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Dopamine antagonist binding: A significant decrease with morphine dependence in the rat striatum
Life Sciences, Vol . 23, pp . 637-642 Printed in the U .S .A .
Pergamon Prese
DOPAMI ;IE ANTAGONIST BINDING : A SIGNIFICANT DECREASE WITH MORPHINE DEPENDENCE IN THE RAT STRIATUM Surendra K . Puri, Theodore C . Spaulding and Charles R . Mantione Hoechst-Roussel Pharmaceuticals Inc ., Route 202-206 North, Somerville, N .J . 08876
Summa~ (sH)-Spiroperidol specific binding was determined in striatal tissue of rats which received a single dose of, or made dependent on morphine . Acute morphine (30 mg/kg i .p .) did not alter (3H) spiroperidol specific binding . However, morphine-dependent rats with two 50 mg pellets when withdrawn for 24 or 48 hours, significantly decreased the binding and increased Kd . Binding sites were reduced with a decrease in Kd in rats implanted with four 50 mg pellets or receiving high doses of morphine . These results indicate that binding characteristics of (sH)-spiroperidol depend on the relative dose of morphine used to induce dependence . Low dose dependence (2 pellets) results in a decrease in binding affinity while high dose dependence (4 pellets or chronic injection) results in an increase of ( 3 H)-spiroperidol affinity in the presence of fewer binding sites . Chronic morphine administration induces supersensitivity to some of the behavioral and biochemical effects of dopamine receptor stimulation . Apomorphine, a dopamine agonist, was more potent in inducing aggression (1), in enhancing stereotypy behavior (2), jumping behavior (3) and in decreasing dopamine turnover in morphine withdrawn rats (4,5) . Enhanced sensitivity to L-DOPA (6) and rotation behavior during naloxone precipitated abstinence after alteration of dopaminergic activity(7,8) have also been reported . These results suggest that chronic administration of morphine leads to the supersensitivity of dopamine receptors . However, contradiction exists to this hypothesis . Apomorphine was less potent in inducing stereotyped behavior (4) and in increasing striatal cyclic adenosine 3'5' monophosphate (5) in morphine withdrawn rats . Although basal adenylate cyclase activity is higher, dopamine sensitivity was decreased by chronic mo rphine administration (9,10) . In the present experiments, we utilized the ( H)-spiroperidol binding assay to test the changes in the sensitivity of dopamine receptors in morphine dependent animals . Methods Male Sprague Dawley rats (180-270 gms) were made dependent by implantation of 50 mg morphine base pellets or by chronic injection of rtarphine sulfate . Either two or four pellets were implanted subcutaneously for 72 hours prior to their removal . Controls were implanted with placebo pellets . In another study morphine sulfate was injected i .p . 3 times a day in dosing increments of 30 mg/ kg/day until a dose of 300 mg/kg/day was reached (9) . The dependent animals were divided into three groups based on the schedule of morphine treatment . Schedule I - 2 pellets, Schedule II - 4 pellets, Schedule III - i .p . injectionas described above . Weight loss during spontaneous withdrawal or the obser0300-9653/78/0814-0637$02 .00/0 Copyright (c) 1978 Pergamon Preea
63 8
DA Receptors and Morphine Dependence
Vol . 23, No . 6, 1978
vance of other abstinence signs (11) were used to assess dependence . Abstinence was monitored 24 and 48 hours after withdrawal . Control rats gained weight after the last treatment or pellet removal, while rats implanted with 2 or 4 morphine pellets, or chronically injected with morphine, lost a significant amount of weight . Further, in rats with morphine-pellet or injected chronically, withdrawal characteristics such as hyperirritability, gnawing, escape attempts and wet dog shakes was observed . (sH)-Spiroperidol specific binding assay procedure was carried out in control and 24 or 48 hours morphine-withdrawn rats . The (sH)-spiroperidol binding was basically carried out as described by Fields et al (12) and Creese et al (13) . Specific (aH)-spiroperidol binding was detérmined as the differencebeween total binding and nonspecific binding obtained in absence and presence of 10 uM d-butaclamol, and expressed as pmoles of (sH)-spiroperidol bound per mg protein or g of wet tissue . Protein was determined by the method of Bradford (14) . Results ( 3 H)-Spiroperidol specific binding, in vitro , was determined in rat striata one hour after morphine sulfate (30 mg/kg, i .p .) or haloperidol (5 mg/kg i .p .) administration . Morphine sulfate did not affect binding, while haloperidol produced about 56% decrease (Table I) . These results suggest that unlike haloperidol, morphine does not compete for dopamine receptor sites . Table I Effect of Acute Administration of Morphine or Haloperidol on (s H)-Spiroperidol Specific Binding in the Rat Striatum Treatment (mg/kg i .p .)
Pretreatment Interval (min)
Saline
(sH)-Spiroperidol Binding mean (pmoles/G of tissue) ± S .D . (N)
60
21 .45 + 2 .74 (4)
Morphine (30)
60
24 .21 ± 3 .75 (4)
Haloperidol
60
(5)
9 .56 ± 0 .83 (4) I
. 1 Significantly different fromcontrôl~P<0 .05~ Animals were made dependent by three different schedules of morphine treatment and were depelleted or withdrawn for 24 or 48 hours before binding assays . Rats treated on Schedule I (2 pellets) showed a significant decrease in the binding of ( 3 H)-spiroperidol, in 24 and 48 hours withdrawn rats when low ligand concentrations (0 .4 and 0 .8 nM) were used . At high ligand concentrations (1 .6 and 3 .2 nM), the binding in morphine-dependent animals was not significantly different from control animals (Table II) . These results show that the affinity of dopamine receptors is decreased after morphine withdrawal in dependent rats . In rats made dependent by Schedule II and III treatments, (sH)-spiroperidol binding was significantly reduced in withdrawn groups with higher ligand concentrations but the binding with lower concentrations was not affected (Table III and IV) . These results indicate that high dose (4 pellets) or chronic administration of morphine significantly decrease the number of binding sites . From the data, binding constants (Ko , nM ; Rp , fmoles/mg protein) were determined by Scatchard analysis . There was a significant increase (three fold) of
Vol . 23, No . 6, 1978
DA Receptors and Morphine Dependence
639
apparent Kd values in Schedule I treated groups in 24 as well as 48 hours after withdrawal along with a slight increase in total binding density (Kd , control 0 .58 vs . 1 .75 and 1 .37 in morphine-withdrawn ; Ro , control - 844 vs . 1050 and 990 in morphine-withdrawn) . In Schedule II . treated 24 hrs withdrawn rats, there was not only a reduction in apparent Kd (control 0 .71 ; 24 hr 0 .48) but also a decrease in receptor density (control 763 ; 24 hr 548) . However, in 48 hours withdrawn rats, the apparent Kd value returned to control levels (control 0 .71 ; 24 hr 0.71) and the total number of binding sites showed a trend towards normality (control 763 ; 24 hr 678), Schedule III treated rats withdrawn for 48 hours showed a decrease in apparent Kd (control 0 .71 ; 48 hr 0 .63) and receptor density (control 763 ; 48 hr 602) . Table II Striatal (sH)-Spiroperidol Binding During Withdrawal in Rats Made Dependent on Morphine by Schedule I Treatmentl Ligand Concentration . (nM) 0 .4 0 .8
1 .6 3,2
(aH)-Spiroperidol Binding Mean (fmoles/mg Protein) ± S.D . (N) Control
406 _+ 9 (5)
504 _+ 41 (5) 593 _+ 45 (5) 771 + 76 (5)
Morphine Withdrawn (hrs) 24 hrs 48 hrs
222 _+ 35 (5) 2
208 _+ 17 (5) 2
416 _+ 20 (5) 2 639 + 20 (5) 750 + 19 (5)
413 _+ 39 (5) 543 _+ 57 (5) 641 ± 6 (5)
1 . 2 pellets = 100 mg morphine base for 3 days . 2 . Significantly different from control (P<0 .05) Table III
Striatal (sH)-Spiroperidol Binding During Withdrawal in Rats Made Dependent on Morphine by Schedule II Treatmentl Ligand Concentration (nM) 0 .2 0 .4 0.8 1 .6
(sH)-Spiroperidol Binding Mean (fmoles/mg Protein) + S .D . (N) Control 158 + 6 (5)
292 + 15 (5) 423 + 10 (5) 489 ± 28 (5)
Mor ine 2 rs 157 + 3 (5) 258 + (5) 360 + 20 (5) 2 398 + 26 (5)2
it
raven
1. 4 péllets = 200 mg for 3 days 2 . Significantly different from control group (P<0 .05)
rs 8
rs
141 ± 3 (5)
261 ± 11 (5)
373 ± 13 (5) 2 410 ± 33 (5) 2
Discussion Both behavioral and neurochemical finding previously reported (1,4,15) suggest that acute effect of morphine resembles the dopamine blocking action of haloperidol . These studies suggested that morphine may inhibit dopamine neurotransmission . Consequently, continuous blockade of dopaminergic neurotransmission during chronic administration of morphine may cause supersensitivity of dopamine receptors . Therefore, in this study, we investigated the character-
64 0
DA Receptors and Morphine Dependence
Vol . 23, No . 6, 1978
istics of dopamine receptors after acute and chronic morphine administration using (sH)-spiroperidol specific binding procedure . Although, a single injection of haloperidol significantly decreased (sH)-spiroperidol specific binding, morphine even in a high dose (30 mg/k ) did not affect spiroperidol binding . In vitro addition of morphine (0 .1 mM~ also did not affect (sH)-spiroperidol bin is ng. Thts suggests that morphine does not directly affect dopamine receptors in the striatum . Our observation is supported by the fact that dopaminesensitive adenylate which is used as a model for dopamine receptor function is not affected by in vitro addition of morphine (16) . Also, in vivo administration of morphinë produces a dose-dependent increase in adenylâte cyclase activity without affecting dopamine sensitivity (9,17,18) . Therefore, the dopamine deficiency syndrome one sees with acute morphine may be due to other than dopamine-receptor blockade . Table IV Striatal (sH)-Spiroperidol Binding During Withdrawal in R~ts Made Dependent on Morphine by Schedule III Treatment Ligand Concentration (nM) 0 .4
0 .8 1 .6 3 .2
( 3 H)-Spiroperidol Binding Mean (fmoles/mg Protein) ± S.D . (N)
Contro 271 _+ 8 (5) 406 + 11 (5)
518 + 26 (5) 614 ± 35 (5)
Mor hine-W thdrawn 234 _+ 7 (5)
rs
374 + 7 (5)
448 ± 6 (5) 2
470 ± 12 (5)2
T. Totaldösé = 2 .~5 g`kg ôvér T2dâys 2. Significantly different from control group (P<0 .05)
In this study, we utilized three different schedules of treatment to induce physical dependence . These treatment represent minimal (2 pellets), intermediate (4 pellets), and maximal (chronic administration) levels of dependence . At minimal dependence, there was a decrease in the dopamine receptor affinity as suggested by a 3 fold increase in apparent Kd with a slight increase in number of binding sites . However, intermediate and maximal levels of dependence resulted in a significant decrease in the number of binding sites . This suggests that morphine-dependence may significantly reduce the Striatal dopamine receptor population . The changes observed in the binding characteristics of ( 3H)-spiroperidol in morphine-dependent rats are due to chronic administration of morphine rather than the presence of residual morphine . This conclusion is supported by the observation that in vitro addition of morphine to the membrane fraction obtained from withdrawn animals did not affect spiroperidol specific binding . An interesting observation was seen in rats implanted with 4 pellets . The apparent Kd was reduced in 24 hours withdrawn rats, but returned to control levels within 48 hours . The receptor density showed a similar trend . In maximally dependent rats, there was an increased affinity (decreased Kd) and a marked reduction in receptor population even after 48 hours of withdrawal . The apparent change in the receptor affinity in morphine dependent animals may explain the behavioral supersensitivity to dopamine agonists . Although the ( 3 H)-dihydroxmorphine specific binding is not altered in whole brain (19), a significant increase of Striatal binding has been reported (2) . Therefore, the changes in dopamine receptor activity observed in morphine dependent animals reflect the changes in other neuronal activity such as opiate
Vol . 23, No . 6, 1978
DA Receptors and Morphine Dependence
641
pathways . The paradoxical effects of an increase in dopamine affinity with simultaneous decrease in binding sites after morphine dependence warrant further experiments . REFERENCES 1. 2. 3. 4. 5. 6. 7. 8. 9. 10 . 11 . 12 . 13 . 14 . 15 . 16 . 17 . 18 . 19 . 20 .
S .K . PURI and H . LAL, Psychopharmacologia 32 : 113-120 (1973) B . COX, M . ARY and P . LOMAX, J . PHARMAC . EXP . THERAP . 196 : 637-641 (1976) . A . HERZ, J . BLASIG and R . PAPESCHI, Psychopharmacoloqia39 : 121-143 (1974) . K . KUSCHINSKY, Psychopharmacologia 42 : 225-229 (1974) . S .K . PURI, L . VOLICER and H . LAL, Neûropharmacology _16 : 205-209 (1977) . E . EIDELBERG and E . ERSPAMER, J . Pharmaco . Exp . Ther 192 : 50-57 (1975) . B . HINE, E . FRIEDMAN, M . TORRELIO and S . GERSHON, Neuröp~armacology 14 : 607-610 (1975) . J .M . MORIHISA and S .D . GLICK, Brain Res . 123 : 180-187 (1977) . S .K . PURI, L . VOLICER and J . COCHIN, J . Neûrochem . 27 : 1551-1554 (1976) . Z . MERAKI, R .L . SINGHAL, P .D . HRDINA, G .M . LING, Life Sci . _16 : 1889-1894 (1975) . W .L . DEWEY, G .A . PATRICK and L .S . HARRIS, Ço mm . on Prob . of- Dru~ p . 729-772 (1975) . J .M . FIELDS, T .D . REISINE AND H .I . YAMAMURA, Brain Res . 136 : 578-584 (1977) . J . F . LEYSEN, W . GOMMEREN and P .M . LADURON, Biochem . Phârmacol . _27 : 307-316 (1978) . M .M . BRADFORD, Anal . Biochem . 72 : 248-250 (1976) . H . LAL, G . GIANUTSOS and S .K . PÜRI, Life Sci . 17 : 29-34 (1975) . S . K . PURI, J . COCHIN and L . VOLIGER, Life Sci .~f6 : 759-768 (1975) . D .H . CLOUET, G .J .GOLD and K . IWATSUBO, Br . J . P~rmaco . 54 : 541-548 (1975) . K . IWATSUBO and D .H . CLOUET, Biochem . Pharmaco . 24 : 1499-1503 (1974) . W .A . KLEE AND R .S . STREATY, Nature 248 : (5443) 663 (1974) . W .L . DEWEY and T .T . CHAU-PHAM ., 0 fates and Endo enous 0 ioid Pe tides . p . 403-406 . Elsevier/Nort h Holland Biomedical Press, Amsterdam 1 6) .
Pergamon Prese
DOPAMI ;IE ANTAGONIST BINDING : A SIGNIFICANT DECREASE WITH MORPHINE DEPENDENCE IN THE RAT STRIATUM Surendra K . Puri, Theodore C . Spaulding and Charles R . Mantione Hoechst-Roussel Pharmaceuticals Inc ., Route 202-206 North, Somerville, N .J . 08876
Summa~ (sH)-Spiroperidol specific binding was determined in striatal tissue of rats which received a single dose of, or made dependent on morphine . Acute morphine (30 mg/kg i .p .) did not alter (3H) spiroperidol specific binding . However, morphine-dependent rats with two 50 mg pellets when withdrawn for 24 or 48 hours, significantly decreased the binding and increased Kd . Binding sites were reduced with a decrease in Kd in rats implanted with four 50 mg pellets or receiving high doses of morphine . These results indicate that binding characteristics of (sH)-spiroperidol depend on the relative dose of morphine used to induce dependence . Low dose dependence (2 pellets) results in a decrease in binding affinity while high dose dependence (4 pellets or chronic injection) results in an increase of ( 3 H)-spiroperidol affinity in the presence of fewer binding sites . Chronic morphine administration induces supersensitivity to some of the behavioral and biochemical effects of dopamine receptor stimulation . Apomorphine, a dopamine agonist, was more potent in inducing aggression (1), in enhancing stereotypy behavior (2), jumping behavior (3) and in decreasing dopamine turnover in morphine withdrawn rats (4,5) . Enhanced sensitivity to L-DOPA (6) and rotation behavior during naloxone precipitated abstinence after alteration of dopaminergic activity(7,8) have also been reported . These results suggest that chronic administration of morphine leads to the supersensitivity of dopamine receptors . However, contradiction exists to this hypothesis . Apomorphine was less potent in inducing stereotyped behavior (4) and in increasing striatal cyclic adenosine 3'5' monophosphate (5) in morphine withdrawn rats . Although basal adenylate cyclase activity is higher, dopamine sensitivity was decreased by chronic mo rphine administration (9,10) . In the present experiments, we utilized the ( H)-spiroperidol binding assay to test the changes in the sensitivity of dopamine receptors in morphine dependent animals . Methods Male Sprague Dawley rats (180-270 gms) were made dependent by implantation of 50 mg morphine base pellets or by chronic injection of rtarphine sulfate . Either two or four pellets were implanted subcutaneously for 72 hours prior to their removal . Controls were implanted with placebo pellets . In another study morphine sulfate was injected i .p . 3 times a day in dosing increments of 30 mg/ kg/day until a dose of 300 mg/kg/day was reached (9) . The dependent animals were divided into three groups based on the schedule of morphine treatment . Schedule I - 2 pellets, Schedule II - 4 pellets, Schedule III - i .p . injectionas described above . Weight loss during spontaneous withdrawal or the obser0300-9653/78/0814-0637$02 .00/0 Copyright (c) 1978 Pergamon Preea
63 8
DA Receptors and Morphine Dependence
Vol . 23, No . 6, 1978
vance of other abstinence signs (11) were used to assess dependence . Abstinence was monitored 24 and 48 hours after withdrawal . Control rats gained weight after the last treatment or pellet removal, while rats implanted with 2 or 4 morphine pellets, or chronically injected with morphine, lost a significant amount of weight . Further, in rats with morphine-pellet or injected chronically, withdrawal characteristics such as hyperirritability, gnawing, escape attempts and wet dog shakes was observed . (sH)-Spiroperidol specific binding assay procedure was carried out in control and 24 or 48 hours morphine-withdrawn rats . The (sH)-spiroperidol binding was basically carried out as described by Fields et al (12) and Creese et al (13) . Specific (aH)-spiroperidol binding was detérmined as the differencebeween total binding and nonspecific binding obtained in absence and presence of 10 uM d-butaclamol, and expressed as pmoles of (sH)-spiroperidol bound per mg protein or g of wet tissue . Protein was determined by the method of Bradford (14) . Results ( 3 H)-Spiroperidol specific binding, in vitro , was determined in rat striata one hour after morphine sulfate (30 mg/kg, i .p .) or haloperidol (5 mg/kg i .p .) administration . Morphine sulfate did not affect binding, while haloperidol produced about 56% decrease (Table I) . These results suggest that unlike haloperidol, morphine does not compete for dopamine receptor sites . Table I Effect of Acute Administration of Morphine or Haloperidol on (s H)-Spiroperidol Specific Binding in the Rat Striatum Treatment (mg/kg i .p .)
Pretreatment Interval (min)
Saline
(sH)-Spiroperidol Binding mean (pmoles/G of tissue) ± S .D . (N)
60
21 .45 + 2 .74 (4)
Morphine (30)
60
24 .21 ± 3 .75 (4)
Haloperidol
60
(5)
9 .56 ± 0 .83 (4) I
. 1 Significantly different fromcontrôl~P<0 .05~ Animals were made dependent by three different schedules of morphine treatment and were depelleted or withdrawn for 24 or 48 hours before binding assays . Rats treated on Schedule I (2 pellets) showed a significant decrease in the binding of ( 3 H)-spiroperidol, in 24 and 48 hours withdrawn rats when low ligand concentrations (0 .4 and 0 .8 nM) were used . At high ligand concentrations (1 .6 and 3 .2 nM), the binding in morphine-dependent animals was not significantly different from control animals (Table II) . These results show that the affinity of dopamine receptors is decreased after morphine withdrawal in dependent rats . In rats made dependent by Schedule II and III treatments, (sH)-spiroperidol binding was significantly reduced in withdrawn groups with higher ligand concentrations but the binding with lower concentrations was not affected (Table III and IV) . These results indicate that high dose (4 pellets) or chronic administration of morphine significantly decrease the number of binding sites . From the data, binding constants (Ko , nM ; Rp , fmoles/mg protein) were determined by Scatchard analysis . There was a significant increase (three fold) of
Vol . 23, No . 6, 1978
DA Receptors and Morphine Dependence
639
apparent Kd values in Schedule I treated groups in 24 as well as 48 hours after withdrawal along with a slight increase in total binding density (Kd , control 0 .58 vs . 1 .75 and 1 .37 in morphine-withdrawn ; Ro , control - 844 vs . 1050 and 990 in morphine-withdrawn) . In Schedule II . treated 24 hrs withdrawn rats, there was not only a reduction in apparent Kd (control 0 .71 ; 24 hr 0 .48) but also a decrease in receptor density (control 763 ; 24 hr 548) . However, in 48 hours withdrawn rats, the apparent Kd value returned to control levels (control 0 .71 ; 24 hr 0.71) and the total number of binding sites showed a trend towards normality (control 763 ; 24 hr 678), Schedule III treated rats withdrawn for 48 hours showed a decrease in apparent Kd (control 0 .71 ; 48 hr 0 .63) and receptor density (control 763 ; 48 hr 602) . Table II Striatal (sH)-Spiroperidol Binding During Withdrawal in Rats Made Dependent on Morphine by Schedule I Treatmentl Ligand Concentration . (nM) 0 .4 0 .8
1 .6 3,2
(aH)-Spiroperidol Binding Mean (fmoles/mg Protein) ± S.D . (N) Control
406 _+ 9 (5)
504 _+ 41 (5) 593 _+ 45 (5) 771 + 76 (5)
Morphine Withdrawn (hrs) 24 hrs 48 hrs
222 _+ 35 (5) 2
208 _+ 17 (5) 2
416 _+ 20 (5) 2 639 + 20 (5) 750 + 19 (5)
413 _+ 39 (5) 543 _+ 57 (5) 641 ± 6 (5)
1 . 2 pellets = 100 mg morphine base for 3 days . 2 . Significantly different from control (P<0 .05) Table III
Striatal (sH)-Spiroperidol Binding During Withdrawal in Rats Made Dependent on Morphine by Schedule II Treatmentl Ligand Concentration (nM) 0 .2 0 .4 0.8 1 .6
(sH)-Spiroperidol Binding Mean (fmoles/mg Protein) + S .D . (N) Control 158 + 6 (5)
292 + 15 (5) 423 + 10 (5) 489 ± 28 (5)
Mor ine 2 rs 157 + 3 (5) 258 + (5) 360 + 20 (5) 2 398 + 26 (5)2
it
raven
1. 4 péllets = 200 mg for 3 days 2 . Significantly different from control group (P<0 .05)
rs 8
rs
141 ± 3 (5)
261 ± 11 (5)
373 ± 13 (5) 2 410 ± 33 (5) 2
Discussion Both behavioral and neurochemical finding previously reported (1,4,15) suggest that acute effect of morphine resembles the dopamine blocking action of haloperidol . These studies suggested that morphine may inhibit dopamine neurotransmission . Consequently, continuous blockade of dopaminergic neurotransmission during chronic administration of morphine may cause supersensitivity of dopamine receptors . Therefore, in this study, we investigated the character-
64 0
DA Receptors and Morphine Dependence
Vol . 23, No . 6, 1978
istics of dopamine receptors after acute and chronic morphine administration using (sH)-spiroperidol specific binding procedure . Although, a single injection of haloperidol significantly decreased (sH)-spiroperidol specific binding, morphine even in a high dose (30 mg/k ) did not affect spiroperidol binding . In vitro addition of morphine (0 .1 mM~ also did not affect (sH)-spiroperidol bin is ng. Thts suggests that morphine does not directly affect dopamine receptors in the striatum . Our observation is supported by the fact that dopaminesensitive adenylate which is used as a model for dopamine receptor function is not affected by in vitro addition of morphine (16) . Also, in vivo administration of morphinë produces a dose-dependent increase in adenylâte cyclase activity without affecting dopamine sensitivity (9,17,18) . Therefore, the dopamine deficiency syndrome one sees with acute morphine may be due to other than dopamine-receptor blockade . Table IV Striatal (sH)-Spiroperidol Binding During Withdrawal in R~ts Made Dependent on Morphine by Schedule III Treatment Ligand Concentration (nM) 0 .4
0 .8 1 .6 3 .2
( 3 H)-Spiroperidol Binding Mean (fmoles/mg Protein) ± S.D . (N)
Contro 271 _+ 8 (5) 406 + 11 (5)
518 + 26 (5) 614 ± 35 (5)
Mor hine-W thdrawn 234 _+ 7 (5)
rs
374 + 7 (5)
448 ± 6 (5) 2
470 ± 12 (5)2
T. Totaldösé = 2 .~5 g`kg ôvér T2dâys 2. Significantly different from control group (P<0 .05)
In this study, we utilized three different schedules of treatment to induce physical dependence . These treatment represent minimal (2 pellets), intermediate (4 pellets), and maximal (chronic administration) levels of dependence . At minimal dependence, there was a decrease in the dopamine receptor affinity as suggested by a 3 fold increase in apparent Kd with a slight increase in number of binding sites . However, intermediate and maximal levels of dependence resulted in a significant decrease in the number of binding sites . This suggests that morphine-dependence may significantly reduce the Striatal dopamine receptor population . The changes observed in the binding characteristics of ( 3H)-spiroperidol in morphine-dependent rats are due to chronic administration of morphine rather than the presence of residual morphine . This conclusion is supported by the observation that in vitro addition of morphine to the membrane fraction obtained from withdrawn animals did not affect spiroperidol specific binding . An interesting observation was seen in rats implanted with 4 pellets . The apparent Kd was reduced in 24 hours withdrawn rats, but returned to control levels within 48 hours . The receptor density showed a similar trend . In maximally dependent rats, there was an increased affinity (decreased Kd) and a marked reduction in receptor population even after 48 hours of withdrawal . The apparent change in the receptor affinity in morphine dependent animals may explain the behavioral supersensitivity to dopamine agonists . Although the ( 3 H)-dihydroxmorphine specific binding is not altered in whole brain (19), a significant increase of Striatal binding has been reported (2) . Therefore, the changes in dopamine receptor activity observed in morphine dependent animals reflect the changes in other neuronal activity such as opiate
Vol . 23, No . 6, 1978
DA Receptors and Morphine Dependence
641
pathways . The paradoxical effects of an increase in dopamine affinity with simultaneous decrease in binding sites after morphine dependence warrant further experiments . REFERENCES 1. 2. 3. 4. 5. 6. 7. 8. 9. 10 . 11 . 12 . 13 . 14 . 15 . 16 . 17 . 18 . 19 . 20 .
S .K . PURI and H . LAL, Psychopharmacologia 32 : 113-120 (1973) B . COX, M . ARY and P . LOMAX, J . PHARMAC . EXP . THERAP . 196 : 637-641 (1976) . A . HERZ, J . BLASIG and R . PAPESCHI, Psychopharmacoloqia39 : 121-143 (1974) . K . KUSCHINSKY, Psychopharmacologia 42 : 225-229 (1974) . S .K . PURI, L . VOLICER and H . LAL, Neûropharmacology _16 : 205-209 (1977) . E . EIDELBERG and E . ERSPAMER, J . Pharmaco . Exp . Ther 192 : 50-57 (1975) . B . HINE, E . FRIEDMAN, M . TORRELIO and S . GERSHON, Neuröp~armacology 14 : 607-610 (1975) . J .M . MORIHISA and S .D . GLICK, Brain Res . 123 : 180-187 (1977) . S .K . PURI, L . VOLICER and J . COCHIN, J . Neûrochem . 27 : 1551-1554 (1976) . Z . MERAKI, R .L . SINGHAL, P .D . HRDINA, G .M . LING, Life Sci . _16 : 1889-1894 (1975) . W .L . DEWEY, G .A . PATRICK and L .S . HARRIS, Ço mm . on Prob . of- Dru~ p . 729-772 (1975) . J .M . FIELDS, T .D . REISINE AND H .I . YAMAMURA, Brain Res . 136 : 578-584 (1977) . J . F . LEYSEN, W . GOMMEREN and P .M . LADURON, Biochem . Phârmacol . _27 : 307-316 (1978) . M .M . BRADFORD, Anal . Biochem . 72 : 248-250 (1976) . H . LAL, G . GIANUTSOS and S .K . PÜRI, Life Sci . 17 : 29-34 (1975) . S . K . PURI, J . COCHIN and L . VOLIGER, Life Sci .~f6 : 759-768 (1975) . D .H . CLOUET, G .J .GOLD and K . IWATSUBO, Br . J . P~rmaco . 54 : 541-548 (1975) . K . IWATSUBO and D .H . CLOUET, Biochem . Pharmaco . 24 : 1499-1503 (1974) . W .A . KLEE AND R .S . STREATY, Nature 248 : (5443) 663 (1974) . W .L . DEWEY and T .T . CHAU-PHAM ., 0 fates and Endo enous 0 ioid Pe tides . p . 403-406 . Elsevier/Nort h Holland Biomedical Press, Amsterdam 1 6) .