz), 6.24 (d, 1H, J ) 1.8 Hz), 6.57 (dd, 1H, J ) 1.8 and 8.1
Hz), 6.71 (d, 1H, J ) 8.1 Hz), 7.06 (d, 1H, J ) 7.6 Hz), 7.21 (d,
1H, J ) 7.6 Hz), 7.33 (t, 1H, J ) 7.6 Hz). Anal. (C23H32NO2I)
C, H, N.
1-(3,4-Dimethoxy-benzyl)-8-methoxy-2,2-dimethyl-
1,2,3,4-tetrahydroisoquinolinium iodide (13d). Compound
13d was prepared according to the same chemical procedure
as described for compound 13a using compound 12d as the
starting material. Recrystallization from EtOH/Et2O gave 13d
as a white solid; yield, 88%; mp 194-195 °C. IR (KBr): 1515,
1271 cm-1. 1H NMR (CDCl3): δ 3.04 (dd, 1H, J ) 6.0 and 18.6
Hz), 3.21-3.31 (m, 2H), 3.37 (s, 3H), 3.53-3.59 (m, 4H), 3.73
(s, 3H), 3.73 (m, 1H), 3.76-3.85 (m, 7H), 5.02 (t, 1H, J ) 4.0
Hz), 6.54 (s, 1H), 6.77-6.81 (m, 3H), 6.85 (d, 1H, J ) 8.0 Hz),
7.33 (t, 1H, J ) 8.0 Hz). Anal. (C21H28NO3I) C, H, N.
8.2 Hz), 7.06 (t, 1H, J ) 7.9 Hz), 7.45 (d, 1H, J ) 7.9 Hz).
Anal. (C20H25NO2ClI) C, H, N.
7-(3,4-Dimethoxy-benzyl)-6,6-dimethyl-4,5,6,7-tetrahy-
drothieno[2,3-c]pyridinium iodide (13i). Compound 13i
was prepared according to the same chemical procedure as
described for compound 13a using compound 12i as the
starting material. Recrystallization from EtOH/Et2O gave 13i
as a white solid; yield, 80%; mp 216-217 °C. IR (KBr): 1518,
1
1264 cm-1. H NMR (DMSO): δ 2.93 (dd, 1H, J ) 11.0 and
13.6 Hz), 3.00-3.14 (m, 5H), 3.32 (s, 3H), 3.70-3.87 (m, 9H),
4.88 (d, 1H, J ) 9.2 Hz), 6.89-6.98 (m, 4H), 7.42 (d, 1H, J )
5.1 Hz). Anal. (C18H24NO2SI) C, H, N.
7-(3,4-Dimethoxy-benzyl)-2,6,6-trimethyl-4,5,6,7-tet-
rahydrothieno[2,3-c]pyridinium iodide (13j). Compound
13j was prepared according to the same chemical procedure
as described for compound 13a using compound 12j as the
starting material. Recrystallization from EtOH/Et2O gave 13j
as a white solid; yield, 83%; mp 139-140 °C. IR (KBr): 1519,
1266 cm-1 1H NMR (CDCl3): δ 2.32 (s, 3H), 2.96-3.03 (m,
.
3H), 3.52 (s, 3H), 3.65-3.68 (m, 4H), 3.87 (s, 3H), 3.92 (s, 3H),
4.00 (m, 1H), 4.17 (m, 1H), 5.58 (dd, 1H, J ) 3.7 and 9.1 Hz),
6.47 (s, 1H), 6.80-6.86 (m, 2H), 7.00 (s, 1H). Anal. (C19H26-
NO2SI) C, H, N.
8-Bromo-1-(3,4-dimethoxy-benzyl)-2,2-dimethyl-1,2,3,4-
tetrahydroisoquinolinium iodide (13e). Compound 13e
was prepared according to the same chemical procedure as
described for compound 13a using compound 12e as the
starting material. Recrystallization from EtOH/Et2O gave 13e
as a white solid; yield, 82%; mp 216-217 °C. IR (KBr): 1517,
Radioligand Binding Studies and Data Analysis. Syn-
aptosomes Preparation. Rats (male Wistar, (250 g) were
killed by decapitation, and the brains were quickly removed
and kept on ice during dissection. The crude cortex was
dispersed in 0.32 M sucrose by using a Potter homogenizer.
After a first centrifugation at 1500g for 10 min, the superna-
tant was centrifuged at 25000g for 10 min. The resulting pellet
was dispersed in 5 mL of 0.32 M sucrose to be aliquoted. The
protein concentration was determined by the method of
Hartree with bovine serum albumin as a standard.36
Binding Experiments. The buffer consisted of a 10 mM
Tris-HCl (pH 7.5) solution containing 5.4 mM KCl and 0.1%
bovine serum albumin. The radioligand was 125I-apamin
(Perkin-Elmer, Specific activity 81.4 TBq mmol-1). Glass fiber
filters (Whatman GF/C) used in these experiments were coated
for 1 h in 0.5% polyethylenimine and then washed with 2.5
mL of the ice-cold buffer just before use. Binding experiments
were always terminated as follows. Aliquots were filtered
under reduced pressure through Whatman filters. Filters were
rapidly washed twice with 2.5 mL of buffer. The radioactivity
remaining on the filter was evaluated with a Packard Tri-Carb
1600TR liquid scintillation analyzer with an efficacy of 69%.
125I-apamin binding to the filters was also estimated in the
absence of synaptosomes. This binding was also subtracted
from the total binding. Curve fitting was carried out using
GraphPad Prism.
Saturation Binding Experiments. Synaptosomes (0.2 mg
of protein/mL) were incubated with increasing concentrations
of 125I-apamin (25 µL) with 975 µL of incubation buffer for 1 h
at 0 °C. Samples were then filtered on Whatman GF/C filter,
and the radioactivity was measured as described above.
Nonspecific binding was determined in parallel experiments
in the presence of an excess of unlabeled apamin (0.1 µM) and
subtracted from the total binding to obtain the specific binding.
Competition Experiments between 125I-Apamin and
Drugs. Synaptosomes (0.2 mg of protein/mL) were incubated
for 1 h at 0 °C with (10 pM of 125I-apamin (25 µL) and nine
concentrations of drugs (10-4-10-7 M). Nonspecific binding
was determined in the presence of an excess of unlabeled
apamin (0.1 µM). Samples were then filtered on a Whatman
filter, and the radioactivity was measured as described above.
Electrophysiological Experiments. The procedure is
largely described in previous papers.19,30 Briefly, male Wistar
rats (150-200 g) were anaesthetized with chloral hydrate (400
mg/kg IP) and decapitated. The brain was excised quickly and
placed in cold (∼4 °C) artificial cerebrospinal fluid (ACSF) at
the following composition (in mM): NaCl, 126; KCl, 2.5; NaH2-
PO4, 1.2; MgCl2, 1.2; CaCl2, 2.4; glucose, 11; NaHCO3, 18;
saturated with 95% O2 and 5% CO2 (pH 7.4). A block of tissue
containing the midbrain was cut in horizontal slices (thickness
350 µm) in a Vibratome (Lancer). The slice containing the
1
1262 cm-1. H NMR (DMSO): δ 3.02 (s, 3H), 3.08 (dd, 1H, J
) 5.1 and 15.6 Hz), 3.23-3.29 (m, 5H), 3.51 (dd, 1H, J ) 6.9
and 15.6 Hz), 3.57-3.63 (m, 4H), 3.72 (s, 3H), 3.88 (m, 1H),
5.14 (t, 1H, J ) 5.8 Hz), 6.62 (d, 1H, J ) 1.5 Hz), 6.80 (dd, 1H,
J ) 1.5 and 8.2 Hz), 6.85 (d, 1H, J ) 8.2 Hz), 7.33 (t, 1H, J )
7.7 Hz), 7.38 (d, 1H, J ) 7.7 Hz), 7.57 (d, 1H, J ) 7.7 Hz).
Anal. (C20H25NO2BrI) C, H, N.
8-Chloro-1-(3,4-dimethoxy-benzyl)-2,2-dimethyl-1,2,3,4-
tetrahydroisoquinolinium iodide (13f). Compound 13f was
prepared according to the same chemical procedure as de-
scribed for compound 13a using compound 12f as the starting
material. Recrystallization from EtOH/Et2O gave 13f as a
white solid; yield, 77%; mp 220-221 °C. IR (KBr): 1517, 1264
1
cm-1. H NMR (DMSO): δ 3.02-3.08 (m, 4H), 3.20-3.29 (m,
5H), 3.53 (dd, 1H, J ) 6.5 and 15.3 Hz), 3.61-3.64 (m, 4H),
3.71 (s, 3H), 3.87 (m, 1H), 5.21 (t, 1H, J ) 5.9 Hz), 6.58 (d,
1H, J ) 1.5 Hz), 6.77 (dd, 1H, J ) 1.5 and 8.2 Hz), 6.85 (d,
1H, J ) 8.2 Hz), 7.34-7.43 (m, 3H). Anal. (C20H25NO2ClI) C,
H, N.
5-Bromo-1-(3,4-dimethoxy-benzyl)-2,2-dimethyl-1,2,3,4-
tetrahydroisoquinolinium iodide (13g). Compound 13g
was prepared according to the same chemical procedure as
described for compound 13a using compound 12g as the
starting material. Recrystallization from EtOH/Et2O gave 13g
as a white solid; yield, 83%; mp 166-167 °C. IR (KBr): 1518,
1
1267 cm-1. H NMR (DMSO): δ 2.87 (dd, 1H, J ) 10.1 and
13.1 Hz), 3.02-3.08 (m, 4H), 3.18 (dd, 1H, J ) 6.0 and 18.5
Hz), 3.41 (s, 3H), 3.60 (dd, 1H, J ) 3.1 and 13.1 Hz), 3.65 (s,
3H), 3.72-3.75 (m, 4H), 3.84 (m, 1H), 4.87 (dd, 1H, J ) 3.1
and 10.1 Hz), 6.29 (d, 1H, J ) 7.8 Hz), 6.55 (dd, 1H, J ) 1.6
and 8.2 Hz), 6.63 (d, 1H, J ) 1.6 Hz), 6.84 (d, 1H, J ) 8.2 Hz),
6.98 (t, 1H, J ) 7.8 Hz), 7.61 (d, 1H, J ) 7.8 Hz). Anal. (C20H25-
NO2BrI) C, H, N.
5-Chloro-1-(3,4-dimethoxy-benzyl)-2,2-dimethyl-1,2,3,4-
tetrahydroisoquinolinium iodide (13h). Compound 13h
was prepared according to the same chemical procedure as
described for compound 13a using compound 12h as the
starting material. Recrystallization from MeOH/MeCOOEt
gave 13b as a white solid; yield, 85%; mp 136-138 °C. IR
1
(KBr): 1518, 1265 cm-1. H NMR (DMSO): δ 2.87 (dd, 1H, J
) 10.1 and 13.1 Hz), 3.07-3.11 (m, 4H), 3.22 (dd, 1H, J ) 6.0
and 18.5 Hz), 3.41 (s, 3H), 3.60 (dd, 1H, J ) 3.1 and 13.1 Hz),
3.65 (s, 3H), 3.72-3.78 (m, 4H), 3.87 (m, 1H), 4.88 (dd, 1H, J
) 3.1 and 10.1 Hz), 6.25 (d, 1H, J ) 7.9 Hz), 6.56 (dd, 1H, J
) 1.4 and 8.2 Hz), 6.64 (d, 1H, J ) 1.4 Hz), 6.84 (d, 1H, J )

N-Methyl-laudanosine and N-Methyl-noscapine
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 15 4981
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region of interest was placed on a nylon mesh in a recording
chamber (volume 500 µL). The tissue was completely immersed
in a continuously flowing (∼2 mL/min) ACSF, heated at 35
°C. Most recordings were made from dopaminergic neurons
located in the substantia nigra pars compacta. Intracellular
recordings were performed using glass microelectrodes filled
with 2 M KCl (resistance 70-150 MΩ). All recordings were
made in the bridge balance mode, using a npi SEC1L amplifier
(Tamm, Germany). The accuracy of the bridge was checked
throughout the experiment. Membrane potentials and injected
currents were recorded on a Gould TA240 chart recorder and
on a Fluke Combiscope oscilloscope. The Flukeview software
was used for off-line analysis in most cases. Drug effects on
the prominent apamin sensitive AHP in dopaminergic neurons
were quantified as the percent reduction of the surface area
of the AHP (in mV s), which was blocked by a maximally active
concentration of apamin (300 nM). Averages of four sweeps
were considered in all cases. The spontaneous firing of the
neurons was usually reduced by constant current injection
(-20 to -100 pA) in order to increase the amplitude of the
AHP. Because the amplitude of the AHP is very sensitive to
the firing rate, care was taken to compare all AHPs of one
cell at the same firing rate. All drugs were applied by
superfusion; complete exchange of the bath solution occurred
within 2-3 min. Curve fitting was carried out using GraphPad
Prism and the standard equation: E ) Emax/[1 + (IC50/x)h],
where x is the concentration of the drug and h is the Hill
coefficient. Numerical values are expressed as means ( SEM.
Apamin (Sigma) and all other drugs were dissolved in water.
Acknowledgment. The technical assistance of Y.
Abrassart, S. Counerotte, L. Massotte, and J.-C. Van
Heugen is gratefully acknowledged. A.G. is a Research
Fellow of the “Fonds pour la formation a` la Recherche
Industrielle et Agricole (F.R.I.A.)”, and J.-F. L. is a
Senior Research Associate of the “Fonds National de la
Recherche Scientifique de Belgique (F.N.R.S.)”. This
work was financially supported in part by F.N.R.S.
Grants 3.4525.98 and 9.4560.03 and by grants from the
“Fonds Spe´ciaux pour la Recherche” 2002 and 2003 of
the University of Lie`ge.
Supporting Information Available: Elemental analysis
data. This material is available free of charge via the Internet
at http://pubs.acs.org.
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