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Circling of mice bearing unilateral striatal lesions: Development of increased response to d-amphetamine
Pergamon Press
Life Sciences Vol . 21, pp " 563-568 Printed in the II .S .A.
CIRCLING OF MICE BEARING IINIIATERAL 3TRIATAL LESIONS : DEVELOPMEIPP OF INCRFASED RESPONSE TO DAMPBETAMINE Stanley D. 8chols Marten-National Laboratories Division of Richardson-Marten Inc . Cincinnati, Ohio 45215 (Received in final form July 14, 1977) SUNß~lARY Mice leaioned in one sidn of the striatum with 6-hydrorydopaminn respond to d-amphetamine by circling ipsivnreivaly. This report communicates the finding that upon repeated dosing with 4 mg/kg s .c . at weekly intervals the mice respond more and morn strongly . Conditioning appears to contribute little to this phenomenon . Mice unilaterally lesioned in the corpus striatum by way of micro-injected 6-hydroaydopamine respond to amphetamine and similar agents by circling towards the lnsioned sidn (1) . Ea initial ezparimenta with this model in our labora tory, which involved counting of the circling of such mice in response to a dose of d-amphetamine on too occasions, the mice consistently circled more on the second occasion than on the first . The present experiments were performed to dietiaguiah the effects of rnpeatnd exposure to amphetamine from the effects of time, which itself affects the sensitivity of mica leeipned ae these were (2) . MSfHODS Male Charles River CD-1 mice were anesthetized aith methoayflurane and leeioned according to the method of Von Voigtlandnr and Moors (1) . This method consists essentially of injecting 4 ~tl of as aqueous solution of 16 Leg of 6-hydröaydopamina HBr plus 0 .8 ~~g of ascorbic acid etsreotaaically into the caudate-putamen nucleus on one side . Nine days later, the mice ante injected with apomorphine HC1, 0 .2 mg/k8' s .c . The mica which circled contravereivelq ten times or more between too and four minutes after injection acre considered to be successfully leeionnd . Taenty such mice were used in this study. They were assigned to two groups of ten each eo that the two groups sere ss nearly matched as possible regarding their performance in the above teat with apomorphinn . Tao of the chronic amphetamine group died, so their antemortem data have been eu laded. Their lose did not materially lessen the match of the two groups . One group received d-amphetamine sulfate, 4 mg/kg e.c ., five times at weekly intervals . The other group of mice a~s iajectad with salins (10 ml/ kg e.c .) on the first four of the above injection days, and with amphetamine, 4 mg/kg s .c . on the fifth. Both groups then received a saline injection .on Day 5 of the fifth week . The experimental procedure consisted of placing each mouse in a clean glass bowl and attaching a harness made of a pipe cleaner. After at leset 563
564
Increased Response to Amphetamine
Vol . 21, No . 4, 1977
five minutes, the mouse was injected and replaced in the bowl . Circling ipsiversive and contravereive to the lesion were separately counted via automatic counters from minutes 2 through 30 after amphetamine . Following saline, circling was counted from minutes 2 through 30 on Week 1 and on Day 5 of the fifth week ; otheraiee, saline-treated mice were treated as above, exenpt that their harnesses warn not co~ected to the counters . Statistical analysis was by way of "Student's" t-test -- paired when the performance of a group of mice on two occasions was compared, and unpaired when one group of mica was compared with the other. RESULTS Figure 1 shoal the mean ipsiversive and contraveraive turns observed over minutes 2 - 30 after injection. Folloaring the first injection of saline, the mean résponses of the chronic saline group were 43 ipsiversive and 3 contra veraiva . The mean ipsiversive responses of the chronic amphetamine group were 140 on the first amphetamine day, 203 on the second, and 232 on the final amphetamine day. The responses of both the second and the last days exceeded that of the first at the P a .02 level ; those of the second and the last days did not differ at the P ~ .1 level . Upon receiving amphetamine, the chronic online group of mice responded with a mean of 139 ipsiversive turns : virtually equal to the response of the chronic amphetamine group to its first dose of amphetamine.
Ip~lwnlw 200 1J è 100 0
E
FIG . 1 . Total ipsiversive and contravnreiva turns over minutes 2 - 30 follaaing s .c . injection of d-amphetamine, 4 mg/kg (a) or saline (s) . The letters at the base of the ipsiversive bare indicate the compound injected . The white bars represent the chronic amphetamine group; the cross-hatched bare represent the chronic saline group . Statistical aignificances ; * P < .05 ** P < .02
Figure 2 shoos the time course of the turning on each amphetamine day. It ie evident that turning has a faster onset after repeated dosing with amphetamine than when initially given . As shown in Figure 1, after the amphetamine phase of the experiments was concluded, both groups of mica responded almost identically to saline . The ipsiversive turning of both groups was slightly less than the ipsiversive turning of the chronic saline group before exposure to amphetamine .
Vol . 21, No . 4, .1977
Increased Response to Amphetamine
565
FIG . 2 Time course of ipeiveraive turning following amphetamine injection . The solid line represents the chronic amphetamine group ; the. dotted line represents the chronic saline group .
DISCUSSION If total circling war the half-hour after dosing is used as a response measure, the response to a constant down of d-amphetamine increases with reThe virtual equality of the responses of the two groups peated administration . of mica to their respective first doses of amphetamine implies that there was little change in sensitivity due to the passage of time in mice not receiving d-amphetamine . Ezamiâation of the time course curves (Figure 2) shows that most of the increase results from a change in time course of .raspoaea : when amphetamine is first given, the rata of turning begins low, then slowly incraeses for about 20 minutes after injection. By the second administration, turning begins at a considerably higher rain, and on Wneke 3 - 5, the rata of turning is about constant for the entire observation time . Ordinary tolerance to various effects of amphetamine has long been known, and there are a fee reports of increasing response with repeated administraSuch increasing eeneitiv tion, which is the opposite of ordinary tolerance. ity upon repeated dosing has been noted in the cases of locomotor activity of mice (3,4) and of stereotypy in mice (4), rats (5,6), and guinea pigs (7) . Changes is distribution or metabolism of amphetamine do not appear to account for such increasing sensitivity to amphetamine in mice . Brian and others (B) administered d-amphetamine, 10 mg/kg i .p . t~ri.ce daily to mice for up to twenty days . The time courses of amphetamine concentration in the brain did not differ between acutely and chronically treated mice . Their methods, capable of detecting 5 ng/g of pare-hydroxyamphatamina or of par~-hydroxynorephridine in brain tissue, detected neither in acute or chronic groups . Thane metabolites of amphetamine are thought to account for some long-term affects of amphetamine . Likewise, Hitsemann and others (4) found that previous implantation of amphetamine-bearing pellets in mice made no difference in the concentration of amphetamine following a subsequent amphetamine injection . This group likewise failed to detect either pare-hydroxyamphetamine or pare-hydroxynorephedrine in .the brains of implanted mice . The fact that the two groups of mice responded almost identically to
566
Increased Response to Amphetamine
Vol . 21, No . 4, 1977
saline after the amphetamine phase of the experiments had ended and that thann responses did not exceed the response of the one group to saline before the amphetamine phase, suggests that conditioning contributed little to the increase in response to amphetamine. Mice in which contravereive turning is induced by apomorphine do not show the increasing response to,a constant dose prnsanted above (unpublished results from this laboratory) . This makes it unlikely that the phenomenon re ported above ie simply a non-specific effect of "practice" in circling . It seems possible that the slowness of turning to reach plateau rate after a single dose of amphetamine might be due to interference by some competing activity which began soon after injection, then diminished to allow accelerated turning . If ordinary tolerance to amphetamine developed with respect to the competing activity, that would account for the present results without heightened sensitivity to amphetamine. The only competing activities observed vrere sniffing and grooming, both of which sometimes appeared to be mildly stereotypic . To account for the apparent reverse on sensitivity, these activities would have to reach maxi mum intensity by four minutes after injection, then gradually decrease, and the maximum intensity would have to diminish with successive Injections . Though not systematically observed in the present experiment, sniffing and grooming stereotypies did not appear to diminish with successive injections . In unleaioned guinea pigs given d-amphetamine at similar doses and by the same route as in the present experiments, such stereotypies reached maxim~mm intensity only 30 minutes or more after the first injection, and following subsequent injections, they reached plateau intensity more quickly, and the plateau intensity was higher (7) . The fact that such etereotypiee behaved vary much ae did turning in the present experim~t, rather than ae would the hypothesized interfering stereotypy, done not favor competing stereotypy as an explanation of the present results . There is a substantial body of evidence which suggests that changes in activity of the nigroetriatal dopaminergic system brings about changes in the functional characteristics of that system . The presumed hypersensitivity that results from the destruction of the dopaminergic terminals in rodents lesioned as ware those used in this study is a prime example . Diminution of the eyntheais of dopamine following a moderate dose of amphetamine has been observed (9,10) . After repeated dosing with methamphatamine, it still appears to be detectable for a weak or more (11) . It sees possible that reduced synthesis and, by inference, release of dopamine might induce hypersensitivity of the corresponding postsynaptic receptors . However, it seems unlikely that a single, moderate dose of amphetamine would thereby induce sufficient hypersenaitivity a week afterwards to explain the results present here . Further understanding of the phenomenon awaits further research .
ACRNOWIBDGEMENT I wish to thank Mr . David Prince for preparation of the mice . REFERENCES 1. 2. 3.
P. F . VON VOIGTLANDER and K. E. MOORE, Neurooharmac . 12 :451-462, 1973 . J. E . THORNBIIRG and R. E. MOORE, JPET . 192:42-49, 1975 . P. H. SHORT and L. SCH[JSTER, PsvchovharmacoloAV 48 :59-67, 1976 .
Vol . 21, No . 4, 1977 4. 5. 6. 7. 8. 9. 10 . 11 .
Increased Reapoase to Amphetamine
567
R . J . H rr~cmu~ , H . H . IAH, F . B . GRAVES, and E . F . DOMII~iO, Pavchovharnoa coloxis (Berl) 30 :227-240, 1973 . L . MAGOS, Europ , J . Phaxmacol . 6 :200-201, 1969 . M, M, RILBEY and S. H . ELLII~IWC)OD, JR,, In Cocainn and Other StimuLnte, E . H, E111nw~ood and M . M . Rilbey, nda ., Plenum, NY, 1976 . H . L . IQ+AWANS and D, I . MARGOLIN, Arch . Gen . Psvchiat . 32 :725-732, 1975 . J . F . BRIEDT, J, E . PEACHEY, B . J . ROGERS, and J . C . RTrNEY, J . Pharm . Pharmac . 29 :49-50, 1977 . R . RQCZEIQSRI and D, S . SEGAL, Eu~o~, J . Phsrmacol . 30 :244-251, 1975 . J . E . HARRIS and R . J . BALDFSSARINx,J. Pharm . Pharmacol . 25 :755-757, 1973 . F . J, ROGAN, W . R . NICHOLS and J . W. GIBB, EurOp . J . Pharmacol . 36 :363371, 1976 .
Life Sciences Vol . 21, pp " 563-568 Printed in the II .S .A.
CIRCLING OF MICE BEARING IINIIATERAL 3TRIATAL LESIONS : DEVELOPMEIPP OF INCRFASED RESPONSE TO DAMPBETAMINE Stanley D. 8chols Marten-National Laboratories Division of Richardson-Marten Inc . Cincinnati, Ohio 45215 (Received in final form July 14, 1977) SUNß~lARY Mice leaioned in one sidn of the striatum with 6-hydrorydopaminn respond to d-amphetamine by circling ipsivnreivaly. This report communicates the finding that upon repeated dosing with 4 mg/kg s .c . at weekly intervals the mice respond more and morn strongly . Conditioning appears to contribute little to this phenomenon . Mice unilaterally lesioned in the corpus striatum by way of micro-injected 6-hydroaydopamine respond to amphetamine and similar agents by circling towards the lnsioned sidn (1) . Ea initial ezparimenta with this model in our labora tory, which involved counting of the circling of such mice in response to a dose of d-amphetamine on too occasions, the mice consistently circled more on the second occasion than on the first . The present experiments were performed to dietiaguiah the effects of rnpeatnd exposure to amphetamine from the effects of time, which itself affects the sensitivity of mica leeipned ae these were (2) . MSfHODS Male Charles River CD-1 mice were anesthetized aith methoayflurane and leeioned according to the method of Von Voigtlandnr and Moors (1) . This method consists essentially of injecting 4 ~tl of as aqueous solution of 16 Leg of 6-hydröaydopamina HBr plus 0 .8 ~~g of ascorbic acid etsreotaaically into the caudate-putamen nucleus on one side . Nine days later, the mice ante injected with apomorphine HC1, 0 .2 mg/k8' s .c . The mica which circled contravereivelq ten times or more between too and four minutes after injection acre considered to be successfully leeionnd . Taenty such mice were used in this study. They were assigned to two groups of ten each eo that the two groups sere ss nearly matched as possible regarding their performance in the above teat with apomorphinn . Tao of the chronic amphetamine group died, so their antemortem data have been eu laded. Their lose did not materially lessen the match of the two groups . One group received d-amphetamine sulfate, 4 mg/kg e.c ., five times at weekly intervals . The other group of mice a~s iajectad with salins (10 ml/ kg e.c .) on the first four of the above injection days, and with amphetamine, 4 mg/kg s .c . on the fifth. Both groups then received a saline injection .on Day 5 of the fifth week . The experimental procedure consisted of placing each mouse in a clean glass bowl and attaching a harness made of a pipe cleaner. After at leset 563
564
Increased Response to Amphetamine
Vol . 21, No . 4, 1977
five minutes, the mouse was injected and replaced in the bowl . Circling ipsiversive and contravereive to the lesion were separately counted via automatic counters from minutes 2 through 30 after amphetamine . Following saline, circling was counted from minutes 2 through 30 on Week 1 and on Day 5 of the fifth week ; otheraiee, saline-treated mice were treated as above, exenpt that their harnesses warn not co~ected to the counters . Statistical analysis was by way of "Student's" t-test -- paired when the performance of a group of mice on two occasions was compared, and unpaired when one group of mica was compared with the other. RESULTS Figure 1 shoal the mean ipsiversive and contraveraive turns observed over minutes 2 - 30 after injection. Folloaring the first injection of saline, the mean résponses of the chronic saline group were 43 ipsiversive and 3 contra veraiva . The mean ipsiversive responses of the chronic amphetamine group were 140 on the first amphetamine day, 203 on the second, and 232 on the final amphetamine day. The responses of both the second and the last days exceeded that of the first at the P a .02 level ; those of the second and the last days did not differ at the P ~ .1 level . Upon receiving amphetamine, the chronic online group of mice responded with a mean of 139 ipsiversive turns : virtually equal to the response of the chronic amphetamine group to its first dose of amphetamine.
Ip~lwnlw 200 1J è 100 0
E
FIG . 1 . Total ipsiversive and contravnreiva turns over minutes 2 - 30 follaaing s .c . injection of d-amphetamine, 4 mg/kg (a) or saline (s) . The letters at the base of the ipsiversive bare indicate the compound injected . The white bars represent the chronic amphetamine group; the cross-hatched bare represent the chronic saline group . Statistical aignificances ; * P < .05 ** P < .02
Figure 2 shoos the time course of the turning on each amphetamine day. It ie evident that turning has a faster onset after repeated dosing with amphetamine than when initially given . As shown in Figure 1, after the amphetamine phase of the experiments was concluded, both groups of mica responded almost identically to saline . The ipsiversive turning of both groups was slightly less than the ipsiversive turning of the chronic saline group before exposure to amphetamine .
Vol . 21, No . 4, .1977
Increased Response to Amphetamine
565
FIG . 2 Time course of ipeiveraive turning following amphetamine injection . The solid line represents the chronic amphetamine group ; the. dotted line represents the chronic saline group .
DISCUSSION If total circling war the half-hour after dosing is used as a response measure, the response to a constant down of d-amphetamine increases with reThe virtual equality of the responses of the two groups peated administration . of mica to their respective first doses of amphetamine implies that there was little change in sensitivity due to the passage of time in mice not receiving d-amphetamine . Ezamiâation of the time course curves (Figure 2) shows that most of the increase results from a change in time course of .raspoaea : when amphetamine is first given, the rata of turning begins low, then slowly incraeses for about 20 minutes after injection. By the second administration, turning begins at a considerably higher rain, and on Wneke 3 - 5, the rata of turning is about constant for the entire observation time . Ordinary tolerance to various effects of amphetamine has long been known, and there are a fee reports of increasing response with repeated administraSuch increasing eeneitiv tion, which is the opposite of ordinary tolerance. ity upon repeated dosing has been noted in the cases of locomotor activity of mice (3,4) and of stereotypy in mice (4), rats (5,6), and guinea pigs (7) . Changes is distribution or metabolism of amphetamine do not appear to account for such increasing sensitivity to amphetamine in mice . Brian and others (B) administered d-amphetamine, 10 mg/kg i .p . t~ri.ce daily to mice for up to twenty days . The time courses of amphetamine concentration in the brain did not differ between acutely and chronically treated mice . Their methods, capable of detecting 5 ng/g of pare-hydroxyamphatamina or of par~-hydroxynorephridine in brain tissue, detected neither in acute or chronic groups . Thane metabolites of amphetamine are thought to account for some long-term affects of amphetamine . Likewise, Hitsemann and others (4) found that previous implantation of amphetamine-bearing pellets in mice made no difference in the concentration of amphetamine following a subsequent amphetamine injection . This group likewise failed to detect either pare-hydroxyamphetamine or pare-hydroxynorephedrine in .the brains of implanted mice . The fact that the two groups of mice responded almost identically to
566
Increased Response to Amphetamine
Vol . 21, No . 4, 1977
saline after the amphetamine phase of the experiments had ended and that thann responses did not exceed the response of the one group to saline before the amphetamine phase, suggests that conditioning contributed little to the increase in response to amphetamine. Mice in which contravereive turning is induced by apomorphine do not show the increasing response to,a constant dose prnsanted above (unpublished results from this laboratory) . This makes it unlikely that the phenomenon re ported above ie simply a non-specific effect of "practice" in circling . It seems possible that the slowness of turning to reach plateau rate after a single dose of amphetamine might be due to interference by some competing activity which began soon after injection, then diminished to allow accelerated turning . If ordinary tolerance to amphetamine developed with respect to the competing activity, that would account for the present results without heightened sensitivity to amphetamine. The only competing activities observed vrere sniffing and grooming, both of which sometimes appeared to be mildly stereotypic . To account for the apparent reverse on sensitivity, these activities would have to reach maxi mum intensity by four minutes after injection, then gradually decrease, and the maximum intensity would have to diminish with successive Injections . Though not systematically observed in the present experiment, sniffing and grooming stereotypies did not appear to diminish with successive injections . In unleaioned guinea pigs given d-amphetamine at similar doses and by the same route as in the present experiments, such stereotypies reached maxim~mm intensity only 30 minutes or more after the first injection, and following subsequent injections, they reached plateau intensity more quickly, and the plateau intensity was higher (7) . The fact that such etereotypiee behaved vary much ae did turning in the present experim~t, rather than ae would the hypothesized interfering stereotypy, done not favor competing stereotypy as an explanation of the present results . There is a substantial body of evidence which suggests that changes in activity of the nigroetriatal dopaminergic system brings about changes in the functional characteristics of that system . The presumed hypersensitivity that results from the destruction of the dopaminergic terminals in rodents lesioned as ware those used in this study is a prime example . Diminution of the eyntheais of dopamine following a moderate dose of amphetamine has been observed (9,10) . After repeated dosing with methamphatamine, it still appears to be detectable for a weak or more (11) . It sees possible that reduced synthesis and, by inference, release of dopamine might induce hypersensitivity of the corresponding postsynaptic receptors . However, it seems unlikely that a single, moderate dose of amphetamine would thereby induce sufficient hypersenaitivity a week afterwards to explain the results present here . Further understanding of the phenomenon awaits further research .
ACRNOWIBDGEMENT I wish to thank Mr . David Prince for preparation of the mice . REFERENCES 1. 2. 3.
P. F . VON VOIGTLANDER and K. E. MOORE, Neurooharmac . 12 :451-462, 1973 . J. E . THORNBIIRG and R. E. MOORE, JPET . 192:42-49, 1975 . P. H. SHORT and L. SCH[JSTER, PsvchovharmacoloAV 48 :59-67, 1976 .
Vol . 21, No . 4, 1977 4. 5. 6. 7. 8. 9. 10 . 11 .
Increased Reapoase to Amphetamine
567
R . J . H rr~cmu~ , H . H . IAH, F . B . GRAVES, and E . F . DOMII~iO, Pavchovharnoa coloxis (Berl) 30 :227-240, 1973 . L . MAGOS, Europ , J . Phaxmacol . 6 :200-201, 1969 . M, M, RILBEY and S. H . ELLII~IWC)OD, JR,, In Cocainn and Other StimuLnte, E . H, E111nw~ood and M . M . Rilbey, nda ., Plenum, NY, 1976 . H . L . IQ+AWANS and D, I . MARGOLIN, Arch . Gen . Psvchiat . 32 :725-732, 1975 . J . F . BRIEDT, J, E . PEACHEY, B . J . ROGERS, and J . C . RTrNEY, J . Pharm . Pharmac . 29 :49-50, 1977 . R . RQCZEIQSRI and D, S . SEGAL, Eu~o~, J . Phsrmacol . 30 :244-251, 1975 . J . E . HARRIS and R . J . BALDFSSARINx,J. Pharm . Pharmacol . 25 :755-757, 1973 . F . J, ROGAN, W . R . NICHOLS and J . W. GIBB, EurOp . J . Pharmacol . 36 :363371, 1976 .