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Tissue distribution of chlorimipramine and its demethylated metabolite after a single dose in the rat
Pharmacological Research Communications, Vol. 16, No. 2, 1984
207
TISSUE DISTRIBUTION OF CHLORIMIPRAMINE AND ITS DEMETHYLATEDMETABOLITE AFTER A SINGLE DOSEIN THE RAT.
L. DELLA CORTEand G.P. SGARAGLI. I s t i t u t o Interfacolt~ di Farmacologia e Tossicologia, Viale G.B. Morgagni, 65 - 50134 Firenze, Italy and Istituto di Scienze Farmacologiche, Via Cassia Sud, Villa Betlem - 53100 Siena, Italy. Receivedm final ~ r m 2 6 Sep~mber 1983
SUMMARY The pattern of distribution of chlorimipramine (CI) and of i t s demethyl-derivative (DMCI) in different organs (l~ng, liver, kidney, heart and spleen) 24 h after a single dose of CI was examined and related to the amounts of the most representative classes of lipids present in t i s sues. The findings here reported
show that lung and liver have the
highest capacity to accumulate Cl while DMCI was preferentially accumulated by lung, spleen and kidney. The capacity of the examined tissues to accumulate CI and DMCI did not relate to their lipid content.
INTRODUCTION After the pioneristic work of M.H. Bickel and H.J. Weder (1968) on the kinetic of distribution of imipramine and of i t s major metabolites, several reports have clearly indicated that most of the mammalian organs share the capacity of storing t r i c y c l i c antidepressant drugs and their l,ipophilic metabolites. Binding to non specific sites has been suggested to be responsible for the high and persistent concentrations of imipramine and its lipophilic metabolites in certain organs. The endoplasmic reticulum has been reported by Gillette (1965) as the most important cell constituent involved in this phenomenon. More recent studies reported the interaction of imipramine and chlorpromazine with biomembranes
0031-6989/84/020207--07/S03.00/0
© 1984 The Italian Pharmacological Society
Pharmacological Research Communications, Vol. 16, No. 2, 1984
208
(Kwant and Seeman, 1969; Bickel and Steele, 1974; Birkett, 1974; Cater eta].,
]974; Bickel and Weder, ]976; Leterrier et a l . , 1976; Di France-
sco and Bickel, 1977). However, i t is s t i l l a matter of debate whether the protein or the lipid mojety of biomembranes play the major role in this interaction. Recently a study has been carried out in this laboratory to investigate whether the tissue distribution of CI, a t r i c y c l i c antidepressant drug and of i t s demethyl derivative could be related to the amounts of the most representative tissue lipid classes. Results so far obtained are reported in this paper.
METHODS Male Sprague-Dawley rats 200-250 g were used throughout. CI-HCI, dissolved in 2 ml water, was administered orally through a gastric tube in a single dose of 90 mg.Kg-l b.w. to six groups of 5 animals. In addition three groups of 5 animals, which received water only, were used as controls for the
measurementof tissue lipid levels. All the
animals were killed, 24 h after treatment, by exsanguination under l i g h t diethyl ether anaesthesia. In one group, used for the measurement of CI and DMCI tissue levels,
blood samples(5 ml)werewithdrawn by heart
puncture using heparinized syringes.
Blood samples were transferred into
tubes and immediately centrifuged at ]ow speed to separate plasma from red cells. After k i l l i n g the animals by exsanguination the organs (lung, liver, kidney and spleen) were removed, weighed as quickly as possible, minced and then suspended with 3 volumes of an ice-cold medium containing 17O mM NaCl, 3 mM KCI, IO mM Na2HP~4.2H2O and 2 mM KH2PO4 adjusted to pH 7.4. The suspension was homogenized in an Ultra Turrax homogenizer at 30% f u l l power, using 4x15 sec strokes. In the other groups, where tissue Content of lipids was measured, tissue homogenates were obtained from the pooled organs of 5 animals. Determination of CI and DMCI. The method has been described previously (Sgaragli et a l . , ]983).
Pharmacological Research Communications, Vol. 16, No. 2 , 1984
209
Determination of ] i p i d s . Lipids were extracted from lung, liver, kidney, spleen and heart homogenates (4.5 ml) by the method of Folch et al. (1957). The extracted lipids (100-400 mg) were loaded on s i l i c i c acid columns (12 mm internal diameter, 120 mm height)and fractionated into neutral lipids and phospholipids by elution with lO0 ml chloroform and methanol, respectively. The total phospholipids were determined by multiplying x 25 the phospholipidphosphorus assayed following the method of Martin and Doty (1949) after digestion with sulfuric acid-perchloric acid 3:2. The colorimetric method of Cramer and Isaksson (1959) was used to determine total cholesterol in the neutral l i pid fraction. Total glycerides were determined by assaying glycerol in the neutral lipid fraction according to thR procedure of Carlson (1963). Statistical methods. The difference between means was analyzed by the Student's two-tailed t-test for independent samples (not significant, P~O.05).
RESULTS AND DISCUSSION Tissue levels of Cl and DMCI were measured 24 h after the administration of a single dose of CI. At this time tissue levels of both compounds had entered the phase of slow decay, thus showing much less v a r i a b i l i t y than those measured at earlier times (Della Corte et a l . , 1981). Results are shown in table I. In all analyzed tissues most of the drug was detected as its N-demethyl-derivative. ried
The DMCI/CI ratio, in fact, was 4 in plasma and var-
between 1.5 in l i v e r and 8 in the other analyzed tissues.
CI and
DMCI were preferentially accumulated by lungs reaching levels of 99 and 198 times higher than those found in plasma, respectively.
Am-
ong other tissues, liver preferentially accumulated CI while spleen and kidney exhibited a relatively high capacity for storing both compounds. In order to shed some light on the mechanism of Cl and DMCi preferential accumulation by lung, tissue contents of various l i p i d classes were also measured (see table l ) . The organ most rich in l i pids was kidney followed by lung> liver'7 heart and spleen.
Total
phospholipids were present in the highest amount in kidney followed by l i v e r > heart ~ lung and spleen. Glycerides were most abundant in
DMCI
8.97 +2.88 4.41 +0.67 26.7 +5.7
1.20 +0.22
2.78 +0.46
3.60 +0.90
36.4+3.8 .
45.4+3.0 .
20.9+I.I .
41.4+4.1
26.7+1.1
total lipid
19.6+4.0 .
31.2+3.0 .
14.5+I.6 .
26.6+3.5
18.3+2.2
.
.
.
7.8+0.3
6.6+0.5
1.4+0.2
6.0+0.9
3.4+0.2
.
.
.
I. I P I D total phospholipid glycerides a
3.2+0.9
3.3+0.5
2.4+0.3
4.0+0.2
1.6+0.2
total cholesterol
CI and DMCI f i g u r e s r e p r e s e n t mean values (~ s . e . ) d e r i v e d from 5 a n i m a l s , L i p i d f i g u r e s r e p r e s e n t mean values (~ s . e . ) of five_ pooled homogenates each d e r i v e d from -I f i v e t r e a t e d a n i m a l s ; they are expressed as mg.g wet t i s s u e except a, expressed as ~moles.g wet t i s s u e . T o t a l l i p i d values were determined g r a v i m e t r i c a l l y on p o r t i o n s of the l i p i d e x t r a c t s evaporated at room t e m p e r a t u r e under N2 stream.
Lung
Liver
Spleen
7.70 +2.98
0.92 +0.17
2.35 +0.67
0.32 +0.07
Heart
Kidney
0.140 +0.031
0.037 +0.062
(pg.g-I wet tissue)
CI
Rat tissue levels of C], D~CI and l i p i d s 24 h a f t e r a single oral dose of CI (90 mg.Kg b.w.)
Plasma
Tissue
Table I .
a
{a {b
(:
0
Pharmacological Research Communications, Vol. 16, No. 2, 1984
lung followed by kidney ~ h~art"> l i v e r and spleen. Total cholesterol was most abundant in kidney and lung. The relationship between CI and DMCI levels and tissue lipid contents was studied. The correlation between tissue levels of CI and DMCI and of l i p i d classes was not significant. The observation that rat lung has the highest capacity for storing CI and DMCI is in agreement with data reported on the tissue distribution of imipramine (Bickel
and Weder, 196B) and chlorproma-
zine (Wechsler and Roizin, 1960). Junod (1972) observed that, in isolated and perfused rat lung, the accumulation of imipramine had the characteristic of a saturable phenomenon, was insensitive to temperature
or Na+ concentration changes, and was antagonized by chlor-
promazine. Orton and coworkers (1973) also observed that amines like amphetamine and imipramine were rapidly concentrated in the lungs and, at least in the case of imipramine, the process was saturable. Only a minimal fraction of the accumulated imipramine could be recovered in the surfactant and in alveolar macrophages. As outlined by Junod (1976), one remarkable feature of lung anatomy is its huge capillary surface, thus the concentration of these drugs in this organ could be explained in terms of a simple binding to cell membranes starting from a purely quantitative basis of membrane surface oistribution. Binding of amphiphilic cationic drugs to cell membranes has been studied in a variety of cellular and subcellular preparations, namely red cells (Kwant and Seeman, 1969), sarcoplasmic ret'fculum (Balzer et al., 1968), liver mitochondria (Huunan-Sepp~l~, 1972), platelets (Boullin and O'Brien, 1968), brain synaptosomes (Weinstein et a l . , 1971), brain membranes (Raisman et a l . , 1979) and lung mitochondrial or microsomal preparations (Minkin et a l . , 1979). There have been a few constant findings in all these systems: the presence of binding sites with high substrate concentration (IO-4M), and a relative insensitivity to a decrease in temperature have been evidentiated. In conclusion, i t would appear from the results under discussion that the lipids here examined do not play an important role in the mechanism of CI and DMCI tissue accumulation. The observation that imi-
21 1
212
Pharmacological Research Communications, VoL 16. No. 2, 1984
pramine binding to rabbit skeletal muscle homogenate was unaffected by prior l i p i d extraction (Fichtl et a l . , 1980) is in agreement with our conclusion.
ACKNOWLEDGMENTS
This research project has been supported by CNR - Roma (contr. n. 82.02043.04.
Dr. I. Megazzini is gratefully aknowledged for her
assistance in the experimental work.
REFERENCES - Balzer H., Makin~se M., Fiehn W. and Hasselbach W., Naunyn-Schmiedeberg's Arch. Pharmak. Exp. Pathol., 260, 456 (1968). -
Bickel M.H. and Steele J.W., Chem. Biol. Interact., _8, 151 (1974). Bicke] M.H. and Weder H.J., Arch. Int. Pharmacodyn., 17.3, 433 (1968).
- Bickel M.H. and Weder H.J., Psychopharmac. Co~un., _2, 231 (1976). - BirkeLt D.J., Clin. Exp. Pharmac., I_, 415 (1974). -
Boullin D.J. and O'Brien R.A., J. Pharm. Pharn,ac., 2__00,583 (1968).
- Carlsen L.A.,J. Atheroscler. Res., 3_, 334 (1963). - Cater B.R., Chapman D., Hawes S.M. and Saville J.,Biochim. Biophys. Acta, 363, 54 (1974). - Cramer K. and Isaksson B., Scand. J. Clin.
Invest. I I ,
213 (1959).
- Della Corte L., Giovannini M.G., Martini P., Megazzini I . ,
Puliti
R.,
Sgaragli G.P. and Wondrak G., Br. J. Pharmac., 72, 184P (1981).
- Di Francesco C. and Bickel M.H., Chem. Biol. Interact., 7_66,335 (1977). - Dittmer J.C. and Lester R.L., J. Lipid Res., --5, 126 (1964). - Fichtl B., Bondy B. and Kurz H., J. Pharmac. Exp. Ther., 215, 248 (1980). - Folch T., Lees M. and Sloane-Stanley G.M., J. Biol. Chem., 22___66,497 (1957). - Gillette J.R., in Drugs and Enzymes, ed. Brodie B.B. and Gillette J.R., p. 9, Pergamon Press, Oxford (1965). -
-
Huunan-Sepp~'l~" A., Acta Chem. Scand., 266, 2713 (1972). Junod A., Pharmac. Thero B, 2, 511 (1976).
Pharmacological Research Communications, Vol. 16, No. 2, 1984
-
Kwant W.O. and Seeman P., Biochim. Biophys. Acta, 18_3, 530 (1969).
- Leterrier F., Mendyk A. and Viret J., Biochem. Pharmac., 2_55, 2469 (1976). - Martin J.B. and Doty B.M., Anal. Chem., 21, 965 (1949). - Minkin R.F., l l e t t K.F. and Madsen B.W., Biochem. Pharmac., 23, 2275 (1979). - Orton T.C., Anderson M.V., Pickett R.D., Eling T.E. and Fouts J.R., J. Pharmac. Exp. Ther., 186, 482 (1973). -
Raisman R., Briley M. and Langer S.Z., Nature, 281, 148 (1979).
-
Sgaragli G.P., Della Corte L. and Gremigni D., Pharmac. Res. Comm., ~5, 231 (1983).
- Wechsler M.B. and Roizin L., J. Ment. Sci., 106, 1501 (1960). -
Weinstein H., Varon S. and Roberts E., Biochem. Pharmac., 20, 103 (1971).
2 1 3
207
TISSUE DISTRIBUTION OF CHLORIMIPRAMINE AND ITS DEMETHYLATEDMETABOLITE AFTER A SINGLE DOSEIN THE RAT.
L. DELLA CORTEand G.P. SGARAGLI. I s t i t u t o Interfacolt~ di Farmacologia e Tossicologia, Viale G.B. Morgagni, 65 - 50134 Firenze, Italy and Istituto di Scienze Farmacologiche, Via Cassia Sud, Villa Betlem - 53100 Siena, Italy. Receivedm final ~ r m 2 6 Sep~mber 1983
SUMMARY The pattern of distribution of chlorimipramine (CI) and of i t s demethyl-derivative (DMCI) in different organs (l~ng, liver, kidney, heart and spleen) 24 h after a single dose of CI was examined and related to the amounts of the most representative classes of lipids present in t i s sues. The findings here reported
show that lung and liver have the
highest capacity to accumulate Cl while DMCI was preferentially accumulated by lung, spleen and kidney. The capacity of the examined tissues to accumulate CI and DMCI did not relate to their lipid content.
INTRODUCTION After the pioneristic work of M.H. Bickel and H.J. Weder (1968) on the kinetic of distribution of imipramine and of i t s major metabolites, several reports have clearly indicated that most of the mammalian organs share the capacity of storing t r i c y c l i c antidepressant drugs and their l,ipophilic metabolites. Binding to non specific sites has been suggested to be responsible for the high and persistent concentrations of imipramine and its lipophilic metabolites in certain organs. The endoplasmic reticulum has been reported by Gillette (1965) as the most important cell constituent involved in this phenomenon. More recent studies reported the interaction of imipramine and chlorpromazine with biomembranes
0031-6989/84/020207--07/S03.00/0
© 1984 The Italian Pharmacological Society
Pharmacological Research Communications, Vol. 16, No. 2, 1984
208
(Kwant and Seeman, 1969; Bickel and Steele, 1974; Birkett, 1974; Cater eta].,
]974; Bickel and Weder, ]976; Leterrier et a l . , 1976; Di France-
sco and Bickel, 1977). However, i t is s t i l l a matter of debate whether the protein or the lipid mojety of biomembranes play the major role in this interaction. Recently a study has been carried out in this laboratory to investigate whether the tissue distribution of CI, a t r i c y c l i c antidepressant drug and of i t s demethyl derivative could be related to the amounts of the most representative tissue lipid classes. Results so far obtained are reported in this paper.
METHODS Male Sprague-Dawley rats 200-250 g were used throughout. CI-HCI, dissolved in 2 ml water, was administered orally through a gastric tube in a single dose of 90 mg.Kg-l b.w. to six groups of 5 animals. In addition three groups of 5 animals, which received water only, were used as controls for the
measurementof tissue lipid levels. All the
animals were killed, 24 h after treatment, by exsanguination under l i g h t diethyl ether anaesthesia. In one group, used for the measurement of CI and DMCI tissue levels,
blood samples(5 ml)werewithdrawn by heart
puncture using heparinized syringes.
Blood samples were transferred into
tubes and immediately centrifuged at ]ow speed to separate plasma from red cells. After k i l l i n g the animals by exsanguination the organs (lung, liver, kidney and spleen) were removed, weighed as quickly as possible, minced and then suspended with 3 volumes of an ice-cold medium containing 17O mM NaCl, 3 mM KCI, IO mM Na2HP~4.2H2O and 2 mM KH2PO4 adjusted to pH 7.4. The suspension was homogenized in an Ultra Turrax homogenizer at 30% f u l l power, using 4x15 sec strokes. In the other groups, where tissue Content of lipids was measured, tissue homogenates were obtained from the pooled organs of 5 animals. Determination of CI and DMCI. The method has been described previously (Sgaragli et a l . , ]983).
Pharmacological Research Communications, Vol. 16, No. 2 , 1984
209
Determination of ] i p i d s . Lipids were extracted from lung, liver, kidney, spleen and heart homogenates (4.5 ml) by the method of Folch et al. (1957). The extracted lipids (100-400 mg) were loaded on s i l i c i c acid columns (12 mm internal diameter, 120 mm height)and fractionated into neutral lipids and phospholipids by elution with lO0 ml chloroform and methanol, respectively. The total phospholipids were determined by multiplying x 25 the phospholipidphosphorus assayed following the method of Martin and Doty (1949) after digestion with sulfuric acid-perchloric acid 3:2. The colorimetric method of Cramer and Isaksson (1959) was used to determine total cholesterol in the neutral l i pid fraction. Total glycerides were determined by assaying glycerol in the neutral lipid fraction according to thR procedure of Carlson (1963). Statistical methods. The difference between means was analyzed by the Student's two-tailed t-test for independent samples (not significant, P~O.05).
RESULTS AND DISCUSSION Tissue levels of Cl and DMCI were measured 24 h after the administration of a single dose of CI. At this time tissue levels of both compounds had entered the phase of slow decay, thus showing much less v a r i a b i l i t y than those measured at earlier times (Della Corte et a l . , 1981). Results are shown in table I. In all analyzed tissues most of the drug was detected as its N-demethyl-derivative. ried
The DMCI/CI ratio, in fact, was 4 in plasma and var-
between 1.5 in l i v e r and 8 in the other analyzed tissues.
CI and
DMCI were preferentially accumulated by lungs reaching levels of 99 and 198 times higher than those found in plasma, respectively.
Am-
ong other tissues, liver preferentially accumulated CI while spleen and kidney exhibited a relatively high capacity for storing both compounds. In order to shed some light on the mechanism of Cl and DMCi preferential accumulation by lung, tissue contents of various l i p i d classes were also measured (see table l ) . The organ most rich in l i pids was kidney followed by lung> liver'7 heart and spleen.
Total
phospholipids were present in the highest amount in kidney followed by l i v e r > heart ~ lung and spleen. Glycerides were most abundant in
DMCI
8.97 +2.88 4.41 +0.67 26.7 +5.7
1.20 +0.22
2.78 +0.46
3.60 +0.90
36.4+3.8 .
45.4+3.0 .
20.9+I.I .
41.4+4.1
26.7+1.1
total lipid
19.6+4.0 .
31.2+3.0 .
14.5+I.6 .
26.6+3.5
18.3+2.2
.
.
.
7.8+0.3
6.6+0.5
1.4+0.2
6.0+0.9
3.4+0.2
.
.
.
I. I P I D total phospholipid glycerides a
3.2+0.9
3.3+0.5
2.4+0.3
4.0+0.2
1.6+0.2
total cholesterol
CI and DMCI f i g u r e s r e p r e s e n t mean values (~ s . e . ) d e r i v e d from 5 a n i m a l s , L i p i d f i g u r e s r e p r e s e n t mean values (~ s . e . ) of five_ pooled homogenates each d e r i v e d from -I f i v e t r e a t e d a n i m a l s ; they are expressed as mg.g wet t i s s u e except a, expressed as ~moles.g wet t i s s u e . T o t a l l i p i d values were determined g r a v i m e t r i c a l l y on p o r t i o n s of the l i p i d e x t r a c t s evaporated at room t e m p e r a t u r e under N2 stream.
Lung
Liver
Spleen
7.70 +2.98
0.92 +0.17
2.35 +0.67
0.32 +0.07
Heart
Kidney
0.140 +0.031
0.037 +0.062
(pg.g-I wet tissue)
CI
Rat tissue levels of C], D~CI and l i p i d s 24 h a f t e r a single oral dose of CI (90 mg.Kg b.w.)
Plasma
Tissue
Table I .
a
{a {b
(:
0
Pharmacological Research Communications, Vol. 16, No. 2, 1984
lung followed by kidney ~ h~art"> l i v e r and spleen. Total cholesterol was most abundant in kidney and lung. The relationship between CI and DMCI levels and tissue lipid contents was studied. The correlation between tissue levels of CI and DMCI and of l i p i d classes was not significant. The observation that rat lung has the highest capacity for storing CI and DMCI is in agreement with data reported on the tissue distribution of imipramine (Bickel
and Weder, 196B) and chlorproma-
zine (Wechsler and Roizin, 1960). Junod (1972) observed that, in isolated and perfused rat lung, the accumulation of imipramine had the characteristic of a saturable phenomenon, was insensitive to temperature
or Na+ concentration changes, and was antagonized by chlor-
promazine. Orton and coworkers (1973) also observed that amines like amphetamine and imipramine were rapidly concentrated in the lungs and, at least in the case of imipramine, the process was saturable. Only a minimal fraction of the accumulated imipramine could be recovered in the surfactant and in alveolar macrophages. As outlined by Junod (1976), one remarkable feature of lung anatomy is its huge capillary surface, thus the concentration of these drugs in this organ could be explained in terms of a simple binding to cell membranes starting from a purely quantitative basis of membrane surface oistribution. Binding of amphiphilic cationic drugs to cell membranes has been studied in a variety of cellular and subcellular preparations, namely red cells (Kwant and Seeman, 1969), sarcoplasmic ret'fculum (Balzer et al., 1968), liver mitochondria (Huunan-Sepp~l~, 1972), platelets (Boullin and O'Brien, 1968), brain synaptosomes (Weinstein et a l . , 1971), brain membranes (Raisman et a l . , 1979) and lung mitochondrial or microsomal preparations (Minkin et a l . , 1979). There have been a few constant findings in all these systems: the presence of binding sites with high substrate concentration (IO-4M), and a relative insensitivity to a decrease in temperature have been evidentiated. In conclusion, i t would appear from the results under discussion that the lipids here examined do not play an important role in the mechanism of CI and DMCI tissue accumulation. The observation that imi-
21 1
212
Pharmacological Research Communications, VoL 16. No. 2, 1984
pramine binding to rabbit skeletal muscle homogenate was unaffected by prior l i p i d extraction (Fichtl et a l . , 1980) is in agreement with our conclusion.
ACKNOWLEDGMENTS
This research project has been supported by CNR - Roma (contr. n. 82.02043.04.
Dr. I. Megazzini is gratefully aknowledged for her
assistance in the experimental work.
REFERENCES - Balzer H., Makin~se M., Fiehn W. and Hasselbach W., Naunyn-Schmiedeberg's Arch. Pharmak. Exp. Pathol., 260, 456 (1968). -
Bickel M.H. and Steele J.W., Chem. Biol. Interact., _8, 151 (1974). Bicke] M.H. and Weder H.J., Arch. Int. Pharmacodyn., 17.3, 433 (1968).
- Bickel M.H. and Weder H.J., Psychopharmac. Co~un., _2, 231 (1976). - BirkeLt D.J., Clin. Exp. Pharmac., I_, 415 (1974). -
Boullin D.J. and O'Brien R.A., J. Pharm. Pharn,ac., 2__00,583 (1968).
- Carlsen L.A.,J. Atheroscler. Res., 3_, 334 (1963). - Cater B.R., Chapman D., Hawes S.M. and Saville J.,Biochim. Biophys. Acta, 363, 54 (1974). - Cramer K. and Isaksson B., Scand. J. Clin.
Invest. I I ,
213 (1959).
- Della Corte L., Giovannini M.G., Martini P., Megazzini I . ,
Puliti
R.,
Sgaragli G.P. and Wondrak G., Br. J. Pharmac., 72, 184P (1981).
- Di Francesco C. and Bickel M.H., Chem. Biol. Interact., 7_66,335 (1977). - Dittmer J.C. and Lester R.L., J. Lipid Res., --5, 126 (1964). - Fichtl B., Bondy B. and Kurz H., J. Pharmac. Exp. Ther., 215, 248 (1980). - Folch T., Lees M. and Sloane-Stanley G.M., J. Biol. Chem., 22___66,497 (1957). - Gillette J.R., in Drugs and Enzymes, ed. Brodie B.B. and Gillette J.R., p. 9, Pergamon Press, Oxford (1965). -
-
Huunan-Sepp~'l~" A., Acta Chem. Scand., 266, 2713 (1972). Junod A., Pharmac. Thero B, 2, 511 (1976).
Pharmacological Research Communications, Vol. 16, No. 2, 1984
-
Kwant W.O. and Seeman P., Biochim. Biophys. Acta, 18_3, 530 (1969).
- Leterrier F., Mendyk A. and Viret J., Biochem. Pharmac., 2_55, 2469 (1976). - Martin J.B. and Doty B.M., Anal. Chem., 21, 965 (1949). - Minkin R.F., l l e t t K.F. and Madsen B.W., Biochem. Pharmac., 23, 2275 (1979). - Orton T.C., Anderson M.V., Pickett R.D., Eling T.E. and Fouts J.R., J. Pharmac. Exp. Ther., 186, 482 (1973). -
Raisman R., Briley M. and Langer S.Z., Nature, 281, 148 (1979).
-
Sgaragli G.P., Della Corte L. and Gremigni D., Pharmac. Res. Comm., ~5, 231 (1983).
- Wechsler M.B. and Roizin L., J. Ment. Sci., 106, 1501 (1960). -
Weinstein H., Varon S. and Roberts E., Biochem. Pharmac., 20, 103 (1971).
2 1 3